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Special  Beport  Series.  No.  121. 


^ribg  (ffouncil 

MEDICAL   RESEARCH 
COUNCIL 


3  9424  00444  9358 

Borna  DiSBSSrtHH^ffizootic  Encephalo- 
Myelitis  of  Sheep  and  Cattle 

BY 

S.  NICOLAU,  M.D.,  D.Sc.,  and 
I.  A.  GALLOWAY,  B.Sc,  M.R.C.V.S. 


"B.C.  LIBRARy 


SF799 


LONDON 
IS  MAJESTY'S  STATIONERY  OFFICE 
1928 

Price  5s.  net 


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U.B.C.  Library  \ 

Report  Series,  No.  121.  j 

CAT  HO.  P^'nX:.,!±i J 


^  ACC.  NO.  '}i  9^^^—  J 

MEDICAL   RES ^^^^^^f^f^—--  — 
COUNCIL 


Borna  Disease  and  Enzootic  Encephalo- 
Myelitis  of  Sheep  and  Cattle 


BY 


S.  NICOLAU,  M.D.,  D.Sc,  and 
I.  A.  GALLOWAY,  B.Sc,  M.R.C.V.S. 


LONDON 

PUBLISHED  BY  HIS  MAJESTY'S  STATIONERY  OFFICE 

1928 

45-5-2I. 


MEDICAL  RESEARCH  COUNCIL 

The  Right  Hon.  Earl  of  Balfour.  K.G..  O.M..  F.R.S.  {Chairmnn). 
The  Right  Hon.  Lord  Mildmay  of  Flete.  P.C.  (Treasurer). 
The  Right  Hon.  William  Graham,  LL.B.,  M.P. 
Sir  Hugh  K.  Anderson,  M.D.,  F.R.S. 
Professor  E.  P.  Cathcart,  C.B.E.,  M.D.,  D.Sc,  F.R.S. 
Professor  G.  Dreyer,  C'.B.E..  M.D.,  F.R.S. 
Professor  T.  R.  Elliott,  C.B.E.,  D.S.O.,  M.D.,  F.R.S. 
Sir  Archibald  E.  Garrod,  K.C.M.G.,  D.M.,  F.R.S. 
Sir  Frederick  G.  Hopkins,  D.Sc,  F.R.S. 
Sir  Charles  J.  Martin,  C.M.G.,  D.Sc,  F.R.S. 
Sir  Charles  S.  Sherrington,  O.M.,  G.B.E.,  F.R.S. 
Sir  Walter  M.  Fletcher,  K.B.E.,  M.D.,  ScD.,  F.R.S.  {Secretary 


PREFACE 

The  present  report,  which  was  received  for  publication  in  October, 
1927,  gives  the  results  of  investigations  made  by  Dr.  Xicolau  and  Mr. 
Galloway,  working  as  guests  in  the  National  Institute  of  Medical 
Research.  These  studies  of  the  virus  which  is  responsible  for  a  danger- 
ous communicable  disease  in  horses,  cattle  and  sheep  have  happily  no 
immediate  practical  apphcation  in  this  country.  Borna  disease  and 
its  congeners  are  at  present  only  known  in  continental  Europe  and  in 
America.  Our  present  immimity,  however,  may  only  be  temporary, 
and  in  any  case  it  is  highly  important  that  we  should  have  the  fullest 
knowledge  of  these  epizootic  diseases  of  other  countries. 

The  scientific  advantages  gained  by  close  association  between 
studies  of  disease  in  animals  and  studies  of  human  disease  have  long 
been  obvious.  The  potential  value  to  medical  science  of  the  work  now 
presented  Ues  in  two  directions.  The  accurate  experimental  study 
of  the  '  virus  '  which  is  the  causal  agent  in  this  disease  is  part  of  the 
general  study  of  disease  viruses,  and  in  this  field  of  work  great  gain 
must  come  from  the  free  exchange  of  ideas,  methods  and  results 
among  workers  in  different  parts  of  it.  Besides  this,  however,  Borna 
disease  has  special  points  of  interest  to  students  of  human  neurology. 
The  infective  virus  produces  changes  in  the  brain  and  spinal  cord,  the 
so-called  encephalo-myehtis,  which  throw  light  upon  analogous  forms 
of  encephahtis  and  myehtis  which  occur  in  sporadic  or  epidemic  form 
in  human  beings. 

It  will  be  seen  that  the  authors  have  extended  or  confirmed  the 
observations  of  many  previous  workers,  and  have  gained  new  know- 
ledge at  several  points  of  detail  by  their  experimental  work.  Fresh 
studies  have  been  made  of  the  immunity  reactions  of  the  virus,  and 
it  has  been  shown  that  animals  may  be  successfully  immunized 
against  it. 

Dr.  Nicolau  was  enabled  to  conduct  this  work  in  the  National 
Institute  by  a  grant  from  the  Roumanian  Government.  Mr.  Galloway 
co-operated  with  him  in  the  course  of  other  work  upon  foot  and  mouth 
disease  in  cattle,  which  he  is  doing  on  behalf  of  the  Ministry  of  Agri- 
culture and  Fisheries.  The  council  are  indebted  to  the  Ministry  for 
the  faciUties  they  have  given  for  this  co-operation. 

28th  July,  1928. 
Medical  Research  Council, 
15  York  Buildings, 
London,  W.C.  2. 


1.)  Ps.  28720.  Wt.  15076.  536/970.  1250.  9/28.  O.U.P. 


BORNA  DISEASE  AND  ENZOOTIC  ENCEPHALO- 
MYELITIS OF  SHEEP  AND  CATTLE 

By  S.  Nicolau.  M.D.,  D.Sc,  and  I.  A.  Galloway,  B.Sc, 
M.R.C.V.S. 

(Xational  Ivstilute  for  Medical  Research.  Hampstead.) 

CONTENTS 

PAOE 

1.  HiSTORiCAi,  AXD  General       .........       7 

I.  Enzootic  Encephalo-myelitis  of  the  Horse  ......        7 

II.  Enzootic  Encephalo-myelitis  of  Cattle         ......        9 

III.  Enzootic  Encephalo-myelitis  of  Sheep         ......      10 

IV.  Malignant  Catarrhal  Fever  of  Cattle  .  .  .  .  .  .11 

V.  Summary         .  .  .  .  .  .  .  .  .  .  .11 

2.  Pbopekties  of  the  Virus       .........     12 

I.  In\-isibility      ...........  12 

II.  Filterability:  Effect  of  Dilution 12 

III.  Centrifugalization    ..........  15 

IV.  Resistance  to  Glycerine    .........  17 

V.  Sensitiveness  to  Heat  and  Desiccation         ......  20 

\^.  Action  of  Utra-violet  Light 20 

VII.  Action  of  Hexamethylenetetramine  (Urotropine)  .  .  .  .21 

VIII.  Action  of  Chloroform  and  Ether         .......  21 

IX.  Action  of  Formalin            .........  22 

X.  Culture 22 

XI.  Summary         ...........  22 

3.  Tkassmission  of  Eqiixe  Strain  to  Babbit  and  from  Rabbit  to  Sheep 

A>'D  VICE  versa  ..........     23 

4.  ExpERniEXTAL  Disease  rx  the  Rabbit    .......     2t 

I.  Experimental  Transmission  of  the  Disease  to  the  Rabbit        .  .  .24 

II.  Authors'  Observations      .........     24 

III.  Symptomatology  of  the  Disease  in  the  Rabbit    .  .  .  .  .27 

IV.  Routes  by  which  Rabbits  can  be  Infected  .  .  .  .  .  .29 

5.  AriHORS'  EXPEKDIESTS  ON  THE  TbASSMISSIOS  OF  THE  DISEASE  TO  MONKEYS, 

AND  SvilPTOMS  OCCUKKING  IN  THESE  ANIMALS  .  .  .  .  .34 

6.  Pathogenicity  of  the  Virus  of  Enzootic  Encephalo-myelitis  fob  the 

Guinea-pig,  Rat,  Mouse,  and  Fowl    .......  39 

I.  Guinea-pig       ...........  39 

II.  Rat 43 

III.  Mouse 44 

IV.  Fowl 44 

7.  Animals  which  have  bees  found  to  be  Resistant  to  Infection  with  the 

Virus  of  Enzootic  Encephalo-myelitis     ......  44 

I.  Dog 44 

II.  Pigeon    ............  44 

III.  Ferret 44 

8.  Distribution  OF  the  Virus  of  Borsa  Disease  IN  THE  Animal  Body  .         .  44 

I.  Passage  of  Virus  through  the  Placenta        .  .  .  .  .  .44 

II.  Distribution  of  the  Virus  in  Various  Organs  and  Tissues        .  .  .45 

III.  Presence  of  Virus  in  the  Peripheral  Nerves  of  Rabbits  inoculated  Intra- 

cerebrally  ..........     45 

IV.  Distribution  of  the  Virus  in  various  Organs  and  Tissues  of  the  Monkey  .     48 


6  CONTENTS 

PAGE 
9.   EUMUJATION  OF  THE  VlRUS  FROM  THE  AsTMAL  OrGASISM    .  .  .  .49 

10.    HiSTOPATHOLOGY  OF  BoRSA  DISEASE  .......       50 

I.  Horse 50 

II.  Cattle 50 

III.  Sheep 51 

IV.  Experimental  Borna  Disease  in  Horses  and  Sheep        .  .  .  .51 
V.  Rabbits  infected  Experimentally        .......     51 

\"I.  -Authors'  Observations      .  ........     52 

A.  Babbit 52 

(1)  ilacroscopical  and  Microscopical  Findings  in  Diverse  Organs     52 

(2)  Lesions  in  the  Central  Servoiis  System  .  .  .  .53 

(a)  The  Brain 53 

(b)  The  Midbrain  and  Medulla  Oblongata  .  .  .58 

(c)  The  Cerebellum 59 

(d)  The  Spinal  Cord 59 

(3)  Lejiions  in  the  Peripheral  Xerrous  System  .  .  .60 

(a)  The  Posterior  Xerie  Boots tiO 

(b)  The  Spinal  Ganglia tiO 

(c)  The  Peripheral  Xerves       .  .  .  .  .  .1)1 

(4)  Summary  and  Discussion  .  .  .  .  .  .62 

B.  The  Guinea-pig 64 

C.  The  Bat  and  Mouse  ........      64 

D.  The  Monkey (H 

( 1 )  Lesions  in  the  Central  Xenons  System  .  .  .  .65 

(a)  Brain      .........  65 

(b)  Cerebellum       ........  66 

(c)  Medulla  Oblongata    .......  66 

(d)  .Spina/  Cord 66 

(e)  Sciatic  Xerre   ........  67 

(f)  Brachial  Xerie 68 

(2)  Summary 68 

11.  hl.MlNITY 69 

1.  Attempts  at  eonferrinn  Immunity  t<i  Rabbits  by  Imipulations  of  Xon- 

attenuated  Virus        .........      69 

A.  Intravenous  Inoculation     ........      69 

B.  Corneal  Inoculation  .  .  .  .  .  .  .71 

C.  Intrateslicular  JniKulalion  .  .  .  .  .  .  .71 

II.  Attempts  at  conferring  Immunity  to  Rabljits  by  Imxiilatiunof  Attouuatod 

Vims       ...........     72 

A.  Virus  inactivated  by  Klher  .......      72 

B.  I'lru*  killed  by  Chloroform  .  .72 
C    Virus  treated  with  Formal           .......      73 

D.  Virv^  inactivated  by  I'ltraviolet  Light  .  .  .  .  .73 

III.  Search  for  Virucidal  .Antibodies  in  the  Serum  of  Immunised  Animals  73 

IV.  Experiments  on  Cross  Immunity  lx>tween  the  Strain  of  Kquine  and  tliat 

of  Ovine  Origin  .........      75 

V.  Experiments   on   Cross    Immunity    between    the    Virus    of    Kncephnlo- 

myelitis  and  other  Viruses  <if  the  KillerpassiuK  (;rcm|i  .  .77 

.•\.  Herpes  ...........      77 

B.  Babies 77 

C.  Poliomyelitis   ..........      7S 

VI.  Summary       ...........      82 

12.  ClIEMOTIIEpArV       ...........      83 

Refekences  ...........     85 

DeSCRIITION    of    I'HOTOGKAIMIS    ANB    CoLilL' KKII    I'LATHS     .  ...        87 


].   HISTOEICAL  AND  GENEEAL 

I.  Enzootic  Excephalo-myelitis  of  the  Horse  (Borna 
Disease). 

The  disease  has  various  designations:  Enzootic  Encephalo- 
myelitis. Meningo-encephalo-myehtis  of  the  Horse,  Nevraxite  en- 
zootique;  Mai  d'Azeau  (after  the  region  where  a  severe  epizootic 
occurred  in  Belgium  in  1909);  Bornasche  Krankheit,  Genickstarre, 
Gehirm-uekenmarksentziindung.  Gehirnruckenmarkseuche,  Nerven- 
fieber ;  Encephahtis  Lymphocytaria  Equi. 

It  is  an  infectious  malady  with  a  mortahty  rate  reaching  90  per 
cent,  in  some  epizootics.  The  characteristic  symptoms  are  due  to 
lesions  in  the  nervous  system  both  central  and  peripheral,  which  are 
produced  by  an  ultravisible  and  filterable  virus. 

The  name  Borna  disease,  by  which  the  disease  is  more  commonly 
known,  originated  from  the  locality  in  Saxony  where  a  severe  epizootic 
occurred  in  the  years  1894  to  1896. 

Borna  disease  has  been  known  for  more  than  a  century.  It  was 
first  described  by  Worz  in  Wlirtemburg  in  1813.  Subsequently  it  was 
reported  from  Germany,  North  America,  the  Argentine,  and  Hungary. 
Since  1900  it  has  appeared  in  Eussia.  in  the  region  of  the  river  Don, 
in  Belgium.  France,  Italy,  Eoumania.  Germany,  and  South  America. 
The  epizootics  have  been  of  varying  severity.  In  1896  the  disease 
.  occurred  in  several  districts  of  Germany,  where  1,198  horses  were 
reported  to  have  contracted  it ;  in  the  epizootic  which  occurred  in  the 
valley  of  the  Colorado  and  the  Brazos  in  America  about  4,000  horses 
and  mules  succumbed.  In  other  areas  the  disease  has  appeared 
sporadically. 

Climatic  conditions  appear  to  have  an  influence  on  the  spread  of 
the  disease,  the  incidence  of  which  is  greater  when  abnormally  warm 
and  moist  conditions  obtain.  The  diagram  pubhshed  by  Zwick, 
Seifried,  and  Witte  (1926)  shows  that  epizootics  generally  begin  in 
the  month  of  May  and  decUne  later,  the  cases  again  becoming  sporadic 
during  the  winter  months. 

The  period  of  incubation  in  the  spontaneously  contracted  disease 
is  difficult  to  estimate  and  is  therefore  not  definitely  known.  Noack 
(1908)  considers  9  days  to  be  an  average. 

Joest  (1926)  suggests  that  the  portal  of  entry  of  the  virus  is  through 
the  nose,  but  the  possibihty  of  ingestion  being  the  mode  of  infection 
cannot  be  excluded. 

The  onset  of  the  disease  may  be  sudden,  but  some  authors  have 
reported  that  fatigue,  gastro-intestinal  disturbances,  and  symptoms 
of  affection  of  the  upper  respiratory  passages  precede  the  onset  of 
characteristic  nervous  symptoms  by  as  much  as  14  days. 

The  first  symptom  which  usually  draws  attention  to  the  infected 
animal  is  lassitude:  the  horse  is  easily  fatigued  and  appears  depressed 
and  indifferent  to  external  impressions.  A  period  of  excitation  follows, 
which  may  last  with  intermittence  till  the  end  of  the  disease.   Tonic 


8  BOENA  DISEASE  AND  • 

contractions  of  diverse  groups  of  muscles  occur  and  there  is  difficulty 
in  mastication  and  deglutition.  During  the  stage  of  excitation  external 
stimuh  produce  exacerbation  of  the  spasms  in  certain  groups  of 
muscles;  champing  of  the  jaws  is  a  common  symptom,  and  sahva 
flows  from  the  commissures  of  the  mouth.  The  pupils  are  imequal  in 
size.  Soon  paresis  or  paralysis  sets  in.  aft'ecting  the  hmdquarters,  the 
muscles  of  the  tail,  muscles  of  mastication,  muscles  of  the  tongue 
and  of  the  back  to  a  varying  degree.  Paraplegia  or  hemiplegia  are 
not  uncommon. 

In  some  cases  the  symptoms  of  encephahtis  are  dominant,  in  some 
those  of  acute  myeUtis  are  more  evident,  while  in  others  symptoms 
characteristic  of  affection  of  both  brain  and  cord  coexist.  The  tem- 
perature generally  remains  normal  throughout,  and  according  to 
Hutyra  and  Marek  (1922)  persistent  fever  indicates  secondary  com- 
phcatious  of  a  septic  nature. 

The  frequency  of  the  respiration  remains  httle  changed,  except 
during  periods  of  excitation,  or  when  the  nucleus  of  origin  of  the 
vagus  nerve  is  involved.  Towards  the  end  of  the  disease,  however, 
the  respirations  become  superficial  and  approach  the  Cheyne-Stokes 
type. 

The  examination  of  the  blood  and  urine  does  not  indicate  any  con- 
stant change.  The  cerebrospinal  fluid  shows  lymphocj-tosis  occa- 
sionally. The  duration  of  the  disease  varies  from  a  few  days  to 
6  weeks.i 

Aetiology.  As  has  been  the  case  with  a  number  of  diseases  subse- 
quently proved  to  be  due  to  a  filterable  virus,  the  disease  was  at  first 
attributed  to  the  pathogenic  action  of  various  cocci.  Siedanigrotzky 
and  Schlegel  (1896)  isolated  a  Gram-positive  diplococcus  and  Johne 
(1896)  a  Gram-negative  one.  Other  observers.  Ostertag  (1900), 
Christiani  (1909),  Marcq  (1909),  Lohr  (1910),  Lessage  and  Frisson 
(1912),  have  found  streptococci  or  diplococci,  each  organism  isolated 
being  considered  by  the  investigator  concerned  as  the  aetiological 
agent.  More  recently  Kraus,  Kantor,  Fischer,  and  Quiroga  (1920) 
isolated  a  diplococcus  similar  to  that  found  by  Siedanigrotzky  and 
Schlegel,  which  they  beHeved  to  be  the  aetiological  agent  of  Boma 
disease  until  the  appearance  of  the  work  of  Moussu  and  Marchand 
in  France,  and  Zwick,  Seifried,  and  Witte  in  Germany,  ijidicating  that 
encephalo-myehtis  of  the  horse  is  produced  by  an  ultravisible  and 
hlterable  virus.  Although  the  vims  isolated  by  Moussu  and  Marchand 
appears  to  differ  from  that  isolated  by  Zwick  and  iiis  collaborators, 
it  is  possible  that  this  may  be  explained  by  the  fact  that  two  strains 
of  the  same  virus  were  being  dealt  with  (see  note  on  p.  28).  Beck  and 
Frolibiise  (1926)  and  Ernst  and  Hahn  (1926)  also  .showed  tiiat  a  virus 
isolated  from  the  brain  of  horses  affected  with  eneephalo-niyelitis  was 
capable  of  infecting  rabbits. 

Joest  in  collaboration  with  Degen  (1909)  determined  the  constant 

'  In  thp  ppiziMttics  dea<:nljed  by  K.  Mnuiuu  (1920)  thr  diiratiun  of  llic  diM'OHc  was 
very  Hlicirt.  According  to  tliiit  author,  in  those  cjuok  Hhuwin^  Hyniptonm  uf  »n  pn- 
cephulitic  type  °  the  evolution  in  rapid.  In  the  coaeg  we  have  i«tudip<l  deuth  intervrnrd 
in  20.  32,  and  37  hours.' 


ENZOOTIC  ENCEPHALO-MYELITIS  9 

presence  of  certain  intranuclear  '  inclusions '  in  the  ganglion  cells  of 
the  Amnion's  horn  which  they  considered  as  reactions  of  the  cell  to 
the  parasite  and  'similar  to  Chlamydozoa'.  In  the  opinion  of  Joest 
these  chlamydozoa,  or  at  least  certain  forms  of  their  cycle  of  develop- 
ment, are  capable  of  passing  through  bacteriological  filters. 

Siedamgrotzky  and  Schlegel  (1896),  Lohr  (1910),  and  Ostertag 
(1900  et  seq.)  made  frequent  attempts  to  infect  laboratory  animals 
by  various  methods  of  inoculation  or  by  feeding  them  with  cerebro- 
spinal fluid,  blood,  emulsions  of  spleen,  liver,  kidney,  or  bone  marrow 
from  horses  affected  Avith  the  disease.   The  results  were  negative. 

In  resume  the  work  of  Moussu  and  Marchand  (1924  et  seq.),  Zwick, 
Seifried.  and  Witte  (1926).  Beck  and  Frohbose  (1926),  Ernst  and 
Hahu  (1926-7).  has  shown  definitely  that  the  aetiological  factor  is 
a  filterable  ultravisible  virus,  and  the  results  obtained  by  Kraus  and 
his  collaborators  (1920)  and  other  investigators  with  their  cultures  of 
diplococci  or  diplo-streptococci  must  now  be  considered  as  due  to 
what  Nicolle  called  microbes  de  sortie. 

II.   Enzootic  Encephalo-myelitis  of  Cattle. 

The  epizootiology  and  symptoms  of  encephalo-myelitis  of  cattle  are 
similar  to  Borna  disease  of  the  horse.  Hutyra  and  Marek  (1922)  refer 
to  outbreaks  of  disease  amongst  cattle  described  bv  Meyer  (1867), 
Schmidt  (1888),  Utz  (1896),  Eoder  (1896),  and  Manfredi  d'Ercole 
(1896)  which  may  be  attributed  to  enzootic  encephalo-myehtis. 
Proger  (1896)  recorded  the  coexistence  of  this  disease  with  Borna 
disease,  and  as  Ernst  and  Hahn  (1927)  have  suggested  that  the 
outbreaks  of  disease  in  Hungary  referred  to  as  '  cerebrospinal  menin- 
gitis' may  also  have  been  encephalo-myehtis.  G.  Moussu,  quoted  by 
E.  Moussu  (1926),  in  1906  studied  an  enzootic  on  a  farm  in  the  region 
of  Ome  which  killed  10  cattle  in  a  few  weeks.  The  disease  had  a 
similar  onset ;  periods  of  excitation  were  observed  followed  by  depres- 
sion with  champing  of  the  jaws,  salivation,  loss  of  vision,  and 
muscular  twitcliings.  The  possibihty  of  intoxication  was  excluded, 
and  the  symptoms  were  suggestive  of  Borna  disease.  Moussu  also  con- 
siders that  the  cases  of  'cerebrospinal  meningitis'  of  cattle  reported 
by  Kragerud  and  Gunderson  (1921)  were  the  same  as  encephalo- 
myelitis of  the  horse,  and  the  symptoms  described  by  Causel  (1924) 
in  an  enzootic  of  what  he  termed  'infectious  bulbar  paralysis' 
amongst  cattle  are  consistent  with  the  view  that  the  animals  suffered 
from  encephalo-myelitis. 

The  symptoms  of  encephalo-myehtis  in  cattle  are  similar  to  those 
of  Borna  disease  in  the  horse. 

Aetiology.  Moussu  (1926),  who  described  31  cases  of  encephalo- 
myehtis  of  cattle  on  18  different  farms  in  France,  attempted  to 
transmit  the  disease  to  laboratory  animals  without  success. 

Ernst  and  Hahn  (1927)  found  lesions  in  the  brain  of  cattle  dead  of 
encephalo-myehtis  analogous  to  those  in  the  brain  of  horses  dead  of 
Borna  disease;  in  the  lesions  the  corpuscles  of  Joest-Degen  were  seen. 
With  the  virus  which  they  recovered  they  succeeded  in  producing 


10  BOENA  DISEASE  AND 

symptoms  in  rabbits  similar  to  those  produced  by  Zwick  with  a  virus 

from  horses. 

The  general  aspect  of  the  lesions  in  the  brain  of  rabbits  infected 
with  the  two  viruses  and  the  similar  presence  of  the  corpuscles  of 
Joest-Degen  suggests  that  varieties  of  the  same  virus  are  the  cause 
of  encephalo-myelitis  of  horses  and  cattle. 

III.   Enzootic  Encephalo-myelitis  of  Sheep. 

Eichbaum,  Stohr.  and  Wilke  (1865,  1866)  recorded  an  enzootic  of 
encephalo-myelitis  among  sheep,  and  Roloff  (1868).  who  examined 
the  brain  of  animals  which  succumbed  to  a  similar  epizootic,  found 
perivascular  infiltration  in  the  pia  mater.  Schmidt  (1870),  about  the 
same  time,  described  an  enzootic  of  a  similar  nature  in  Prussia.  Later, 
Popow  (1882)  and  Wischnikowitsch  (1889)  described  the  disease  in 
Kussia.  Prietsch  (1896)  referred  to  an  enzootic  among  sheep,  and 
suggested  the  possibiHty  that  the  source  of  infection  was  the  water 
in  troughs  contaminated  by  the  virus  of  Boriia  disease  of  horses. 
Walther  (1899)  reported  an  enzootic  in  two  Hocks  of  sheep  in  the  district 
of  Borna  at  a  time  when  equine  encephalo-myelitis  was  prevalent ;  the 
two  diseases,  equine  and  ovine,  bore  many  resemblances  to  one  another. 
Savigne  and  Leblanc  (1897)  also  have  described  an  enzootic  in  France. 

The  descriptions  of  these  authors  ditfer  essentially  and  may  not  all 
refer  to  the  same  disease.  More  precise  accounts  of  ovine  encephalo- 
myeUtis  have  been  pubhshed  within  recent  years  by  Spiegl  (1922), 
Priemer  (1925),  Beck  and  Frohbiise  (1926),  Moussu  (1926),  Miessner 
(1926),  and  others,  from  which  it  appears  that  in  the  spontaneous 
disease  of  the  sheep  the  same  succession  of  symptoms  occur  as  in 
horses  and  cattle.  The  evolution  of  the  disease  may  take  as  long  as 
from  2  to  12  days.  Death  generally  supervenes.  The  temperature  is 
variable.  In  certain  cases  the  temperature  may  rise  to  41°  C.,  while 
in  others  no  pyrexia  is  observed.  According  to  Moussu  (1926)  the 
incubation  period  averages  27  days. 

Aetiologn.  Beck  (1925)  studied  the  microscopic  lesions  in  tlic  l)rain 
of  sheep  dead  of  the  disease,  and  emphasized  their  resemblance  to 
those  found  in  the  i)rain  of  horses  dead  of  Horna  disease.  In  the 
ganglion  cells  of  the  .\nimon'8  horn  of  sheep  the  characteristic  oxy- 
))hilic  intranuclear  corpuscles  of  Joest  and  Degen  were  found.  In 
collaboration  with  Frolil)ose,  Beck  (1926)  transmitted  the  disease  to 
rabbits,  and  from  the  similarity  in  the  symptoms  and  in  the  lesions 
produced  by  the  virus  isolated  l)y  them  fri)m  sheep  with  that  isolated 
by  Zwick  from  horses,  considered  that  the  two  diseases  were  identical. 

.\l)ont  the  same  time  Moussu  and  Marphand  also  passed  tlie  disease 
to  rabbits  and  transmitted  it  from  sliee))  to  sheep.  Later  Miessner 
(1926)  and  Ernst  and  Halm  (1927)  contirmed  the  transmissibility  of 
the  disease  of  sheep  to  raijbits. 

We  shall  describe  later  how  the  experimental  study  of  enzootic 
encephalo-myelitis  of  sheep  has  shown  that  th(>  disease  is  produced 
by  a,  virus  similar,  if  not  identical,  with  that  which  produces  liornu 
dJBeuBe  in  horses. 


ENZOOTIC  ENCEPHALO-MYELITIS  11 

IV.   Malignant  Catarrhai,  Fever  of  Cattle. 

Glamser  (1926)  and  Dobberstein  (192.5)  found  perivascular  and 
parenchymatous  infiltration  as  well  as  alterations  in  the  ganglion 
cells  of  the  brain  of  cattle  dead  of  a  disease  which  they  called  malig- 
nant catarrhal  fever.  These  lesions  were  similar  to  those  described 
in  Borna  disease.  Ernst  and  Hahn  (1927)  also  draw  attention  to  the 
similarity  of  the  lesions  in  the  brain  of  cattle  dead  of  this  disease  with 
those  found  in  encephalo-myehtis  of  the  horse,  and  in  3  out  of  5  cases 
they  observed  the  intranuclear  corpuscles  of  Joest-Degen  in  the 
ganglion  cells  of  the  Amnion's  horn.  Emulsions  from  the  brain  of  one 
of  these  cases  inoculated  intracerebrally  into  rabbits  produced  a 
disease  similar  to  experimental  Borna  disease  and  transmissible  from 
rabbit  to  rabbit.  They  concluded  that  malignant  catarrhal  fever  of 
cattle  is  produced  by  a  virus  which  approaches  very  closely,  if  it  be 
not  identical  with,  that  producing  encephalo-myelitis  in  horses. 

We  might  here  mention  that  Ernst  and  Hahn  (1927)  also  made  an 
observation  indicating  that  deer  are  susceptible  to  Borna  disease. 
A  sick  deer  was  killed  by  a  hunter  under  curious  circumstances.  The 
animal  allowed  the  hunter  to  approach  very  closely,  drew  away  in 
fear,  and  then  rushed  on  him  suddenly.  The  ears  were  seen  to  twitch 
and  the  animal  turned  round  in  a  circle  until  the  hunter,  probably 
more  frightened  than  the  animal,  killed  it.  The  head  was  brought  to 
Munich.  The  brain  showed  the  presence  of  lesions  similar  to  those  of 
Borna  disease  of  the  horse,  and  the  intranuclear  corpuscles  of  Joest- 
Degen  were  observed.  Their  attempts  at  transmission  of  the  disease 
to  laboratory  animals  had  given  no  results  at  the  time  of  publication 
of  these  observations. 

V.  Summary. 
From    the    foregoing   resume    of    the    literature    of    spontaneous 
encephalo-myelitis  in  different  species,  the  following  conclusions  may 
be  drawn : 

(1)  The  enzootic  encephalo-myelitis  of  horses  and  cattle  and  of 
sheep  appears  to  be  the  same  disease.  The  symptomatology  and  the 
lesions  found  in  the  central  nervous  system  are  analogous,  and  the 
intranuclear  corpuscles  of  Joest-Degen  occur  in  the  large  ganglion 
cells  of  the  Amnion's  horn  in  all  three  species  suffering  from  the 
disease  in  question. 

(2)  From  cases  of  all  three  diseases  a  virus  has  been  recovered 
and  shown  to  be  responsible  for  the  disease. 

(3)  From  the  observations  of  Ernst  and  Hahn  it  would  seem  not 
improbable  that,  if  the  animals  had  not  in  addition  to  malignant 
catarrhal  fever  a  concomitant  infection  with  Borna  disease,  some 
of  the  cases  described  as  malignant  catarrhal  fever  of  cattle  were  en- 
cephalo-myehtis. 

(4)  Deer  appear  to  suffer  from  a  similar  disease  spontaneously. 

(5)  The  transmission  of  the  disease  under  natural  conditions  is 
probably  by  the  respiratory  tract  or  by  ingestion. 


12  BORNA  DISEASE  AND 

2.   PROPERTIES  OF  THE  VIRUS 
I.  Invisibility. 

The  virus  in  ultravisible.  Various  methods  of  staining  have  been 
used  to  discover  a  parasite,  but  without  success.  Methods  of  im- 
pregnation with  silver  have  not  revealed  the  presence  of  parasites 
in  the  brain  of  animals  dead  of  the  experimental  disease  in  our  hands. 
The  existence  of  the  virus  in  the  brain  is,  however,  associated  with 
the  presence  of  intranuclear  corpuscles,  first  described  by  Joest  and 
Degen  (1909).  The  interpretation  of  the  nature  of  these  corpuscles 
which  we  favour  is  discussed  in  the  chapter  dealing  with  the  histo- 
pathology  of  the  disease,  and  we  are  persuaded  that  they  are  of  the 
same  nature  as  the  similar '  inclusions '  found  in  other  diseases  produced 
by  filterable  viruses,  such  as  fowl  plague,  fowl  pox,  distemper,  and 
'Virus  III'  disease  of  rabbits. 

II.   Filterability:  Effect  of  Dilution. 

Filtration  of  emulsions  containing  viruses  through  filters  which 
hold  back  bacteria  generally  greatly  diminishes  the  concentration 
of  the  virus.  There  are  many  reasons  for  tliis,  apart  from  the  size 
of  the  virus.  If  the  virus  is  contained  in  or  on  cellular  particles  these 
may  be  retained,  or  the  virus  itself  may  be  adsorbed  on  the  filter. 
In  such  adsorption  the  hydrogen-ion  concentration  and  the  electrical 
charge  carried  by  the  virus  exert  a  decisive  influence.  The  pressure 
under  which  the  filtration  is  carried  out  is  also  a  factor  of  importance. 

Further,  the  sensitiveness  of  the  tissue  into  which  a  filtrate  is 
inoculated  may  influence  the  decision  whether  a  virus  is  filterable  or 
not.  When  a  sensitive  reagent  is  employed,  the  passage  of  an 
extremely  small  quantity  may  be  detected.  For  instance,  the  dose  of 
neurovaccinia  required  to  infect  a  rabbit  if  inoculated  intracerebrally 
is  1/lOOth  to  l/lOOOth  of  that  required  to  produce  pustules  on  apphca- 
tion  to  the  scarified  skin.  An  experimenter  using  the  former  method 
might  conclude  that  the  virus  passed  a  i)acteria-proof  filter,  one  using 
the  latter  that  it  did  not.  The  sensitiveness  of  tlie  tissue  into  which 
a  filtrate  is  inoculated  is  therefore  a  factor  of  capital  importance  often 
neglected  in  interpretations  as  to  the  filterability  or  noii-tilti-rability 
of  the  virus. 

The  experiments  of  Moussu  (1926),  Zwick,  Seifried,  and  Witte 
(1920),  and  Ernst  and  Hahn  (1927)  show  that  the  virus  of  IJorna 
disease  can  pass  through  ordinary  bacteriological  filters,  but  tliat 
filtration  of  the  virus  is  not  easily  effected.  Zwick  carried  out  more 
than  thirty  filtration  experiments  with  different  filters  and  inoculati»d 
more  than  100  rabliits  with  the  various  tiltrates  i)efore  he  succeeded 
in  infecting  tlie  animals  with  a  filtrate. 

We  liave  carried  out  two  filtration  ex])eriments,  using  the  follow- 
ing technique:  from  the  brain  of  Rabbit  275,  which  died  on  the  82n(l 
day  after  cerei>ral  inoculation  and  exiiibited  characteristic  lesions 
throughout   the  nervous  system,  an  eniidsion  1  :  :{(•  was  made  with 


ENZOOTIC  ENCEPHALO-MYELITIS  13 

physiological  saline.  The  emulsion  was  placed  in  the  ice-chest  for  three 
hours  to  allow  the  coarser  particles  to  deposit.  The  supernatant  fluid 
was  filtered  by  aspiration  under  low  pressure  through  a  Handler  filter. 

Experiment  1. 

The  filtrate  was  inoculated  intracerebrally  into  Babbits  218a,  168a, 
228a,  and  201a,  on  the  15.3.27. 

During  a  period  of  174  days  following  the  inoculation  Eabbit  218a 
did  not  show  any  symptoms,  and  its  weight  increased  in  an  even 
curve  by  730  gms.  After  this  period  had  elapsed  the  resistance  of 
the  animal  was  tested  with  fresh  passage  virus  by  intracerebral  inocu- 
lation. It  died  in  37  days  after  showing  typical  symptoms  of  the 
disease.  On  microscopic  examination  of  sections  of  the  central 
nervous  system  characteristic  lesions  were  found. 

Rabbit  168a,  weighing  700  gms..  and  Rabbit  228a,  weighing  2,080 
gms.,  also  remained  well  for  174  days  following  the  inoculation,  and 
their  weights  increased  to  1,330  gms.  and  2,640  gms.  respectively. 
They  were  both  reinoculated  intracerebrally  with  fresh  passage  virus 
on  September  5, 1927  (174th  day  since  inoculation),  and  proved  not  to 
be  refractory  to  infection:  Eabbit  168a  dying  14  days  and  Eabbit 
228a  37  days  after  inoculation  with  the  test  dose.  The  control 
rabbit,  25a,  inoculated  intracerebrally  on  the  same  date  as  the  others, 
died  48  days  after  infection,  and  typical  lesions  were  found  in  the 
central  nervous  system, 

Rabbit  201a  was  inoculated  intracerebrally  with  filtrate  on  the 
15.3.27. 


Observations. 

Dale. 

Weight  in  gms. 

Clinical  observations. 

15.3.27 

1,500 

20.3.27 

1,620 

Normal. 

25.3.27 

1,660 

Normal. 

31.3.27 

1,.520 

Animal  appeared  to  be  ill.  Rabbit  found 

2.4.27 

dead  18  days  after  inoculation. 

On  the  autopsy  no  abnormal  condition  of  the  organs  was  observed  and 
the  cultures  of  the  brain  remained  sterile.  On  microscopic  examina- 
tion slight  infiltration  and  perivascular  lesions  were  found  in  the  brain, 
mid-brain,  and  spinal  cord,  and,  in  addition,  a  marked  pathological 
'  satellitism '  of  the  nerve-cells.  Passage  was  made  with  the  brain  of 
this  rabbit  to  a  healthy  rabbit.  No.  260a,  with  the  results  given  in 
detail  below: 

Rabbit  260a,  weighing  1,740  gms.,  was  inoculated  intracerebrally 
with  an  emulsion  of  the  brain  of  Rabbit  201a. 

It  developed  a  disease  of  a  recurrent  nature  and  succumbed  after 
the  second  crisis,  161  days  after  inoculation. 

Autopsy.   All  organs  macroscopically  normal. 

Cultures  in  broth  from  the  brain  remained  sterile. 

Microscopic  examination.  Intense  and  characteristic  lesions  were 
found  in  the  brain  and  in  the  cord.  The  intranuclear  corpuscles  of 
Joest-Degen  were  also  found. 


14  BORNA  DISEASE  AND 


Observations. 

Daft. 

Weight  in  gins. 

2.4.27 

1,700 

19.4.27 

1.800 

28.4.27 

1,900 

5.5.27 

1,920 

12.5.27 

1,600 

16.5.27 

1,570 

Clinical  observations. 
Nothing  abnormal  in  animal's  condition. 


Depressed,  posterior  paresis. 

Placed  on  his  flank  the  animal  made 

several    vain   efforts    to    recover     its 

normal  position. 
Condition  ameliorated — slight  paresis  of 

hindquarters. 


30.5.27 

1.600 

Much  improved. 

13.6.27 

1,640 

Normal. 

22.6.27 

1,740 

,, 

30.6.27 

1,770 

15.7.27 

1,800 

1.8.27 

1,700 

8.8.27 

1,540 

Animal  ill. 

15.8.27 

1,300 

Paresis  of  hindquarters. 

24.8.27 

1,180 

„                     »* 

31.8.27 

1,000 

,• 

9.9.27 

980 

Head  depressed,  paresis 

t\'pical  symptoms  of  the  disease. 
12.9.27  820  Placed  on  the  flank,  the  rabbit  made  vain 

efforts     at     recovering     the     normal 

position. 
13.9.27  Found  dead  161  days  after  inoculation. 

Experiment  2. 

A  second  filtration  experiment  was  carried  out  with  the  same 
technique.  The  filtrate  through  a  Mandler  filter  was  inoculated  into 
the  brain  of  four  rabbits.  150.\.  15*2.a.  "iKU.  aiul  •231.\. 

Babbit  231a  died  accidentally  5  days  after  inoculation. 

Rabbit  210.\  remained  unaffected  and  gained  progressively  in 
weight. 

The  protocols  of  Rabbits  150.\  and  152.\  are  given  below. 


Rabbit  150  a. 

Rabbit  152a. 

Weight  In 

Clinical  obseria- 

Weight  in      Cli 

in ical  observa- 

Dale.                gms. 

tions. 

gms. 

tions. 

15.3.27               1.800 

2,200 

20.3.27               1,880 

2.440 

Normal. 

31.3.27               1,860 

2.44<J 

15.4.27               2,260 

2,.">00 

28.4.27               2.320 

2,660 

5.5.27               2..520 

2,680 

12.5.27               2,420 

Slight  paresis  lie- 
hind. 

2,620 

21.5.27              2,340 

2,680 

30.5.27              2„340 

Condition  im- 
proved.- 

2,620 

•• 

5.6.27               2,460 

Normal. 

2,650 

,. 

13.6.27               2,M(» 

2.790 

,, 

29.6.27               2.700 

., 

2.700 

15.7.27               2,9<W 

2,650 

1.8.27              2,970 

2.«Mt 

29.8.27             3.000 

,. 

2,tU><) 

Therefore  only  Rabbit  1.50.\  showed  sliglit  transitory  .symptoms 
which  ultimately  passed  away  completely.  These  symptoms  may  have 
been  due  to  the  inoculation  of  a  small  dose  of  virus. 


ENZOOTIC  ENCEPHALO-MYELITIS  15 

In  order  to  determine  whether  the  3  rabbits,  150a,  152a,  and  210a, 
had  developed  any  degree  of  immunity  as  a  result  of  inoculation  with 
the  filtrate,  167  days  after  the  first  injection  they  received  intra- 
cerebrally  a  virulent  emulsion  of  brain  at  the  same  time  as  a  normal 
rabbit,  25a.  which  served  as  a  control.  Babbit  150a  died  on  the  49th 
day.  Rabbit  152a  on  the  40th  day,  and  Rabbit  210a  on  the  40th  day, 
and  the  control  rabbit  on  the  48th  day  after  inoculation  of  the  test 
dose.  Typical  lesions  were  found  in  all  cases  on  microscopic  examina- 
tion of  sections  of  brain  and  spinal  cord. 

Conclusions.  The  results  in  our  few  experiments  support  the  con- 
clusion of  Zwick,  Seifried,  and  Witte  (1927)  that  the  virus  of  Borna 
disease  can  pass,  though  with  great  difficulty,  through  bacteriological 
filters  which  retain  ordinary  bacteria,  and  that  the  concentration  of 
virus  in  the  filtrate  is  much  reduced. 

Zwick  succeeded  in  obtaining  an  active  filtrate  after  filtering  a 
virulent  emulsion  of  brain  through  a  Zsigmondy  Bachmann  collodion 
membrane,  of  which  the  size  of  the  pore  was  estimated  to  be  0-75 /u,. 

Effect  of  dilution.  Few  titration  experiments  have  been  made. 
Zwick  records  that  a  virulent  emulsion  of  brain  is  still  capable  of  pro- 
ducing the  disease  in  a  dilution  of  1  :  10,000. 

III.  C'entrifugalization. 
Experiment  1. 
A  homogeneous  emulsion  of  virulent  brain  was  made,  and  after 
the  larger  particles  had  been  allowed  to  deposit,  the  supernatant  fluid 
was  pipetted  and  centrifugaHzed  for  5  minutes  at  5,400  revolutions  per 
minute.  The  supernatant  fluid  after  centrifugahzation  was  carefully 
pipetted  off  and  inoculated  intracerebrally  into  three  rabbits  weighing 
between  1,300  and  1,500  gms.  These  three  rabbits  fell  ill,  showed 
typical  symptoms  and  died,  39,  48,  and  90  days  respectively  after  the 
inoculation.  The  characteristic  lesions  of  Borna  disease  were  found 
in  the  central  nervous  system  of  all  three. 

(a)  Rabbit  322b.  Weight  1,540  gms. 

(b)  Rabbit  321b.  Weight  1,300  gms. 

(c)  Rabbit  320b.  Weight  1,500  gms. 

The  inoculation  was  made  on  15.3.27.  The  protocol  of  the  three 
rabbits  is  given  below: 

Rabbit  322b. 


Date. 

W 

ight  in  gms. 

Clinical  observations. 

15.3.27 

1,540 

23.3.27 

1,680 

Normal. 

25.3.27 

1,700 

31.3.27 

1,700 

10.4.27 

1,620 

^, 

15.4.27 

1,580 

„ 

19.4.27 

1,420 

Paresis  of  hind  quarters. 

28.4.27 

1,220 

Typical  symptoms  of  the  disease. 

2.5.27 

— 

Found  dead  48  days  after  inoculation. 

Culture  of  the  bra 

n.     Negative 

Microscopic 

exam 

'nation.    Intense  and  characteristic  lesions  in  the  central  nervous 

system. 

16  BOENA  DISEASE  AND 

Rabbit  321b. 

Date.  Weight  in  gms.                              Clinieal  obseri-ations. 

15.3.27  1,300 

23.3.27  1,280               Normal. 

25.5.27  1,380 

31.3.27  1,420 

10.4.27  1,480 

15.4.27  1,600 

19.4.27  1,640 

28.4.27  2,000 

10.5.27  2,100 

21.5.27  2,320 

30.5.27  2,080                Paresis  of  hindquart<'rs. 

5.6.27  1 ,800              Typical  symptoms  of  disease. 

11.6.27  1,580              Coma.  Killed  90  days  after  infection. 
Cultures  of  the  brain.  Negative. 
Microscopic  ejcamination.  Intense  and  characteristic  lesions  in  the  central  nervous 

system. 

Bahhit  320b. 

Clinical  obsenations. 


Date. 

Weight  in  gm 

15.3.27 

1,500 

23.3.27 

1,600 

25.3.27 

1,620 

31.3.27 

1,440 

10.4.27 

1,320 

15.4.27 

1,320 

19.4.27 

1,140 

23.4.27 

1,020 

Paresis  ? 

Typical  symptoms  of  the  disease. 

Died  39  days  after  infection. 
Cultures  from  the  brain.   Negative. 

Microscopic  examination.    Mild,  but  characteristic  lesions  in  the  central  nervous 
system. 

Experiment  2. 
A  virulent  emulsion  of  brain  was  allowed  to  deposit,  and  the  super- 
natant fltiid  centrifugalized  as  in  the  last  experiment  (5,400  revs,  per 
minute)  for  15  minutes.  Two  rabbits  were  inoculated  intracerebrally 
with  the  supernatant  fluid.  Both  these  rabbits  developed  the  disease 
typically  with  paralysis,  and  died  28  and  45  days  respectively  after 
the  inoculation,  showing  lesions  of  a  characteristic  nature  in  the 
central  nervous  system. 

(a)  Rabbit  324b.   Weight  1,150  gms. 

(b)  Rabbit  326"b.   Weight  1,150  gms. 
The  inoculation  was  made  on  15.3.27. 

'Die  protocol  oj  these  two  rabbits  is  recorded  below. 

Rabbit  324b. 

Clinical  ol/seri'ations. 
Normal. 


DaU. 

Weight  in  gms. 

15.3.27 

l.l.W 

2(1.3.27 

1..360 

25..3.27 

1.400 

31. .3.27 

1,330 

11.4.27 

1.140 

1.5.4.27 

l.ltM) 

19.4.27 

I,l(i0 

28.4.27 

1.080 

29.4.27 

— 

Commencement  of  symptoms. 

Paralysis  of  hindquarters. 

Found  dead  45  clays  after  in(H'ulation. 
Cultures  from  the  brain.   Negative. 

Microscopic  examination.  The  brain  and  spinal  cord  showed  the  presence  of  typical 
lesions. 


ENZOOTIC  ENCEPHALO-MYELITIS  17 


Rabbit  326b. 

Date. 

W 

eight  in  gms. 

Clinical  observations. 

15.3.27 

1,150 

20.3.27 

1,360 

25.3.27 

1,440 

31.3.27 

1,380 

11.4.27 

1,020 

Paresi.s  of  hindquarters. 

12.4.27 

— 

Found  dead  28  day.s  after  inoculation. 

Microscopic  examination.  The  brain  and  -spinal  cord  showed  the  presence  of  typical 
lesions. 

Conclusion.  Centrifugalization  for  even  15  minutes  at  5,400  rev. 
per  minute  does  not  deprive  the  supernatant  fluid  of  virulence.  This 
fact,  in  addition  to  the  properties  of  filterabihty  and  invisibility  of  the 
pathogenic  agent  suggests  that  the  size  of  the  infective  element  is 
excessively  small.  It  is  affected  by  centrifugalization  in  the  same  way 
as  other  filterable  viruses  such  as  those  of  foot-and-mouth  disease, 
rabies,  herpes,  vaccinia,  and  pohomyeHtis. 


IV.   Eesistance  to  Glycerine. 

Moussu  (1926)  found  that  a  portion  of  brain  preserved  its  virulence 
at  room  temperature  (July-August,  Alfort)  for  18  days,  but  that  its 
pathogenic  action  was  lost  after  32  days.  According  to  Zwick  the 
brain  of  a  rabbit  preserved  its  virulence  in  glycerine  for  from  4  to  5 
months,  and  in  our  experiments  glycerinated  virus  kept  in  the  cold 
room  at  4°  C.  was  still  virulent  after  113,  135,  and  in  one  case  161 


To  find  the  best  conditions  for  keeping  the  virus  in  the  cold  room 
the  following  solutions  were  tried.  (1)  Pure  glycerine,  (2)  pure  gly- 
cerine covered  with  a  layer  of  sterile  paraffin  oil,  (3)  glycerine  mixed 
with  an  equal  part  of  sterile  physiological  saline,  (4)  glycerine  mixed 
with  an  equal  part  of  phosphate  saline  M/25,  pH.  7.6.  The  brain  of 
Rabbit  77a  (dead  of  Borna  disease  48  days  after  inoculation  intra- 
cerebrally  with  typical  lesions  in  the  C.N.S.)  was  taken  aseptically 
and  divided  into  four  equal  portions,  and  one  of  the  portions  placed 
in  each  of  the  four  media  referred  to  above.  At  the  end  of  a  certain 
time  a  fragment  of  each  portion  was  taken  and  an  emulsion  of  it 
inoculated  intracerebrally  into  rabbits  to  test  its  virulence.  A  table 
of  results  is  given  on  page  18. 

The  results  recorded  in  the  table  on  p.  18  show  that  the  virus  may 
remain  virulent  in  the  cold  room  at  4°  C.  at  least  113  days  in  a 
medium  consisting  of  pure  glycerine,  glycerine  diluted  to  50  per  cent, 
with  physiological  sahne,  or  glycerine  diluted  to  50  per  cent,  with 
phosphate  saline  M/25,  pH.  7.6.  In  two  further  experiments  under 
similar  conditions  the  virus  preserved  in  glycerine  remained  virulent 
for  135  days  and  161  days  respectively. 

(1)  An  emulsion  of  the  brain  of  Rabbit  25  (dead  of  Borna  disease 
35  days  after  inoculation)  which  had  been  kept  in  glycerine  in  the 
ice-chest  for  135  days  was  inoculated  intracerebrally  into  Rabbit  145a. 


18 


BORXA  DISEASE  AND 


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ENZOOTIC  ENCEPHALO-MYELITIS  19 

Rabbit  14o.\.   Weight  2,500  gms. 

Clinical  observations. 


Date. 

Weight  in  gms 

13.6.27 

2,500 

22.6.27 

2,500 

29.6.27 

2,500 

7.7.27 

2,320 

14.7.27 

1,900 

19.7.27 

1.550 

20.7.27 

— 

First  symptoms  of  the  disease. 
Typical  symptoms  of  the  disease  in  an  advanced 
stage. 

Died  47  days  after  inoculation. 

Cultures  from  brain.  Negative. 

Microscopic  examination.  Brain  and  cord  showed  presence  of  intense  lesions. 

(2)  Virus  (brain  of  Rabbit  100  dead  of  enzootic  encephalo-myelitis 
on  the  37th  day  after  inoculation)  kept  in  glycerine  161  days  in  the 
chest,  was  inoculated  Lntracerebrally  into  Rabbit  142\  on  the  3.6.27. 

Rabbit  142a.   Weight  2,440  gms. 

Date.  Weight  in  gms.  Clinical  observations. 

13.6.27  2,500  Normal. 

22.6.27  2,620 

29.6.27  2,440 

7.7.27  2,040  Typical  symptoms  commencing. 

14.7.27  1,850 

16.7.27  1,600  Died  43  days  after  inoculation. 

Cultures  from  brain.     Negative. 
Microscopically.    Characteristic  and  intense  lesions  were  present  in  the  central 

nervous  system. 

We  have  observed  that  certain  of  the  rabbits  infected  lntracere- 
brally with  virus  kept  in  glycerine  succumbed  to  the  disease  at  an 
earher  date  than  rabbits  inoculated  with  an  emulsion  of  fresh  virulent 
brain  as  is  indicated  by  the  following  experiment. 

(3)  Rabbit  275  was  inoculated  lntracerebrally  ^ith  a  virus  kept  in 
pure  glycerine  for  48  days  on  10.2.27. 

Clinical  observations. 
No  abnormal  symptoms  observed. 


Date. 

Weight  in  gms 

10.2.27 

2,100 

19.2.27 

2,150 

23.2.27 

2,040 

27.2.27 

1,940 

5.3.27 

1,700 

7.5.27 

— 

9.3.27 

1,520 

12.3.27 

— 

13.3.27 

1,260 

14.3.27 

1,220 

Commencing  paresis  of  the  hind  quarters. 

Paresis  of  hindquarters  more  advanced. 

Animal  remained  in  comer  of  the  cage  hunched  up, 

depressed.  The  paresis  was  still  present. 
Intense  .salivation.   Paralysis  of  the  hind  quarters. 
Died  32  dajs  after  the  inoculation. 

The  microscopic  examination  of  sections  of  brain  and  cord  of  Rabbit  142a  revealed 
the  presence  of  intense  and  characteristic  lesions. 

The  following  table,  which  shows  the  chain  of  the  series  of  passages 
of  the  \'irus  from  rabbit  to  rabbit,  of  which  Rabbit  275  forms  a  con- 
necting link,  demonstrates  the  fact  that  although  it  weighed  more 
than  2,000  gms.  it  died  at  an  earher  date  than  the  other  rabbits  of 
the  same  series. 

b2 


20  BORNA   DISEASE  AND 


Inoculated  iniracerehrally 

Interval  between  inocula- 

Xo. of  Rabbit. 

with 

tion  and  death. 

100 

Fresh  rinis. 

37  days. 

77a 

Fresh  virus  from  Rabbit  100. 

48      „ 

275 

Virus  from  Rabbit  77a  kept  in 
glycerine  for  48  days. 

32     „ 

211 

Fresh  virus  from  Rabbit  275. 

42      „ 

212 

„               „               „ 

*4      „ 

A  similar  observation  has  been  made  a  number  of  times  in  the 
course  of  our  experiments,  and  tliis  is  recorded  as  a  typical  example. 
Levaditi,  Harvier,  and  Nicolau  record  similar  findings  for  the  virus 
of  herpes,  and  this  has  been  confirmed  more  recently  by  Perdrau. 

Y.   Sensitiveness  to  Heat  and  Desiccation. 

(a)  Heat.  Zwick  and  his  collaborators  found  that  cerebral  emul- 
sions heated  for  5,  10,  15,  20,  and  25  minutes  respectively  at  50°  C. 
preserved  their  virulence  for  the  rabbit.  In  some  instances  a  similar 
emulsion  heated  for  30  minutes  at  50°  C.  became  avirulent.  Heated 
for  30  minutes  at  57°  C.  or  10  minutes  at  70°  C.  in  the  water-bath  the 
virulence  of  the  emulsion  was  destroyed. 

(b)  Desiccation.  Zwick  found  that  a  virulent  emulsion  of  brain 
dried  for  6  to  10  hours  at  30°  C.  proved  to  be  avirulent  when  inoculated 
1  or  10  days  after  such  desiccation. 

VI.   Action  of  Ultra-violet  Light.^ 

We  proceeded  in  the  followmg  manner.  A  homogeneous  emulsion 
of  virulent  brain  was  centrifugaUzed  for  5  minutes.  The  supernatant 
riuid  was  carefully  pipetted  into  a  small  quartz  flask  and  exposed  for 
5  minutes  to  the  rays  from  a  mercury  arc.  Two  mercury  vapour 
lamps  (K.B.B.  type,  25  amperes,  210  volts  D.C.)  were  employed 
8  inches  distant.  The  flasks  were  slowly  rotated  during  the  exposure 
so  that  a  fresh  thin  film  of  fluid  was  constantly  exposed  to  the  lamp. 
The  flasks  dipped  periodically  into  cold  water  in  a  trough  to  prevent 
overheating  during  the  experiment.  The  irradiated  fluid  was  inocu- 
lated into  the  brain  of  a  rabbit.  At  the  same  time  a  portion  of  the 
non-irradiated  emulsion  was  inoculated  intracerebrally  into  two 
rabbits,  which  served  as  controls.  The  protocols  of  these  experiments 
are  recorded  on  page  21. 

The  rabbit  inoculated  intracerebrally  with  the  virus  which  had 
been  subjected  to  the  rays  of  the  mercury  arc,  did  not  show  any 
symptoms  during  three  months,  wliile  the  controls  died  after  23  and 
34  days  respectively,  showing  that  the  virus  subjected  to  the  action 
of  ultra-violet  rays  (radiations  of  wave-lengths  5,720-2,320  A.U.)  is 
killed  in  a  maximum  of  5  minutes. 

Kaljbit  1G3a  was  reinoculated,  along  with  a  control  rabbit, 
Xo.  25a,  with  fresh  passage  virus  86  days  later  and  died  on  the 

'  Wc  are  indfbtml  to  Dr.  Kidinow  of  the  Depart  menf  of  Applied  Physiology  ( Katiunal 
Institute  for  Medical  R<-Bearch)  for  his  collaboration  in  these  exp<Tinient8. 


ENZOOTIC  6NCEPHAL0-MYELITIS 


21 


38th  day.    The  control  rabbit  succumbed  to  the  injection  on  the 
48th  day. 


Irradiated  emulsion. 

Controls — non-irradiated  emulsion. 

Habbit  163a. 

Rabbit  165a. 

Rabbit  166a. 

Weight,  2,120 

gms. 

Weight,  2,010  gms. 

Weight,  1,900  gms. 

13.6.27.  1,940  ems. 

Nonnal. 

13.6.27.  1,950  gms.  Normal. 

13.6.27.  1,780  gms. 

22.6.27.  1,950     „ 

22.6.27.  1,900     „ 

22.6.27.  1,500     „ 

29.6.27.  2,000     „ 

" 

29.6.27.  2,000     „ 

29.6.27.  1,400  gms.  Typical 
symptoms. 

7.7.27.  2,160     „ 

„ 

7.7.27.  1,490  gms.  Typical 

3.7.27.  Found  dead  23  days 

symptoms. 

after  inoculation. 

14.7.27.  2,300     „ 

14.7.27.    1,000   gms.    Rabbit 

Microscopic   examination.     In- 

1.8.27. 2,650     „ 

,, 

dying,  killed  34  days  after 

tense  lesions  in  the  central 

15.8.27.  2,800     „ 

,, 

infection. 

nervous  system  of  a  charac- 

28.8.2T. 2,800     „ 

,, 

Microscopic  examination.   In- 

teristic type. 

5.9.27.  2,800     „ 

" 

tense  characteristic  lesions 
in     the     central     nervous 
system. 

VII.  The  Action  of  Hexamethylenetetramine  (Urotropinb). 
The  experiments  of  Moussu  (1926)  show  that  when  equal  quantities 
of  a  virulent  emulsion  of  brain  and  a  10  percent,  solution  of  urotropine 
are  mixed  and  kept  at  room  temperature  for  12  hours,  the  virus  can 
still  be  demonstrated  in  the  mixture  after  this  time. 

VIII.   Action  of  Chloroform  and  Ether. 

(a)  Action  oj  Chloroform.  A  thick  emulsion  of  the  brain  of  a  rabbit 
dead  of  experimental  Borna  disease  (Rabbit  182a,  dead  on  the  40th 
day  after  cerebral  infection)  was  mixed  with  five  volumes  of  chloro- 
form, and  the  mixture  kept  for  18  hours  at  room  temperature.  The 
fluid  part  of  the  mixture  was  removed  by  evaporation  in  a  vacuum 
over  sulphuric  acid.  The  dried  brain  residue  was  powdered  in  a  mortar 
and  suspended  in  physiological  saUne.  This  suspension  was  then  in- 
oculated intracranially  into  two  rabbits  (234a  and  235a).  Rabbit 
219a,  which  was  inoculated  with  an  emulsion  of  the  same  brain  not 
mixed  with  chloroform,  served  as  control. 

Five  days  later  both  Rabbits  234a  and  235a  received  a  further 
inoculation  with  the  virus  which  had  undergone  similar  treatment 
with  chloroform. 


Control  rabbits — vims  not 

Virus  treated  with  dhlaroform. 

treated  with  chloroform. 

Rabbit  235a.  2,600  gms. 

SabbU  234a.  2,470  gms. 

Rabbit  219a.  2,000  gms. 

5.8.27. 

Inoculated. 

5.8.27.  Inoculated. 

5.8.27.  Inoculated. 

10.8.27. 

Second   inoculation 

10.8.27.  Second     inoculation 

15.8.27.  2,100  gms.  Normal. 

15.8.27. 

2,300  gms.  Normal. 

15.8.27.  2,450  gms.  Normal. 

24.8.27.  1,789  gms.     Com- 
mencement of  disease. 

24.8.27. 

2,350     „ 

24.8.27.  2,.500     „ 

29.8.27.  1,580  gms.  Typical 
disease. 

29.8.27. 

2,400     „ 

29.8.27.  2,550     „ 

2.9.27.  Found  dead  31  days 

9.9.27. 

2,500     „ 

31.8.27.  Eeinoculated  with 

after  infection. 

15.9.27. 

2,700     „ 

fresh  virulent  virus. 

Pa.ssage  of  brain  to  fresh  rabbit 

28.9.27. 

2,880     „ 

9.9.27.  2,600  gms.  Normal. 

is  positive. 

15.9.27.  2,700     „ 

Microscopic  examination  of  sec- 

28.9.27. 2,670     „        „ 

tions  of  brain  and  cord  re- 
vealed intense  and  charac- 
teristic lesions. 

22  BOEXA  DISEASE  AND 

These  experiments  show  that  the  virus  is  inactivated  by  contact  -vnth 
chloroform  for  18  homrs  at  room  temperature. 

(b)  Action  of  Ether.  The  teclinique  -was  similar  to  that  employed  in 
the  above  experiment  ■with  chloroform.  Eabbit  231a,  which  received 
an  intracerebral  inoculation  of  ether-treated  brain  emulsion,  was 
kept  mider  observation  for  5  months.  It  never  showed  any  symp- 
toms of  Boma  cUsease  and  gained  780  gms.  in  weight.  The  control 
rabbit,  219a,  inoculated  with  non-treated  brain  died  in  31  days  of  a 
typical  infection. 

Note. — The  experiments  on  the  effect  of  chloroform  and  ether  on  the  virus  are 
only  preliminaiT.  Obviously  there  are  certain  details  in  the  technique  used  which 
will  have  to  be  controlled,  and  improved  methods  are  now  being  employed. 

IX.  Action  of  Formalin. 
About  one  gramme  of  the  brain  of  Eabbit  3o5a  (wliich  died  27  days 
after  intracerebral  inoculation  with  the  passage  virus  of  Boma 
disease)  was  emulsified  in  15  c.cms.  of  a  solution  of  formalin  in 
physiological  saline  (2  :  1,000).  The  emulsion  was  rendered  as  homo- 
geneous as  possible,  and  was  then  placed  at  the  temperature  of  the 
laboratory  for  18  hours.  Subsequently  rabbits  were  inoculated  intra- 
cerebrally  with  the  formohzed  emulsion.  An  emulsion  of  the  brain  of 
Eabbit  3o5a  in  a  similar  dilution  not  treated  with  formaUn  was 
inoculated  as  a  control  into  the  brain  of  a  rabbit. 


Protocols.  The  inoculations  were  made  10.8.27. 

Virus  treated  urilh  formalin  iaoeulaUd  iniracerebraUy. 

Control — virus  not  treated  vilh 
formalin. 

Rabbit  44k.  2,710  gms. 

Sabbit45\.  S,0OO  gms. 

Rabbit  356a.  2,100  gms. 

15.8.27.  2,600  gms.  Noimal. 
23.8.27.  2,700     „ 
30.8.27.  2,t>«)     „ 
5.9.27.  2,750     „ 

11.9.27.  2,740     „ 

15.9.27.  2,800     „ 

22.9.27.  2,720     „ 

6.10.27.  2,740  „ 
12.10.27.  2,800  „ 
21.10.27.  2,780    „          „ 

15.8.27.  1,800  gms.  Nonnal. 
23.8.27.  1,750     ., 
30.8.27.  1,650     „ 
5.9.27.  1,800     „ 

11.9.27.  1,950     „ 

15.9.27.  2,010     „ 

22.9.27.  1,980     „ 

6.10.27.  2,000  „ 
12.10.27.  2,000  „ 
21.10.27.  2,010     „ 

15.8.27.  2,(60  gms.  Normal. 
23.8.27.  2,150     „ 
30.8.27.  1,900     „ 
5.9.27.  1,780  gni.s.  Disease 

commencing. 
11.9.27.  1,550  gms.  Typical 

dUease. 
14.9.27.  Found  dead  30th  day. 
Typical  microscopic  lesions  in 

central  nervous  system. 

Conclusion.  The  conclusion  arrived  at  is  that  formalin  in  a  con- 
centration of  0-2  per  cent,  inactivates  the  virus  after  18  hours' contact 
at  room  temperature. 

X.   Culture. 

All  attempts  at  cultivation  of  the  virus  liavc  remained  ncfjative 
up  to  the  present. 

XI.  Summary. 

From  the  observations  of  the  authors  quoted  and  our  own,  it 
appears  that  the  virus  of  Boma  disease  possesses  the  properties  common 
to  those  of  vaccinia,  herpes,  rabies,  and  poliomyelitis  whicli  Levaditi 
has  grouped  together  under  the  name  '  ectodernioses  neurotropes*. 
Under  favourable  conditions  it  filters  through  bacteria-proof  filters, 


ENZOOTIC  ENCEPHALO-MYELITIS  23 

although  most  of  the  virus  is  held  back,  and  through  a  collodion 
ultrafilter  which  will  allow  colloidal  particles  of  large  dimensions  to 
pass.  The  infectivity  of  the  supernatant  fluid  cannot  be  removed  by 
centrifugalization  for  15  minutes  at  5,400  revs,  per  minute.  It  is 
resistant  to  the  action  of  glycerine,  but  sensitive  to  desiccation,  ultra- 
violet hght  and  heat.  It  is  destroyed  by  ether,  chloroform,  and  for- 
mahn.  It  has  not  been  propagated  outside  the  body. 

3.   TRANSMISSION  OF  EQUINE  STRAIN  TO  RABBIT 
AND  FROM  RABBIT  TO  SHEEP,  AND  VICE  VERSA 

Moussu  (1926)  inoculated  an  emulsion  of  the  brain  of  a  rabbit  pre- 
viously infected  with  the  virus  from  a  horse  into  the  anterior  chamber 
of  the  eye  of  a  horse.  The  animal  developed  symptoms  3  days  after 
the  inoculation  and  died  in  8  days.  The  lesions  found  in  the  brain 
were  very  intense,  infiltrative,  and  haemorrhagic.  A  rabbit  inoculated 
with  an  emulsion  from  the  brain  of  this  horse  died  4  days  later.i 
This  same  author  failed  to  infect  horses  with  virulent  material  from 
rabbits  by  subcutaneous  inoculation  or  per  os.  Zwick  and  his  col- 
laborators (1926)  inoculated  a  horse  intracerebrally  with  the  brain  of 
a  rabbit  suffering  from  experimental  Borna  disease.  The  virus  had 
been  passaged  in  this  species  of  animal  nine  times.  The  horse  fell 
ill  53  days  after  the  inoculation.  For  11  days  it  showed  the  typical 
symptoms  of  encephalo-myehtis  and  death  followed  64  days  after 
the  inoculation.  The  lesions  of  the  nervous  system  were  characteristic 
of  Borna  disease.  The  intranuclear  corpuscles  of  Joest-Degen  were 
demonstrated  in  the  ganghon  cells  of  the  Ammon's  horn. 

We  have  referred  previously  to  the  experiments  of  Beck  and  Froh- 
bose  (1926),  Moussu  (1926),  Miessner  (1926),  and  Ernst  and  Hahn 
(1927),  wliich  showed  that  the  virus  of  encephalo-myelitis  originating 
from  sheep  can  be  transmitted  to  rabbits  by  experimental  inocula- 
tion. Moussu  and  Marchaud  (1924)  succeeded  in  passing  the  disease 
from  sheep  to  sheep.  Zwick  and  his  collaborators  (1926)  failed  to 
transmit  the  disease  to  adult  sheep  by  intracerebral  inoculation  with 
a  strain  derived  from  a  horse  and  passed  through  rabbits.  Using  the 
same  strain  of  virus  they  succeeded,  however,  in  conferring  the  disease 
on  young  lambs.  Death  followed  88  days  after  the  inoculation  and 
typical  cerebral  lesions  were  revealed. 

Direct  inoculation  from  horse  to  lamb  was  also  successful.  The 
lamb  showed  characteristic  symptoms  and  died  92  days  after  infec- 
tion. Inoculation  of  the  brain  of  this  lamb  to  a  rabbit  gave  a  positive 
result,  but  inoculation  of  the  cord  gave  a  negative  result.  Beck  and 
Frohbose  (1926)  did  not  succeed  in  infecting  the  horse  by  the  intra- 
cerebral route  with  virus  from  sheep  dead  from  the  spontaneous 

'  The  experimenta  of  Moussu  and  Marchand  have  been  criticized  since  his  inocu- 
lated animals  succumbed  very  early,  and  also  because  the  lesions  of  the  brain  were 
surprisingly  acute  when  compared  with  those  found  in  the  classical  disease  occurring 
spontaneously  or  in  animals  infected  with  the  viruses  isolated  by  the  German  school. 
We  had  the  intention  of  comparing  their  strain  with  those  of  Zwick  and  Miessner,  but 
Moussu  has  informed  us  that  his  strain  is  not  now  available. 


24  BOEXA  DISEASE  AND 

disease.   On  the  other  hand,  they  succeeded  in  infecting  sheep  with 
virus  obtained  from  horses. 

From  the  foregoing  resume  of  the  hterature  the  following  conclu- 
sions may  be  drawn: 

(1)  The  virus  originating  from  horses  passaged  through  rabbits  can 
be  transmitted  back  to  the  horse. 

(2)  The  virus  taken  directly  from  the  horse  or  subsequently  pas- 
saged through  rabbits  is  pathogenic  for  lambs. 

(3)  Attempts  at  transmitting  the  disease  from  sheep  to  horses  have 
so  far  been  unsuccessful. 

4.  EXPEEIMENTAL  DISEASE  IN  THE  EABBIT 

I.   Experimental  Transmission  of  the  Disease  to  the 

Rabbit 

Moussu  (1926)  inoculated  an  emulsion  of  the  brain  of  a  horse  dead 
of  encephalo-m5-ehtis  into  the  anterior  chamber  of  the  eye  of  the 
rabbit.  In  one  experiment  three  animals  were  inoculated  by  this 
route.  One  died  on  the  9th  day;  the  two  others  survived;  the  rabbit 
which  died  constituted  the  head  of  the  series  of  passages  that  the 
author  continued  until  he  obtained  a  'fixed'  virus  which  killed  the 
rabbits  infected  by  the  intraocular  route  in  4  to  6  days.  In  another 
experiment  using  similar  material  one  out  of  live  animals  inoculated 
intraocularly  died  11  days  after  receiving  the  injection;  the  other  four 
survived.  In  the  majority  of  cases  a  marked  excitability  was  a 
characteristic  symptom.  Moussu  states,  however,  that  certain 
rabbits  die  'following  an  infection  with  a  slower  evolution,  lasting 
more  than  a  fortnight'. 

Zwick  inoculated  rabbits  intracerebrally  with  the  cerebral  sub- 
stance (Ammon's  horn,  caudate  nucleus,  and  cerebral  cortex)  taken 
from  the  brain  of  a  horse  dead  of  Borna  disease.  He  observed  typical 
symptoms  of  the  disease  after  a  period  of  about  4  weeks,  and  the 
lesions  were  analogous  to  those  described  previously  in  the  horse. 
Passage  from  rabbit  to  rabbit  was  effected,  the  period  of  incubation 
after  inoculation  being  about  3  weeks. 

Zwick  and  his  collaborators  (1926)  succeeded  in  infecting  rabbits 
with  the  virus  from  16  out  of  21  cases  of  horses  dead  of  Borna  disease 
which  was  verified  histologically.  The  incubation  period  of  the  disease 
in  such  rabbits  infected  by  the  intracerebral  route  was  from  3  to  4 
weeks.  Death  ensued  8  to  14  days  after  the  appearance  of  the 
characteristic  symptoms. 

Beck  and  Frohbose  (1926)  also  infected  rabbits  with  the  virus  from 
horses  and  sheep.  Miessner  (1926)  with  sheep  virus,  and  Ernst  and 
Hahn  (1927)  with  viruses  from  horses,  sheep,  and  cattle. 

II.  Authors'  Observations. 

The  virus  of  encephalo-myelitis  of  equine  or  ovine  origin  isolated 

by  tlie  (terman  workers  does  not  become  'fixed'  when  passaged 

through  rabbits.  The  period  of  incubation  varies  from  15  to  50  days, 

and  w('  have  also  observed  recurrent  forms  of  the  disease  in  rabbits. 


ENZOOTIC  ENCEPHALO-MYELITIS 


25 


We  have  inoculated  more  than  200  rabbits  by  the  intracerebral 
route  with  the  virus  originating  from  horses  or  sheep.  In  Table  I 
the  period  of  incubation  and  the  time  between  the  onset  of  the  disease 
and  death  is  recorded  for  a  total  of  50  rabbits  used  for  passaging 
a  virus  originally  obtained  from  a  horse  and  sent  to  us  by  Professor 
Zwick.  Table  I. 


Number 

Commence- 

Duration 

of 

Weight  in 

ment  of 

of  the 

Lesions  in 

rabbit. 

gms. 

disease. 

disease. 

9  Death. 

C.N.S. 

25 

2,160 

21st  day 

14  days 

35th  day 

Intense. 

22 

2,000 

20tli    „ 

4     „ 

24th    „ 

Positive. 

24 

1,420 

21st    „ 

19     „ 

40th    „ 

,, 

IOd 

2,500 

27th   „ 

10     „ 

37th    „ 

Intense. 

llD 

1,000 

17th    „ 

10     „ 

27th    „ 

,, 

*93a 

1,520 

7th    „ 

1     „ 

8th    „ 

Slight.' 

68a 

1,150 

20th    „ 

5     „ 

25th    „ 

Ditense. 

50a 

1,620 

40th    „ 

13     „ 

53rd     „ 

,, 

77a 

1,570 

38th    „ 

10     „ 

48th     „ 

,, 

80 

1,360 

21st    „ 

7     „ 

28th    „ 

,, 

5lA 

1,350 

20th    „ 

8     „ 

28th    „ 

,, 

34 

2,000 

43rd    „ 

14     „ 

57th    „ 

„ 

298 

820 

25th   „ 

14     „ 

39th    „ 

,, 

275 

2,100 

25th    „ 

7     „ 

32nd   „ 

,, 

202 

1,050 

29th    „ 

3     „ 

32nd    „ 

„ 

282 

1.040 

27th    „ 

4     „ 

31at     „ 

,, 

244 

1,200 

33rd    „ 

12     „ 

45th    „ 

„ 

261 

1,280 

23rd    „ 

8     „ 

31st     „ 

„ 

232 

1,340 

20th    „ 

11     „ 

31st     „ 

,, 

219 

1,820 

16th   „ 

5     „ 

21st     „ 

„ 

295 

1,300 

9th   „ 

10     „ 

19th    „ 

Very  intense. 

211 

1,580 

31st    „ 

11     „ 

42nd    „ 

j^ 

212 

1,400 

31st    „ 

13     „ 

44th    „ 

^, 

220 

1,500 

31st    „ 

8     „ 

39th    „ 

,, 

222 

1,540 

35th    „ 

13     „ 

48th    „ 

,, 

224 

1,150 

32nd  „ 

13     „ 

45th    „ 

,, 

226 

1,150 

24th    „ 

4     „ 

28th    „ 

Positive. 

70 

1,700 

20th    „ 

2     ,, 

22nd    „ 

Intense. 

56 

1,400 

20th    „ 

6     „ 

26th    „ 

,, 

61 

1,500 

24th    „ 

6     „ 

30th    „ 

251 

1,280 

18th    „ 

9     „ 

27th    „ 

237 

1,680 

18th    „ 

13     „ 

31st     „ 

255 

1,920 

35th    „ 

7     „ 

42nd    „ 

256 

1,860 

32nd  „ 

15     „ 

47th    „ 

30 

1,120 

28th    „ 

8     „ 

36th    „ 

44 

1,350 

26th   „ 

7      „ 

33rd    „ 

58 

1,000 

27th    „ 

7     „ 

34  th    „ 

Positive. 

*291 

950 

No  sympt. 

— 

15th    „ 

Slight.' 

85b 

1,500 

32nd  „ 

6     „ 

38th    „ 

Very  intense. 

*43 

1,020 

No  svmpt. 

— 

12th    „ 

Slight. 

60 

1,800 

30tii   „ 

7     „ 

37th    „ 

Intense. 

62 

1,610 

22nd  „ 

17     „ 

39th    „ 

,^ 

271 

1,220 

31st    „ 

10     „ 

41st     „ 

„ 

•80 

820 

No  sympt. 

— 

7th    „ 

Slight. 

269 

2,200 

19th   „ 

2     „ 

21st    „ 

Positive. 

•775 

1,780 

No  sympt. 

— 

13th    „ 

Positive.' 

296 

2,900 

28th   „ 

7     „ 

33rd    „ 

Intense. 

277 

2,100 

13th    „ 

11     ,. 

24th    „ 

„ 

78s 

1,700 

24th    „ 

8     „ 

32nd    „ 

^^ 

116a 

1,700 

18th    „ 

4     „ 

22nd    „ 

" 

'  When  the  brain  of  these  rabbits  was  passaged,  positive  results  were  obtained. 
Rabbits  inoculated  with  passage  of  virus  from  the  brain  of  Rabbit  No.  93a  died  in 


26  BOEXA  DISEASE  AND 

Of  50  rabbits  inoculated  in  the  brain : 
23  died  between  21  and  33  days  after  inoculation. 
6  died  in  less  than  21  days. 
21  died  in  more  than  33  days. 
Only  exceptionally  did  the  rabbit  die  in  less  than  3  weeks  when 
infected  by  the  intracerebral  route.    The  detailed  histopathological 
study  of  each  case  showed  that  the  rabbits  dead  7  to  8  days  after 
inoculation  had  minimal  infiltrative  lesions  in  the  central  nervous 
system.  The  presence  of  virus  in  the  brain  was  proved  b}'  subsequent 
passage  and  in  all  cases  complete  autopsies  were  made  to  exclude  the 
possibihty  of  death  from  other  causes. 

The  animals  which  died  between  21  days  and  57  days  presented  the 
characteristic  lesions  in  the  nervous  system,  which  are  described  in  full 
in  the  chapter  dealing  with  the  histopathology  of  the  disease.  The 
intensity  of  the  lesions  was  not  in  direct  relationship  with  the  duration 
of  the  malady.  As  has  also  been  observed  by  Zwick  the  incubation 
period  was  longer  in  larger  animals.  Generally,  rabbits  weighing  less 
than  1,500  gms.  were  more  susceptible  to  infection  than  older  rabbits. 
Excluding  the  five  animals  in  Table  I  marked  with  an  asterisk,  all 
of  which  died  in  15  days  or  under,  the  average  time  which  elapsed 
between  the  intracerebral  inoculation  and  the  death  of  the  animals 
in  our  experiments  was  20  daj's  in  rabbits  of  less  than  1,500  gms.  and 
86  days  in  rabbits  over  this  weight  at  the  time  of  infection. 


Table  II. 


Number 

Commence- 

of 

Weight  in 

ment  of 

Duration 

Lesions  in 

rtMit. 

gnu. 

disease. 

of  disease. 

Death. 

C.NJS. 

164 

1,220 

21st  day 

8  days 

29th  day 

Intense. 

18 

1,240 

26th    „ 

9     „■ 

35th    .. 

246 

1,380 

28th    „ 

10     „ 

38th    ., 

„ 

234 

1,420 

7th    „ 

— 

7th    „ 

Slight. 

243 

1.420 

20th    „ 

2     „ 

22nd    „ 

Intense. 

235 

1,500 

22nd  „ 

9     „ 

33rd    „ 

„ 

262 

2,220 

40th   „ 

8     „ 

48th    „ 

„ 

<l30 

3,640 

23rd   „ 

12     „ 

35th    „ 

., 

10 

2,140 

18th   „ 

9     „ 

27th    „ 

„ 

91 

1,620 

19th   „ 

8     „ 

27th    „ 

„ 

274 

1,770 

18th   „ 

10     „ 

28th    „ 

„ 

28 

1,900 

23nl   „ 

14     „ 

37th    „ 

„ 

86 

1,800 

20th   „ 

12     „ 

32nd    „ 

„ 

63 

1,600 

20th   „ 

8     ., 

28th    „ 

„ 

162a 

2,180 

2Ut    „ 

12     ,. 

33rd    „ 

„ 

167a 

2,420 

22nd   „ 

7     ., 

29th    ,. 

178 

1,770 

25th    „ 

9     „ 

34th    ,. 

„ 

207a 

840 

19th   ,. 

2    .. 

2lBt      „ 

Well  marked. 

181a 

2.100 

26th   ., 

6     ,. 

32nd    „ 

„ 

69 

1,670 

26th   „ 

7     ., 

33rd    ., 

„ 

180a 

1,890 

27th   .. 

12     „ 

30th    „ 

.. 

170a 

2,220 

26th   .. 

6     „ 

32nd    „ 

•• 

Table  II  sots  forth  similar  observations  in  the  case  of  22  rabbits 

53  days;  from  No.  291.  in  38  days;  from  No.  77s  in  21  days.     Intense  lesions  were 
found  ill  the  C.N. 8.  of  these  latter  rabbits. 


ENZOOTIC  ENCEPHALO-MYELITIS  27 

inoculated  with  a  strain  of  ovine  origin  kindly  sent  to  us  by  Professor 
Miessner.  The  average  time  between  inoculation  and  death  was  33 
days  for  the  15  rabbits  weighing  more  than  1,500  gms.  and  28  days 
for  the  6  rabbits  weighing  less  than  1,500  gms. 

III.  Symptomatology  of  the  Disease  ix  the  Eabbit. 

Obsen-ations  have  been  made  on  over  200  rabbits.  During  the 
first  2  to  4  days  following  the  injection  the  weight  of  the  animal 
decreases  shghtly,  to  return  later  to  the  normal.  Once  the  period  of 
traumatic  shock  has  subsided  the  animal  puts  on  weight  and  no  mor- 
bid symptoms  are  seen  during  15  to  20  days.  Subsequently  it  becomes 
slow  in  its  movements  and  appears  depressed;  the  weight  decreases 
progressively  and  the  first  characteristic  symptom  develops.  When 
the  rabbit  is  placed  on  its  side  it  makes  efforts  to  recover  its  feet, 
beating  the  air  with  its  hind  legs  before  eventually  recovering  the 
normal  position.  Wliile  the  depression  referred  to  above  suggests 
modifications  in  the  meninges  and  cerebrum,  the  symptoms  described 
later  point  to  changes  in  the  spinal  cord. 

The  animal  assumes  a  characteristic  attitude  in  the  cage  with 
the  head  in  the  angle  formed  by  two  walls;  it  appears  somnolent 
and  the  somnolence  lasts  till  the  end  of  the  disease.  The  symptoms 
of  nen'ous  origin  become  mtensified  gradually;  among  these  are 
those  of  amaurosis.  When  the  animal  is  allowed  to  run  towards 
an  object  it  runs  into  it  as  if  it  had  not  seen  it.  Grinding  of 
the  teeth  is  observed,  sometimes  with  increased  sahvation.  There 
is  paresis  of  the  ears,  which  fall  to  the  right  and  left  of  the  head. 
The  head  itself  is  depressed.  When  the  animal  is  placed  at  the  edge 
of  a  table  it  hangs  its  head  over  the  edge  below  the  level  of  the 
rest  of  the  body.  Trismus  may  occur.  The  symptoms  of  a  myelitic 
character  become  exacerbated.  Taken  from  the  cage  and  placed  on 
its  side,  the  rabbit  makes  vain  efforts  to  rise.  At  this  stage  its  position 
at  rest  is  characteristic;  it  remains  hunched  up  in  a  corner,  the  head 
is  dropped  as  if  it  was  no  longer  capable  of  supporting  it  (Fig.  1), 
and  sometimes  the  back  is  humped.  The  muscles  of  the  back  be- 
come soft  and  flaccid.  Attempts  to  resist  with  the  hind  legs  when 
the  animal  is  held  by  the  skin  of  the  back  are  feeble  or  absent.  Finally 
paralysis  of  the  hind  quarters  occurs,  which  spreads  later  to  the 
fore-hmbs.  The  animal  ceases  to  feed,  either  from  loss  of  appetite 
or  difiicultj'  in  deglutition,  and  loses  weight'.  In  certain  cases 
the  loss  of  weight  may  be  the  dominant  feature  of  the  disease.  Very 
often  rabbits  at  the  end  of  the  disease  have  lost  nearly  half  of  their 
original  weight  (Charts  I  and  II). 

We  have  never  observed  excitement  in  our  experimental  animals, 
but  always  depression. 

The  study  of  the  blood  has  given  inconstant  results.  In  certain 
animals  a  sHght  hyperleucocytosis  has  been  observed  with  a  shght 
increase  in  polymorphonuclears.  In  others  the  leucocytosis  16,000 
to  18,000  per  c.mm.  was  accompanied  by  lymphocytosis.    In  the 


28 


BOENA  DISEASE  AND 


r 

Gms, 
2000 

\ 

\ 

V 

I 

\ 

\ 

N 

1400 

\ 

1 

I 

1200 

OiED  ON  30TH  DAy\ 

8       10      12      14      16      J8      20     22      24     26 
Chart  I. 


i 

- 

A 

A 

I 

\ 

,i 

\r 

v\ 

,^ 

\ 

VI 

n 

A 

1  '• 

\ 

/"* 

V 

/ 

\ 

Gms. 

1 

\ 

^J> 

-. 

—o 

_.<y 

■'^ 

\ 

\ 

\ 

1800 

! 

1600 

EMPERATURE 

\ 

1400 

ON57-I 
1 

PHD 

ENZOOTIC  ENCEPHALO-MYELITIS 


29 


terminal  stage  a  marked  increase  in  polymorphonuclears  is  the  rule. 
The  number  of  erythrocytes  remains  unchanged,  and  they  show  no 
morphological  changes. 

Chart  III  shows  the  parallelism  between  the  augmentation  of  the 
number  of  leucocytes  per  c.mm.  and  the  number  of  lymphocytes 
obtained  from  the  leucocytic  count.  This  modification  of  the  number 
of  leucocytes  is  not  constant. 

No  fever  exists  during  the  course  of  the  disease  in  the  rabbit.  This 
fact  was  noted  constantly  when  the  temperature  of  a  series  of  rabbits 


i 

\  i 

I 

1 

?. 

\; 

", 

/ 

\ 

h 

f 

', 

1 

\ 

^ 

\ 

\\ 

\ 

1 

! 

/ 

,' 

v 

I 

1     ' 

1 

. 

• 

/ 

^ 

V 

1 

\ 

j    1 

K 

/ 

'/ 

^> 

1 

k 

\ 

■ 

1/ 

\ 

' 

i  / 

\ 

V 

\l 

V 

1 

LEUCOCYTES 

\ 

DIED 

\ 

o 

, 

1    1    1    1    1    1 

b 

10      14      18      22     26 


10      14      18      22     26     ^      3 


inoculated  intracerebrally  was  taken  daily  at  the  same  hour.  Death 
takes  place  in  coma — the  temperature  being  hypothermic  (35°  C- 
34°  C. :  see  Chart  II). 


IV.   Routes  by  which  Rabbits  can  be  Infected. 

Intracerebral. 

Intracerebral  inoculation  produces  the  disease  in  a  constant  manner, 
followed  by  death. 

Intratliecal. 

Beck  (1925)  infected  by  introducing  virus  intrathecally,  and  we 
have  also  succeeded  in  infecting  rabbits  by  this  route. 

Experiment  1 .  Rabbit  207,  weighing  780  gms.  was  inoculated  intra- 
thecally in  the  lumbar  region  with  0-5  c.cm.  of  a  virulent  emulsion 
of  brain   diluted  1  :  10  in  physiological  saline.    Forty- three  days 


30  BORXA  DISEASE  AND 

after  the  inoculation  the  condition  of  the  animal  was  such  that  when 
taken  out  of  the  cage  it  walked  with  its  legs  spread  out  from  the 
body.  Paresis  of  the  hiiid  quarters  was  well  marked,  and  increased 
gradually.  The  animal  wasted  and  died  66  days  after  the  inoculation. 
Examination  of  the  brain  revealed  the  presence  of  characteristic 
lesions,  and  a  passage  of  this  brain  to  a  healthy  rabbit  gave  a 
positive  result. 

Sciatic  Nerve. 

Introduction  of  several  drops  of  a  virulent  emulsion  of  the  virus 
into  the  sciatic  nerve  did  not  infect  animals  with  encephalo-mj'eUtis 
in  the  experiments  of  Moussu  (1926),  and  Zwick  and  his  collaborators 
(1926).  We  have,  however,  succeeded  several  times  in  producing 
Boma  disease  in  rabbits  inoculated  by  this  route. 

Experiment  2.  The  right  sciatic  nerve  of  Eabbit  208,  weighing 
850  gms.,  was  exposed  by  incision  and  2  or  3  drops  of  a  virulent 
emulsion  of  the  brain  of  a  rabbit  infected  with  encephalo-myehtis 
was  injected  into  the  substance  of  the  nerve-trimk.  The  point  of  intro- 
duction of  the  needle  was  seared  to  prevent  the  exit  of  fluid  into  the 
surrounding  tissues.  The  operation  was  carried  out  aseptically  and 
the  incision  healed  by  first  intention.  The  animal  showed  no  morbid 
symptoms  for  35  days  and  put  on  weight,  reaching  1,350  gms. 
Subsequently  it  became  prostrate,  wasted  progressively,  and  showed 
marked  inco-ordination,  which  became  accentuated  later;  the  animal 
died  on  the  48th  day.  The  brain  was  proved  to  be  bacteriologically 
sterile,  and  no  lesions  were  found  which  might  ser^-e  to  explain  the 
cause  of  death,  other  than  those  in  the  central  and  peripheral  nervous 
system.  These  were  of  an  intense  character,  and  were  found  in  the 
brain,  in  the  spinal  cord  (cervical,  thoracic,  dorsal,  and  lumbar 
regions),  and  also  in  the  inoculated  nerve. 

Emulsions  from  the  brain  and  also  the  dorso-lumbar  part  of  the 
spinal  cord  were  inoculated  into  the  brain  of  fresh  rabbits.  These 
inoculations  produced  the  disease,  showing  that  the  virus  was  present 
both  in  the  brain  and  cord  of  rabbits  inoculated  into  a  peripheral 
ner%'e. 

Babbit  270  was  also  inoculated  into  the  sciatic  nerve  by  the  same 
method  as  recorded  above  for  Rabbit  208.  This  rabbit  (270)  showed, 
65  days  after  the  inoculation,  paralysis  of  the  leg  into  the  sciatic  nerve 
of  which  virus  had  been  inoculated.  Paralysis  of  the  hind  quarters 
followed,  with  grinding  of  the  teeth,  and  other  typical  symptoms. 
The  animal  was  found  dead  on  the  78th  day.  Lesions  wore  demon- 
strated throughout  the  nervous  system  (brain,  mesencephalon,  cord 
in  all  regions,  inoculated  nerve,  the  sciatic  of  the  opposite  side  (non- 
inoculated),  as  well  as  in  the  brachial  nerves). 

The  detailed  description  of  these  lesions  will  be  given  later. 

Rabbit  27t>  was  inoculated  into  the  right  sciatic  with  the  same 
technique  as  before.  The  leg  on  the  side  of  inoculated  nerve  became 
useless  after  28  days,  and  the  animal  died  8  days  later  with  lesions  in 
the  central  and  peripheral  nervous  system. 


ENZOOTIC  ENCEPHALO-MYELITIS  31 

Anterior  Chamber  of  Eye. 

Moussu  (1926)  and  also  Zwick  (1926)  have  shown  that  it  is  possible 
to  infect  the  rabbit  by  inoculation  of  a  virulent  emulsion  of  the  brain 
of  the  horse  into  the  anterior  chamber  of  the  eye,  and  we  have  con- 
firmed the  possibility  of  infection  by  this  route. 

Rabbit  206  received  several  drops  of  the  supernatant  fluid  from  a 
virulent  emulsion  of  brain  into  the  anterior  chamber.  The  point  of 
inoculation  was  carefully  seared.  During  the  period  immediately  fol- 
lowing the  inoculation  a  coagulum  of  fibrin  could  be  seen  in  the  eye, 
but  this  was  absorbed  later.  The  animal  died  23  days  after  the  inocu- 
lation, and  lesions  characteristic  of  Borna  disease  were  foimd  in  the 
central  nervous  system;  these,  however,  were  not  very  acute. 

The  control  rabbit  inoculated  intracerebrally  with  the  same  emul- 
sion died  in  the  average  time  with  well-marked  lesions  in  the  central 
nervous  system. 

Rabbits  86s  and  89s  inoculated  in  the  anterior  chamber  with  an 
emulsion  containing  virus  fell  ill  29  and  36  days  respectively  after 
infection,  and  died  on  the  34th  and  51st  day  with  typical  symptoms 
in  the  central  nervous  system.  The  control  rabbit  (intracerebral 
route)  died  37  days  after  inoculation. 

A  fourth  rabbit  inoculated  intraocularly  with  the  virus  survived 
without  having  shown  any  symptoms. 

Thus,  of  four  rabbits  inoculated  in  the  anterior  chamber  three 
became  infected  and  died  of  the  disease,  while  the  fourth  showed  no 
morbid  symptoms  and  survived.  These  results  are  in  accordance  with 
those  of  Zwick  who  had  five  positive  results  in  six  attempts  to  infect 
rabbits  by  the  intraocular  route. 

No  macroscopic  modification  of  the  cornea  followed  the  introduc- 
tion of  the  virus,  but  microscopically  there  was  slight  infiltration  with 
lymphocytes  between  the  corneal  laminae.  In  one  case  an  interstitial 
infiltration  with  mononuclear  cells  of  the  optic  nerve  was  found. 

Corneal  Scarification. 

Zwick  (1926)  has  stated  thatmfection  by  corneal  scarification  causes 
the  disease  only  rarely.  We  have  inoculated  four  rabbits  by  scarifica- 
tion of  the  cornea;  none  became  infected,  nor  was  there  any  visible 
keratitis.  However,  one  of  the  rabbits  (Rabbit  81s)  which  was  sub- 
sequently reinoculated  intracerebrally  with  a  virulent  emulsion  of 
brain  110  days  later  remained  well,  whereas  a  control  rabbit  which 
had  received  a  cerebral  inoculation  with  the  same  emulsion  died 
48  days  later  after  showing  typical  symptoms  of  the  disease,  and  with 
the  characteristic  lesions.  The  corneal  inoculation  may  have  rendered 
the  rabbit  refractory  to  infection  by  the  intracerebral  route. 

Conjunctival  Sac. 

Instillation  of  a  virulent  emulsion  into  the  conjunctival  sac  of  the 
eye  produced  no  effect. 

Nasal  Mucosa. 

The  nasal  mucosa  appears  to  be  a  possible  portal  of  entry  of  the 
virus,  and  Joest  (1927)  has  suggested  that  natural  contagion  in  the 


32  BOENA  DISEASE  AND 

horse  is  effected  by  this  route.  The  results  of  attempts  by  Zwick 
(1926),  Beck  and  Frohbose  (1926)  to  infect  rabbits  by  this  route  have, 
however,  been  inconstant. 

Scarified  Skin. 

Zwick  apphed  a  virulent  emulsion  of  brain  to  the  scarified  skin 
without  result.  Our  results  ^  confirm  those  of  Zwick.  As,  however, 
the  experiments  of  Flexner  and  Amos  (1917)  with  pohomyeHtis,  and 
of  Levaditi  and  Nicolau  (1922,  1923)  with  herpes  and  neurovaccinia 
show  that  previous  injection  of  substances  hke  physiological  saline, 
bouillon,  or  normal  serum  into  the  brain  may  increase  susceptibiUty, 
we  introduced  physiological  saline  either  into  the  brain  or  intra- 
thecally  in  rabbits  which  had  received  an  apphcation  of  the  virus  on 
to  the  depilated,  shaved,  and  scarified  skin.  The  animals  prepared  by 
the  inoculation  of  sahne  intrathecaUy  did  not  show  any  symptoms  of 
the  disease,  while  those  which  had  been  subjected  to  an  irritation  of 
the  brain  \vith  saline  subsequently  contracted  the  disease,  and  died. 
Typical  lesions  were  found  in  the  central  nervous  system,  and  passage 
of  the  brain  to  new  rabbits  by  the  mtracerebral  route  gave  positive 
results.  This  experiment  shows  that  when  the  nervous  system  is  in 
a  state  of  special  receptivity  due  to  the  diminution  of  the  normal 
power  of  defence,  infection  can  take  place  from  the  skin.  The  virus 
probably  reached  the  brain  by  way  of  the  intercostal  nerves,  thus 
being  protected  from  the  action  of  leucocytes  circulating  in  the  blood. 
This  interpretation  is  supported  by  the  following  experiments.  Four 
rabbits  were  inoculated  with  1-5  c.cms.  of  a  centrifugalized  emulsion 
of  virulent  brain  into  the  marginal  vein  of  the  ear.  Two  of  the  rabbits 
inoculated  intravenously  received  simultaneously  0-3  c.cm.  of  physio- 
logical saline  into  the  brain,  but  neither  of  these  animals  contracted 
the  disease,  nor  did  the  two  which  received  only  the  intravenous 
inoculation.  Two  months  later  the  immunity  of  these  four  rabbits 
was  tested  by  intracerebral  inoculation,  and  all  proved  susceptible  to 
infection.  The  virulence  of  tiie  enmlsion  used  to  infect  the  four  rabbits 
intravenously  was  proved  by  the  intracerebral  inoculation  of  two 
controls.  So  that  it  would  appear  that  the  virus  was  destroyed  in  the 
blood-stream,  or  in  the  tissues  before  it  reached  the  brain. 

Our  further  attempts  to  infect  by  the  intravenous  route  gave  us 
negative  results,  but  Zwick  (1926)  succeeded  exceptionally  when  4 
intravenous  injections  were  given  at  intervals.  Positive  results  have 
also  been  obtained  by  Ernst  and  Hahn  (1927).  Usually,  however, 
they  found  that  repeated  inoculations  by  the  intravenous  route 
instead  of  conferring  the  malady  produced  solid  resistance.  This  will 
be  discussed  in  the  chapter  dealing  with  immunitj'. 

Subcutaneous. 

Subcutaneous  inoculation  may,  exceptionally,  lead  to  a  fatal  en- 
cephalo-myelitis.  Zwick  and  his  collaborators  (1926),  who  employed 
repeated  injections  of  the  virus,  produced  the  disease  with  a  greater 

'  At  no  time  did  the  skin  Dhow  any  macroscopic  changes  which  cuuld  be  attributed 
to  the  viruB. 


ENZOOTIC  EXCEPHALO-MYELITIS  33 

frequency.  They  obtained  similar  results  by  inoculation  of  the  virus 
intraperitonealhj;  intramuscular  injections  with  tlie  virus  did  not 
confer  the  disease  m  their  experiments. 

Intratestwular. 

According  to  the  latter  investigatoi-s  the  introduction  of  virus  by 
the  iuti-atesticular  route  did  not  produce  infection  in  rabbits,  but  in 
our  experiments  it  has  done  so. 

Babbit  209,  weighing  1,850  gms.,  was  inoculated  mider  anaesthesia 
into  both  testicles  \\ith  a  virulent  emulsion. of  the  brains  of  four 
rabbits  dead  of  experimental  Borna.  The  dilution  of  the  emulsion  of 
brain  substance  in  physiological  saline  was  1  :  20.  During  a  period  of 
43  daj's  the  animal  showed  no  morbid  symptoms  and  its  weight 
increased  to  2, 540  gms.;  then,  without  other  symptoms,  wasting 
commenced;  19  days  later  the  weight  had  fallen  to  1,860  gms.  (680 
gms.  loss),  and  inco-ordination  with  shght  paresis  of  the  hind  quarters 
was  noticed.  Paresis  became  accentuated  and  other  symptoms  charac- 
teristic of  the  disease  became  manifest.  On  the  71st  day  after  the 
inoculation.  20  days  after  the  first  loss  of  weight  was  recorded  and 
8  days  after  the  first  chnical  symptom,  the  animal  died,  weighing 
only  1.4S0  gms.  The  lesions  found  in  the  central  nerv^ous  system 
were  characteristic  and  were  especially  intense  in  the  lumbar  region. 
The  topogi'aphy  of  the  lesions  in  the  cord  indicated  that  the  virus 
had  spread  from  the  point  of  inoculation  to  the  bram  through  the  cord 
centripetally.  The  intranuclear  inclusions  of  Joest-Degeu  were  found. 

The  passage  of  the  brain  and  cord  of  Rabbit  209  to  fresh  animals 
gave  positive  results,  indicating  the  presence  of  virus  in  both.  The 
control  rabbit  inoculated  intracerebrally  with  the  emulsion  of  brain 
which  served  to  infect  Rabbit  209  died  32  days  after  inoculation. 

Rabbit  289.  weighing  1,720  gms.,  was  inoculated  into  the  right 
testicle  with  a  \'irulent  emulsion  of  brain.  During  60  days  it  put  on 
weight,  reaching  2.320  gms.  Tliis  weight  was  maintained  for  14  days, 
when  wasting  began.  On  the  90th  day  after  inoculation  paresis  of  the 
hind  quarters  was  observed.  The  animal  died  105  days  after  inocula- 
tion. 

Rabbit  273  inoculated  into  the  right  testicle  at  the  same  time  showed 
no  symptoms  and  sm^vived,  while  the  control  rabbit  inoculated  intra- 
cerebrally died  after  37  days  with  the  typical  symptoms  and  lesions 
characteristic  of  the  disease. 

Intratracheal. 

We  have  made  two  unsuccessful  attempts  to  infect  rabbits  by 
intratracheal  inoculation.  In  the  first  attempt  two  rabbits  received 
each  0-3  c.cm.  of  a  thick  emulsion  of  brain  containing  virus  into  the 
trachea,  which  had  been  exposed  by  incision;  both  these  rabbits  sm-- 
vived  without  having  shown  any  symptoms,  while  the  control  which 
had  received  an  intracerebral  inoculation  with  the  same  virus  suc- 
cumbed to  the  infection.  In  the  second  experiment  four  j'oung  rabbits, 
between  670  and  860  gms.,  were  inoculated.  Each  received  0-5  c.cm. 
of  a  thick  emulsion  of  the  brain  of  a  rabbit,  dead  of  Borna  disease, 

6409  „ 


34  BOENA  DISEASE  AND 

diluted  1  :  5.  Kept  under  observation  more  than  six  months  they 
maintained  their  normal  state  of  health,  more  than  doubling  their 
weight.  The  controls  of  this  experiment  inoculated  intracerebrally 
died  of  a  typical  encephalo-myehtis  on  the  4'2nd  and  iith  day  after 
injection. 

Per  Os. 

Attempts  at  infecting  rabbits  per  os  are  of  special  interest  for  the 
interpretation  of  natural  infection  in  horses,  cattle,  and  sheep.  Zwick 
and  his  collaborators  (1926)  succeeded  in  infecting  rabbits  by  mixing 
virulent  brain  with  the  food.  He  refers  to  this  as  infection  by  the 
intestinal  route,  but  as  the  virus  was  administered  by  the  mouth  with 
the  food,  the  pre-existence  of  small  traumatic  lesions  in  the  mouth 
might  permit  the  implantation  of  the  virus.  As  infection  by  the  nasal 
mucosa  has  been  shown  to  be  possil)le,  one  cannot  exclude  the  possi- 
biUty  that  infection  took  place  by  the  buccal  mucous  membrane, 
especially  when  one  considers  the  existence  of  nervous  tissue  imme- 
diately below  the  mucous  membrane  covering  the  tongue  (Manouehan 
and  Viala,  1926).  Supposing  the  virus  to  have  been  implanted  in  such 
nervous  tissue,  it  is  quite  easy  to  conceive  how  it  might  ultimately 
reach  the  brain. 

Attempts  to  infect  rabbits  by  cohabitation  have  been  unsuccessful. 

5.  AUTHORS  EXPERDIENTS  ON  THE  TRANSMISSION  OP 
THE  DISEASE  TO  MONKEYS  {MACACUS  RHESUS), 
AND  SYMPTOMS  OCCURRING  IN  THESE  ANIMALS 

Monkey  >/.  1.  A  fine  specimen  of  Macacus  rhesus  weighing  3,800 
gms.  was  kept  under  observation  during  16  days  prior  to  inoculation. 
The  animal's  temperature  varied  very  little. 

On  15.3.27  it  was  inoculated  intracerebrally  under  anaesthesia 
with  1-5  c.cms.  of  a  vindent  emulsion  of  the  brain  of  a  rabbit  diluted 
1  :  5.  Two  control  rabbits  were  inoculated  intracerebrally  at  the  same 
time  as  the  monkey;  they  developed  typical  symptoms  on  the  28th 
and  31st  day,  and  died  of  Borna  disease  on  the  42nd  and  44th  day. 

Between  15.S.27  and  11.5.27,  a  period  of  57  days,  the  monkey 
showed  no  symptoms  and  the  tenij)erature  remained  normal. 

15.3.27.  Weight  3,800  gms.  Temperature  38-9'  C.  Received  inoculation  w-ith  the 
brain  emulsion  from  Rabbit  275. 

11.5.27.  Fifty-seven  days  after  inoculation  the  monkey  appeared  depressed.  Tem- 
perature 38-4°  C   Slight  diarrhoea. 

12.5.27.   Same  condition. 

14.5.27.  Temperature  38-5  C.  No  diarrhoea.  L<'»«  Uvely  than  usual,  appearetl 
to  prefer  to  remain  with  the  back  to  the  light  (photophobia  ?). 

16.5.27.    Condition  unchanged. 

16.5.27.  Photophobia  well  marked.  The  monkey  hid  its  head  under  the  straw 
of  the  cage  and  would  not  move  when  disturbed.  It  allowed  itself  to 
Im?  caught  easily  and  dcfende<l  itself  when  npproache*!  almost  exclusively 
with  the  left  hand  and  foot.  Slight  [Hire.sis  of  the  right  arm  was  detc^-tcii. 
The  pupils  were  e<iual  anil  reacte<l  normally  to  light. 

18.5.27.  Weight  3,280  gms.  Temi)erature  .■J8-2'' ('.  Animal  fe<-dlng  normally.  It 
remainol  in  a  comer  of  the  cage  with  the  head  elroppe-d  like  that  of  u 


ENZOOTIC  ENCEPHALO-MYELITIS  35 

rabbit  ill  from  Borna  disease.  It  allowed  itself  to  be  caught  easily,  offer- 
ing but  little  resistance  and  with  the  left  arm  only.  It  was  found  possible 
to  introduce  the  thermometer  into  the  rectum  without  holding  the  legs, 
which  fell  practically  inert.  When  the  monkey  was  put  on  the  ground  it 
moved  much  more  slowly  than  normally,  dragging  the  right  leg,  which 
showed  paresis.  Paresis  was  less  evident  in  the  left  leg.  The  animal  could 
grip  the  cage  with  the  left  hand,  the  only  limb  which  preserved  its  normal 
function.  The  right  leg  and  arm  did  not  grip  or  gripped  only  badly.  The 
animal  attempted  to  climb,  but  fell  exhausted  by  the  effort.  It  was 
roused  with  difficulty.  There  appeared  to  be  no  trouble  in  preserving 
equilibrium.  The  animal  appeared  to  be  able  to  see.  The  diarrhoea  was 
replaced  by  constipation.  *' 

19.5.27.  Temperature  38-5°  C.  Same  symptoms  as  on  the  preceding  day.  On 
opening  the  cage  the  monkey  did  not  move  or  try  to  get  out.  When 
taken  out  and  left  to  run  it  fell  on  the  right  side,  of  which  the  paralysis 
was  more  accentuated.  When  the  monkey  was  held  by  the  skin  of  the 
neck,  the  legs,  which  showed  a  marked  flaccidity,  fell  inert  without  any 
resistance  (Fig.  2).  If  a  finger  was  presented,  the  monkey  gripped  it  with 
the  left  hand  only,  the  right  hand  showed  paralysis  of  the  flexors  of  the 
digits. 

20.5.27.   Temperature  38-5°  C.  Same  condition. 

21.5.27.  Temperature  38-1"  C.  Weight  3,280  gms.  Animal  still  feeding.  When 
taken  out  of  the  cage  it  was  found  to  tire  quickly,  and  after  a  feeble 
effort  at  escaping  it  remained  on  the  ground  immobile  for  several 
minutes. 

23.5.27.  Temperature  380°  C.  Motor  disturbances  were  accentuated.  Monkey 
remained  hunched  up  in  a  corner  of  the  cage  (Fig.  3). 

24.5.27.   Temperature  37-5°  C.   Same  condition. 

25.5.27.  Temperature  37-3°  C.  Complete  paralysis  of  the  legs.  When  making 
movements,  it  supported  itself  especially  with  the  left  arm  and  dragged 
the  paralysed  legs.  For  the  most  part  it  preferred  to  remain  hunched  up 
in  the  corner  of  the  cage. 

26.5.27.    Found  procumbent.   Weight  3,180  gms. 

27.5.27.  Temperature  36-5°  C.  Animal  procumbent,  the  respirations  were  irregu- 
lar and  infrequent.  Incontinence  of  urine,  no  grinding  of  the  teeth,  no 
ocular  symptoms,  no  hj'persahvation,  both  legs  paralysed.  As  the  animal 
was  dving  it  was  killed  by  means  of  chloroform  at  6  o'clock  in  the  evening 
73  days  after  the  inoculation,  16  days  after  the  first  symptoms  were 
observed. 

Autopsy  of  Monkey  1. 

The  dura  mater,  pia  mater,  and  the  brain  substance  appeared  normal. 

The  cord  was  slightly  hyperaeraic,  but  no  haemorrhagic  areas  were 

found. 
The  buccal  epithelium  and  tongue  showed  desquamation  of  the  epi- 
thelium and  redness. 
The  parotid  gland.   Normal  in  aspect. 
Lung.    Normal. 

Spleeji,  liver,  and  gall  bladder.    Normal. 

Kidney.   Congested  in  both  the  cortical  and  medullary  zones. 
Adrenal.    Normal. 

Peritoneal  cavity,  bladder,  and  intestines.    Normal. 
Cultures  made  from  the  brain,  spleen,  and  blood  proved  to  be  bac- 

teriologically  sterile. 
XoU.  A  study  of  the  blood,  made  during  the  course  of  the  infection,  indicated  no 
decided  changes;  the  leucocytic  formula  remained  normal,  except  for  a  slight 
increase  in  the  number  of  polymorphonuclears  in  the  last  stages  of  the  disease. 
c2 


36  BOKNA  DISEASE  AND 

Passages  into  rabbits  made  with  emulsions  from  various  parts  of 
the  central  nervous  system  of  this  monkey  gave  positive  results 
(see  p.  48). 

An  emulsion  of  the  brain  of  M.  1  was  likewise  inoculated  into  two 
monkeys  of  the  same  species  by  the  intracerebral  route  (Monkev  M.  2 
and  M."  B)  on  the  28.5.27. 

Monkey  M.  3  [Macacus  rhesus).  Weighed  3,150  gms. 

During  a  period  of  71  daj-s  after  the  inoculation,  the  animal 
remained  free  from  all  morbid  symptoms,  and  the  temperature 
remained  normal.  On  the  72nd  day  the  monkey  was  found  procum- 
bent, although  the  day  before  there  was  no  obvious  illness.  It  was 
paralysed  in  the  legs  and  arms,  and  it  could  not  assume  an  upright 
position.  SUght  ptosis  of  the  right  eyeUd  was  observed.  The  tem- 
perature was  subnormal.  The  follow"ing  day  tiie  respirations  were 
irregular,  gasping,  and  moist  rales  were  heard.  .\s  the  animal  was 
in  convulsions  it  was  killed.  The  findings  on  autopsy  were  the  same 
as  for  Monkey  M.  1.  Rabbits  inoculated  with  emulsions  of  various 
parts  of  the  nervous  sytem  died  of  Boma  disease  witli  characteristic 
lesions. 

Monkey  M.  2  (Macacus  rhesus).  The  evolution  of  the  disease  in  this 
monkey  was  entirely  different. 

28.5.27.  Animal  inoculated.  Weight  3,200  gms.  Temperature  38-5°  C.  During 
a  period  of  33  days  after  the  inoculation  the  blood,  the  body-weight,  and 
the  temjxrature  curve  showed  no  marked  changes,  and  there  was  no  evi- 
dence of  any  s_\-mptoms. 

30.ti.27.  Temperature  38-2' C.  Weight  3.100  gms.  When  a  stick  was  given  to  the 
animal  it  could  not  seize  it  with  the  right  hand,  and  if  irritated  it 
defended  itself  with  the  left  hand.  The  monkej-  was  able  to  run  and  climb 
normally. 

1.7.27  and  2.7.27.   No  change  in  condition. 

3.7.27.    In  addition  to  the  paresis  of  the  right  arm  the  animal  showtnl  lassitude 
and  appeared  less  agile. 
From  the  3.7.27  to  the  1 1.7.27.  the  condition  remained  unchanged. 

14.7.27.  Forty-seven  days  after  the  inoculation.  Weight  2.790  gms.  When  taken 
out  of  the  cage  the  animal  ran  ami  dimlH.'d  with  difticulty.  The  paresis 
of  the  right  arm  was  amcliorate<l.  but  there  was  paresis  of  the  legs,  more 
accentuated  on  the  left  side.   The  left  arm  also  showetl  slight  paresis. 

18.7.27.  The  left  eye  was  closed  completely,  due  to  ptosis  of  the  upper  eyelid 
(Fig.  4).  The  face  was  drawn  to  the  right  side.  The  paresis  of  the  arm 
and  leg  on  the  left  side  was  now  easily  discernible.  If  the  animal  when 
seated  was  gently  pushed  it  fell  on  to  the  li-ft  side.  It  uttcre<l  a  plain- 
tive cry  from  time  to  time.  No  lesions  could  be  seen  on  the  cornea  or 
conjunctiva. 

20.7.27.  Marke<l  excitation  was  observed.  The  animal  appeared  to  have  hallu- 
cinations. It  threw  itself  at  imaginary  objects,  attempted  to  bite  fre- 
({uently,  and  struggled  and  fell  as  if  in  a,  fit.  There  was  nearly  complete 
paralysis  of  the  left  side  and  pto.sis  of  the  left  eyelid. 

22.7.27.  The  symptoms  appeare*!  to  be  ameliorated.  The  animal  was  much 
calmer  and  the  eye  could  be  o|)ene<l  |>artially. 

25.7.27.  Eye  nearly  completely  open.  Animal  still  aggmisive,  uttered  cries  from 
time  to  time,  paralysis  less  noticeable. 


ENZOOTIC  ENCEPHALO-MYELITIS  37 

28.7.27.  Eye  appeared  normal,  and  the  condition  of  the  monkey  practically 
normal. 
1.8.27.  Ptosis  of  the  right  eyeUd.  Slight  paresis  of  the  hind  quarters,  which, 
however,  was  not  sufficient  to  prevent  the  monkey  from  climbing.  It 
became  fatigued  easily,  however,  and  remained  in  a  corner  of  the  cage 
crying  out  from  time  to  time. 
3.8.27.    Paralysis  of  the  right  side  of  the  face  (see  Fig.  5)  was  observed.    Animal 

could  .still  run  and  climb. 
5.8.27.   Same  condition.   Weight  3,170  gms. 
8.8.27.    Paralysis  of  face  diminished.   Animal  irascible. 

12.8.27.  Spasmodic  contractions  of  muscular  groups  of  the  back  and  shoulders 
were  the  only  signs  of  the  animal  being  other  than  normal. 

13.8.27.  Left  eye  deviated  to  the  internal  canthus.  Tendency  to  remain 
hunched  up. 

14.8.27.  Animal  decidedly  ill.  Head  hanging  over  on  to  right  shoulder :  internal 
strabismus  still  present  and  left  pupil  dilated.  Right  eye  mobile.  Pupil 
of  this  eye  reacted  to  light.  Nystagmus  present.  The  animal  had  periods 
of  excitement. 

15.8.27.  Monkey  uttering  cries  with  a  low  feeble  raucous  voice.  Mouth  opened 
after  shomng  dragging  to  the  right  side.  Internal  strabismus  on  the  left 
side,  pupil  on  this  side  also  dilated.  Nystagmus  exacerbated.  Paralysis 
of  muscles  of  left  shoulder,  head  falling  over  on  to  the  right  shoulder. 

17.8.27  and  21.8.27.    Condition  unchanged. 

27.8.27.  Until  27.8.27  animal  appeared  normal.  On  this  date  the  animal  had 
complete  aphonia  (probably  paralysis  of  the  recurrent  nerve).  The  head 
hung  over  the  right  side  and  the  monkey  showed  signs  of  cerebral 
disorder.  Strabismus  was  less,  face  was  drawn  to  the  right  side,  and 
there  was  frequent  spasmodic  contraction  of  the  facial  muscles  ('tic). 
The  pupils  were  unequal  in  size.  The  animal  carried  out  movements 
of  mastication  without  the  teeth  coming  in  apposition,  for  this  reason 
it  fed  with  difficulty.  Deglutition  was  not  carried  out  easily,  and  the 
animal  appeared  to  have  difficulty  in  orientation.  Taken  out  of  the  cage 
it  could  not  co-ordinate  its  movements  in  the  direction  desired. 

29.8.  27.  Weight  2,900  gms.  Animal  still  made  movements  of  mastication  con- 
tinually, and  twitching  of  the  muscles  of  the  mouth  was  present.  There 
was  internal  strabismus,  as  weU  as  aphonia  and  weakness  of  the  muscles 
of  the  neck  on  the  left  side.  The  monkey  remained  hunched  up,  or  had 
periods  of  excitement  which  were  increased  by  noises,  movements,  &c. 

31.8.27.   Condition  unchanged. 
2.9.27.   Weight  2,820  gms.  Spastic  contractions  of  divers  muscular  groups  were 
produced  by  noises.    The  tongue  was  drawn  to  the  right  side.    The 
animal  fed  no  more  owing  to  the  impossibility  of  swallowing.    'Tic'  of 
the  mouth  persisted,  also  the  strabismus  and  the  inequahty  of  the  pupils. 
4.9.27.   The  left  eye  appeared  practically  normal  and  the  pupils  equal  in  size. 
The  animal,  however,  still  trembled  at  times   and  bit   at   imaginary 
objects;  frequent  'tic'  was  observed  with  aphonia.    The  animal,  how- 
ever, fed  well. 
9.9.27.   The  condition  of  the  animal  had  not  changed. 

16.9.27.  The  cry  appeared  more  normal,  ocular  disturbances  were  absent, 
although  'tic'  and  champing  of  the  jaws  was  still  observed.  The  head 
was  hung  over  the  right  shoulder,  and  the  animal  showed  increased 
salivation. 

20.9.27.  The  condition  of  the  animal  was  unchanged;  to  a  certain  degree  the 
syndrome  in  this  monkey  might  be  compared  to  that  exhibited  by  a  man 
affected  with  post-encephaUtic  Parkinsonism. 

22.9.27.    Same  condition.   Weight  3,000  gms. 

5.10.27.  Up  to  this  date  the  monkey  remained  in  a  similar  state,  this  being  the 
130th  day  since  the  inoculation.  At  this  time  the  animal  was  inoculated 
with  the  virus  of  polio-myelitis.  The  result  is  given  in  the  chapter 
dealing  with  immunitv. 


38  BOENA   DISEASE  AND 

These  three  monkeys  showed  three  different  forms  of  the  disease. 
In  all  three  cases  the  incubation  period  was  very  long,  but  the  dura- 
tion of  the  morbid  symptoms  varied.  Monkey  3  had  an  acute  attack 
which  lasted  only  2  days.  In  Monkey  1  the  evolution  was  slower,  the 
time  between  the  onset  of  symptoms  and  death  being  16  daj's. 
Monkey  2  had  recurrent  attacks  during  a  period  of  97  days  (see  sub- 
sequent history,  p.  80). 

The  virus  after  passage  through  monkeys  liad  not  lost  its  \irulence 
for  the  rabbit  or  guinea-pig. 

In  the  chapter  dealing  with  histopathology  the  lesions  in  the 
nervous  sytem  will  be  described  in  full.  We  may,  however,  anticipate 
here  by  the  statement  that  we  found  lesions  of  an  intense  character, 
in  the  brain,  and  of  a  more  discrete  character  in  the  cord.  Lesions 
were  also  found  in  the  spinal  ganglia,  posterior  nerve-roots,  and  peri- 
pheral nerves.  Both  clinically  and  pathologically  the  disease  may 
be  described  as  an  encephalo-myelitis  complicated  with  a  ganglio- 
radiculitis  and  peripheral  neuritis.  The  virus  when  introduced  into 
the  brain  evidently  spreads  not  only  to  the  cord,  but  also  to  the 
peripheral  nerves,  for  not  only  have  we  found  lesions  in  these,  but 
we  have  also  been  able  to  demonstrate  the  presence  of  virus. 

Discussion. 

The  pathogenicity  of  the  virus  for  the  monkey,  and  the  clinical 
features  presented  in  this  animal,  raise  the  question  of  the  relation 
between  enzootic  encephalo-myelitis  of  domestic  animals  and  polio 
myelitis  in  man.  There  are  marked  resemblances  in  the  clinical 
aspects  as  well  as  in  the  alterations  in  the  cord  and  spinal  ganglia  in 
the  two  diseases.  The  former  virus  is,  however,  pathogenic  for  the 
rabbit,  while  the  latter  is  generally  considered  not  to  be  so.  The 
incubation  period  of  experimental  encephalo-myeUtis  in  the  monkey 
is  longer  than  in  the  disease  produced  by  the  virus  of  poho-myelitis. 

Another  question  is  whether  the  virus  of  encephalo-myelitis  is 
pathogenic  for  man,  and  if  such  is  the  case,  whether  some  luunan 
disease  of  the  nervous  system  at  present  of  unknown  origin  may 
possibly  be  due  to  it.  It  is  a  common  observation  in  clinical  medicine 
that  exposure  to  cold  may  determine  an  attack  of  facial  neuritis  or 
sciatica.  Conceivably,  such  attacks  might  be  the  expression  of  an 
unrecognized  infection  by  some  virus,  akin  to  enzootic  encephalo- 
myelitis, which  in  the  lirst  place  caused  only  shght.  if  any,  general 
symptoms  of  disease  and  then  proceeded  down  into  the  peripheral 
nerves,  where  it  remained  latent  until  the  added  factor  of  cold  deter- 
mined the  local  incidence  of  paralysis  or  pain.  Similar  views  may  be 
argued  with  regard  to  herpes  zoster,  recurrent  herpes,  or  the  so-called 
peripheral  forms  of  epidemic  encephalitis.  An  analogous  action  of 
cold  was  recorded  liy  Pasteur,  when  certain  rabbits  that  were  resistant 
to  the  inoculation  of  an  atteiuiated  ral)ies  virus,  at  once  showed 
paralytic  symptoms  after  exposure  to  severe  cold. 

The  curious  epidemic  at  Lille,  studied   i)y   David  and   Dekester 


ENZOOTIC  ENCEPHALO-MYELITIS  39 

(1926),  has  suggested  that  a  certain  form  of  sciatica,  at  least,  is  an 
infectious  disease;  and  there  are  many  human  cases  on  record  of 
myelitis  associated  with  peripheral  neuritis,  apparently  of  a  contagious 
type,  in  some  of  which,  as  in  the  instance  of  the  child  described  by 
Pehu  and  Dechaume  (1927),  there  were  in  the  peripheral  nerves  in- 
flammatory lesions  closely  resembling  those  described  by  us  in 
monkeys  and  rabbits  infected  with  Borna  disease. 

We  give  below  a  resume  ot  Pehu  and  Dechaume"s  case  in  some  detail 
because  the  symptoms  resemble  in  certain  respects  those  which  our 
Monkey  M.  2  showed  after  inoculation  with  the  virus  of  Borna  disease. 
The  child,  20  months  old,  was  in  perfect  health  up  to  the  day  when 
it  showed  some  lassitude  three  months  before  going  to  hospital.  The 
temperature  did  not  exceed  36-5°  C.  Two  months  later  it  could  no 
longer  walk  and  had  lost  power  in  the  arms.  At  the  time  of  entering 
the  hospital  it  showed  flaccidity  and  paralysis  in  the  lower  extremities 
without  Babinski's  sign.  There  was  also  slight  paresis  of  one  arm. 
The  following  day  the  child  collapsed  although  not  losing  conscious- 
ness. There  were  no  cerebral  symptoms,  nor  vomiting,  neither  did 
somnolence  exist,  but  the  pulse  was  rapid  and  irregular.  The  condi- 
tion lasted  11  days  and  the  child  died  suddenly  without  convulsions. 
The  case  was  diagnosed  as  a  peripheral  form  of  epidemic  encephalitis, 
referred  to  as  pseudo-myelitic. 

On  autopsy  there  were  no  macroscopic  changes.  On  microscopic 
examination  of  sections,  discrete  lesions,  for  the  most  part  exudative, 
were  found  in  the  cord,  especially  in  the  lumbar  region.  Perivascular 
infiltrations  occurred  in  the  brain.  No  neuronophagia  was  recorded, 
and  the  spinal  ganglia  were  not  examined.  The  lesions  found  in  sec- 
tions of  the  median  nerves,  sciatic  nerve,  and  posterior  tibial  nerves 
were  comparable  to  those  described  by  us  in  the  sciatic  and  brachial 
nerves  of  rabbits  and  monkeys  inoculated  intracerebrally  with  the 
virus  of  Borna  disease  (see  pages  61  and  67). 

Pehu  and  Dechaume  suggested  that  the  presence  of  lesions  in  the 
peripheral  nerves  might  be  coexistent  with  the  presence  of  virus.  In 
the  case  of  Borna  disease  we  have  proved  that  the  presence  of  virus 
is  coexistent  with  the  existence  of  lesions  in  the  peripheral  nerves 
(see  page  45). 

6.  PATHOGENICITY  OF  THE  VIRUS  OF  ENZOOTIC  EN- 
CEPHALO-MYELITIS  FOR  THE  GUINEA-PIG,  RAT, 
MOUSE,  AND  FOWL 

I.   Guinea-pig. 

The  introduction  of  virus  intracerebrally  into  guinea-pigs  may  pro- 
duce the  disease.  The  incubation  period  varies  in  individual  cases  and 
death  is  inconstant.  Zwick  and  his  collaborators  (1926)  were  the  first 
to  transmit  the  disease  to  guinea-pigs,  and  to  make  passages  in  series 
from  brain  to  brain.  Some  guinea-pigs  proved  to  be  resistant  to  infec- 
tion.   According  to  their  experiments  death  followed  infection  in 


40  BORNA  DISEASE  AND 

from  3  weeks  to  13  months.  They  also  succeeded  in  infecting  rabbits 

with  the  virus  passaged  through  guinea-pigs. 

In  Table  III  we  give  the  period  of  incubation  and  the  duration  of 
the  disease  in  a  batch  of  guinea-pigs  which  in  our  experiments  proved 
to  be  susceptible  to  the  virus  inoculated  intracranially. 

Table  III. 


No.  of  guinea- 

Weight  in 

Commencement 

Lesions  in 

pig. 

gms. 

of  disease. 

Death. 

brain. 

85E 

470 

73rd  day 

83rd  day 

Intense. 

84k 

400 

107th    „ 

132nd    ,. 

.. 

94e 

500 

23rd    .. 

114th     .. 

95k 

500 

58th     ., 

65th     ,. 

Discrete. 

93k 

480 

40th     ., 

.58th     .. 

.\verai^. 

98k 

450 

57th     .. 

64th     ,. 

99k 

580 

18th     „ 

19th     „ 

Intense. 

1()0k 

560 

58th     „ 

182nd    „ 

We  have  also  made  experiments  to  determine  the  relative  suscepti- 
biUty  of  guinea-pigs  to  infection.  Forty-five  guinea-pigs  of  about  the 
same  size  (400-GOO  gms.)  were  divided  into  tliree  lots  of  15. 

Lot  A  were  injected  with  an  emulsion  of  the  brain  of  a  rabbit  dead 
of  Boma  disease  diluted  1  :  10. 

Lot  B  were  injected  with  the  same  emulsion  diluted  1  :  100. 

Lot  C  were  inoculated  with  the  emulsion  diluted  1  :  1.000. 

The  results  were  as  follows : 

In  Lot  A  all  the  guinea-pigs  died  after  showing  typical  symptoms 
of  the  disease.  52,  58.  74.  133,  134,  139,  139,  141,  141,  143.  148,  150, 
153,  15(5.  and  170  days  after  the  inoculation.  Lesions  characteristic 
of  enzootic  encephalo-myelitis  were  found  in  sections  of  the  brain  and 
spinal  cord  of  these  animals;  moreover,  the  corpuscles  of  Joest- 
Degen  were  demonstrated. 

Tablk  IV. 


Lot  A. 


Dilution  of 

emulsion  of 

brain  inocu- 

A'o. of 

Animal 

lated. 

gutnea-pig. 

died. 

1:10 

lA 

2a 
3a 
4a 

62nd  day 
SSth     „ 
74th     „ 
ISSrd    .. 

Intense  lesioiu  in  rentral  nervnuii  Nyato 

n. 

5a 

134th     .. 

Very  intense  le«iiin  in  C'.N.S. 

Ha 

139th    .. 

Discretei  lesions  in  C'.N.S. 

7a 

139tb     „ 

Intense  lesions  in  C.N.S. 

8a 

14Ut     „ 

P,                                             M 

0A 

14l8t       ., 

Mild  lesions  in  C.N.S. 

10a 

143rd    „ 

Intense  lesions  in  C'.N.S. 

■ 

llA 

U8th     „ 

M                        ,. 

12a 

150th     „ 

13a 

163rd     .. 

Very  intense  Icoions  in  C.N.S. 

14a 

166th     ., 

Intense  li«ii>ns  in  I'.N.S. 

15a 

170th     „ 

ENZOOTIC  ENCEPHALO-MYELITIS 


41 


In  Lot  B  two  of  the  guinea-pigs  succumbed  to  an  intercurrent 
infection.  The  thirteen  others  died  58,  105,  130,  130,  135,  137,  138, 
140.  140,  140,  147,  149,  and  153  days  after  infection. 

Table  V. 
LotB. 


Dilution  of 

emulsion  of 

brain  inocu- 

So. of 

A  n  imal 

lated. 

gmnea-pig. 

died. 

Observations. 

1:100 

IB 

58th  day 

Intense  lesions  in  central  nervous  system. 

jj 

2b 

105th     „ 

Slight  lesions  in  C.N.S. 

,, 

3b 

130th     „ 

Very  intense  lesions  in  C.N.vS. 

„ 

4b 

1.30th     „ 

Intense  lesions  in  C.N.S. 

OB 

135th     „ 

».                    ,» 

„ 

6b 

137th     „ 

i»                    »» 

J, 

7b 

138th     „ 

„                    » 

„ 

8b 

140th     „ 

,,                            n 

,j 

9b 

140th     „ 

Slight  lesions  in  C.N.S. 

„ 

10b 

140th     „ 

Intense  lesions  in  C.N.S. 

„ 

11b 

147th     „ 

..                    »» 

J, 

12b 

149th     „ 

»                    „ 

„ 

13b 

153rd     „ 

.,                    ,. 

„ 

14b 

5th     „ 

Accidental  death. 

•• 

15b 

12th     „ 

„ 

In  Lot  C  five  of  the  animals  died  of  an  intercurrent  infection,  the 
ten  remaining  died  70,  90,  96,  101,  140,  141,  141.  147,  149,  and  150 
days  after  intracerebral  inoculation. 


Table  VI. 

Lot  C. 

Dilution  of 

emulsion  of 

brain  inocu- 

No. of 

Animal 

lated. 

guinea-pig. 

died. 

Observations. 

1  :  1,000 

Ic 

70th  day 

Discrete  lesions  in  central  nervous  system. 

2c 

90th    „ 

Intense  lesions  in  C.N.S. 

3c 

96th    „ 

,,                    », 

4c 

101st     „ 

„                    >> 

, 

i5c 

140th    „ 

,»                    ,, 

J 

6c 

141st     „ 

>•                    „ 

7c 

141st     „ 

,»                    .. 

8c 

147th    „ 

„                    ,. 

9c 

149th    „ 

Very  intense  lesions  in  C.N.S. 

^ 

10c 

150th    „ 

»,                »•                ,» 

lie 

5th    „ 

Accidental  death. 

12c 

8th    „ 

.♦               », 

13c 

8th    „ 

>,               „ 

, 

14c 

8th    „ 

..               •, 

15c 

nth    „ 

„ 

As  in  the  case  of  the  guinea-pigs  of  Lot  A,  the  symptoms  in  guinea- 
pigs  of  Lot  B  and  C  were  characteristic,  and  sections  made  from  the 


42 


BORNA  DISEASE  AND 


central  nervous  system  showed  the  typical  changes  produced  by  the 
virus  of  Boma  disease  in  other  animals. 

These  results,  while  demonstrating  the  variation  in  individual  sus 
ceptibility,  also  point  to  the  fact  that  resistance  to  infection  in  the 
guinea-pig  is  not  so  marked  as  it  appeared  to  be  from  the  results 
obtained  by  Zwick,  since  of  thirty-eight  gvunea-pigs  inoculated  in  our 
experiments  thirty-eight  succumbed  to  the  disease  (the  seven  guinea- 
pigs  dead  from  other  causes  are  not  mcluded). 

The  virus  passaged  through  guinea-pigs  still  preserved  its  patho- 
genicity for  the  rabbit. 

The  disease  in  the  guinea-pig  is  similar  to  that  of  the  rabbit.  After 
a  variable  period  the  animal  appears  depressed,  there  is  marked 
somnolence,  and  abstention  from  food.  Characteristic  nervous  symp- 
toms follow,  those  indicating  affection  of  the  cord  being  especially 
well  marked.  The  syndrome  is  as  described  in  the  rabbit.  The  bind 
legs  become  paralysed  (Fig.  6)  and  the  fore  legs  are  mvolved  later 
(Fig.  7).  The  loss  of  weight  is  less  marked  than  in  the  case  of  the 
rabbit. 

From  the  four  following  experiments  the  susceptibility  of  the 
guinea-pig  after  the  virus  is  inoculated  would  appear  to  be  diminished 
by  a  simultaneous  inoculation  of  the  same  material  intramus- 
cularly. 

An  emulsion  of  virulent  brain  originating  from  a  rabbit  dead  of 
experimental  Boma  disease  was  inoculated  into  the  brain  of  eight 
guinea-pigs,  and  at  the  same  time  1  c.cm.  of  the  same  emulsion  was 
inoculated  into  the  quadriceps  group  of  muscles  of  four  of  them. 
The  results  of  these  four  experiments  are  tabulated  below: 

Table  VII. 


No.  of 
guinea-pig. 

<.I5k 
97  b 

Ouinea-pig 
died. 

65th  day 

13l8t      „ 

Letions  in  the  brain. 

I.  Inoculated  into  the  brain 
Inoculated  into  the  brain 
and  muscle. 

Discrete. 
Intense. 

II.   Inoculated  into  the  brain 
Inoculated  into  the  brain 
and  muscle 

«3k 

91 B 

58th     „ 
116th     „ 

Of  average  intensity. 

III.    Inoculated  into  the  brain 
IniM'ulated  into  the  brain 
and  muscle 

99k 

9«B 

19th  day 
Survived 

Intense. 

IV'.   Inoculated  into  the  brain 
Inoculated  into  the  brain 

98k 
2UUu 

64th  dav 
184th    ., 

Of  average  intensity. 
Intense. 

iiiid  muscle 

This  observation  is  comparablt^  to  that  of  Ernst  and  Halm  (1927), 
who  found  that  when  rabbits  inoculated  intracercbrally  witli  viru- 
lent emulsion  received,  either  at  the  same  time  or  subsequently, 
injections  of  virus  into  the  veins,  they  did  not  develop  a  fatal 
cnophalitis. 


ENZOOTIC  ENCEPHALO-MYELITIS  43 

Attempts  at  infecting  the  guinea-pig  by  intradermal  inoculation  of 
an  emulsion  of  virulent  brain  into  the  metatarsal  pad  (following  the 
technique  used  by  Waldmann  and  Pape  (1921)  in  foot-and-mouth 
disease,  and  by  Gildemeister  and  Herzberg  (1925)  in  experimental 
herpes)  did  not  succeed.  The  guinea-pigs  inoculated  varied  in  weight 
from  100  gms.  to  750  gms.,  and  were  kept  under  observation  for  over 
seven  months,  but  no  symptoms  were  seen  at  any  time  during  this 
period. 

II.  Eat. 

Zwick,  Seifried,  and  Witte  (1926)  infected  rats  with  the  virus  of 
Borna  disease  by  intracerebral  inoculation.  Death  supervened  40,  53, 
and  62  days  respectively,  after  infection.  Some  rats  showed  no 
symptoms  and  survived.  The  virus  passaged  through  the  rat  had 
not  lost  its  pathogenicity  for  the  rabbit. 

In-  our  experiments  large  rats  appeared  to  be  more  susceptible  to 
the  disease  than  young  animals.  Four  rats  (three  old  and  one  yoimg) 
were  inoculated  intracerebrally  with  an  emulsion  of  the  brain  of  rat 
No.  I  which  died  67  days  after  infection.  (Typical  lesions  of  Borna 
disease  were  found  m  sections  of  the  brain  of  rat  No.  1.)  The  three 
large  rats  died  22,  37,  and  74  days  respectively  after  inoculation. 
They  all  developed  typical  symptoms,  and  sections  of  the  brain 
showed  the  presence  of  characteristic  lesions  microscopically.  The 
young  rats  kept  under  observation  for  six  and  a  half  months 
remained  perfectly  normal. 

The  control  rabbit  inoculated  with  the  same  emulsion  of  brain 
from  Eat  1  died  on  the  27th  day  of  a  typical  infection. 

Subsequently  four  rats  (two  large  and  two  small)  were  inoculated 
with  an  emulsion  of  one  of  the  brains  of  one  of  the  large  rats  men- 
tioned above  (that  dead  on  the  37th  day).  The  two  older  rats  died 
on  the  40th  and  82nd  day  after  infection,  while  the  two  younger 
animals  survived  124  days,  succumbing  later  to  an  intercurrent 
infection.  In  all,  we  have  inoculated  twenty-eight  rats;  of  these  only 
the  older  rats  contracted  the  disease. 

The  symptoms  in  the  rat  are  similar  to  those  in  the  guinea-pig. 
They  commence  with  motor  disturbances,  inco-ordination,  and  difB- 
culty  in  maintaining  equiUbrium.  Paralysis,  coma,  cachexia,  and 
death  follow  later. 

Up  to  the  time  of  writing  we  have  succeeded  in  making  at  least 
four  passages  in  this  species.  The  rats  in  the  series  died  67,  37,  82, 
and  47  days  respectively,  after  inoculation,  showing  that  the  course 
of  the  disease  in  the  rat  is  as  variable  as  in  the  guinea-pig.  It  would 
appear  that  virus  passaged  through  rats  when  inoculated  intra- 
cerebrally into  rabbits  produced  the  disease  after  a  shorter  in- 
cubation period  than  when  the  virus  was  passaged  in  series  through 
rabbits. 

In  sections  of  the  brain  of  our  experimental  rats  the  corpuscles  of 
Joest-Degen  were  found.  Zwick  (1926),  however,  failed  to  find  them 
in  the  brain  of  rats  inoculated  with  the  virus  of  Borna  disease. 


U  BORNA  DISEASE  AND 

III.  Mouse. 

We  have  been  able  to  infect  mice  by  the  intracerebral  route,  but 
this  species  of  rodent  is  apparently  less  susceptible  to  the  infection. 
As  in  the  rat,  age  appears  to  have  an  important  bearing  on  the  sus- 
ceptibihty  of  the  mouse  to  the  disease.  Mice  weighing  more  than 
20  gms.  generally  contracted  the  disease  and  died,  while  smaller  mice 
survived  without  showing  symptoms.  In  our  experiments  mice  died 
on  the  37th,  52nd,  81st,  and  126th  day  respectively,  after  inoculation. 

These  mice  wasted  considerably,  walked  with  tortoise-hke  move- 
ments, and  showed  other  motor  disturbances.  Typical  lesions  were 
demonstrated  in  the  brains  of  the  mice  and  the  intranuclear  '  inclu- 
sions '  of  Joest-Degen  were  present  in  the  Ammon's  horn. 

lY.  Fowl. 
Zwick,  Seifried,  and  Witte  (1926)  found  the  fowl  to  be  susceptible 
to  intracerebral  inoculation.    In  one  case  the  incubation  period  was 
37  days,  and  death  followed  15  days  later.    Passage  from  fowl  to 
rabbit  gave  a  positive  result. 

7.  ANIMALS  WHICH  HAVE  BEEN  FOUND  TO   BE  RESIS- 

TANT TO  INFECTION  WITH  THE  VIRUS  OF  ENZOOTIC 
ECEPHALO-MYELITIS 

I.  Dog. 

According  to  Zwick,  Seifried,  and  Witte  (1926)  the  dog  appears  to 
be  resistant  to  infection  with  the  virus  of  Boriui  disease.  This  fact 
obviates  up  to  a  certain  point  confusion  with  the  virus  of  rabies.  A 
greater  number  of  experiments  require  to  be  done,  however,  before 
the  dog  can  be  definitely  classed  among  the  animals  resistant  to 
infection. 

II.  Pigeon. 

These  same  authors  demonstrated  that  the  pigeon  is  resistant  to 
intracerebral  infection  with  the  virus. 

III.  Ferret. 

We  inoculated  six  ferrets,  three  young  and  three  adults,  by  the 
intracerebral  route  and  kept  them  under  observation  for  seven 
months,  but  no  morbid  symptoms  developed. 

8.  DISTRIBUTION  OF  THE  VIRUS  OF  BOUNA  DISEASE  IN 

THE  ANIMAL  BODY 

I.   Passage  of  Virus  through  the  Placenta. 
Ernst  and  Hahn  (1927)  showed  tliat  the  virus  is  capable  of  passing 
the  placenta  of  the  mare  and   infecting  the   foetus  during   intra- 
ut(!rine  hfe.    In  two  cases  tht>  virus  was  demonstrated  by  inoculation 


ENZOOTIC  ENCEPHALO-MYELITIS  45 

of  rabbits  iutracerebrally  with  the  brain  of  foals  born  of  mothers  ill 
with  enzootic  encephalo-myeUtis.  Further,  they  demonstrated  lesions 
characteristic  of  Boma  in  the  sections  of  the  brain  of  both  the  foals 
and  the  mothers  in  these  cases. 

II.   Distribution  of  the  Virus  in  Various  Organs  and 

Tissues 

Zwick,  Seifried,  and  Witte  (192G)  tested  four  samples  of  blood  from 
infected  rabbits,  three  samples  of  blood,  two  of  spleen,  two  of  kidney, 
and  two  of  Uver,  from  horses  ill  from  Boma  disease,  but  failed  to 
find  the  virus.  Ernst  and  Halm  (1927),  on  the  other  hand,  proved  the 
blood  to  contain  the  virus  during  some  stages  of  the  illness  of  a  rabbit 
suffering  from  Borna  disease. 

Similar  apparently  contradictory  results  have  been  obtained  in 
experimental  infections  produced  by  other  filterable  viruses  where  the 
virus  may  sometimes  be  found  in  the  blood,  e.g.  rabies,  vaccinia, 
herpes,  and  foot-and-mouth  disease. 

Ernst  and  Hahii  (1927)  foimd  the  vitreous  body  of  the  eye  infective 
after  a  rabbit  had  been  inoculated  intracerebrally.  The  virus  has 
also  been  demonstrated  by  Zwick  and  his  collaborators  (1926)  in  the 
submaxillary  salivary  gland  in  inoculated  rabbits. 

III.    Presence  of  Virus  in  the  Peripheral  Nerves  of 
Eabbits  Inoculated  Intracerebrally. 

The  present  writers  have  demonstrated  the  virus  in  the  peripheral 
nerves  of  rabbits  infected  by  the  intracerebral  route  in  which  in- 
filtrating lesions  in  the  nerve  occurred. 

Experiment  1.  A  portion  of  both  sciatic  nerves  taken  from  1  cm. 
below  their  emergence  from  the  greater  sciatic  foramen  to  the  popli- 
teal region  was  removed  aseptically  from  Rabbit  130a,  which  died 
50  days  after  intracerebral  inoculation.  An  emulsion  of  these  two 
portions  of  sciatic  nerve  was  made  in  physiological  saline  and  inocu- 
lated into  the  brain  of  Rabbits  213a  and  214a. 

Eahbit  213a. 
Weight  2,000  gins. 
14.7.27.   Intracerebral  inoculation  with  emulsion  of  sciatic  nerve. 
1.8.27.   Animal  normal.   2,000  gms. 
7.8.27.   Animal  normal. 
15.8.27.   Commencement  of  paresis  of  the  hind  quarters.   2,000  gms. 
24.8.27.   Typical  symptoms  of  the  disea.se.    1,600  gms. 
29.8.27.   Animal  very  ill.   1,350  gms. 
29.8.27.    Died  the  46th  day  after  the  inoculation. 

Autopsy.   All  organs  macroseopically  normal.   Cultures  from  the  brain  negative. 
Sections.   Intense  lesions  of  a  characteristic  type  in  the  central  nervous  system. 
The  corpuscles  of  .Joest-Degen  were  also  demonstrated. 


46  BOENA  DISEASE  AND 

Babbit  214a. 
Weight  2,100  gms. 
14.7.27.   Intracerebral  inoculation  with  au  emulsion  of  the  sciatic  nerve. 
1.8.27.    Normal.   2.250  gms. 
7.8.27.    Animal  normal. 

15.8.27.   Tj-pical  sj-mptoms  of  the  disease.    Head  depressed;  placed  on  its  side 
the    animal    showed    the    characteristic     myelitic    syndrome. 
1,800  gms. 
20.8.27.   Animal  very  ill.   1,600  gms. 
22.8.27.   Found  dead  39  days  after  the  inoculation. 
Autopsy.   No  lesions  in  organs.   Cultures  from  the  brain  negative. 
Sections.    Intense  lesions  characteristic  of  Boma  disease  were  demonstrated 

throughout  the  central  nervous  system. 
Passage.   The  brain  of  this  rabbit  «ri.s  passaged  to  Rabbit  291  x. 

Babbit  291.K. 

24.8.27.    Intracerebral  inoculation  with  an  emulsion  of  the  brain  of  Rabbit  214a. 
15.9.27.   T^-pical  symptoms  of  the  disease.    1,590  gms. 
21.9.27.    Found  dead  the  28th  day.    1,150  gms. 

Characteristic  intense  lesions  were  found  in  the  central  nervous  system,  and  the 
corpuscles  of  Joest-Degen  were  demonstrated. 

The  two  rabbits  died  after  showing  typical  symptoms  of  the 
disease.  Lesions  of  a  characteristic  type  were  demonstrated  through- 
out their  central  nervous  system,  and  moreover,  the  virus  was  demon- 
strated in  their  brain. 

Experiment  2.  In  a  second  experiment  the  virus  was  sought  for  also 
in  the  brachial  nerve  of  a  rabbit  which  had  succumbed  31  days  after 
inoculation  into  the  brain.  The  nerves  were  removed  aseptically  and 
emulsified  in  sterile  mortars.  The  emulsions  were  then  inoculated 
intracerebrally  into  rabbits.  The  results  of  the  inoculation  are  given 
below. 

(1)  Brachial  Xene. 

Rabbit  36k.    Weight  2,870  gms. 

15.9.27.   Inoculated  intracerebrallv  with  an  emulsion  of  the  brachial  nerve  of 

Rabbit  252a. 
22.9.27.   No  symptoms.   2,850  gms. 
28.9.27.   Animal  normal.   2,700  grms. 
6.10.27.   Animal  normal.  2,620  gms. 
10.10.27.   Commencement  of  paresis.   2,350  gms. 
12.10.27.    Tj'pical  symptoms  of  the  disease.   2,150  gms. 
14.10.27.   Animal  die<l  36th  day  after  inoculation.   1,800  gms. 
Autopsy.   All  organs  appeared  normal. 
Cultures  oj  the  brain.   Negative. 

Microscopic  examination  of  sections  of  the  brain  and  other  parts  of  the  central 
nervous  system  showed  the  presence  of  typical  lesions. 

(2)  Sciatic  Nerve. 

Rabbit  .J.5a.    Wright  1,200  gms. 

15.9.27.    Inoculated  intracerebrallv  with  nn  emuUion  of  the  sciatic  nerve  of 

Rabbit  252a. 
22.9.27.    .\nimal  normal.    1,050  gms. 
28.9.27.    Animal  normal.    1,020  gms. 
6.10.27.    Animal  normal.    1,100  gms. 


ENZOOTIC  ENCEPHALO-MYELITIS  47 

10.10.27.   Commencement  of  paresis.    1,150  gms. 
12.10.27.   Typical  symptoms  of  the  disease.   1,050  gms. 
14.10.27.   Tj-pical  symptoms  of  the  disease.   960  gms. 
15.10.27.    Found  dead  37  days  after  infection. 
AtUopsy.  All  organs  normal. 
Cultures  of  the  brain.  Xegative. 

Typical  lesions  were  found  on  microscopical  examination  of  sections  from  the 
central  nervous  system. 

This  second  experiment  shows  that  when  the  virus  is  introduced  into 
the  animal  organism  by  the  intracerebral  route,  it  may  subsequently 
be  found  in  the  brachial  as  well  as  the  sciatic  nerves. 

In  a  third  experiment  Babbits  27a  and  30a  were  inoculated  intra- 
cerebrally  with  an  emulsion  of  the  sciatic  nerve  taken  as  in  ex- 
periment 1. 

Rabbit  27a. 
Rabbit  27a.    Weight  1,690  gms. 

6.9.27.  Date  of  inoculation. 

15.9.27.  Animal  normal.   1,750  gms. 

22.9.27.  Animal  normal.    1,900  gms. 

28.9.27.  Animal  normal.   1,870  gms. 

6.10.27.  Animal  normal.    1,880  gms. 

10.10.27.  Commencement  of  the  disease.    1,850  gms. 

12.10.27.  Tj-pical  sj-mptoms  of  the  disease.    1,700  gms. 

15.10.27.  Animal  in  agonal  stage  of  death.    1,450  gms.    Found  dead  later,  39th 

day  after  infection. 

Autopsy.  Xo  macroscopic  lesions. 

Rabbit  30a. 
Rabbit  30a.    Weight  IJ.jO  gms. 

6.9.27.  Date  of  inoculation. 

15.9.27.  Animal  normal.    1,750  gms. 

22.9.27.  Animal  normal.    1,820  gms. 

28.9.27.  Animal  normal.    1,900  gms. 

6.10.27.  Animal  normal.  2,000  gms. 

10.10.27.  Slight  paresis  of  hind  quarters.   2,050  gms. 

12.10.27.  Slight  paresis  of  hind  quarters.   2,000  gms. 

21.10.27.  Typical  symptoms  of  the  disease  present.   1,900  gms. 

All  these  eight  rabbits  inoculated  intracerebrally  with  emulsions 
of  either  the  brachial  nerve  or  sciatic  nerve  of  rabbits  dead  of  experi- 
mental enzootic  encephaUtis  contracted  the  disease  and  died.  The 
presence  of  lesions,  and  of  the  corpuscles  of  Joest-Degen  in  the 
central  nervous  system  of  these  eight  rabbits,  as  well  as  the  positive 
passage  made  ■«ith  the  brain  of  one  of  them,  indicates  that  the  virus 
of  Boma  disease  generahzes  into  the  peripheral  nervous  system 
centrifugally.  The  lesions  occurring  in  the  peripheral  nerves  are 
described  later  (see  p.  61). 


48 


BOENA  DISEASE  AND 


IV. 


Distribution  of  the  Virus  in  various  Organs  and 
Tissues  of  the  Monkey. 


We  have  studied  the  distribution  of  the  virus  in  the  animal  organ- 
ism of  Monkey  M.  1  (Macacus  rJtesus)  (see  p.  34)  which  died  of 
Boma  disease  73  days  after  infection. 

Experiment  1.  Two  rabbits  were  inoculated  intracerebrally  with 
emulsions  of  the  cerebrum,  medulla  oblongata,  spinal  cord  (dorso- 
lumbar),  parotid,  spleen,  testicle,  blood,  and  adrenal  glands. 

The  results  are  set  forth  in  Table  VIII. 


Organ. 


Cerebrum 

Medulla 
oblongata 

Spinal  cord 
I'arotid  gland 

Spleen 


Adrenal 
gland 


Heart  Bluod 
(defibrinated) 


No.  of 
rabbit. 


USA 

126a 


117a 
123  a 


109a 
124a 


121a 
130a 


125a 
127a 


131a 
132a 


IIQa 
133a 


120a 
112a 


Weight 
in  gnus. 


1,700 
2,100 

2,280 
1,200 

1,«40 
l,tMO 

2,060 
1,860 

2,400 

1,000 

2,100 
1,700 

2,000 
1,100 

2,060 

1,800 


Table  VIII. 

First  appearance 
of  disease. 

22nd  day 
23rd     „ 

19th     „ 
22nd    „ 

20th     „ 
20th     „ 

Dead  accidentally 
34th  day 


Death  due  in  other 
causes  14th  day 
Survived 

Survived 
Survived 

30th  day 
26th     „ 

Death  due  to  other 

oausea 
Survived 


1 

Death. 

Lesions. 

37th  day 
30th    ., 

+ 
+ 

27th    „ 
37th    „ 

+ 
+ 

27th    „ 
30th    „ 

+ 
+ 

12th    „ 
51st     „ 

0 

+ 

— 

0 

40th    „ 
55th    „ 

+ 

+ 

0 

Passage. 


The  virus  was  demonstrated  in  the  brain,  medulla  oblongata, 
spinal  cord,  parotid,  and  adrenal  glands;  it  was  not  demonstrated  in 
the  spleen,  testicle,  or  blood. 

The  observations  were  repeated  with  the  organs  of  Monkey  M.  3 
(see  p.  36),  and  the  results  are  given  in  Table  IX. 


ENZOOTIC  ENCEPHALO-MYELITIS 


49 


Table  IX. 


Organ. 
Cerebrum 

Spinal  cord 

Parotid 
gland 

Adrenal 
gland 

Ovary 


Bone  mar- 
row 


No.  of 
rabbit. 

252a 

258a 

WeigU 
in  gms. 

1,370 

1,850 

Kidney 


Heart  blood 
defibrinated 


Lung 

Mesenteric 
gland 


255a 
261a 


265a 
266a 


249a 
267a 


259a 
271a 


262a 
264a 


250a 
268a 


254a 
270a 


269a 
272a 


251a 
263a 


273a 
274a 


First 
appearance 
of  disease. 


1,400 
3,020 


1,950 
3,390 


2,070 
2,790 


2,520 
1,690 


1,750 
1,850 


2,350 
2,320 


720 
2,150 


1,750 
1,740 


3,290 
1,970 


750 
1,540 


20th  day 
20th    „ 


20th 
20th 


Survived 


Survived 


Slst  day 
26th    „ 


30th 
28th 


Accidentally 
11th  day 

± 

15th    „ 

-U 

17th    „ 

— 

Accidentally 
10th  day 

8th  day 

- 

6th    „ 

Accidentally 
3rd  day 

- 

No  passage 
made. 


Negative. 


Negative. 


In  this  monkey  the  virus  was  found  in  the  brain  and  spinal  cord 
only,  and  could  not  be  demonstrated  in  the  ovary,  spleen,  bone 
marrow,  Uver,  kidney,  the  blood,  the  lung,  the  mesenteric  glands, 
adrenal  glands,  or  parotid  gland. 


9.    ELIMINATION  OF  THE  VIRUS  FROM  THE  ANIMAL 
ORGANISM 

As  is  the  case  with  the  viruses  of  herpes,  rabies,  &c.,  the  pathogenic 
agent  of  enzootic  encephalo-myehtis  is  ehminated  by  the  saliva  and 
nasal  secretions.  The  results  of  the  German  school  (Zwick,  Seifried, 
and  Witte,  1926;  Ernst  and  Hahn,  1926)  are  in  agreement  with  regard 
to  this  point.  The  urine  of  animals  ill  from  the  disease  has  always 
proved  avirulent. 

6409  n 


50  BOENA  DISEASE  AND 

10.    HISTOPATHOLOGY  OF  BORNA  DISEASE 
I.  Horse. 

Siedamgrotzky  and  Schlegel  (1896)  described  the  disease  as  a 
'serous  leptomeningitis'.  Both  Johne  (1896)  and  Ostertag  (1900  et 
seq.)  failed  to  find  lesions  in  the  brain  or  its  coverings,  and  considered 
the  disease  to  be  an  intoxication  of  the  central  nervous  system  by 
bacterial  toxins.  Dexler  (1900)  refers  to  Borna  disease  as  a  dis- 
seminated encephalo-niyehtis  with  leucocytic  inliltration  around  the 
vessels  and  in  the  nerve  substance.  Oppenheiui  (1907)  considered  it 
to  be  an  acute  localized  meningo-encephalitis  of  a  non-purulent 
nature,  the  meninges  being  more  especially  atl'ected.  Joest  and  Degen 
(1909),  who  made  a  more  detailed  study  of  the  histo-pathology  of 
Borna  disease  of  the  horse,  regarded  it  as  an  acute  meningo-encephalo- 
myehtis,  non-punilent  in  character,  with  perivascular  infiltrations 
by  lymphocytes  in  the  cord  and  brain.  Cellular  'inclusions'  were  also 
found  in  the  gangUon  cells  of  the  Amnion's  horn,  in  the  hippocampus, 
and  sometimes  also  in  other  regions  of  the  brain.  These  'inclusions' 
they  described  are  within  the  nucleus,  round  in  shape,  and  arranged 
sometimes  in  pairs.  They  stain  red  with  ^lann's  or  Lentz's  stain,  and 
have  often  an  unstained  halo  around  them.  Their  dimensions  vary 
from  the  hmit  of  visibihfy  to  the  size  of  a  nucleolus.  Heydt  (1914) 
was  able  to  confirm  the  findings  of  Joest  and  Degen  in  every  detail. 
Moussu  and  Marehand  (1924)  (see  also  thesis  of  Moussu,  1926)  did  not 
find  the  intranuclear  corpuscles  described  by  Joest  and  Degen. 

The  cases  investigated  by  Moussu  and  Marehand  were  of  an  acute 
haemorrhagic  type  and  polymorphonuclear  leucocytes  were  found  in 
the  lesions  in  large  numbers.  They  described  an  agglomeration  of 
infiltrating  mononuclear  cells  surrounding  the  nerve-cells  in  the  brain, 
j)roducing  in  certain  cases  deformation  of  these  cells  by  compression 
of  the  cell  membrane.  These  authors  did  not  record  actual  neurono- 
phagia. 

Zwick,  Seifried,  and  Witte  (1924),  Beck  and  Frohbose  (1926).  and 
Enist  and  Hahn  (1927)  confirmed  the  work  of  Joest  and  Degen  (1909) 
Zwick  and  his  collaborators  emphasize  the  fact  that  the  (lerman 
workers  have  never  found  degenerative  lesions  of  the  neuron  nor 
neuronophagia  in  any  region  of  the  nervous  system  where  they  were 
searched  for. 

II.  Cattle. 

The  similarity  of  the  disease  in  cattle  and  horses,  as  indicated  by 
a  study  of  the  lesions  in  the  central  nervous  system  with  the  jjresence 
of  virus  in  these  lesions,  lias  been  mentioned  in'  Ernst  and  Halm 
(1927).  'i'hey  demonstrated  the  corpuscles  of  Joest-Degen  in  tlie 
brain  of  cattle  dead  of  the  disease.  A  detailed  description  of  the  lesions 
has  also  been  given  in  the  thesis  of  Moussu  (1926).  He  states  that 
'the  ]iroc(ss  in  the  central  nervous  system  in  enzootic  encejihaio- 
myelitis  (»f  cattle  consists  of  a  diffuse  jjolio-encephalitis  with  a  jire- 
domiuance  of  lesions  in  the  cerebral  cortex,  of  wiiicii  the  deeper  layers 


ENZOOTIC  ENCEPHALO-MYELITIS  51 

are  more  particularly  affected.  There  are  similar  lesions  in  the  basal 
gangha  of  the  cerebral  hemisphere,  in  the  peduncle,  in  the  medulla 
oblongata,  and  even  in  the  spinal  cord ;  but  these  lesions  are  always 
less  marked  than  those  in  the  cerebral  cortex.  This  encephalitis  is 
accompanied  sometimes  by  perivascular  lesions  and  diffuse  capillary 
haemorrhages.  It  is  easy  to  understand  that  these  lesions  may  lead 
to  rapid  death  when  affecting  the  medulla  oblongata'. 

III.  Sheep. 

Priemer  (1925),  Beck  (1925),  Moussu  (1926),  Miessner  (1926),  and 
Ernst  and  Hahn  (1927)  made  a  study  of  the  lesions  in  the  central 
nervous  system  in  sheep  dead  of  the  disease.  They  came  to  the  con- 
clusion that  the  tissue  changes  were  analogous  to  those  produced  by 
the  virus  of  the  equine  type.  Beck  was  the  first  to  demonstrate  the 
intranuclear  corpuscles  of  .loest-Degen  in  the  nerve-cells  of  the  brain 
of  sheep  dying  from  the  spontaneous  disease. 

No  observer  has  described  any  departure  from  the  normal  in  the 
histology  of  other  organs  either  in  sheep,  horses,  or  cattle. 

IV.  Experimental  Borna  Disease  in  Horses  and  Sheep. 

The  lesions  described  in  the  nervous  system  of  horses  and  sheep 
infected  experimentally  are  identical  with  those  found  in  the  spon- 
taneous disease. 

V.    Rabbits  Infected  Experimentally. 

The  first  description  of  lesions  in  the  central  nervous  system  in 
rabbits  infected  with  the  virus  of  encephalo-myeHtis  was  given  by 
Moussu  and  Marchand  (1924).  These  authors  were  apparently  work- 
ing with  a  virus  which  differed  from  those  isolated  by  workers  in 
Germany;  and.  while  this  difference  may  possibly  be  accounted  for 
by  an  increased  virulence,  their  description  of  the  lesions  has  remained 
unique  and  unconfirmed  up  to  the  present.   It  is  as  follows: 

'In  rabbits  which  die  soon  after  the  inoculation,  the  alterations  are 
those  of  an  acute  meningo-encephalitis  with  a  predominance  of  lesions 
in  the  pia  mater.  The  meninges  are  infiltrated  with  immature  cells  of 
which  a  large  number  contain  eosinophile  granules.  These  same  cells 
can  be  found  around  intracerebral  vessels  (cortical  or  subcortical). 
There  are  alterations  of  the  choroid  plexus,  and  epithelium  of  the 
ventricles;  there  is  infiltration  of  the  "subependymal  zone  and  the 
pyramidal  zones  are  much  altered.  The  lesions  of  the  cerebellum  are 
identical  with  those  of  the  brain,  while  the  alterations  in  the  vessels 
of  this  region  are  also  as  intense  as  in  the  meninges. 

No  bacteria  were  found,  nor  intranuclear  inclusions.  In  animals 
which  died  at  a  later  stage  the  lesions  were  less  intense.  The  lesions 
predominate  in  the  anterior  region  of  the  brain  and  are  those  of  a 
subacute  encephalo-myelitis.  The  inflammatory  lesions  of  the  men- 
inges are  only  observed  in  the  septum  and  in  the  spaces  between  the 
convolutions.  The  same  may  be  said  of  perivascular  infiltrations. 
d2 


52  BORN  A  DISEASE  AND 

Embryonic  cells  ("cellules  embryomiaires ")  are  present,  containing 
eosinophile  granules.  There  is  an  indammation  of  the  ependyma  with 
subependymal  lesions. 

In  the  olfactory  lobe  one  finds  small  indammatory  areas.  The 
cellular  lesions  are  well  marked,  but  less  intense  than  in  the  preceding 
form.  In  the  cerebellum  one  tinds  several  areas  of  periarteritis  situated 
in  the  white  matter.   No  bacteria  or  inclusions  can  be  found.' 

Zwick.  Seifried,  and  Witte  (1926),  like  Beck  and  Frohbose,  Ernst 
and  Hahn  (1927),  and  Miessner  (1927),  who  have  been  interested, 
in  passing,  in  the  liistopathology  of  the  central  nervous  system  of 
rabbits  infected  experimentally,  devote  only  a  few  lines  to  the 
question.  In  general,  the  summary  of  their  findings  may  be  given 
as  follows:  Macroscopically,  apart  from  the  brain  and  cord,  which 
appeared  to  be  hyperaemic,  all  the  organs  preserved  their  normal 
aspect.^  A  microscopic  study  of  the  lesions  in  the  brain  revealed  a 
shght  meningitis  with  mononuclear  cells;  more  or  less  infiltration  of 
the  cerebral  cortex  and  the  Amnion's  horn  with  lymphocj'tes ;  peri- 
vascular infiltrations,  especially  in  the  small  and  middle-size  vessels; 
and  the  presence  of  intranuclear  corpuscles  in  the  large  ganglion  cells 
of  the  Amnion's  horn.  Zwick  and  his  collaborators,  as  well  as  other 
investigators  who  have  studied  the  disease  in  Germany,  have  never 
observed  degenerative  processes  in  the  nerve-cells  nor  recorded 
neuronophagia.  Zwick  found  perivascular  infiltrations  in  the  spinal 
cord  in  cases  which  had  paresis  or  paralysis. 

The  summary  given  above  records  briefly  the  obser\atioiis  made 
by  other  workers  whose  attention  has  been  directed  particularly  to 
the  brain,  while  the  participation  of  the  spinal  cord  in  the  patho- 
logical process  has  been  referred  to  only  exceptionally.  There  is  no 
pubhshed  work  on  lesions  of  the  nerve-roots,  spinal  ganglia,  ami  peri- 
pheral nerves. 

VI.  Authors'  Observations. 

A.  Rabbit. 

(1)  Macroscopical  and  Microscopical  Findings  in  Diverse  Organs. 

Our  observations  have  been  made  on  animals  infected  either  with 
the  strain  of  virus  originating  from  horses  (Zwick)  or  that  originating 
from  sheep  (Miessner).  On  post-mortem  examination  in  the  majority 
of  cases,  a  congestion  of  the_ meninges  which  may  sometimes  be  in- 
tense is  found.  In  other  cases  the  aspect  of  the  nervous  system  may  be 
normal. 

Sometimes    the    stomach    presents    the    lenticular    haeniorrhnges 

'  Kriwt  and  Halm  ( 1027)  described  haemorrhagic  lenticular  fi>rnmtioii8  in  the  inucoaa 
of  the  Htciinacli  of  rahhitH  dead  of  Horna  disease.  We  have  fimiid  these  lentiotilar  for- 
mations (see  Fiji.  !•)  in  the  majority  of  caaes  in  which  rahlills  had  a  prolonKed  jiaralytio 
phase  and  a  long  agonal  Ht«ge.  lln<ler  the  microscope  they  appeared  to  be  haemor- 
rhages fdlliiwini;  ii|K>n  an  autodigestion  of  the  stomach  mucosa.  They  did  nut  appcur 
to  have  any  dilinite  struct  ure,  nor  are  they  s|KH'ilie  forex|)i'rinientjil  enzootic  eucephalo- 
myelitih,  kIiI'  <-  ue  havr  found  IlKin  aJMO  In  lier|>etic  encephalilla  and  oilier  morbid  oon- 
ditionji. 


ENZOOTIC  ENCEPHALO-MYELITIS  53 

referred  to  above,  but  we  bave  found  tbat  tbese  are  not  a  specific 
reaction  to  tbe  virus  (Fig.  9).  In  the  larger  number  of  cases  examined 
in  detail  (more  than  thirty  rabbits)  the  kidneys  showed  a  marked 
congestion.  This  hyperaemia  was  not  limited  to  the  cortical  zone, 
but  affected  the  medullary  zone  to  the  same  extent.  Sections  from 
such  kidneys  showed  that  there  were  small  multiple  haemorrhages  in 
the  region  of  the  glomerulus,  as  well  as  in  the  collecting  and  con- 
voluted tubules.  These  extravasations  formed  sometimes  actual 
haemorrhagic  areas.  The  epithehum  hning  the  renal  tubules  was 
normal,  there  were  no  infiltrative  processes  peri-  or  intratubular. 
The  condition  may  be  described  as  renal  congestion;  not  a  true 
nephritis. 

On  microscopic  examination  the  parotid  showed  occasionally  small 
areas  of  infiltration  composed  of  lymphocytic  elements  surrounding 
certain  of  the  striated  canaliculi;  at  the  same  time  the  cytoplasm  of 
certain  of  the  cells  of  the  acini  had  become  oxyphilic  when  stained 
by  Mann,  while  the  nucleus  appeared  oedematous  and  took  up  an 
abnormal  eccentric  position  in  the  cells.  Although  many  sections  of 
the  parotid  of  rabbits  were  examined,  these  lesions  were  found  only 
occasionally.  Without  presuming  that  they  were  produced  by  the 
action  of  the  virus,  it  should  be  mentioned  that  they  coincided  with 
the  presence  of  virus  in  this  organ.  In  rabies  and  distemper,  oxy- 
philic corpuscles,  intra-  or  extra-cellular,  staining  red  by  Mann's 
method,  have  been  described  as  concomitant  with  the  presence  of 
virus  in  the  parotid.  No  actual  corpuscles  have  been  found  by  us  in 
the  parotid  of  animals  infected  with  the  virus  of  enzootic  encephalo- 
myehtis. 

In  the  medullary  zone  of  the  adrenal  small  accumulations  of  lym- 
phocytic elements  were  occasionally  seen.  The  lung,  the  liver,  the 
spleen,  the  testicle,  and  the  ovary  appear  macro-  and  microscopically 
normal. 

Characteristic  lesions  of  an  intense  nature  are  found  only  in  the 
nervous  system.  We  may  class  these  in  two  categories,  (1)  infiltrative 
and  (2)  degenerative.  Both  types  of  lesions  may  be  met  with  in  the 
brain,  mesencephalon,  cerebellum,  spinal  cord,  and  spinal  ganglia. 
In  the  nerve-roots  and  peripheral  nerves  (sciatic  and  brachial  nerves 
examined)  only  infiltrative  lesions  have  been  found. 

(2)  Lesions  iii  the  Central  Nervous  System. 
(a)  The  Brain.  The  pia  mater  is  infiltrated  with  mononuclear 
leucocytes,  varying  in  individual  cases.  In  some  areas  only  a  trace 
of  this  infiltrative  process  may  be  seen,  while  in  others  three  or  four 
layers  of  infiltrative  cells  occur.  They  are  especially  marked  in  the 
region  of  the  meningeal  vessels  as  well  as  in  the  spaces  between  the 
convolutions,  and  may  form  actual  meningeal  plaques.  The  infiltrating 
elements  are  lymphocytes,  plasma  cells,  and  large  mononuclears. 
Vessels  of  the  pia  mater  are  often  surrounded  by  '  cuffs '  constituted 
by  mononuclear  leucocytes  (Fig.  10).  The  most  intense  lesions  of 
the  meninges  are  generally  found  at  the  base  of  the  brain.  In  sections 


54  BOENA  DISEASE  AND 

cut  at  right  angles  to  the  surface  of  the  brain  vessels  passing  from  the 
meninges  into  the  cortical  substance  surrounded  by  lymphocytic 
'cufling'  have  often  the  aspect  of  septa  (Fig.  11).  In  the  cortex  diffuse 
infiltration  of  lymphocytes  accompanied  by  proUferation  and  mobili- 
zation of  the  neurogUal  cells  occurs.  Especially  in  the  liippocampus 
(the  so-called  'elective  zone'  in  herpetic  encephahtis  of  the  rabbit), 
the  neuron  degenerates,  the  nucleus  swells,  the  chromatin  becomes 
rarefied,  collects  towards  the  periphery  of  the  nucleus,  and  in  its 
place  appear  small  oxyphihc  globules  which  may  be  at  the  hmit  of 
visibihty  or  may  reach  the  size  of  a  nucleolus.  Some  of  the  neurogUal 
cells  of  this  region  appear  to  undergo  the  same  degenerative  process. 
It  is  similar  to,  although  less  intense  than  that  described  by  Levaditi, 
Harvier,  and  Nicolau  (1922)  in  experimental  herpetic  encephalitis 
in  the  rabbit,  leading  to  the  formation  of  encephalitic  neuro- 
corpuscles,  and  is  not  of  a  specific  nature.  The  oxyphilic  degenera- 
tion of  the  nucleus  may  lead  to  the  formation  of  larger  corpuscles 
surrounded  by  a  halo,  morphologically  identical  with  those  described 
as  specific  in  Borna  disease  by  Joest  and  Degeu  (1909).  The  nucleus 
may  react  in  the  same  waj'  to  other  causes.  Even  the  halo  is  not 
wanting  in  the  figures  given  by  Levaditi,  Harvier,  and  Nicolau.  We 
regard  this  phenomenon  as  possibly  the  result  of  the  action  of  a 
karyotropic  virus,  the  degenerated  karyoplasm  fusing  round  the 
pathogenic  agents  These  nuclear  lesions  may  be  found  in  all  regions  of 
the  brain,  although  the  German  writers  have  described  their  presence 
only  in  the  Cornu  Ammonis,  in  which,  indeed,  they  are  more  con- 
stantly found.  They  may  be  single  or  in  pairs  surrounded  or  not  by 
a  ciiaracteristic  halo.  When  sections  are  stained  with  Mann's  stain 
or  toluidin  blue-eosin,  the  intranuclear  corpuscles  are  stained  rose  or 
red.  while  the  nucleolus  is  more  of  a  violet  tint.  They  occur  also  in 
the  pyramidal  cells  of  the  cerebral  cortex  and  even  in  the  neuroglial 
cells  (large  granulo-adipose  cells  of  the  hippocampus). 

In  the  Ammon's  horn  newly  formed  capillaries  are  sometimes  seen. 
The  vessels  of  this  region  appear  dilated,  gorged  with  l)lood,  and  their 
adventitia  infiltrated  with  several  layers  of  mononuclear  leucocytes. 
Plasma  cells  are  abundant  in  the  process  of  perivascular  infiltration. 
We  have  never  found  such  a  large  number  in  rabies  of  the  dog  or 
rabbit,  in  pohomyehtis  of  monkeys,  in  human  encephalitis,  in  chronic 
herpetic  encephalitis  of  rabbits,  or  in  the  vascular  lesions  produced 
by  the  presence  of  the  so-called  '  Encephalitozoon  cuniculi '.  Small 
lymphocytes  may  also  be  found  in  large  numbers  in  these  perivascular 
infiltrations,  but  large  mononuclears  cells  are  also  present  to  a  less 
extent.  The  presence  of  polymorphonuclear  leucocytes  is  excep- 
tional. 

Sometimes  in  the  thickness  of  the  'cuffing'  are  degenerated  lympho- 
cytes or  plasma  cells.  The  nucleus  of  these  degenerated  cells  has 
become  intensely  oxyphilic.  It  is  reduced  in  size  and  condensed  in 
'blocks'  without  any  definite  structure.  When  Mann's  stain  is  used, 
the  protojilasm,  in  the  case  of  the  degenerated  plasma  cells,  stains 
rose;    the  unchanged  elements  stain  bhir.    Here  and  tliere  in  the 


ENZOOTIC  ENCEPHALO-MYELITIS  55 

mass  of  the  infiltrating  cells  rare  fragments  of  degenerated  chromatin 
can  be  found,  probably  arising  from  degenerated  mononuclear 
leucocytes  or  pyknosed  polymorphonuclear  cells.  The  intensity  of 
the  infiltration  varies.  Sometimes  the  perivascular  'cuffing'  is  con- 
stituted by  one,  two,  or  three  layers,  while  in  other  cases  a  massive 
nodule  resembling  a  gumma  may  he  found  with  a  small  vessel  in 
the  centre.  These  alterations  occur  not  only  in  the  vessels  of  the 
Ammon's  horn,  but  in  all  the  regions  of  the  central  nervous  system 
where  'cuffing'  may  be  found. 

In  the  Cornu  Ammonis  areas  of  mononuclear  infiltration  are  seen 
between  the  ganglion  cells,  in  the  row  of  fusiform  cells,  or  in  the  chain 
of  small  granular  cells  (Fig.  12).  Neuroghal  cells  of  neoformation 
may  also  participate  in  this  infiltrative  process. 

We  have  never  found  true  neuronophagia  at  this  site,  but  we  have 
encountered  a  curious  phenomenon,  the  exact  nature  of  which  we 
iiad  some  trouble  m  determining.  In  a  section  of  brain  examined 
under  the  oil  immersion  we  found  in  one  of  the  large  ganghon  cells 
of  this  region  stained  with  toluidin  blue-eosin  a  species  of  '  cyst '  in 
the  interior  of  the  protoplasm  (Figs.  13,  14.  15,  16).  The  rest  of  the 
cell  preserved  its  normal  aspect.  The  'cyst'  compressed  the  nucleus 
and  pushed  it  towards  the  periphery  of  the  cell,  forming  a  marked 
depression  in  the  nuclear  membrane.  The  diameter  of  this  '  cyst '  was 
about  7 IX.  It  was  marked  off  from  the  cytoplasm  of  the  neuron  by 
a  membrane,  and  contained  six  basiphilic  granules  of  equal  size, 
placed  symmetrically  at  its  periphery. 

On  subsequent  minute  examination  of  the  cells  of  the  Ammon's 
horn  we  found  these  '  cysts '  again  in  several  preparations.  They  have 
been  found  only  in  this  region  and  always  in  the  protoplasm  of  the 
cell.  Their  diameter  varied  from  5  /x  to  9  /x.  During  the  search  we  have 
seen  occasionally  plasma  cells,  the  nucleus  of  which  had  undergone 
degeneration,  which  in  its  appearance  recalled  this  intracellular 
'cyst';  the  karyoplasm  was  condensed  into  several  small  round  in- 
tensely chromatophihc  granules,  apparently  attached  at  equal  dis- 
tances to  the  nuclear  membrane.  The  presence  of  a  protoplasmic 
circle  around  this  formation  removed  from  our  minds  the  supposition 
that  we  were  deahng  with  a  '  micros poridian  cyst '.  and  showed  dis- 
tinctly that  it  was  a  degenerated  infiltrating  cell  (PI.  I,  Fig.  3).  The 
staining  reactions  indicated  that  the  cystic  formations  were  the 
degenerated  nuclei  of  the  plasma  cells  which  had  penetrated  into  the 
interior  of  the  large  cells  of  the  Ammon's  horn.  We  have  found  such 
bodies  in  approximately  5  per  cent,  of  cases  examined.  They  have 
never  been  met  with  in  the  brain  of  normal  rabbits,  or  in  the  brain  of 
rabbits  which  have  succumbed  to  infection  with  the  viruses  of  herpes, 
rabies,  or  vaccinia. 

We  have  described  these  formations  in  detail,  since  the  elucidation 
of  their  nature  required  extended  observations.  Many  control 
animals  were  examined  to  ensure  that  we  were  not  dealing  with  a 
spontaneous  '  microsporiduin  '  disease  of  the  rabbit. 

Pathological  changes  other  than  the  above  are  found  more  con- 


56  BOENA   DISEASE  AND 

stantlv  in  the  Amnion's  horn,  especially  in  cases  showing  intense 
intiltrative  lesions.  For  instance,  a  number  of  the  nerve-cells  may 
show  degenerative  changes  characterized  by  the  following  appear- 
ances. The  nucleus  appears  oedematous,  the  chromatin  is  fragmented, 
while  the  protoplasm  shows  more  or  less  advanced  tigrolysis.  The 
protoplasm  also  contains  vacuoles  and  the  cellular  membrane  is 
denticulated.  In  some  cells,  also,  of  the  Ammon's  horn,  the  nucleus, 
and  sometimes  too  the  protoplasm,  becomes  oxyphiUc.  The  fusiform 
cells  which  are  found  in  the  upper  part  of  the  row  of  large  ganghon 
cells  show  intense  nuclear  degeneration:  the  karyoplasm  is  con- 
densed in  a  'block'  and  stains  red  with  Mann"s  stain.  When  a  pre- 
paration so  stained  was  decolorized  gradually,  and  examined  after 
each  stage  of  the  process  until  these  degenerated  nuclei  liecame  a  pale 
rose  colour,  it  was  found  that  they  had  no  definite  structure,  but 
consisted  of  a  round  mass  of  homogeneous  condensed  chromatin  much 
smaller  in  size  than  the  unchanged  nuclei.  This  type  of  nuclear 
degeneration  has  also  been  met  with  in  the  mesencephalon  (PI.  Ill, 
Fig.  4). 

We  have  ahready  referred  above  to  the  fact  that  oxyphilic  corpuscles 
which  may  or  may  not  be  surrounded  by  halos  are  found  in  the 
nucleus  of  some  of  the  cells  of  the  Amnions  horn.  They  can  be 
distinguished  from  the  nucleolus  by  their  different  staining  reactions 
(PI.  I,  Fig.  2).  These  intranuclear  corpuscles— the  specific  'inclu- 
sions' of  the  German  workers — may  be  single  or  in  twos  or  threes, 
varying  in  size.  Sometimes  they  may  be  at  the  hmits  of  visibility, 
or  they  may  be  as  large  as  2  /x  or  3  fi.  These  corpuscles  occur  else- 
where. They  may  be  found  in  the  cytoplasm  of  the  cell  and  are 
possibly  expelled  intra  vitani.  but  as  it  is  possible  for  the  nucleolus  of 
a  cell  to  be  dislodged  by  the  microtome  knife,  the  same  factor  might 
carry  the  intranuclear  corpuscle  into  the  cvtoplasm. 

Our  opinion  is  that  the  intranuclear  'inclusion'  in  Horna  disease  is 
possibly  a  reaction  of  the  karyoplasm  against  the  pathological  agent 
which  penetrates  the  interior  of  the  nucleus.  Possibly  the  chromatin 
masses  around  the  infective  virus  elements.  This  is  suggested  by  the 
staining  reactions,  since  the  condensed  mass  of  chromatin  wliich 
forms  the  corpuscle  undergoes  degeneration  from  the  centre  towards 
the  periphen,-  (the  centre  appears  oxyphilic  and  the  periphery  basi- 
philic  in  certain  corpuscles). 

Around  the  lateral  ventricles  well-marked  infiltrations  are  found, 
these  being  in  some  cases  very  intense.  The  choroid  plexus  is  also 
infiltrated.  The  epitheUuni  of  the  ventricle  and  of  the  ependyma  is 
unchanged. 

Occasionally  in  the  parenchyma  in  the  region  of  tht!  ventricle,  or 
even  in  the  cerebral  cortex  itself  in  the  superficial  areas,  karyo- 
kinetic  figures  may  be  seen.  Probably  this  karyokinesis  is  in  niol)ilized 
cells  of  the  vascular  endothelium  which  have  penetrated  into  the 
ner\'e  substance.  In  certain  preparations  we  have  seen  2,  8,  or  even 
4  karyokinetic  figures.  The  mitosis  sometimes  undergoes  oxyphilic 
degeneration. 


ENZOOTIC  ENCEPHALO-MYELITIS  57 

There  are  lesions  affecting  the  area  above  and  below  the  ependyma 
and  also  the  surrounding  zone:  a  mobihzation  of  the  neurogUal 
elements  takes  place,  while  at  the  same  time  lymphocytic  elements 
are  found  in  the  immediate  proximity  of  the  neuron.  Up  to  a  certain 
point  this  phenomenon  is  comparable  with  the  'satellitism'  described 
by  Metchnikofi'  in  senihty.  Certain  of  the  nerve-cells  are  surrounded 
on  all  sides  by  'satelhte'  cells  and  cells  of  infiltration,  which  occa- 
sionally penetrate  the  interior  of  the  cell.  Six,  eight,  and  even  ten 
neurons  may  be  seen  '  besieged '  in  one  microscopic  field.  The  cells 
which  come  in  immediate  contact  with  the  nerve-cell  push  in  the  cellular 
membrane  and  form  '  cups '  in  the  periphery  of  the  cytoplasm,  giving 
the  nerve-cell  a  denticulated  border.  This  phenomenon  is  more  com- 
monly met  with  in  sections  from  rabbits  dying  within  the  first  20  days 
after  inoculation.  The  intensity  of  'satellitism'.  is  in  inverse  propor- 
tion to  the  meningeal  and  perivascular  lesions.  When  the  meningitis 
and  perivascular  cuffing  are  at  a  minimum  '  satelhtism '  may  represent 
the  only  departure  from  the  normal  discovered  in  the  brain. 

From  the  examination  of  a  large  number  of  preparations  we  are  led 
to  beheve  that  'satelhtism'  is  a  stage  which  may  either  disappear 
during  the  evolution  of  the  disease,  resulting  in  a  quasi-normal 
state,  or  become  intensified  and  be  followed  by  neuronophagia.  We 
have  found  neuronophagia  present  in  sections  of  brain  showing 
marked  'satellitism'  (Fig.  17).  Our  conception  of  the  various  stages 
of  the  struggle  against  the  virus  is  as  follows:  When  the  nervous 
system  is  invaded  by  the  virus  the  neuroghal  elements  and  mono- 
nuclear lymphocytes  are  attracted  to  the  parasitized  neuron.  If  the 
neuron  succeeds  in  freeing  itself  from  the  virus,  the  local  reaction 
ceases  at  this  stage,  and  the  resorption  of  the  satellite  elements 
follows:  but  if  the  neuron  dies  in  the  struggle  against  the  virus  after 
undergoing  intense  degeneration,  it  is  invaded  by  the  satelhte  cells 
and  rare  polymorphonuclear  leucocytes ;  the  process  has  now  reached 
the  stage  of  neuronophagia.  When  the  struggle  between  the  neuron 
and  the  virus  terminates  without  neuronophagia  taking  place  the 
animal  survives  until  the  meningeal  lesions,  perivascular  infiltrations, 
and  infiltration  of  the  Amnion's  horn  become  incompatible  with  life. 
In  this  case  death  takes  place  at  a  later  stage,  i.e.  in  from  25  to  50  days. 

These  conclusions  as  to  the  evolution  of  the  morbid  process  in  the 
brain,  formed  from  observations  on  the  character  and  position  of  the 
lesions  in  a  large  number  of  rabbits  dying  early  or  late  after  inocula- 
tion, have  received  further  support  from  the  study  of  the  histogenesis 
of  the  alterations  in  the  central  nervous  sytem  of  six  rabbits  killed 
at  regular  intervals  after  inoculation,  i.e.  on  the  5th,  10th,  15th, 
20th,  and  27th  day,  and  of  one  which  died  on  the  31st  day.  In  these 
animals  also,  the  microscopical  examination  of  sections  of  various 
parts  of  the  nervous  system  showed  that  the  first  modification  in  the 
central  nervous  sytem  is  the  mobihzation  and  prohferation  of  the 
neuroghal  cells  around  the  neuron — 'satellitism'.  this  being  more 
marked  in  the  pons  and  the  medulla  oblongata.  In  the  process  of 
'satelhtism'  one  finds  not  onlv  neuroglial  cells  but  also  mononuclear 


58  BOENA  DISEASE  AND 

cells  taking  part.  Later,  infiltration  of  the  Cornu  Ammonis,  the 
meninges,  and  the  vascular  tissues  takes  place.  In  the  subjects  of  our 
experiments  the  latter  process  began  to  appear  towards  the  15th  or 
•20th  day  after  the  inoculation. 

The  presence  of  polymorphonuclear  cells  is  quite  exceptional  no 
matter  at  what  stage  of  the  infection  or  from  what  site  one  examines 
sections  of  the  central  nervous  system;  the  infiltrative  lesions  are 
constituted  from  the  beginning  by  mononuclear  cells. 

There  are  notable  differences  between  the  development  of  the 
encephaUtis  of  Boma  disease  in  the  rabbit  and  chronic  herpetic 
encephaUtis  produced  experimentally  in  the  same  species.  In  the 
brain  infected  by  herpes  an  acute  stage  is  observed  in  which  poly- 
morphonuclear leucocytes  take  part,  and  are  fomid  in  large  numbers 
in  the  perivascular  'cuffing'  as  well  as  in  the  nodular  lesions  at  the 
base  of  the  brain  in  the  region  of  the  hippocampus.  If  the  animal 
survives  this  acute  stage  and  recovers  what  have  been  termed  by 
Levaditi  and  Xicolau  (19'2'2)  'lesions  d'imnmnite'  may  be  found. 
These  are  small  nodular  or  diffuse  areas  of  parenchymatous  infiltra- 
tion situated  in  the  hippocampus  (the  'zone  elective')  and  are  pro- 
duced by  mononuclear  cells  that  have  replaced  the  polymorpho- 
nuclear leucocytes  with  which  the  inflammatory  process  commenced. 
In  those  rabbits  which  just  fail  to  resist  the  disease  and  die  in  20  to 
30  days,  the  lesions  are  more  intense,  but,  as  in  the  case  of  the 
'lesions  d'immmiite',  the  infiltration  consists  of  mononuclear  cells 
which  have  taken  the  place  of  the  polymorphonuclear  leucocytes  that 
predominated  in  the  acute  stage  of  the  inflammatory  process.  In  the 
case  of  the  infection  of  the  brain  of  the  rabbit  witli  tiie  virus  of  Boma 
disease,  polymorphonuclear  leucocytes  do  not  plaj'  a  part  in  the  early 
inflammatory  process;  during  this  early  stage,  one  observes  only 
'sateliitism'  of  the  neuron,  while  the  infiltrative  lesions  are  produced 
by  mononuclear  cells  alone  from  the  beginning  of  tiie  process  until 
the  final  stage. 

(b)  The  Mid-brain  and  Medulla  Oblongata.  In  tlie  mesencephalon 
and  the  medulla  ol)l()ngata  similar  infiltrative  and  degenerative 
lesions  occur.  'Cuffing'  of  tlie  vessels  is  frequently  observed.  Certain 
of  the  nerve-cells  appear  to  be  in  a  state  of  advanced  tigrolysis 
(Xissl's  granules  have  disappeared).  Degeneration  both  of  tiie  nucleus 
and  the  cytoplasm  is  frequently  found.  Certain  of  the  cells  appear 
to  have  their  pnjtophism  split  up,  the  nucleus  lieing  jieripheral, 
swollen,  and  completely  degenerated  (Fl.  I,  tig.  1).  Occasionally 
typical  neuronophagia  is  encountered.  Iti  tiiese  regions  also  we  have 
demonstrated  the  presence  of  the  intranuclear  cor])uscies  of  Joest- 
Degen.  The  cells  which  contain  them  generally  preserve  otherwise 
their  normal  aspect ;  the  nuclear  membrane  is  intact,  wliile  the  proto- 
plasm is  structurally  unchanged  and  stains  normally.  The  converse 
is  also  true;  we  liave  never  found  the  corpuscles  of  Joest-Degen  in 
cellH  in  advanced  stages  of  degeneration  or  disintegration.  Negri  liodies 
in  rabies  are  also  oidy  found  in  nerve-ceils  wiiicii  are  otherwise  normal. 
In    tlie    mesencepliaion,    as    mentioned    above,    one     meets     most 


ENZOOTIC  ENCEPHALO-MYELITIS  59 

frequently  with  '  satellitism '  of  the  neuron,  which  in  certain  cases  goes 
so  far  as  to  constitute  true  neuronophagia  (Fig.  18).  In  the  mesen- 
cephalon also  we  may  tiud  small  islands  of  mononuclear  cells  in  the 
parenchyma  without  any  relation  to  the  vessels.  In  certain  cases  we 
found  neuroglial  cells  (granulo-adipose  cells)  showing  nuclear  oxy- 
chromasia  and  occasionally  small  oxyphihc  corpuscles  within  the 
nucleus,  similar  to  the  so-called  'encephahtis  neurocorpuscles '  of 
herpes  described  by  Levaditi.  Harvier,  and  Nicolau  (1922). 

(c)  Cerebellum.  In  the  case  described  the  lesions  in  the  cerebellum 
were  much  more  intense  than  the  average,  since  usually  the  altera- 
tions consisted  only  of  shght  meningitis  and  perivascular  infiltration 
accompanied  by  occasional  'satelhtism'  of  the  cells  of  Purkinje.  In 
the  septum  there  was  a  marked  infiltration  with  mononuclear  cells. 
Massive  perivascular  'cufiing'  was  present,  especially  in  the  white 
substance  between  the  convolutions.  In  the  vessels  themselves, 
which  were  gorged  with  blood,  an  excessive  number  of  mononuclear 
cells  were  found.  There  was  an  intense  infiltration  in  the  granular 
layer,  which  in  some  cases  was  completely  destroyed  and  replaced 
by  areas  formed  exclusively  of  mononuclear  cells.  Here  and  there  in 
the  islands  of  lymphocytes  'basket'  cells  -vsith  a  pale-staining 
degenerated  protoplasm  were  found,  their  border  appearing  irregular. 
Several  of  the  cells  of  Purkinje  appeared  to  be  hyperchromatic,  and 
their  nucleus  was  in  some  cases  eccentric  and  stained  by  acid  stains. 
In  others  the  nucleus  was  not  separated  from  the  rest  of  the  proto- 
plasm since  the  remains  of  the  nuclear  membrane  appeared  to  have 
disappeared.  Other  Purkmje  cells  were  degenerated  and  appeared  as 
cell  shadows.  In  other  parts  the  karyoplasm  of  certain  of  these  cells 
was  condensed  around  the  nucleolus.  Satellitism  of  the  basket  cells 
and  the  cells  of  Purkinje  was  noticeable,  but  true  neuronophagia  was 
not  seen.  The  lesions  were  confined  to  certain  areas;  other  parts  of 
the  cerebellum  were  perfectly  normal. 

In  a  case  where  the  inoculation  of  the  virus  was  made  by  the 
intratesticular  route  the  lesions  in  the  cord  were  especially  well 
marked,  and  in  the  cerebellum  the  satelhtism  of  the  cells  of  Purkinje 
was  occasionally  so  advanced  as  to  constitute  almost  a  true  neurono- 
phagia. 

(d)  The  Spinal  Cord.  Generally  it  may  be  stated  that  the  intensity 
of  the  lesions  found  in  the  brain  or  in  the  spinal  cord  corresponded 
with  the  intensity  of  the  sj^mptoms  observed  during  life.  In  the 
rabbits  we  have  examined  pathological  changes  were  always  present 
in  the  cord  whether  symptoms  of  affection  of  this  part  of  the  central 
nervous  system  were  present  or  not,  but  these  were  much  more 
intense  when  the  symptoms  produced  by  affection  of  that  region 
domuiated  the  cerebral  symptoms. 

As  in  the  case  of  the  brain,  the  meninges  of  the  cord  are  not  as 
a  rule  greatly  affected.  Only  in  isolated  cases  was  a  severe  meningitis 
found. 

The  anterior  and  posterior  septa  may  be  more  or  less  infiltrated 
with  mononuclear  cells.    Perivascular  'cufiing'  is  seen  both  in  the 


60  BORNA  DISEASE  AND 

grey  and  white  matter  of  the  cord.  In  the  anterior  and  posterior 
horns  infiltrations  with  mononuclear  cells  may  be  seen. 

The  most  intense  infiltrations  are  found  in  the  posterior  horn, 
while  in  the  anterior  horn  degenerative  lesions  of  the  neuron  are  more 
common.  The  process  of  degeneration  in  the  nerve-cells  is  the  same 
as  that  in  other  regions  of  the  central  nervous  system:  tigrolysis 
occurs,  nuclear  oxychromasia  exists,  while  the  whole  cell  shows  a 
marked  hyperchromasia.  Vacuolization  of  the  cytoj)lasm,  a  degenera- 
tive process,  may  also  be  seen  (PI.  II,  Fig.  1),  while  in  rare  cases  when 
the  lesions  as  a  whole  have  been  exceptionally  intense,  occasional 
neuronophagia  was  recorded  (Fig.  19).  The  phenomenon  of  'satel- 
litism'  appears  to  be  more  commonly  met  with  in  the  cord  than  in 
the  brain  itself.  The  intranuclear  corpuscles  of  Joest-Degen  found 
in  the  cord  have  generally  been  in  the  nerve-cells  of  the  anterior  horn 
(PL  I.  Fig.  5;  PI.  Ill,  Fig.  2). 

Here  and  there  small  islands  of  lymphocytes  may  be  found  in- 
filtrating both  the  white  and  the  grey  suiistauce,  these  islands  being 
unconnected  with  vessels.  Frequently  it  has  been  observed  that  the 
zone  of  Lissauer  is  the  site  of  a  well-marked  mononuclear  infiltration. 

The  lesions  found  in  the  spinal  cord  are  comparable  with  those  found 
in  poliomyehtis.  The  neuronophagia  so  characteristic  in  the  cord 
of  monkeys  infected  with  the  virus  of  the  latter  disease  (to  which 
rabbits  are  generally  considered  not  to  be  susceptible)  is  also  present 
in  the  cord  of  rabbits  infected  with  the  vn-us  of  Borna  disease,  but 
to  a  less  degree. 

(3)  Lesions  in  tite  Peripheral  Nervous  System. 

(a)  The  Posterior  Nerve-roots  arising  from  the  cells  in  the  zone  of 
Lissauer  have  infiltrative  lesions  which  vary  in  intensity.  This  zone, 
as  has  i)een  stated  above,  is  generally  infiltrated  with  mononuclear 
cells.  The  infiltration  takes  place  i)etween  tlie  nerve  filaments,  and 
consists  of  a  chain  of  lymphocytes.  In  some  cases  only  traces  of  this 
infiltration  can  be  seen,  while  in  other  cases  massive  perivascular 
infiltration  may  occur  (Fig.  20).  The  interstitial  infiltration  in  the 
anterior  nerve-roots  is  very  discrete  or  absent. 

This  process  of  radiculitis  has  not  been  mentioned  by  other  workers 
who  have  studied  the  disease  in  animals  infected  experimentally; 
nor  lias  the  process  of  infiltration  of  the  nerve-roots  been  described 
in  the  spontaneous  disease. 

(b)  The  Spinal  (Uinqlia.  The  most  intense  lesions  in  the  peripheral 
nervous  system  have  been  found  constantly  in  the  spinal  ganglia. 
The  process  of  intiltration  in  the  posterior  nerve-roots  becomes  more 
intense  as  they  enter  the  ganglion,  and  Ix'tweeii  the  nerve-fibres 
which  ])!iss  through  the  substance  of  the  ganglion  a  well-marked 
monomiclear  infiltration  is  seen.  In  the  rest  of  the  ganglion  tbe 
lesions  as  a  rule  are  very  intense. 

The  alterations  in  the  ganglion  and  tbe  various  elements  taking  part 
in  the  infiltrative  and  degenerative  i)rocesses  at  this  site  are  always 
the  same,  no  matter  rn»ni  wbiit  individual  case  or  from  what  region 


ENZOOTIC  ENCEPHALO-MYELITIS  61 

of  the  cord  (cervical,  thoracic,  or  lumbar)  the  ganglion  is  taken 
(Figs.  21,  22).  The  changes  are  as  great  and  the  lesions  of  the  same 
importance  when  the  ganghon  originates  from  a  case  showing  altera- 
tions in  the  cord  which  are  scarcely  discernible  as  from  a  case  where 
such  changes  are  very  pronounced. 

The  capsule  of  the  ganglion  shows  neither  infiltration  nor  degenera- 
tion. In  the  interior  of  the  ganglion  mononuclear  interstitial  infiltra- 
tion is  abundant.  The  small  intraganghonic  vessels  show  perivascular 
'cuffing".  The  infiltrating  mononuclear  elements  are  found  dis- 
seminated between  the  nerve-cells  or  massed  together  forming  actual 
nodules  comparable  with  those  described  by  Van  Gehuchten  and  Nehs 
(1900)  in  rabies  (Fig.  23).  The  mononuclear  cells  may  be  grouped 
together  in  small  islands  between  the  nerve  fascicuh  which  traverse 
the  gangUon.  The  ganghon  cells  themselves  appear  to  be  'choked'  by 
the  infiltrative  process  in  some  microscopic  fields.  In  certain  parts 
these  cells  undergo  profound  changes:  the  nucleus  becomes  oxj'phiHc, 
the  protoplasm  loses  its  granular  nature,  assuming  a  homogeneous 
appearance,  and  becomes  shghtly  oxyphihc  when  stained  with 
toluidin  blue  and  eosin.  We  have  found  that  the  changes  in  the  cells 
of  the  ganglion  are  more  marked  than  in  any  other  region  of  the 
nervous  system;  and  the  intranuclear  corpuscles  are  larger  and  in 
greater  number  here  than  in  any  other  site.  In  some  microscopic 
fields  the  nucleus  of  every  cell  may  contain  one  or  two  corpuscles  of 
Joest-Degen  surrounded  by  a  halo. 

The  most  important  and  frequent  type  of  lesion  in  the  ganghon, 
however,  is  neuronophagia.  Lymphocytes,  plasma  cells,  and  large 
mononuclears  penetrate  the  peripheral  zone  of  the  neuron.  One  often 
finds  a  clear  zone  in  the  protoplasm  around  these  infiltrating  cells 
suggestive  of  the  action  of  a  proteolytic  ferment  hberated  by  the 
invading  cells.  Later  the  mass  of  detritus  of  the  neuron  is  removed 
by  the  macrophages  aided  by  occasional  polymorphonuclearleu- 
cocytes.  The  number  of  infiltrating  cells  increases,  the  whole  body 
of  the  nerve-cell  being  invaded,  and  finally,  in  place  of  the  neuron, 
one  finds  nothing  but  a  nodule  formed  by  mononuclear  cells  (Figs. 
23,  24,  25,  and  26).  The  most  intense  lesions  of  both  an  infiltrative 
and  degenerative  character  are  found  in  the  peripheral  zone  of  the 
ganghon;  this  point  will  be  discussed  again  later. 

(c)  The  Peripheral  Nerves.  In  the  peripheral  nerves  infiltrative 
lesions  are  also  fomid.  A  detailed  study  has  been  made  of  lesions  found 
in  the  sciatic  and  brachial  nerves.  The  technique  employed  in  carry- 
ing out  this  research  was  as  follows : 

All  the  rabbits  of  which  the  sciatic  and  brachial  nerves  were  sec- 
tioned for  histological  examination  had  been  inoculated  intracere- 
brally  with  the  virus  of  Borna  disease.  We  removed  the  terminal 
part  of  the  cord  (sacral)  with  the  roots  of  the  sciatic  nerve  and  their 
various  gangha  together  with  a  portion  of  the  peripheral  parts  of  the 
nerves.  This  whole  was  fixed  in  Duboscq-Brasil-Bouin  fluid.  Longi- 
tudinal sections  were  made  after  the  manner  figured  (Fig.  No.  27). 

We  have  found  lesions  in  all  cases  examined,  these  being  more 


62  BOEXA  DISEASE  AND 

intense  towards  the  origin  of  the  nerve  and  becoming  less  intense 
towards  its  termination.  The  alterations  consist  of  interstitial  or  peri- 
vascular intiltrations  with  mononuclear  cells.  The  nerve-sheath  is. 
as  a  rule,  unaffected.  In  certain  cases  the  infiltrations  appear  to 
'dissect'  the  nerve  filaments  (Fig.  28).  The  whole  process  constitutes 
a  descending  neuritis  produced  by  the  virus  propagatuig  centri- 
fugally.^ 

Kecently  G.  Marinesco  and  S.  Draganesco  (1927)  pubUshed  their 
observations  on  the  pathogenic  process  in  herpes  zoster.  A  complete 
chnical  report  is  given  of  cases  in  wliich  the  locahzation  of  the  lesions 
in  the  nervous  system  suggested  to  the  authors  that  the  infection 
commenced  by  an  ascending  neuritis  followed  by  a  gangho-radicuUtis 
and  myelitis.  'Wohlwill  (1924).  Levaditi  (1926),  "Pette  (1924),  Foerster 
(1924),  and  others  advanced  similar  hypotheses  as  to  the  centripetal 
propagation  of  the  infection.  In  support  of  their  theory  as  to  virus 
ascending  from  the  peripheral  nerves.  Marinesco  and  Draganesco 
refer  to  the  lesions  in  the  corresponding  gangha:  '  In  the  gangUon  the 
most  intense  lesions  were  in  the  peripheral  zone  . . .  this  topography  of 
the  inflammation  explains  the  spread  of  the  infection  hv  the  peri- 
capsular  lymph  vessels,  to  the  interior  of  the  ganghon." 

However,  from  a  comparison  of  the  description  of  the  lesions  pro- 
duced by  the  downward  extension  of  the  virus  in  rabbits  infected 
with  the  virus  of  Borna  disease,  and  those  in  herpes  zoster,  it  will 
be  seen  that  the  histological  pictures  are  identical.  We  have  found 
lesions  in  the  peripheral  nerve  not  only  close  to  the  ganglion  as  de- 
scribed by  Marinesco  in  zoster,  but  also  in  the  terminal  filaments 
farthest  removed  from  the  ganglion,  showing  that  the  virus  in  our 
experiments  diffused  by  centrifugal  propagation.  These  facts  allow 
us  to  assume  that  the  topography  of  the  lesions  is  not  a  criterion  by 
which  to  judge  the  portal  of  entry  of  the  virus  with  a  suflicient  degree 
of  accuracy.  Comparable  lesions  can  be  produced  in  the  central  and 
peripheral  nervous  system  both  by  infection  intracerebrally  or  by 
inoculation  of  the  virus  into  the  sciatic,  i.e.  no  matter  whether  the 
infection  is  ascending  or  descending. 

This  example  of  lesions  being  produced  in  the  peripheral  nervous 
system  after  introduction  of  the  virus  into  the  central  nervous  system 
(brain),  suggests  the  possibiUty  of  infection  being  central  in  origin 
in  the  case  also  of  herpes  zoster.  \  similar  pathogenic  process  is  not 
excluded  in  recurrent  herpes,  peripheral  forms  of  ei)idemic  encepiia- 
iitis,  and  perhaps  also  in  certain  cases  of  sciatica. 

(4)  Sumnuiry  and  Discussion. 

The  inoculation  of  the  virus  of  enzootic  encephalo-niyelitis  intra- 
cerebrally into  rabbits  produces  changes  in  the  nervous  system,  which 
are  those  of  a  meningo-encephalo-myelitis,  a  ganglio-radiculitis.  and 
a  peripheral  interstitial  neuritis. 

The  lesions  in  the  central  nervous  system  as  well  as  in  the  spinal 

■  We  h»ve  ticf-n  able  to  demonittratc  tlio  prcsoiirc  <if  viru«  in  the  pcriplirriil  m>rvc« 
by  inuculatidii  of  their  emuliiionii  into  the  hrnin  of  ralibit«  (Reo  ]i.  40). 


ENZOOTIC  ENCEPHALO-MYELITIS  63 

ganglia  are  both  infiltrative  and  degenerative.  The  meningitis  and  the 
perivascular  and  parenchymatous  infiltrations  are  produced  by  mono- 
nuclear cells. 

Pathological  'satellitism'  of  the  neuron  is  most  pronounced  in  the 
mesencephalon,  medulla  oblongata,  and  spinal  ganglia,  but  may  be 
found  also  in  other  regions  of  the  cord  and  brain.  It  may  in  some 
cases  be  so  advanced  as  to  constitute  true  neuronophagia.  The  latter 
phenomenon  is  most  common  in  the  paravertebral  ganglia. 

The  intranuclear  corpuscles  of  Joest-Degen,  considered  by  the 
present  writers  to  be  evidence  of  an  attempt  at  defence  by  the  nerve- 
cell,  and  referred  to  as  specific  '  inclusions '  by  other  workers,  may  be 
found  in  the  various  regions  of  the  brain,  cord,  and  spinal  gangha. 
They  are  almost  constantly  present  in  the  large  ganglion  cells  of  the 
Cornu  Anmionis  and  the  nerve-cells  in  the  spinal  ganglia.  In  our 
opinion  the  cell  which  reacts  against  the  presence  of  the  virus  by 
the  formation  of  intranuclear  corpuscles  has  formed  a  barrier  to  the 
extension  of  the  destructive  action  of  the  virus  in  'blocking' 
the  infective  elements  within  a  condensation  of  its  chromatin.  It  is 
feasible  to  conceive  that  this  process  removes  the  virus  and  renders  it 
inoffensive:  for  this  reason  the  cell  maintains  its  integrity.  In  those 
cases  where  the  cell  becomes  degenerated  or  neuronophagia  takes 
place,  one  may  suppose  that  the  nucleus  has  been  incapable  of  sur- 
rounding the  infective  particles  by  condensation  of  its  chromatin 
and  thus  limiting  the  extension  of  the  activity  of  the  virus.  This 
failure  to  form  intranuclear  corpuscles  may  be  due  to  the  quality  of 
the  virus  (virulence),  the  quantity  of  the  virus,  or  the  deficiency  in 
the  normal  resisting  power  of  the  neuron,  the  result  being  that  the 
virus  multiphes  and  ultimately  destroys  the  cell.  One  must  recall 
that  the  figure  described  on  p.  56  supports  this  view  as  to  the  method 
of  production  of  the  corpuscles  of  Joest-Degen. 

The  infiltrations  in  the  nerves  are  interstitial  in  character  and  are 
produced  by  the  invading  mononuclear  cells  arranging  themselves  in 
chains  between  the  nerve  filaments.  Perivascular '  cuffing'  also  occurs. 

A  peripheral  interstitial  neuritis  occurs  in  Borna  disease  after  the 
introduction  of  the  virus  into  the  brain,  and  the  authors,  without 
excluding  the  possibility  of  ascending  infections,  have  suggested 
that  herpes  zoster, ^  recurrent  herpes,  the  peripheral  forms  of  epidemic 
encephalitis,  and  perhaps  also  certain  forms  of  sciatica  may  be  the 
secondary  manifestations  of  a  disease,  the  original  focus  of  which  is 
in  the  central  nervous  system. 

The  hypothesis  has  already  been  advanced  (see  p.  88)  that  in  cases 
of  infections  with  these  viruses  central  infection  of  the  brain  takes 
place;  but  the  central  nervous  system,  being  able  to  resist  the  action 
of  the  virus  more  efficaciously,  shows  no  manifest  disturbances,  while 
the  peripheral  nerves,  poor  in  methods  of  defence,  do  not  rid  them- 
selves of  the  infecting  elements  which  proUferate  and  produce  lesions. 

'  The  work  of  Head  and  Campbell  (1900)  on  the  pathology  of  herpes  zoster  also 
suggests  that  zona  is  a  secondary  peripheral  manifestation  of  a  disease  originating 
in  the  central  nervous  system. 


64  BOENA  DISEASE  AND 

B.   The  Guinea-pig. 

Macroscopic  Examination.  The  brain  and  spinal  cord  appeared 
congested.  No  other  organ  showed  pathological  changes  except  the 
stomach,  in  which  occasionally  the  non-specific  lenticular  haemor- 
rhagic  areas,  similar  to  those  described  in  the  rabbit,  were  found. 

Microscopic  Examination.  Lesions  were  fomiil  in  the  central  and 
peripheral  nervous  sytem  and  in  the  kidney.  The  alterations  in  the 
stomach  wall,  when  they  existed,  were  comparable  with  those  found 
in  the  rabbit,  namely,  autodigestion  of  the  mucosa  with  small 
loeahzed  haemorrhage.  As  in  the  case  of  the  rabbit,  the  kidneys 
showed  marked  congestion,  but  no  actual  nephiitis  was  recorded. 

Central  Nerrous  System.  The  lesions  found  in  the  brain  were  similar 
to  those  in  the  rabbit,  except  that  their  intensity  was  less.  The  intra- 
nuclear corpuscles  of  Joest-Degen  were  usually  found  in  the  Cornu 
Ammonis  and  elsewhere  (PI.  I,  i"ig.  4).  Infiltrative  and  degenerative 
lesions  characteristic  of  the  disease  were  found  (PI.  Ill,  Fig.  8). 

The  lesions  in  the  mesencephalon,  cerebellum,  and  spinal  cord  were 
as  in  the  rabbit.  The  intranuclear  corpuscles  of  Joest-Degen  were 
frequenth'  present  in  the  anterior  horn  of  the  spinal  cord. 

Peripheral  Nervous  System.  Infiltrative  processes  with  mono- 
imclear  elements  were  found  in  the  nerve-roots,  but  they  were  not 
so  marked  as  in  the  rabbit.  The  lesions  in  the  spinal  ganglia  were 
similar  in  nature  to  those  described  in  the  rabbit,  but  were  not  so 
acut<».  The  lesions  in  the  peripheral  nerves  were  more  intense  in  that 
portion  nearest  the  ganglia. 

C.   The  Rat  and  Mouse. 

The  organs,  except  the  nervous  system  and  the  kidney,  were 
macroscopically  and  microscopically  normal.  In  the  brain  and  spinal 
cord  the  lesions  found  were  similar  to  those  in  the  rabbit  and  the 
guinea-pig.  The  intranuclear  corpuscles  of  Joest-Degen  were  found 
in  the  brain  of  rats  and  mice.  Infiltrative  lesions  of  a  discrete  nature 
were  also  found  in  the  sciatic  nerve  of  the  rat. 

D.    The  Monkey  (Macacus  rhesus). 

Our  description  of  the  lesions  found  in  the  monkey  is  made  from 
a  study  of  sections  from  different  parts  of  the  nervous  system  of 
Monkey  M.  1  {Macaciui  rhrsu.<>).  The  protocol  of  the  experiment  in 
which  this  animal  was  infected  is  given  on  p.  84.  Similar  lesions  were 
found  in  Monkey  M.  8.  although  the  clinical  pictun*  in  the  case  of  the 
latter  monkey  was  diffrrent  from  that  of  M.  1. 

The  macroscopical  and  microscopical  examination  of  the  spleen, 
liver,  pancreas,  lung,  myocardiuui,  testicle,  ovary,  parotid  gland,  and 
the  mesenteric  and  inguinal  lymph  glands  did  not  reveal  any  patho- 
logical changes  in  these  organs;  the  kidneys  were  iiyperaemic.  In 
the  adrenal  gland  there  was  slight  infiltration  with  lymphocytes  in 
the   medullary   zone,   the   lymphocytes   being  disseminated   in   tlie 


ENZOOTIC  ENCEPHALO-MYELITIS  65 

parenchyma  or  grouped  together  in  small  islands.  The  brain  appeared 
to  be  normal  by  naked-eye  examination. 

(1)  Lesions  in  the  Central  Nervous  System. 

(a)  The  Brain.  Frontal  Lobe.  Meningitis  of  a  mild  character  was 
present  in  some  areas,  becoming  intensified  near  those  vessels  in 
which  slight  perivascular  infiltrations  were  observed.  The  patho- 
logical process  in  the  meninges  might  be  described  as  an  'irritation' 
rather  than  a  true  meningitis,  while  the  perivascular  'cuffing'  con- 
sisted of  three  or  four  layers  of  cells  only  in  the  walls  of  the  vessels  in 
contact  with  the  brain ;  in  the  rest  of  the  vessels  the  process  of  infiltra- 
tion hardly  existed.  Certain  of  the  small  vessels  penetrating  the 
cerebral  parenchyma  from  the  meninges  were  surrounded  by  charac- 
teristic '  cuffing '.  There  was  infiltration  with  mononuclear  cells  in  the 
septum.  The  lesions  in  the  meninges,  the  'cuffing'  (Fig.  30),  and  the 
infiltration  of  the  septum,  were  produced  by  lymphocytes,  plasma 
cells,  and  macrophages  exclusively.  In  the  parenchyma,  and  especially 
in  the  white  matter,  extensive  'cuffing'  of  the  vessels  could  be  seen, 
consisting  of  ten  to  twenty  layers  of  infiltrative  cells.  Some  of  the 
pyramidal  cells  appeared  to  be  degenerated.  Intense  satelhtism  of  the 
neuron  was  present  in  some  instances,  and  in  certain  of  the  cells 
oxyphilic  corpuscles  surrounded  by  a  halo  of  the  type  described  by 
Joest  and  Degen  were  found  (PI.  Ill,  Fig.  1).  The  karyoplasm  was 
rarefied  in  the  greater  number  of  the  cells  containing  'inclusions', 
suggesting  that  the  degenerated  chromatin  was  condensed  in  the 
corpuscles.  In  the  deeper  part  of  the  brain,  both  satelhtism  and  the 
intranuclear  corpuscles  of  Joest- Degen  were  less  frequent  than  in 
the  peripheral  zone. 

Parietal  Lobe.  The  meninges  were  infiltrated  with  mononuclear 
cells  (Fig.  29)  which  formed  plaques  in  certain  regions.  There  was 
discrete  infiltration  of  the  septum.  In  the  brain  substance,  peri- 
vascular infiltrations,  consisting  of  many  layers  of  cells,  were  found 
forming  small  nodules:  that  these  were  perivascular  was  evidenced 
by  the  presence  of  a  small  vessel  in  the  centre  of  the  nodule  (Fig.  31). 
No  neuronophagia  was  recorded  in  this  region  of  the  brain,  although 
the  acute  '  satelhtism '  of  the  neuron  sometimes  suggested  the  pheno- 
menon. Occasional  nerve-cells  in  a  state  of  degeneration  had  eccentric 
nuclei  and  their  protoplasm  was  undergoing  tigrolysis.  No  actual 
parenchymatous  infiltration  could  be  seen,  but  rare  mononuclear 
cells  were  dispersed  in  the  parenchyma.  A  large  number  of  nerve- 
cells  in  this  region  contained  large  oxyphihc  corpuscles  surrounded  by 
halos  within  the  nuclei. 

Occipital  Lobe.  Meningitis  was  rarely  observed  in  this  region  of  the 
brain.  Certain  of  the  meningeal  vessels  had  several  layers  of  mono- 
nuclear cells  on  their  walls  in  contact  with  the  brain.  In  the  septum, 
infiltration  was  not  well  marked,  although  several  venules  were  sur- 
rounded by  'cuffing'.  Eich  perivascular  infiltrations  were  found  in 
the  parenchyma,  more  especially  near  the  large  pyramidal  cells. 
Pathological  'satelhtism'  of  the  neuron  by  neuroghal  elements  and 


66  BOEXA  DISEASE  AND 

occasional  lymphocA'tes  was  not  uncommon  in  the  occipital  lobe: 
these  satelhte  cells  sometimes  produced  marked  depressions  in  the 
protoplasm  of  the  host  cell.  The  intranuclear  corpuscles  were  of  very 
much  smaller  dimensions  and  were  observed  less  frequently  than  in 
the  parietal  lobe. 

Hippocampus.  The  lesions  in  the  meninges  were  similar  to  those 
found  in  the  occipital  lobe.  The  perivascular  'cuffing'  in  the  paren- 
chyma was  poor  in  elements.  There  was  a  slight  infiltration  with 
mononuclear  cells  between  the  large  and  small  pyramidal  cells.  The 
number  of  'inclusions'  was  greater  in  this  region  than  in  the  occi- 
pital lobe. 

Cornu  Ammonis.  In  this  region  of  the  brain  the '  cuffing '  around  the 
vessels  was  so  extensive  as  to  suggest  a  nodule  or  pseudo-gumma :  the 
presence  of  a  small  vessel  in  the  centre  of  the  nodule  was  discerned 
with  difficulty.  In  addition,  there  were  small  groups  of  mononuclear 
cells  bearing  no  relation  to  vessels.  In  certain  cases  'satelhtism' 
was  so  advanced  that  it  could  almost  be  described  as  neuronophagia. 
The  large  ganghon  cells  preserved  their  normal  structxire  and  nearly 
all  contained  within  their  nucleus  well-marked  corpuscles  of  the  Joest- 
Degen  type. 

Basal  Ganglia.  Perivascular  '  cuffing '  was  observed.  In  some  of  the 
degenerated  nerve-cells  the  nucleus  could  not  be  differentiated  from 
the  rest  of  the  protoplasm.  Pathological  'satellitism'  of  the  nerve- 
cells  was  intense.  Intranuclear  corpuscles  were  found  on  occasion, 
not  only  in  the  nucleus  but  even  in  the  cytoplasm.  Small  islets  of 
mononuclear  cells  were  also  found. 

Pons.  The  process  of  perivascular  infiltration  was  abundantly 
present  in  tliis  region. 

Many  of  the  ner\e-cells  were  in  an  advanced  stage  of  degeneration, 
while  the  nuclei  were  swollen  and  unrecognizable  as  such.  The  proto- 
plasm of  the  cells  was  in  a  state  of  tigrolysis  and  its  contour  was 
broken  in  many  places  by  splitting.  The  cells  containing  intranuclear 
corpuscles  were  othenvise  morphologically  normal.  No  areas  of 
infiltration  were  seen. 

(b)  Cerebellum.  There  was  slight  infiltration  of  the  meninges  and 
septum  with  mononuclear  cells:  tiie  parenchyma  was  not  infiltrated. 
The  layer  of  the  small  granular  cells  had  a  normal  aspect.  CiTtain  of 
the  cells  of  Purkinje  were  degenerated.  In  some  of  the  latter  cells 
oxyphihc  corpuscles  with  a  characteristic  halo  were  recorded. 

(c)  Medulla  Oblongata.  Meningeal  changes  were  slight.  Vessels  were 
normal  without  'cuffing'  and  the  parenchyma  was  not  infiltrated. 
Occasional  neurons  showed  evidence  of  commencing  degeneration, 
and  in  certain  of  these  large  oxyphilic  corjjuscles  were  present. 

(dj  Spinal  Cord.  Cervical  liegion.  I'he  meninges  were  normal  and 
no  'cuffing'  was  present  in  them  or  in  the  grey  or  white  matter.  A 
slight  diffuse  inliltration  was  ol)served.  In  the  anterior  horns  non- 
degenerated  neurons  had  intranuclear  corpuscles  (Joest-Degen  type). 
In  certain  regions  satellitism  was  a  marked  feature.  The  zone  of  the 
cells  of  Lissauer  was  infiltrated   with   lympliocytes.    The  posterior 


ENZOOTIC  EXCEPHALO-ifYELITIS  67 

tiCTTe-roofe  showed  interstitial  infiltration  and  sometimes  even 'cuffing' 
in  the  vessels.  The  corresponding  s-pinal  ganglia  showed  intense  inter- 
stitial infiltrations:  only  rare  polymorphonuclear  leucocytes  were 
found.  The  protoplasm  of  certain  of  the  cells  in  the  ganglia  had 
undergone  tigrolysis,  and  in  some  cases  also  had  become  oxyphiKc; 
the  nucleus  of  these  affected  cells  was  eccentric.  Neuronophagia 
was  frequently  seen  and  nodules  of  mononuclear  cells  comparable 
with  those  found  in  rabies  were  not  uncommon.  Large  intranuclear 
corpuscles  were  found  in  nerve-cells  of  the  gangUa  which  otherwise 
preserved  their  morphological  integrity  (Fig.  35). 

Thoracic  Begion.  No  meningitis  was  present,  nor  was  there  any 
evidence  of  'cuffing'.  There  was  sUght  infiltration  of  the  posterior 
horns  and  a  concomitant  degeneration  of  certain  nerve-cells  of  the 
anterior  horns.  In  the  latter  region  also  rare  intranuclear  corpuscles 
were  found  (PI.  II,  Fig.  2).  The  posterior  nerve-roots  showed  inter- 
stitial infiltration  and  '  cuffing '  around  the  vessels. 

Lumbar  Region.  No  perivascular  infiltrations  and  no  meningitis 
could  be  seen.  There  was  a  slight  diffuse  parenchymatous  infiltration 
with  lymphocytes.  Satelhtism  of  the  neuron  was  present,  both  in 
the  anterior  and  the  posterior  horns.  There  were  well-marked  lesions 
in  the  lateral  honis:  the  protoplasm  was  markedly  oxyphilic  and  the 
nuclei  of  the  cells  were  eccentric.  Sometimes  the  sateUite  cells  pene- 
trated the  protoplasm  of  the  degenerated  neurons  and  constituted 
almost  true  neuronophagia.  Certain  small  nerve-cells  of  the  anterior 
bom  showed  neuronophagia.  Intranuclear  corpuscles  were  rare.  Inter- 
stitial infiltrations  and  'cuffings'  were  seen  in  the  posterior  nerve- 
roots  (Fig.  34). 

In  the  spinal  gangha  the  lesions  were  of  a  similar  nature  to  those 
encountered  in  the  upper  regions  of  the  cord,  but  were  more  intense 
(Fig.  86).  Neuronophagia  was  a  constant  feature.  The  nuclei  of  cells 
containing  intranuclear  corpuscles  of  the  Joest-Degen  type  showed 
a  rarefication  of  the  karyoplasm,  but  no  other  morphological  changes 
were  present  in  such  cells  (PL  III,  Fig.  5). 

Sacro-caudal  Region.  In  the  large  nerv^e  fasciculi  rich  infiltration 
with  mononuclear  cells  could  be  seen,  together  with  marked  'cuffing' 
round  the  vessels  (Fig.  33). 

In  the  posterior  roots  of  the  ner^^es  the  interstitial  infiltration  and 
'cuffing'  round  the  vessels  was  rich. 

The  spinal  gangha  showed  similar  lesions  to  those  recorded  in  the 
lumbar  region.  The  capsules  of  such  gangha  were  unaffected,  and  the 
lesions  were  marked  in  the  periphery  of  the  gangUon.  In  the  nerves, 
after  their  exit  from  the  ganghon,  lesions  similar  to  those  seen  in  the 
nerve-roots  were  found.  Massive  infiltration  with  mononuclear  cells 
was  seen  between  the  fascicuh  of  the  nerves  (epmeurium),  in  the 
connective  tissue  around  the  nerve  (perineurium),  and  even  in  the 
endoneurium.  '  Cuffing'  of  the  vessels  was  also  seen. 

(e)  Sciatic  Nerve.  A  study  of  transverse  sections  from  the  sciatic 
ner\"e  after  its  exit  from  the  greater  sciatic  foramen  showed  that  there 
was  a  diffuse  infiltration  between  the  ner\'e  fascicuh;  this  was 
E  2 


68  BORNA  DISEASE  AND 

generally  more  pronounced  in  the  interior  of  the  fascicuh.  In  the 
thickness  of  the  nerve  an  intense  perivascular  infiltration  was  seen 
(Figs.  38  and  39).  Often  the  connective  tissue  of  the  sheath  was 
unchanged,  while  intense  infiltrative  lesions  were  seen  in  the  thick- 
ness of  the  nerve. 

Examination  of  longitudinal  sections  made  from  the  sciatic  nerve 
half-way  between  the  greater  sciatic  foramen  and  the  pophteal  region 
showed  that  interstitial  infiltration  with  mononuclears  existed  along 
with  massive  perivascular  'cuffing',  while  the  nerve-sheath  itself 
appeared  perfectly  normal. 

(f)  Brachial  Nerve.  The  alterations  seen  in  tliis  nerve  were  analogous 
to  those  described  in  the  sciatic  nerve;  even  in  the  lower  third  of  the 
fore  limb  intense  lesions  were  found  (Fig.  37). 

The  infiltrative  lesions  of  the  nerve-roots  and  peripheral  parts  of 
the  nerves,  as  well  as  the  alterations  in  the  paravertebral  ganglia,  show 
that  in  the  monkey,  as  in  the  rabbit  infected  intracerebrally  with  the 
virus,  the  infective  agent  travels  from  the  central  nervous  system  to 
the  periphery  along  the  nerves,  producing  the  lesions  described  above, 
which  constitute  a  ganglio-radicuhtis  and  descendiny  peripheral  inter' 
stitial  neuritis. 

(2)  Summary. 

In  the  brain  there  existed  a  mild  meningitis  with  perivascular 
infiltrations  and  diffuse  parenchymatous  infiltrations,  which  some- 
times formed  actual  nodules;  massive  perivascular  infiltration  was 
also  seen  in  the  parenchyma;  there  was  degeneration  of  nerve-cells 
and  satelhtism.  Intranuclear  corpuscles  of  the  type  described  by 
Joest  and  Degen  were  more  easily  found  and  more  numerous  than  in 
the  rabbit. 

In  the  cerebellwn  there  was  found  a  slight  meningeal  reaction  with 
perivascular  'cuffing',  satelhtism  of  the  cells  of  Purkinje  in  certain 
areas,  and  rare  oxyphilic  corpuscles  (Joest-Degen  type)  in  the  nuclei 
were  recorded. 

In  the  spinal  cord  no  meningitis  was  recorded  and  there  was  an 
absence  of  'cuffing'  in  the  vessels  of  the  coverings  of  the  cord.  In  the 
posterior  horn,  a  diffuse  infiltration  was  observed,  this  being  more 
marked  in  the  lumbar  region.  In  this  region  of  the  cord  also  a  similar 
infiltration  was  seen  in  the  anterior  and  lateral  horns.  In  the  anterior 
horns  degeneration  in  the  nerve-cells  was  recorded,  and  sometimes 
ncuronophagia. 

In  the  peripheral  werroMS  system  an  infiltrative  radiculatis  existed. 

The  spinal  ganglia  were  intensely  affected,  showing  lesions  both 
infiltrative  and  di'generative;  neuronophagia  was  a  connnon  feature. 

A  peripheral  neuritis  {sciatic  and  lirachial  nerves)  was  present,  con- 
sisting of  interstitial  and  jicrivascular  infiltration. 

The  topography  of  the  lesions  as  a  whole  shows  that  the  virus  intro- 
duced into  the  brain  produces  lesions  locally,  spreads  to  the  rest  of 
the  central  nervous  system,  and  finally  travels  down  the  peripheral 
nerves.   The  cells  in  tlie  lesions  are  almost  exclusively  nionoiiuclears. 


ENZOOTIC  ENCEPH.\LO-MYELITIS  69 

11.  IMMUNITY 

All  authors  agree  that  in  the  horse  an  attack  of  enzootic  encephalo- 
myehtis  contracted  spontaneously  does  not  render  this  animal  im- 
mune to  a  second  attack;  with  regard  to  the  disease  in  cattle  and 
sheep,  no  precise  records  are  available  as  to  this  point.  The  rabbit 
appears  to  behave  differently.  Zwick  (1926)  and  his  collaborators 
obser%'ed  that  in  one  case  a  rabbit  which  had  been  infected  experi- 
mentally and  had  shown  symptoms  typical  of  the  disease  ultimately 
recovered  and  resisted  a  second  intracerebral  inoculation.  The  same 
authors  succeeded,  although  not  constantly,  in  producing  a  solid 
immunity  by  repeated  inoculation  of  virus  either  subcutaneously  or 
intravenously.  They  showed  hkewise  that  the  introduction  of  a  large 
quantity  of  virus  intraperitoneally  may  render  rabbits  refractory  to 
subsequent  infection.  Ernst  and  Hahn  (1926)  have  shown  that  the 
inoculation  of  virus  intracerebrally  into  rabbits  does  not  lead  to  the 
development  of  the  disease,  if,  during  the  period  of  incubation,  such 
animals  receive  repeated  inoculations  of  virulent  material  into  the 
veins. 

Zwick  and  his  collaborators  (1926),  in  the  few  experiments  which 
they  record  in  attempts  to  demonstrate  antibodies  in  the  serum  of 
immunized  animals  by  neutraUzation  of  virus  in  vitro,  did  not  obtain 
very  conclusive  results. 

In  passaging  the  strains  of  virus  with  which  we  have  been  working, 
viz.  a  strain  originating  from  horses  (Zmck)  and  a  strain  originating 
from  sheep  (Miessner),  we  have  never  found  any  healthy  iminoculated 
rabbit  refractory  to  infection. 

I.  Attempts  at  Conferring  Immunity  to  Rabbits  by 
Inoculations  of  Non- attenuated  Virus. 

A.  Intravenous  Inoculatian. 

(1)  Four  rabbits  were  inoculated  intravenously  with  1-5  corns,  of 
a  virulent  centrifugahzed  emulsion,  which,  M^hen  inoculated  into  the 
brain  of  a  rabbit,  produced  the  disease  and  death  at  the  end  of  39 
days.  The  rabbits  inoculated  intravenously  did  not  show  any 
symptoms  and  gained  weight.  Between  91  and  206  days  later  they 
were  inoculated  intracerebrally  with  fresh  passage  virus.  Table  X. 
below  shows  that  a  single  intravenous  inoculation  of  virulent  emul- 
sion does  not  produce  immunity  in  rabbits. 

(2)  Another  rabbit.  No.  69,  weight  1,980  gms.,  received  three  inocu- 
lations of  virulent  material  into  the  veins. 

16.5.27.  First  intravenous  inoculation  of  3  c.cms.  of  centrifugalized  emulsion. 

21.5.27.  Animal  normal.   Weight  1,780  gms. 

29.5.27.  Animal  normal.   Weight  1,640  gms. 

30.5.27.  Second  intravenous  inoculation  of  5  c.cms.  of  virulent  emulsion. 

8.6.27.  Animal  normal.   Weight  1,600  gms. 

13.6.27.  Third  intravenous  inoculation  of  1-5  c.cms.  of  virulent  emulsion. 

14.6.27.  Animal  normal.   Weight  1,680  gms. 

22.6.27.  Animal  normal.   Weight  1,670  gms. 


70 


BOEXA  DISEASE  AND 


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ENZOOTIC  ENCEPHALO-MYELITIS  71 

On  23.6.27  the  rabbit  received  a  test  inoculation  of  virulent  material 
at  the  same  time  as  a  control.  The  control  died  after  34  days  with 
symptoms  and  lesions  characteristic  of  the  disease  in  the  central 
nervous  system.  The  rabbit  prepared  by  intravenous  inoculations 
behaved  as  recorded  below. 

3f0.6.27.  Animal  normal.   Weight  1,7.50  gms. 

7.7.27.  Animal  normal.   Weight  1,950  gms. 

10.7.27.  Animal  normal.   Weight  1,680  gms. 

14.7.27.  Commencement  of  the  disease.   Weight  1,580  gms. 

23.7.27.  Typical  symptoms  of  the  disease.   Weight  1,400  gms.  ■• 

26.7.27.  Animal  died  during  the  day.   Weight  1,230  gms. 

Microscopic  examination  of  the  brain  and  spinal  cord  revealed  the 
presence  of  characteristic  lesions. 

In  this  case  three  intravenous  inoculations  did  not  lead  to  immunity 
to  cerebral  infection. 

That  four  intravenous  injections  of  virulent  material  may,  however, 
produce  a  soUd  immunity  is  shown  by  the  experiment  recorded  on 
p.  75,  Babbit  67. 

That  the  intracerebral  inoculation  of  a  filtrate  (Handler  filter)  of 
virulent  emulsion  does  not  lead  to  the  development  of  immunity  may 
be  concluded  from  the  experiments  mentioned  on  p.  13. 

The  effect  of  introducing  a  thick  emulsion  of  virulent  brain  into  the 
trachea  of  rabbits  was  tested,  but  it  did  not  produce  the  disease  nor 
immunity  in  animals  so  treated.  0-5  c.cm.  of  a  virulent  emulsion  of 
brain  was  introduced  mto  the  trachea  of  three  animals.  As  they  did 
not  present  any  morbid  symptoms  for  174  days  they  were  inoculated 
intracerebrally  with  a  virulent  emulsion.  They  all  died  from  enzootic 
encephalo-myehtis  after  37,  27,  and  40  days  respectively.   (See  p.  33.) 

Eabbits  which  survive  corneal  or  intratesticular  inoculations  of 
virulent  material  may  become  refractory  to  the  disease  as  tested  sub- 
sequently by  intracerebral  inoculation. 


B.  Corneal  Inoculation. 

The  experiment  recorded  on  p.  31  (Babbit  81s)  shows  that  infec- 
tion by  the  corneal  route  can  iminunize  the  rabbit  against  a  sub- 
sequent inoculation  by  the  cerebral  route. 


C.  Intratesticular  Inoculation. 

Some  measure  of  immunity  may  also  follow  intratesticular  inocu- 
lation. Two  rabbits  (273  and  289)  were  inoculated  into  the  right 
testicle  with  1  c.cm.  of  a  virulent  cerebral  emulsion.  One  of  these 
rabbits,  No.  289,  died  after  105  days.  Typical  lesions  were  found  in 
sections  of  the  various  parts  of  the  central  nervous  system,  and  pas- 
sage of  its  brain  to  a  fresh  rabbit  gave  a  positive  result. 

The  other  rabbit,  No.  273,  survived,  and  when  inoculated  by  the 


72  BOENA  DISEASE  AND 

intracerebral  route  112  days  later  it  proved  to  be  immune.  The 
control  rabbit  inoculated  by  the  intracerebral  route  died  on  the 
48th  day. 

II.   Attempts  at  Conferring  Immunity  to  Rabbits  by 
Inoculation  of  Attenuated  Virus.^ 

A.  Virus  Inactivated  by  Ether. 

Roux  ^  showed  the  attenuating  action  of  ether  on  the  virus  of 
rabies.  Later  Eemlinger  (1919)  used  an  ether-treated  virus  sub- 
cutaneously  to  produce  an  immunity  in  rabbits  to  intracerebral 
inoculation  ^vith  fixed  virus.  Alvisatos  (1922)  and  Hempt  (1925)  used 
ether-treated  virus  as  a  means  of  vaccinating  man  against  rabies. 

Marie  and  Mutermilch  (1927)  have  shown  that  one  can  immmiize 
rabbits  against  rabies  by  inoculating  intrathocally  virus  treated  with 
ether.  We  have  tried  to  immunize  rabbits  against  the  virus  of  ence- 
phalo-myeUtis,  employing  a  similar  technique,  with  the  difference  that 
the  virus  treated  with  ether  was  inoculated  into  the  brain;  the  proof 
of  the  avirulence  of  the  virus  treated  with  ether  and  the  technique 
employed  has  been  described  on  p.  22. 

The  results  obtained  are  given  below. 

Two  rabbits,  220a  and  217a,  weighing  2,450  and  1,950  gms. 
respectively,  received  two  intracerebral  inoculations  of  0-5  c.cm.  of 
ether-treated  vims  at  an  interval  of  5  days.  Twenty-one  daj'S  after 
the  last  inoculation  they  were  inoculated  intracerebrally  along  with 
a  control  rabbit,  14a,  with  fresh  virus.  Rabbit  217a  died  on  the 
42nd  day,  Rabbit  220a  on  the  17th  day.  and  control  Rabbit  14a  on 
the  30th  day  after  inoculation,  and  typical  microscopic  lesions  were 
found  in  the  central  nervous  system  of  all  three. 

Thus  two  successive  intracerebral  inoculations  into  rabbits  of 
virus  treated  with  ether  did  not  produce  immunity  to  subsequent 
intracerebral  inoculation. 

B,  Virus  killed  by  Chloroform. 

The  proof  that  virus  treated  with  chloroform  is  inactivated  has 
already  been  given  on  p.  21.  In  the  experiment  about  to  be  described, 
rabbits  were  twice  inoculated  intracerebrally  with  virus  treated  with 
chloroform,  the  interval  between  the  inoculations  being  5  days.  The 
animals  were  tested  3  weeks  after  the  second  inoculation,  and  since 
the  experiments  were  made  at  the  same  time  as  the  experiments  with 
ether-treated  virus,  the  control  Rabbit  14a  served  for  both. 

The  rabbits  prepared  with  virus  treated  with  chloroform  succumbed 

'  The  exporimrnU  in  attcmpU  to  immunize  with  virus  trontod  with  ether,  chloro- 
form, or  formalin  recorded  are  preliminary  and  for  orientation ;  it  is  possible  that 
modificationH  in  technique  might  change  the  results  originally  obtained.  There  are  at 
present  under  experiment  other  series  of  animals,  and  the  retiults  obtained  in  these 
■ulwrquent  attempts  will  lie  recorded  at  some  future  date. 

'  Koux's  unpublished  olwervations. 


ENZOOTIC  ENCEPHALO-MYELITIS  73 

to  a  subsequent  intracerebral  infection  after  37  and  42  days  respec- 
tively. Cultures  made  from  these  brains  were  negative,  but  lesions 
characteristic  of  infection  with  Borna  disease  were  found  on  micro- 
scopic examination  of  sections  of  brain,  cord,  and  spinal  ganglia. 

C.    Virus  treated  with  Formol. 

On  p.  22  it  was  shown  that  virus  treated  with  0-2  per  cent,  formalin 
for  1 8  hours  at  room  temperature  failed  to  infect  rabbits  by  the  intra- 
cerebral route. 

The  vaccine  was  prepared  the  day  before  use  by  subjecting  fresh 
virulent  brain  material  to  the  action  of  formahn  in  this  concentration. 
A  batch  of  ten  rabbits  was  prepared  by  subcutaneous  inoculation  of 
this  vaccine:  the  results  of  the  experiment  are  recorded  in  Table  XI. 
The  results  obtained  in  this  experiment  were  not  very  satisfactory.  Of 
the  ten  rabbits  prepared  by  inoculation  of  formolized  virus  two  died 
accidentally,  five  died  following  upon  infection  with  the  test  dose 
given  intracerebrally,  and  three  proved  to  be  immune  to  the  test  dose 
given  intracerebrally. 

D.    Virus  Inactivated  by  Ultra-violet  Light. 

An  experiment  on  this  subject  is  recorded  on  p.  20  in  which  a 
rabbit  which  survived  after  having  received  an  inoculation  of  virus 
exposed  for  5  minutes  to  the  action  of  rays  emitted  by  the  mercury 
arc  proved  to  be  still  susceptible  to  infection  by  the  intracerebral 
inoculation  of  virulent  material. 

III.   Search  foe  Virucidal  Antibodies  in  the  Serum  of 
Immunized  Animals. 

The  serum  of  the  immune  rabbit,  223a  (see  p.  75),  was  taken  and 
mixed  with  equal  parts  of  a  centrifugahzed  virulent  cerebral  emul- 
sion; at  the  same  time  two  further  mixtures  of  the  same  cerebral 
emulsion  were  made,  one  with  equal  parts  of  normal  rabbit  serum 
and  the  other  with  physiological  sahne.  The  three  mixtures  were 
kept  for  2  hours  at  37°  C.  and  were  subsequently  inoculated  intra- 
cerebrally into  rabbits. 

The  results  were:  (1)  the  rabbit  inoculated  with  the  mixture  of 
virus  and  immune  serum  died  on  the  14th  day  after  inoculation; 
its  brain  passaged  to  a  fresh  rabbit  killed  in  35  days;  the  lesions 
found  in  sections  of  the  brain  of  both  of  these  rabbits  were  those  of 
experimental  Borna  disease. 

(2)  The  control  rabbits  inoculated  with  the  mixture  of  normal 
rabbit  serum  and  virus  died  39  and  34  days  respectively  after 
inoculation  of  Borna  Disease,  the  lesions  found  on  microscopical 
examination  of  sections  of  the  nervous  system  being  typical. 

(3)  The  rabbits  inoculated  with  the  mixture  of  virus  and  physio- 
logical sahne  also  succumbed  to  the  inoculation. 


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BOENA   DISEASE  AND 


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ENZOOTIC  ENCEPHALO-MYELITIS  75 

IV.  Experiments  on  Cross  Immunity  between  the  Strain 
OF  Equine  and  that  of  Ovine  Origin. 

Our  experiments  have  been  carried  out  with  two  strains  of  enzootic 
encephalo-myehtis,  the  one  originating  from  the  horse  (Zwick's 
strain),  and  the  other  from  the  sheep  (Miessner's  strain). 

Experiment  1.  Rabbit  223a.  Weight  2,280  gms.  A  diluted  emulsion 
of  virulent  brain  (Zwick  strain)  which  had  been  pulped  and  preserved 
in  glycerine  at  room  temperature  for  several  weeks  was  inoculated 
into  the  brain  on  22.10.26.  The  animal  lost  weight  slightly,  developed 
paresis  of  the  hind  quarters  about  the  20th  day  after  the  infection,  but 
subsequently  recovered,  and  on  15.12.26,  54  days  after  the  inocula- 
tion, it  appeared  perfectly  normal  and  had  a  weight  of  2,400  gms.  On 
this  date  it  was  inoculated  intracerebrally  with  fresh  passage  virus 
(Zwick  strain)  and  survived,  while  the  control  rabbit  (Rabbit  68a, 
weighing  1,150  gms.)  inoculated  by  the  same  route  fell  ill  on  the  20th 
day  following  upon  infection,  and  died  on  the  25th  day.  Characteristic 
lesions  were  found  throughout  the  central  nervous  system  on  micro- 
scopic examination. 

Rabbit  223a  was  therefore  immune  against  infection  with  the  strain 
of  equine  origin.  On  the  12.5.27  (i.e.  202  days  after  the  first  inocula- 
tion, and  148  days  after  the  second  inoculation  with  the  equine  strain, 
it  was  inoculated  intracerebrally  with  the  strain  of  ovine  origin.  At 
the  same  time  a  control  rabbit  was  similarly  inoculated.  Rabbit  223a 
continued  in  health,  and  gained  in  weight.  The  control  showed  symp- 
toms of  Borna  disease  20  days  after  the  inoculation,  and  died  27  days 
after  the  injection.  The  usual  characteristic  lesions  were  found  in 
sections  of  the  central  nervous  system  of  this  control  rabbit,  and  a 
portion  of  the  brain  infected  another  rabbit. 

Experiment  2.  The  fact  that  repeated  inoculations  of  fresh  virus 
into  the  veins  of  rabbits  may  immunize  them  against  subsequent 
intracerebral  inoculation  has  already  been  referred  to  (see  p.  71). 
We  prepared  rabbits  by  vaccinating  in  this  way  with  a  virus  of  ovine 
origin.!  A  fresh  virulent  emulsion  of  brain  was  centrifugaKzed  for 
5  minutes.  The  supernatant  fluid  was  carefully  pipetted  off  and 
inoculated  in  the  marginal  vein  of  the  ear  of  rabbits,  care  being  taken 
that  the  injection  was  carried  out  very  slowly. 

Babbit  67.    Weight  1,580  gms. 
12.5.27.   First  injection  of  2  corns,  of  a  virulent  centrifugalized  emulsion  into  the 

vein. 
16.5.27.    Weight  1,600  gms.    Second  injection  of  3  corns,  of  a  virulent  oentri- 

fugaUzed  emulsion  into  the  vein. 
21.5.27.    Weight  1,680  gms.   Animal  normal. 
30.5.27.   Weight  1,700  gms.   Weight  normal. 
5.6.27.   Weight  1,800  gms.   Animal  normal. 

7.6.27.    Third  injectimi  of  5  o.cms.  of  a  virulent  centrifugalized  emulsion  into  the 
vein. 

'  Several  animals  were  prepared  in  the  same  way,  but  we  give  below  the  protocol 
of  the  rabbit  which  served  for  an  experiment  of  cross  immunity  between  the  equine  and 
ovine  strain. 


76  BORNA  DISEASE  AND 

8.6.27.   Weight  1,690  gms.  Animal  normal. 
13.6.27.   Weight  1,820  gms.    Fourth  injection  of  3  c.cms.  of  a  virulent  centri- 

fugalized  emulsion  into  the  vein. 
22.6.27.   Weight  1,870  gms.  Animal  normal. 

On  23.6.67  the  animal,  together  with  a  control  rabbit,  was 
inoculated  intracerebrally  with  an  emulsion  of  fresh  virus  of  equine 
origin. 

Rabbit  67.    Weight  1,870  gms. 
30.6.27.   Animal  nonnal.   Weight  1,980  gms. 
7.7.27 


l.l.i,!. 

14.7.27. 

2,200 

20.7.27. 

2,150 

25.7.27. 

2.100 

28.7.27. 

2,000 

15.8.27. 

2,100 

29.8.27. 

2,150 

Control. 
Rabbit  178ii.    Weight  1,770  gms. 
30.6.27.   Normal.   Weight  1,820  gms. 
7.7.27.  „  „       1,850    „ 

14.7.27.  „  „       1,800    „ 

18.7.27.   Commencement  of  the  disease.   Weight  1.650  gms. 
20.7.27.   Typical  svmptoms  of  the  disease.   Weight  1,540  gms. 
25.7.27.   Severely  ill.   Weight  1,300  gms. 
26.7.27.   Found  dead  33rd  day  after  inoculation. 
Microscopical  examination  of  sections  of  the  central  nervous  system — charac- 
teristic lesions. 

This  experiment  shows  that  the  virus  of  ovine  origin  immunizes 
against  that  of  equine  origin. 

Experiment  3.  A  rabbit  (81s)  which  had  resisted  inoculation  with 
virulent  virus  of  ovine  origin,  by  scarification  of  the  cornea,  was 
inoculated  intracerebrally  110  days  later  with  fresh  rabl)it  passage 
virus  of  equine  origin.  At  the  same  time  a  control  rabbit  ('25a)  was 
inoculated  intracerebrally  with  the  same  fresh  passage  virus. 
Rabbit  81s  remained  well,  while  the  control,  25a,  showed  typical 
symptoms  of  Borna  disease  32  days  after  inoculation,  and  died  on 
the  4Sth  day.  In  tliis  case  also  the  virus  of  ovine  origin  innnunized 
Rabbit  81s  against  the  pathogenic  action  of  the  virus  of  equine  origin 
introduced  into  the  brain. 

Conclusion.  From  the  above  experiments  the  following  conclusions 
can  be  drawn: 

(1)  Rabbits  which  have  become  resistant  to  the  virus  of  enzootic 
encephalo-myelitis  of  equine  origin  prove  also  to  be  refractory  to 
infection  by  intracerebral  inoculation  of  the  virus  of  ovine  origin; 
the  converse  is  also  true. 

(2)  Rabbits  which  have  become  immunized  against  enzootic 
encephalo-myelitis  keep  this  acquired  inimunity  for  at  least  148  days. 
If  the  immunity  is  reinforced  by  repeated  inoculations  it  may  last 
at  least  203  days. 


ENZOOTIC  EXCEPHALO-MYELITIS 


77 


V.  Experiments  on  Cross  Immunity  between  the  Virus  of 
Enzootic  Encephalo-myelitis  and  other  Viruses  of  the 
Filter-passing  Group. 

A.  Herpes} 

On  the  basis  of  the  following  experiment  we  arrived  at  the  conclu- 
sion pubhshed  as  a  preliminary  note  in  the  British  Journal  of  Experi- 
mental Pathology  (1927)  that  rabbits  immunized  against  the  virus  of 
herpes  are  still  susceptible  to  infection  with  the  virus  of  Borna 
disease. 

Four  large  rabbits,  111a,  183a,  185a,  and  187a  were  immunized 
against  herpes.  On  7.6.27  their  immunity  was  tested  by  the  intra- 
cerebral inoculation  of  fresh  herpetic  virus;  they  resisted  such 
infection,  while  the  control  rabbit,  No.  148a,  infected  by  the  same 
route,  died  on  the  6th  day.  Sixteen  days  later  the  four  surviving 
rabbits  were  inoculated  intracerebrally  \vith  the  virus  of  encephalo- 
myehtis,  together  with  a  control  rabbit.  No.  182a.  All  the  rabbits 
showed  symptoms  typical  of  experimental  Borna  disease  and  died ;  the 
results  are  recorded  below. 


Ho.  of  rabbit. 

Date  of  death. 

CvMure  of 
brain.        \  Lesions. 

lllA 

183a 

185  A 
187  a 
Control  Rabbit 
182a 

Died  on  the  40th  day  after  inoculation 
„      32nd 
„      31st 

„      32nd        

.,       40th 

Negative 

Typical. 

Conclusion.  No  cross  immunity  exists  between  herpes  and  enzootic 
encephalo-myehtis. 

B.  Rabies.^ 

On  9.9.27  two  rabbits  proved  to  have  been  immunized  against 
the  virus  of  Borna  disease  (Babbit  223a,  see  p.  75,  and  Rabbit  67, 
see  p.  75),  together  with,  as  a  control.  Babbit  41a,  were  inoculated 
intracerebrally  with  a  'street'  virus  of  rabies.  The  two  rabbits, 
223a  and  67,  which  were  immune  to  Borna  disease,  became  paralysed 
on  the  8th  day  after  the  inoculation  with  rabies  and  died  on  the  10th 
day;  cultures  from  the  brain  were  negative.  Negri  bodies  were  found 
in  sections  of  the  Ammon's  horn  of  both  rabbits.  The  control  rabbit 
became  paralysed  after  8  days  and  died  on  the  12th  day;  Negri 
bodies  were  found  in  sections  of  the  Ammon's  horn. 

Conclusion.  No  cross  immunity  exists  between  rabies  and  enzootic 
encephalo-myelitis. 

'  We  thank  Dr.  Perdrau,  of  the  National  Institute  for  Medical  Research,  for  kindly 
putting  a  strain  of  this  virus  at  our  disposal. 

'  We  thank  Dr.  Manouelian  of  the  Pasteur  Institute,  Paris,  for  his  kindness  in  putting 
this  strain  at  our  disposal. 


78  BORNA  DISEASE  AND 

C.  Polio-myelitis.^ 
In  the  experiments  with  this  virus  two  strains  have  been  used, 
(1)  a  strain  of  low  virulence  kindly  furnished  by  Professor  Mac- 
intosh, (2)  a  very  virulent  strain  kindly  provided  by  Professor  Petit. 
This  latter  strain,  which  has  been  utihzed  by  us  in  another  series  of 
experiments,  killed  monkeys  by  intracerebral  inoculation  as  in- 
dicated below: 

Table  XIII. 

Monkey  (Macacus  rhesus)  No.  5.  Died  the  9th  day  after  inoculation. 

No.  6.  „       12th      " 

No.  13.  „       nth         

No.  15.  „       11th         

The  lesions  produced  by  this  strain  of  polio-myehtis  in  the  central 
nervous  system  of  monkeys  were  very  intense,  neuronophagia  was 
frequently  found  in  the  anterior  horns  of  the  spinal  cord  as  well  as 
in  the  paravertebral  ganglia. 

The  virus  of  polio-myehtis  taken  from  monkeys  infected  experi- 
mentally is  generally  considered  as  non-pathogenic  for  the  rabbit.- 

In  a  first  series  of  experiments  we  gave  repeated  intracerebral 
inoculations  of  poho-myeUtis  to  young  rabbits;  some  time  afterwards 
the  rabbits  so  prepared  were  infected  by  the  same  route  with  the 
virus  of  enzootic  encephalo-myelitis. 

Table  XIV  gives  the  results  of  this  experiment. 

Thus  the  four  rabbits  having  received  three  successive  intracere- 
bral inoculations  with  the  virus  of  polio-myehtis  and  one  rabbit 
which  received  no  such  inoculation,  proved  to  be  still  susceptible  to 
infection  with  the  virus  of  Borna  disease. 

The  conclusion  that  absolutely  no  cross  immunity  exists  between 
the  virus  of  polio-myehtis  and  that  of  Borna  disease,  however,  is 
weakened  by  the  following  experiment  on  a  monkey. 

Monkey  M.  2  {Macacus  rhesus),  which  had  apparently  almost  com- 
pletely recovered  from  an  intracerebral  inoculation  of  tlie  virus  of 
Borna  disease,  after  having  shown  the  morbid  symptoms  recorded  on 
p.  36,  was  inoculated  into  the  brain  with  a  passage  virus  of  polio- 
myehtis  (strain  Petit).  At  the  same  time  two  controls  of  a  comparable 
size  and  the  same  species  were  similarly  inoculated.  The  subsequent 
history  of  the  two  controls  was  as  follows : 

M.  13  {Macacus  rhesus) 
5.10.27.    Inoculated  intracerobrally  with  the  viru.s  of  polio- mj-elitis. 
10.10.27.    Normal. 

11.10.27.   Paralysed,  found  procumbent. 
12.10.27. 

'  Our  thankn  are  due  to  M.  le  Prof.  A.  Petit,  of  the  I'iMit<<ur  Institute,  and  I'rof.  Miwv 
IntoHh  of  the  Itland  iSutton  Institute  for  Bupplying  us  with  Htrnins  of  polio-myelitiH 
virus. 

•  KruuDoand  Meinirke  (1909)  and  also  Dahni  (l!Ht9),  however,  have  recorded  a  few 
experimentH  Kiving  poxitive  n-Hult.s,  iininK  virus  taki'n  directly  from  human  cases,  hut 
the  general  opinion  held  to-day  is  that  the  virus  of  |>olio-niyeliti«  is  not  (jathogcnic  for 
rahbitii. 


ENZOOTIC  ENCEPHALO-MYELITIS 


79 


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80  BORXA  DISEASE  AND 

13.10.27.   Paralysed,  found  procumbent. 

14.10.27. 

15.10.27. 

16.10.27.   Found  dead,  11th  daj-s  after  inoculation. 

Cultures  of  brain.  Negative. 

Sections.  Lesions  typical  of  polio-myelitis. 

M.  15  {Macacos  rhesus) 
5.10.27.   Inoculated  intracerebrally  with  the  virus  of  polio-myelitis. 
10.10.27.   Normal. 
11.10.27. 
12.10.27. 
13.10.27. 

14.10.27.   Paralysis  of  the  left  arm. 
15.10.27.   Found  procumbent  and  paralysed. 
16.10.27.    Found  dead  11th  day. 
Cultures  of  brain.   Negative. 
Sections.   Lesions  typical  of  polio-myelitis. 

Monkey  M.  2  {Macacos  rhesus),  which  had  survived  the  inoculation 
with  Borna  disease,  when  inoculated  with  polio-myelitis,  behaved  as 
follows : 

5.10.27.  Inoculated  into  the  brain  with  the  virus  of  polio-myelitis  130  days  after 
the  inoculation  with  the  virus  of  Borna  disease. 

10.10.27.   No  added  symptoms  up  to  this  date. 

15.10.27. 

17.10.27. 

19.10.27. 

20.10.27.    Fifteenth  day,  myoclonic  movements. 

21.10.27.    Epileptiform  trembling  movements. 

22.10.27.   Slight  paresis  of  the  hind  quarters. 

23.10.27.    Paresis  more  marked. 

24.10.27.  Total  paralysis  of  the  hind  quarters  and  partial  paralysis  of  the  fore- 
limbs. 

25.10.27.  Could  make  movements  with  the  head,  but  the  total  paralysis  of  the 
arms  and  legs  prevented  it  from  getting  up. 

26.10.27.   Same  condition. 

27.10.27.   Same  condition,  but  still  feeding. 

28.10.27. 

29.10.27.   Animal  in  a  comatose  state. 

30.10.27. 

31.10.27.  Died  26  days  after  the  inoculation  with  the  virus  of  polio-myelitis  and 
156  days  after  the  infection  with  Borna. 

We  have  drawn  attention  in  the  course  of  this  monograph  to  the 
rosomblance  between  the  symptomatology  and  histological  picture 
in  polio-myelitis  in  the  monkey  and  experimental  enzootic  encephalo- 
myelitis in  th(!  same  species.  The  dilferences  i)etween  tiieni  are  the 
longer  period  of  incubation  and  the  slower  evolution  and  less  in- 
tensity of  the  lesions  in  the  nervous  system  in  the  latter  dis(>ase. 

Monkey  M.  2  died  after  a  lapse  of  time  nuich  longer  than  the 
averag(!  for  animals  inoculated  with  polio-myelitis,  the  strain  used 
generally  killing  the  species  of  animal  in  from  !)  to  11  days.  We  had, 
therefore,  to  determiiH?  whether  it  died  from  polio-myelitis  or  from 
a  persisting  infect  ifni  of  I'orna  disease.   On  an  anatomo-puthological 


ENZOOTIC  ENCEPHALO-MYELITIS  81 

study  of  the  various  parts  of  the  nervous  system  of  this  monkey  the 
lesions  of  experimental  enzootic  encephalo-myelitis  were  found,  includ- 
ing the  corpuscles  of  Joest-Degen.  The  general  aspect  of  the  altera- 
tions in  the  lumbar  region  of  the  spinal  cord  indicated  that  these  were 
produced  by  the  virus  of  Borna  disease  and  not  by  the  virus  of  polio- 
myelitis. This  conclusion  received  further  support  when  emulsions 
from  various  parts  of  the  nervous  system  were  passaged  to  fresh 
animals.    The  results  of  these  inoculations  were  as  follows: 

(1)  Eabbit  460a,  weighing  2,320  gms.,  and  Rabbit  462a,  weighing 
1,740  gms.,  were  inoculated  intracerebrally  with  an  emulsion  of  the 
brain  of  Monkey  M.  2.  The  former  died  on  the  37th  day  and  the  latter 
on  the  38th  day  after  infection,  and  lesions  characteristic  of  experi- 
mental Borna  disease  were  found  in  sections  of  the  various  parts  of 
the  nervous  system  of  both  animals.  A  passage  was  made  with  the 
brain  of  Rabbit  462a  to  a  fresh  Rabbit  405a,  weighing  1,330  gms. 
Eabbit  40.5a  succumbed  on  the  42nd  day  of  Borna  disease. 

(2)  Rabbit  464a,  weighing  1,960  gms..  Rabbit  458a,  weighing 
1,760  gms.,  and  Rabbit  459a  weighing  2,500  gms.,  were  inoculated 
intracerebrally  with  an  emulsion  of  the  cervical  region  of  the  spinal 
cord  of  Monkey  M.  2.  Rabbit  464a  died  on  the  34th  day,  458a  on  the 
30th  day,  and  459a  on  the  30th  day  after  inoculation,  and  lesions 
typical  of  Borna  disease  were  found  in  sections  of  the  nervous  system 
of  all  three.  A  passage  was  made  with  the  brain  of  Rabbit  458a  to  a 
fresh  rabbit,  414a,  which  died  on  the  33rd  day  of  experimental  enzootic 
encephalo-myelitis. 

(3)  Eabbit  454a,  weighing  1,960  gms..  Rabbit  455a,  weighing 
1,960  gms.,  and  Rabbit  463a,  weighing  2,300  gms.,  were  inoculated 
intracerebrally  with  an  emulsion  of  the  lumbar  cord  of  Monkey  M.  2. 
The  first  died  on  the  40th  day,  the  second  on  the  35th  day,  and  the 
third  on  the  34th  day  of  experimental  Borna  disease.  Rabbit  415a, 
weighing  1,880  gms.,  when  inoculated  intracerebrally  with  an  emul- 
sion of  the  brain  of  Rabbit  463a,  died  of  Borna  disease  on  the  30th  day. 

The  results  of  these  rabbit  inoculations  proved  that  the  virus  of 
Boma  disease  still  existed  in  the  brain  and  spinal  cord  of  Monkey  M.  2. 

Monkey  M.  20  {Macacus  rhesus)  was  inoculated  with  an  emulsion 
of  a  portion  of  the  brain  and  the  cervical  cord  of  Monkey  M.  2.  It 
remained  healthy  for  a  period  of  60  days,  when  the  commencement  of 
paresis  of  the  hind  quarters  was  first  observed.  Paresis  later  became 
accentuated  and  reached  the  fore  hmbs.  The  animal  died  on  the  68th 
day  after  inoculation,  and  the  lesions  of  experimental  enzootic  ence- 
phalo-myehtis  were  found  in  sections  of  the  central  nervous  system. 
Rabbits  340a  and  341a  were  inoculated  intracerebrally  with  an  emul- 
sion of  the  brain  of  Monkey  M.  20,  and  both  died  of  experimental 
Boma  disease. 

In  resume,  the  activity  of  virus  of  Borna  disease  which  had  become 
dormant  in  the  central  nervous  system  of  Monkey  M.  21 ,  130  days  after 
the  inoculation,  as  indicated  by  the  clinical  picture,  was  revived  by 
the  introduction  of  the  virus  of  polio-myelitis  by  the  intracerebral 
route.   As  a  result  of  the  second  infection  the  monkey  died  and  we 

6409  „ 


82  BORNA  DISEASE  AND 

demonstrated  the  presence  of  the  virus  of  Borna  disease  in  its  brain 
and  spinal  cord,  but  the  virus  of  poho-myehtis  had  been  destroyed. 

Without  wishing  to  attach  too  much  importance  to  this  single 
experiment,  we  are  forced  to  conclude  that  the  monkey  which  sur- 
vived for  130  days  the  infection  with  Borna  disease  showed  an 
exceptional  resistance  of  an  anomalous  character  to  the  virus  of 
polio-myehtis.  This  resistance  might  be  the  result  of  a  certain  degree 
of  immmiity  conferred  by  the  first  infection.  On  the  other  hand,  it 
might  be  explauied  in  another  way.  The  experiments  by  Gilde- 
meister  and  Herzberg  (1925)  indicate  that  guinea-pigs  which  showed 
lesions  on  the  pads  of  the  metatarsus  as  a  result  of  infection  with 
the  virus  of  vaccinia  develop  a  certain  local  resistance  to  infection 
with  herpetic  virus  inoculated  by  the  same  route.  The  authors  con- 
clude that  there  is  a  certain  degree  of  immunity  conferred  by  one 
virus  against  infection  with  the  other.  In  a  similar  way,  certain 
rabbits,  which  after  recovering  from  vaccinal  keratitis,  when  inocu- 
lated some  time  later  on  the  same  cornea  with  the  virus  of  herpetic 
encephaUtis,  sometimes  resist  the  second  infection. 

We  do  not  consider  in  these  circumstances  that  an  inununity 
results  in  the  proper  sense  of  the  word,  but  rather  a  local  mobiUzation 
of  the  elements  of  defence  provoked  by  the  first  virus  inoculated,  thus 
conferring  on  the  tissue  a  certain  degree  of  resistance  of  a  non- 
specific character  which  does  not  exist  normally. 

We  intend  to  repeat  our  experiment.  In  addition,  monkeys  are 
being  immunized  against  polio-myehtis  which  will  be  inoculated  later 
with  the  virus  of  Borna  disease. 

VI.    SlMMARV. 

A  solid  immunity  can  occasionally  be  obtained  against  Borna 
disease  in  the  rabbit  by  inoculating  suitably  attenuated  virus  into 
the  brain. 

Multiple  intravenous  injections,  infection  by  corneal  scarification, 
or  intratesticular  inoculation  with  fresh  virus,  can  also  produce  im- 
munity. 

We  have  not  succeeded  in  producing  inununity  by  inoculating 
virus  killed  by  chloroform,  ether,  or  ultra-violet  light  intracerebrally 
into  rabbits. 

Multiple  inoculations  subcutaneously  of  large  quantities  of  for- 
molized  virus  leads  to  immunity  in  a  Umited  number  of  animals. 

Rabbits  immunized  against  an  equine  strain  of  the  virus  of  Borna 
disease  were  resistant  to  intracerebral  infection  with  an  ovine  strain 
and  vice  versa. 

No  cross  immunity  was  obtained  between  Borna  disease  and  herpes 
or  rabies.  Cross  inununity  between  Borna  disease  and  pt)lio-niyeliti9 
was  not  observed  when  rabbits  were  the  subject  of  experiment,  but 
in  an  experiment  carried  out  on  a  monkey  the  result  suggested  that 
some  resistance  to  the  virus  of  polio-myelitis  may  be  produced  by  a 
previous  attack  of  experimental  Borna  disease. 


ENZOOTIC  ENCEPHALO-MYELITIS  83 

13.   CHEMOTHERAPY 

Various  medicaments,  calomel,  mercuric  chloride,  salvarsan, 
atoxyl,  have  been  tried  in  the  treatment  of  enzootic  encephalo- 
myelitis. The  greatest  success  as  regards  the  treatment  by  drugs 
has  been  claimed  for  the  administration  of  urotropine  (hexamethy- 
lenetetramine).  Moussu  and  Marchand  (1926)  obtained  remarkably 
good  results  in  the  treatment  of  the  disease  in  horses  and  cattle  by 
inoculation  of  urotropine  intravenously.  According  to  these  authors, 
when  15  to  20  gms.  is  administered  on  the  appearance  of  the  first 
symptoms,  the  mortahty  in  epizootics  may  be  lowered  from  80  or  90 
per  cent,  to  25  per  cent.  In  Germany  Ostertag  (1924)  has  generahzed 
the  use  of  urotropine  on  a  large  scale  in  the  treatment  of  enzootic 
encephalo-myehtis.  H.  Bohn  (1927)  inoculated  30  gms.  per  day  for 
4  to  5  days  intravenously  into  affected  horses  with  good  results. 
Trepel  (1926)  advocates  the  use  of  a  total  of  100  gms.  during  the 
course  of  a  few  days.  Grimm  (1927)  obtained  a  smaller  number  of 
recoveries  than  the  authors  cited  above.  All  authors  appear  to  be 
agreed  as  to  the  beneficial  action  of  the  drug  in  Borna  disease  of  the 
horse. 

We  have  carried  out  experiments  to  see  whether  urotropine  was  as 
efficacious  in  treating  the  disease  in  the  rabbit  as  in  the  horse.  We 
commenced  by  determining  the  maximum  intravenous  dose  supported 
by  a  rabbit.  This  appeared  to  be  about  0-5  gms.  per  kgm.,  a  dose 
which  can  be  repeated  at  least  eight  times  at  4  to  5  days'  interval. 
Subsequently  the  experiment  recorded  in  Table  XV  was  carried  out. 

Summary.  The  dose  of  urotropine  we  gave  to  the  rabbit  was 
equivalent,  weight  for  weight,  to  100-200  gms.  for  the  horse  and  to 
a  total  amount  of  600  gms.,  yet  we  were  not  able  to  demonstrate  any 
prophylactic  or  curative  action. 


P  2 


84 


BOENA  DISEASE  AND 


Result. 

5" 

i        i 
.1  >>     ^>. 

1* 

ll 

l" 

"> 

U 

3 
X 

24.7.27. 

Typical  symptoms. 
Weight  1,550  gms. 

Typical  disease. 
Weight  1,000  gms. 

Typi(!al  disease. 
Weight  1,050  gms. 

1 

11 

1  1 

in. 7. 27. 
Urotropine  tnoca- 
laled  into  veins. 

Animal  showing 
symptoms. 
Weight  1,880  gms. 

Weight  1,900  gms. 
Typical  disease. 
Urotropine  O-S  gr. 

Typii^al  (tomraence- 

ment  of  disease. 
Weight  2,100  gms. 
Urotropine  0-8  gr. 

Typical  symptoms 
\Veight  l^fiOOgms. 
No  urotropine 

i- 
■s  2 

U.7.27. 
Urotropine  inocu- 
lated into  veins. 

Weight  1,900  gms. 
Urotropine  0-8  gr. 

Weight  2,000  gm's. 
Urotropine  0-8  gr. 

Weight  2,200  gms. 
Urotropine  0'8  gr. 

Weight  1,800  gms. 
No  urotropine 

1 
.2? 

e^ 

Mx 
c  ° 

u 

Weight  2,0.50  gms. 
Urotropine  0-8  gr. 

Weight  2,300  gms. 
Urotropine  0-8  gr. 

Weight  1,850  gms. 
No  urotropine 

Weight  2,,320  gms. 
Urotropine  1  gr. 

2.7.27. 

Urotropine  inocu- 
lated into  veins. 

Weight  1,950  gms. 
Urotropine  0-8  gr. 

Weight  2,100  gms. 
Urotropine  0-8  gr. 

Weight  2,250  gms. 
Urotropine  0-8  gr. 

E 

^§ 

II 

U 

28.6.27. 
Urotropine  inocu- 
lated into  veins. 

Weight  2,020  gms. 
Urotropine  0-8  gr. 

Weight  2,150  gms. 
Urotropine  0-8  gr. 

Weight  2.200  gms. 
Urotropine  0-8  gr. 

Weight  1,820  gms. 
No  urotropine 

a&) 

3i 
U 

Inoculation  of 
virus  intracere- 
brally. 
23.6.27. 
Urotropine  inocu- 
lated into  veins. 

§4 

a-x 

«.| 

•a    5. 

^  £ 

■n 

Weight  2.100  gms. 
Urotropine  0-8  gr. 

Weight  2,220  gms. 
Urotropine  0-8  gr. 

Virus  into  brain. 
Weight  1,770  gms. 
No  urotropine 

Not  inoculated 
with  vims. 
Weight  2,400  gms. 
Urotropine  i  gr. 

1 

18U 
179a 

Control 
178a 

1< 

ENZOOTIC  ENCEPHALO-MYELITIS  85 

REFERENCES 

Alberttn,  a.  (1926). — Thesis.   Lyons. 

Alvisatos,  G.  p.  (1922).   Deutsche  med.  Wchnschr.,  48,  295. 

Baudoin,  a.,  and  Schaeffer,  H.  (1926).    Paris  Med..  16,  249. 

Beck,  A.  (1925).   Ztschr.f.  Infektioiisk.  d.  Hausliere,  28,  99. 

Beck,  A.,  and  Frohbose  (1926).   Archiv  f.  wissensch.  u.  prald,  Tierh.,  54,  84. 

B^RIEL,  L.  (1926).— i(/o7i  med.,  1,  667. 

BtRiEL,  L.,  and  Devic,  A.  (1926).    Li/on  med.,  1,  671. 

BoHN,  H.  (1927).   Deutsche  tierarztl.  Wchnschr.,  35,  584. 

Causel,  M.  (1924).   Eec.  med.  vet.,  100,  526. 

Christiani  (1909).   Arch.  f.  ivissensch.  v.  prakt.  Tierh.,  35,  253. 

Dahm  (1909).   Miiiichen.  med.  Wchnschr.,  56,  2553. 

David  and  Dekester  (1926).   Bull,  et  mem.  Soc.  med.  d'hop.  de  Paris,  3,  S.,  50,  75. 

Dexler,  H.  (1900).   Ztschr.f.  Thiermed.,  4,  110. 

DiOBY,  K.  (1919).   Immunity  in  Health.    Oxford  Medical  Publication.s. 

Dobberstein,  J.  (1925).   Berl.  tierarztl .  Wchnschr.,  41,  177. 

Eichbaum,  Stohr,  and  VVilke  (186.5-6).    Mitt.  d.  tierdrzt.  Prax.  in  preuss.  Stoat. 

Berlin,  135. 
Ellinoer,  R.  (1927).    Tierarztl.  Rdsch.,  .33,  201. 
Ernst,  W.  (1925).   Munchen.  tierarztl.  Wchnschr.,  76,  477. 
Ernst,  W.,  and  Hahn,  H.  (1926).   Munchen.  tierarztl.  Wchnschr.,  77,  46. 

(1927).   Munchen.  lierdrztl.  Wchnschr.,  IS,  So. 

Flexner,  S.,  and  Amos,  H.  L.  (1917).  J.  Exper.  Med.,  25,  525. 

FoERSTER  cited  by  Marinesco  and  Draganesco. 

Gildemeister,  E.,  and  Herzbero,  K.  (1925).   Deutsche  med.  Wchnschr.,  51,  97. 

Glamser  (1926).    Deutsche  tierdrztl.  H'c/j?(«cAr.,  34,  312. 

Grimm  (1927).   Berl.  tierdrztl.  Wchnschr.  (1927),  43,  153. 

Head  and  Campbell  (1900).   Brain,  23,  353. 

Hempt,  a.  (1925).   Ann.  de  Vlnst.  Pasteur,  39,  6.32. 

Heydt  (1914).   Deutsche  tierarztl.  Wchnschr.,  UQ. 

HuTYRA  and  Marek  (1922).    Spezielh  Pathologic  v.  Therapie  d.  Hausliere,  6th  Ed. 

Jena. 
Joest,  E.  (1921).   Ztschr.f.  Infektionskr.  d.  Haustiere,  21,  97. 

(1926).   Klin.  Wchnschr.,  5,  209. 

Joest,  E.,  and  Deo  en,  K.  (1909).   Ztschr.f.  Infektionskr.  d.  Hausliere,  6,  348. 

(1911).   Ztschr.f.  Infektionskr.  d.  Haustiere,  9,  !. 

(191 1 ).   Deutsche  Ztschr.  f.  Nervenh.,  42,  293. 

JOHNE  (1897,  1).   Deutsche  Ztschr.f.  Thiermed.,  22,  369. 

(1897,  2).    Ber.  u.  d.  Veterinarw.  im  Konigr.  SacA,«. 

Kahlmeter,  G.  (1927).   Acta  med.  Scandinav.,  65,  709. 

Kragerud,  a.,  and  Gunderson,  T.  (1921).   Gorges  offtsielle  statistik.,  7,  15. 

Kbaus,  R.,  Kantor,  L.,  Fischer,  H.,  and  Quirooa,  R.  (1920).   Ztschr.f.  Immuni- 

tdtsforsch.,  30,  121. 
Krause,  p.,  and  Mefnicke,  E.  (1909).   Deutsche  med.  Wchnschr.,  35,  No.  12. 
Lessage,  J.,  and  Frisson,  M.  (1912).  Rev.  gen.  de  med.  vet.,  20,  657. 
Levaditi,  C.  (1926).   L' herpes  et  le  zona.   Ma.s9on  et  Cie,  Paris. 
Levaditi,  C,  and  Nicolau,  S.  (1923).  Ann.  d.  I'lnst.  Pasteur,  37,  1. 

(1922).   Compt.  rend.  Acad.  d.  sc,  174,  778. 

Levaditi,  C,  Harvieb,  P.,  and  Nicolau,  S.  (1921).  Compt.  rend.  Soc.  de  hiol.,  84,  524. 

(1922).   Ann.  d.  I'lnst.  Pasteur,  36,  63  and  105. 

LoHR  (1910).     Beitr.   z.   Bakt.  d.   Gehirnruckenmarksseuche  d.   Pferde.    Inaug.   Diss. 

(Jjeipzig).   Dresden. 
Manooelian.  Y.,  and  Viaia,  J.  (1926).  Compt.  rend.  Acad.  d.  sc,  183,  1357. 

(1927).   Compt.  rend.  Acad.  d.  sc,  184,  630. 

Marchand,  L.,  and  Monssn,  R.  (1924).  Compt.  rend.  Acad.  d.  sc,  178,  149. 

Marcq,  J.  (1909).  ^7!?!.  de  med.  vet.,  58,  11. 

Marie,  A.,  and  Mutermilch,  S.  (1927).   Compt.  rend.  Acad.  d.  sc,  184,  911. 

Marinesco,  G.,  and  Draganesco,  S.  (1927).  Bull.  Acad,  de  med.,  97,  473. 

MiKSSNER,  H.  (1926).   Deutsche  tierarztl.  Wchnschr.,  34,  637. 

MotJSSU,  R.  (1922).    Rec  de  med.  vet.,  98,  261. 

(1926).    Recherches  sur  certain   affections  du  syst.  nerveux  central  des  animaux 

domestiques.   Thesis,  Vigot  Freres,  Paris. 
Moussn,  R.,  and  Marchand,  L.  (1924).  Rec  de  med.  vet.,  100,  5;  65. 
Nicolau,  S.,  and  Galloway,  I.  A.  (1927).  Brit.  J.  Exper.  Path.,  8,  336. 
NoACK  (1908).   Ber.  u.  d.  Veterinarw.  im  Konigr.  Sachs.,  p.  41. 


86  BORNA  DISEASE 

Ofpenbeim,  H.  (1907).   ZUchr.f.  hijrkiionshr.  d.  Hausliere,  2,  148. 
OSTERTAG  (1900).   Berl.  tierarztl.  HcAnxcAr.,  433. 

(19C»6).   Btrl.  tierarztl.  Wchnschr.,  &22. 

(1924).   Berl.  tierarztl.  Wchnschr.,  40,  705. 

PtHT.  M.,  and  Dechafme,  J.  (1927).   Ann.  dc  mirt.,  22,  172. 

Petiot  (1921-2).  Thesis  (Paris). 

Pette  cited  by  Slarienesco  and  Draganesco. 

Popow  (1882),  cited  by  Hutyra  and  Marek. 

Pbiemek  (1925)  cited  in  the  thesis  of  Schmidt.   Leipzig  (1926). 

Prietsch  (1896).   Ber.  u.  d  Yeterindrw.  im  Konigr.  Sachs.,  p.  126. 

Pkoger  (1896).    Ber.  u.  d.  Veterinanr.  im  Konigr.  Sachs.,  p.  105. 

RArTMANS,  H.  (1926).   Tierdrztl.  Bdsch.,  32,  841. 

RA^•r^•A,  A.  (1927).  Presse  mid  ,  35,  646. 

Remusger,  P.  (1919).   Compt.  rend.  Soc.  de  biol.,  82,  52. 

RocH,  M.,  and  Bickel,  G.  (1927).  Schiceiz.  med.  Wchnschr.,  57,  18. 

ROLOFF  (1868).   iiiil.  a.  d.  tierarztl.  Prax.  in  preuss.  Slaal.  Berlin,  135. 

SA\^G^•t,  A.,  and  Leblasc,  P.  (1897).  J.  mid.  lit.  et  de  zooteck.,  1897,  5  S.,  1,  274. 

Schmidt  (1870).  ilag.f.  d.  ges.  Tierh.,  Berlin,  186. 

SiEDAMOROTZKY  and  ScHLEGEL  (1896).  Arch.  f.  tcissensch.  v.  prakt.  Thierh.,  22,  287. 

Skoog,  a.  L.  (1923).  Med.  Clin.  Sorth  Amer.,  7,  1323. 

Spiegl,  a.  (1922).   Zlschr.f.  Infektionskr.  d.  Haustiere,  23,  147. 

Targowla,  J.  (1894).  Rev.  Xeurol,  2,  465. 

Thomas,  A.,  and  Rendu  (1925).  Bev.  Xeurol.,  2,  758. 

Thomas,  H.  M.  (1898).  Phila.  M.  J.,  1,  885. 

Trepel  ( 1926).   Berl.  tierarztl.  HcAn^rAr.,  42,  881. 

Vak  Gehichtes,  a.,  and  Nelis,  G.  (1900).   Presse.  mid.,  8,  113. 

ViETS,  H.  R.  (1927).  Arch.  Xeurol.  and  Psychial.,  17,  794. 

Waldmass,  J.,  and  Pape,  J.  (1921).  Berl.  tierdrzll.  Wchnschr.,  37,  449. 

Walther  (1899).   Ber.  v.  d.  Veterindrtc.  im  Konigr.  Sachs.,  p.  80. 

WiscHNiKowrrcH  (1889)  cited  by  Hutyra  and  Marek. 

VVOHLWILL,  F.  (1924).   Ztschr.f.  d.  ges'  Xeurol.  «.  Psychial.,  89,  171. 

WoRZ  cited  by  Hut\Ta  and  Marek  (see  above). 

ZwiCK.  \V.  (1925).    Berl.  tierarztl.  Wchnschr.,  41,  453. 

(1926).    Seucheubekampfung,  3,  57. 

ZwicK,  W.,  and  Seitried,  0.  (1924).   Berl.  lierdrztl.  Wchnfchr.,  40,  465. 

(1926).   Bert,  tierdrzll.  Wchnschr.,  41,  129. 

ZwiCK,  W.,  Seifried,  0.,  and  Witte,  J.  (1926).   Ztschr.f.  Inftklionskr.  d.  Uaustiere, 
30,42. 


DESCRIPTION  OF  PHOTOGRAPHS  AND  COLOURED 
PLATES 

Fig.  1.  Photograph  of  Rabbit  No.  275.  29  days  after  intracerebral  infection  with 
the  virus  of  Borna  disease,  and  3  days  before  death.  The  characteristic  position  of  the 
head  and  ears  is  depicted. 

Fig.  2.  Photograph  of  7l/aono?(«  rAes«.«  No.  1.  04  days  after  inoculation.  Paralysis 
of  the  hind  quarters;  paresis  of  the  right  arm,  the  hand  not  being  able  to  grip  any  object 
presented  to  it. 

Fig.  3.  Photograph  of  Macaciis  rhesus  No.  1.  69  days  after  inoculation.  Charac- 
teristic '  hunched  up  '  attitude. 

Fig.  4.  Photograph  of  Ma(acus  rhesus  No.  2.  .51  days  after  the  infection  by  the  intra- 
cerebral route.   Facial  paralysis  on  the  left  side. 

Fig.  5.  Photograph  of  Macacus  rhesus  No.  2.  67  days  after  inoculation.  3rd  crisis 
of  the  disease.   Facial  paralysis  on  the  right  side. 

Fig.  6.  Photograph  of  Guinea-pig  9.5k.  60  days  after  intracerebral  inoculation,  and 
5  days  before  death.   Paralysis  of  the  hind  quarters. 

Fig.  7.  Photograph  of  Guinea-pig  1a.  50  days  after  intracerebral  inoculation,  and 
2  days  before  death.   General  paralysis. 

Fig.  8.  Photograph  of  Rat  No.  14.  63  days  after  inoculation  intraeerebrally,  and 
4  days  before  the  death  of  the  animal.  At  this  stage  the  rat  placed  on  the  flank  made 
several  useless  efforts  with  the  fore-legs  to  recover  its  normal  position. 

Fig.  9.  Photograph  of  the  stomach  wall  of  Rabbit  212  which  died  44  days  after 
the  intracerebral  inoculation,  showing  the  haemorrhagic  lenticular  areas.  L  =  greater 
curvature  of  the  stomach;  P  =  lenticular  haemorrhagic  areas. 

Fig.  10.  Microphotograph,  x  160.  Rabbit  209  dead  71  days  after  intratesticular 
inoculation.  Meningitis  with  mononuclear  cells  with  '  cuffing  '  around  the  vessels. 
V  =  lumen  of  blood-vessel;  C  =  cerebral  cortex;  M  =  mononuclear  leucocj'te  of  inflam- 
matory process. 

Fig.  11.  Microphotograph,  x  300.  Rabbit  51a  dead  28  days  after  intracerebral 
infection.  Perivascular  'cuff'  in  the  cerebral  cortex  cut  longitudinally.  V  =  lumen  of 
blood-vessel ;  E  =  endothelium  of  blood-vessel ;  M  =  mononuclear  leucocytes  taking  part 
in  the  infiltrative  process. 

Fig.  12.  Microphotograph,  -^165.  Rabbit  11d  dead  27  days  after  intracerebral 
inoculation.  Section  showing  the  aspect  of  the  Cornu  Amynoyiis.  F  =  area  of  infiltrating 
lymphocytes  between  the  nerve-cells;  L  =  infiltrating  lymphocytes;  C  =  cells  containing 
the  corpuscles  of  Joest-Degen  within  their  nuclei. 

Fig.  13.  Microphotograph,  X  800.  Rabbit  25  dead  35  days  after  cerebral  infection. 
'Cyst'  in  a  nerve-cell  of  the  Ammon's  horn.  C  =  'cyst';  N  =  nucleus;  n=nucleolus. 

Fig.  14.  Microphotograph,  x  1,000.  Rabbit  80a  dead  28  days  after  inoculation 
intraeerebrally.  '  Cyst "  in  the  protoplasm  of  a  nerve-cell  of  the  Ammon's  horn.  C  =  'cyst'; 
N  =  nucleolus. 

Fig.  15.  Microphotograph,  x  900.  Rabbit  1  Id  dead  27  days  after  cerebral  infection. 
'Cyst'  in  the  protoplasm  of  a  nerve-cell  of  the  Cornu  Amman  is. 

Fig.  16.  Microphotograph,  x  1,000.  Rabbit  51a  dead  28  days  after  inoculation  into 
the  brain.   'Cyst'  in  the  protoplasm  of  a  nerve-cell  in  the  Cornu  Arnmonis. 

Fig.  17.  Microphotograph,  x  1,000.  Rabbit  No.  22  dead  24  days  after  cerebral 
infection.  Nerve-cell  in  medulla  oblongata  showing  neuronophagia.  C  =  degenerated 
cytoplasm ;  p  =  polj'morphonuclear  penetrating  neuron ;  m  =  mononuclear  cells. 

Fig.  18.  Microphotograph,  x  230  (enlarged  two  diameters).  Rabbit  295  dead  19  days 
after  intracerebral  inoculation.  Section  in  the  region  of  the  pons  showing  (d)  degener- 
ated nerve-cells,  (s)  ' salellitism'  with  commencing  neuronophagia,  (n)  neuronophagia, 
(1)  lymphocytes  in  the  parenchyma. 


88  DESCEIPTIOX  OF  PHOTOGRAPHS,  ETC. 

Fig.  19.  Microphotograph,  x  1,000.  Rabbit  278  inoculated  iutracerebrally  and 
sacrificed  20  days  later.  Commencing  neuronophagia  in  the  anterior  horn  of  the  spinal 
cord  in  the  lumbar  region.  N  =  nucleus  of  nerve-cell:  P  =  cytoplasm;  M  =  mononuclear 
cells. 

Fig.  20.  Microphotograph,  x  66.  Rabbit  44  dead  33  days  after  intracerebral  inocula- 
tion. Section  of  the  lumbar  region  of  the  spinal  cord.  .S  =  cord;  M  =  perivascular  "cuff' 
in  a  nerve-trunk;  I  =  interstitial  mononuclear  infiltration;  A  =  accumulation  of  mono- 
nuclear leucocytes. 

Fio.  21.  Microphotograph,  x  66.  Rabbit  No.  30  dead  36  days  after  the  inoculation 
given  intracerebrally.  Spinal  ganglion  in  the  lumbar  region.  N  =  ganglion  nerve-oell 
in  a  state  of  neuronophagia;  I  =^ formation  of  a  nodule  of  mononuclear  leucocytes 
which  has  formed  at  the  site  where  a  nerve-cell  is  undergoing  neuronophagia;  V  = 
perivascular  infiltration. 

Fio.  22.  Microphotograph,  x  66.  Rabbit  50a  dead  53  days  after  cerebral  inocula- 
tion. Section  of  the  cord  in  the  lumbar  region  including  a  spinal  ganglion.  V  = 
vacuolization  in  certain  ganglion  cells;  I  =  interstitial  infiltration;  N  —  neuronophagia ; 
P  =  perivascular  infiltration. 

Fio.  23.  Microphotograph,  x  1,000.  Rabbit  164  dead  29  days  after  intracerebral 
inoculation.  Spinal  ganglion  in  the  lumbar  region.  Neuronophagia  with  the  formation 
of  nodules  consisting  of  mononuclear  cells.  R  =  remains  of  a  destroyed  neuron; 
V  =  vacuole  in  a  nerve-cell;  P  =  polymorphonuclear;  L  =  lymphocytes. 

Fig.  24.  Microphotograph,  x  950.  Rabbit  No.  30  dead  36  days  after  intracerebral 
inoculation.  Spinal  ganglion  in  the  lumbar  region.  Degeneration  of  a  ganglion  cell 
and  commencing  neuronophagia.  C  — degenerated  cell  which  has  become  o.xyphilic  and 
is  on  the  way  to  destruction;  P- polymorphonuclear  cells;  L'^  lymphocytes;  M  =  mono- 
nuclears participating  in  the  process  of  neuronophagia. 

Fio.  25.  Microphotograph,  x  1,000.  Rabbit  164  dead  29  days  after  intracerebral 
inoculation.  Spinal  ganglion  in  the  thoracic  region.  N-^ neuronophagia;  G  — particle 
of  chromatin  probably  originating  from  a  pyknosed  polymorphonuclear;  M  -macro- 
phage; C  =  cell — commencing  neuronophagia;  L     lymphocytes. 

Fio.  26.  Microphotograph,  x  8(M).  Rabbit  No.  30  dead  36  days  after  cerebral  in. 
oculation.  Spinal  ganglion  in  the  lumbar  region.  M  ==  mononuclear  cells  infiltrating; 
D  =  commencing  neuronophagia;  V  -  vessel  surrounded  by  perivascular  'cuff'. 

Fio.  27.  Microphotograph,  :■  5.  Rabbit  140a  dead  49  days  after  intracerebral  infec- 
tion with  glycerinated  virus  of  passage.  Section  of  the  terminal  part  of  the  spinal  cord 
with  roots  of  the  sciatic  nerve  and  corresponding  ganglion.  M  =spinal  cord;  R  nerve- 
root;  G ^spinal  ganglion;  .S" sciatic  nerve.  In  sections  cut  in  this  way  the  lesions  in 
the  regions  mentioned  above  can  be  seen  as  a  whole. 

Fio.  28.  Microphotograph,  v  350.  Rabbit  140a.  Longitudinal  section  of  the  sciatic 
nerve  near  the  popliteal  region.  A  perivascular  'cuff'  is  seen.  V  lumen  of  vessel. 
M  =  infiltrating  mononuclear  cells;  L     lymphocytes  in  the  thickness  of  the  nerve. 

Fio.  29.  Microphotograph,  x  l.iO.  MararunrhenunNoA.  Sectiim  of  the  parietal  lobe; 
C  =  cerebral  corte.i;  M  ^  mononuclear  meningitis;  V  blood. vessel  cut  longitudinally 
with  walls  infiltrated  with  mononuclear  cells. 

Fio.  30.  Microphotograph,  :•  120.  Mararu-i  rhesng  No.  3.  Periva.scular  <ufling  in 
the  frontal  lobe  of  the  cerebral  cortex.  V  lumen  of  vessel;  P  =  periva.scular  infiltration. 
Fio.  31.  Microphotograph,  x  150.  Macacii«  rhemis  No.  1.  Section  of  the  parietal 
lobe  of  the  cerebral  cortex.  Pseudo  gumma  (nodule)  prcxiuced  by  mononuclear  colls, 
with  a  small  ves.>«el  in  the  centre  (very  intense  process  of  perivascular  infiltration). 
V      l)lood.ves.sel;  K     nodule  formed  by  monimudear  cells. 

Kio.  32.  Microphotograph,  ^360.  Maranis  rhesun  No.  I.  Section  through  basal 
ganglia.   Small  area  of  monimuclear  cells.    N     neuron;  L     lymphocytes. 

Fio.  33.  Microphotograph,  >  150.  MiirncuA  rhemn'iin.X.  Vascular  infiltration  in  the 
terminal  part  of  the  spinal  cord.  V  lumen  of  blocnl. vessel;  M  muscular  coat; 
I'     infiltrating  mononuclear  cells. 

Fio.  34.  Microphotograph,  >  3tMJ.  Macacun  rhtDU-i  tio.  \.  Posterior  root  of  sciatic 
nerve.  Interstitial  infiltrati(m  and  perivasc'ular  'i^ufliiig'.  V  lumen  of  bloud-vosaci; 
M     mononuclear  cell  in  the  |)rocess  of  |>eriviiscular  'cutting';  in     macrophage. 

Fig.  35.  Microphotograph,  •  "(H).  Marncun  rheaux  No.  3.  Spinal  ganglion  in  cervical 
region.  Corpuscles  (type  Joest-Degen)  surrtiunded  by  a  halo  in  the  nerve-cells  of  the 
ganglion.   C     intranuclear  corpuscle  of  .loest.Degen;  N     nucleolus. 


DESCEIPTION  OF  PHOTOGRAPHS,  ETC.  89 

Fig.  36.  Microphotograph,  ( ;<  3(5,  enlarged  2t  diameters).  Maearus  rhesus  No.  1. 
Section  of  spinal  ganglion  in  lumbar  region  of  spinal  cord  showing  the  profound  changes 
which  predominate  the  peripheral  parts  of  the  nervous  system.  (i)  =  intranuclear  cor- 
puscle (Joest-Degen)  with  surrounding  halo;  n  =  nodule  of  mononuclear  cells  replacing 
destroyed  neuron;  d  =  degenerated  ganglion  cells;  np  =  neuronophagia ;  ic  =  peri- 
cellular infiltration;  it  =  interstitial  infiltration;  c  =  capsule  of  ganglion. 

Fig.  37.  Microphotograph,  x  55.  ilacacus  rhesus  No.  3.  Section  through  the 
brachial  nerve.  Perivascular  'cuffing'  and  interstitial  infiltration.  G  =  nerve  sheath; 
V  =  perivascular  'cuffs';  I  =  interstitial  infiltration. 

Fig.  38.  Microphotograph,  x460.  Macacus  rhesiisNo.  \.  Transverse  section  of  the 
sciatic  nerve  several  centimetres  from  its  exit  from  the  greater  sciatic  foramen — peri- 
vascular 'cuff'.  V  =  lumen  of  blood-vessel;  E  =  vascul^  endothelium;  L  =  lympho- 
cytes. 

Fig.  39.  Microphotograph,  x55,  (photograph  enlarged  2A  diameters).  Macacus 
rhesus  No.  1.  Transverse  section  of  sciatic  nerve  after  its  exit  from  the  greater  sciatic 
foramen.  V  =  large  vessel  with  perivascular  infiltration ;  v  =  small  vessels  showing  same 
phenomenon;  1  =  Ij'mphocytes  (interstitial  infiltration). 

COLOURED  PLATES 

PLATE  I 
Fig.  1.   Staining,  toluidin  blue-eosin.   Obj.  5  mm.,  oe.  2,   x260. 

Rabbit  295  dead  19  days  after  intracerebral  infection.    CelliJar  degeneration  in 
the  medulla  oblongata.    N  =  neuron  in  normal  state;   C  =  degenerated  nerve-cell — ■ 
tigrolysis-oxyphilia — nucleus    eccentric;    E  =  degenerated   neuron;    F  =  fragment    of 
nerve-cell;  L  =  lymphocytes. 
Fig.  2.   Staining,  Mann.    1   12  oil  immersion,  oc.  i. 

Rabbit  25  dead  35  days  after  cerebral  inoculation.   Oxyphilic  corpuscle  surrounded 
by  a  halo  in  a  nerve-cell  in  the  Ammon's  horn.   C  =  corpuscle  (type  Joest-Degen)  sur- 
rounded by  a  halo. 
Fig.  3.   Staining,  Mann.    1/12  oil  immersion,  oc.  4. 

Rabbit  80.i  dead  28  days  after  the  inoculation  into  the  brain.   N  =  nucleus  of  neuron 
repulsed  by  the  'cyst';  n  =  nucleolus;  C  =  'C'yst\  this  'cyst'  is  the  degenerated  nucleus 
of  a  mononuclear  cell  which  has  succeeded  in  penetrating  the  nerve-cell. 
Fig.  4.   Staining,  toluidin  blue-eosin.    1/12  oil  immersion,  oc.  4. 

Guinea-pig  85e  dead  82  days  after  cerebral  infection.   Section  of  medulla  oblongata. 
N  =  nucleolus;  C  =  corpuscle  of  Joest-Degen;  H  =  halo  around  the  corpuscle. 
Fig.  5.   Staining,  Mann.   1   12  oil  immersion,  oe.  4. 

Rabbit  209  dead  71  days  after  the  inoculation  of  the  virus  into  the  testicle.  Neuron 
of  the  anterior  horn  of  the  spinal  cord  in  the  lumbar  region.  C  =  Intranuclear  cor- 
puscle (.Joest-Degen):  N=nucleolus. 

PLATE  II 
Fig.  1.  Staining,  toluidin  blue-eosin.   1/12  oil  immersion,  oc.  1. 

Rabbit  243  dead  22  days  after  inoculation  into  the  brain.    Section  of  the  cord 
anterior  horn  shoiving  degeneration  of  the  nerve-cells.    V  =  small  blood-vessel;  P  = 
protoplasm   of  degenerated  nerve-cells;   D  =  protoplasm  debris;   0  =  vacuole  in  the 
cytoplasm  of  a  degenerated  neuron ;  C  =  cytoplasm ;  L  =  lymphocytes. 
Fio.  2.   Staining,  Mann.    1/12  oil  immersion,  oc.  4. 

Macacus  rhesus  No.  1.  Neurons  of  the  anterior  horn  of  the  thoracic  region  of  the 
cord  containing  corpuscles  (.Joest-Degen  type).  N  =  nucleus  of  the  nerve-cell;  n  = 
nucleolus;  C'  =  corpuscle  (Joest-Degen)  surrounded  by  a  halo.  At  the  periphery  of  the 
corpuscle  a  blue  staining  area  is  seen. 

PLATE  III 
Fig.  1.   Staining,  Mann.    1/12  oil  immersion,  oc.  2. 

Macacus  rhesus  No.  1.  PyTamidal  cells  of  the  frontal  lobe  containing  oxyphilic  cor- 
puscles surrounded  by  a  halo.   N  =  nucleolus;  C  =  corpuscle  (Joest-Degen). 

Fig.  2.  Staining,  Mann.   1/12  oil  immersion,  oc.  4. 

Rabbit  298  dead  39  days  after  intracerebral  inoculation.  Cervical  region  of  the 
cord :  anterior  horn.  Formation  of  intranuclear  corpuscles  in  the  interior  of  the  nucleus 
of  the  neuron.    N  =  nucleolus;  C'  =  oxyphilic  corpuscles;  a  study  of  the  process  of  the 


90  DESCRIPTION  OF  PHOTOGEAPHS,  ETC. 

formation  of  the  corpuscles  of  Joest-Degen  has  shown  that  several  small  oxyphilic 

corpuscles  fuse  together  to  form  one  large  corpuscle  stirrounded  by  a  halo. 

Fig.  3.   Staining,  orange  G.  eosin,  polychrome  blue  (Unna).    1   12  oU  immersion,  oc.  4. 

Guinea-pig  99k  dead  19  days  after  intracerebral  inoculation.    A  lymphocj-te  has 
penetrated  into  the  cytoplasm  of  a  nerve-cell  of  the  Cornu  Amnionic.    N  =  nucleolu8; 
L  =  lymphocytes. 
Fig.  4.  Staining,  Mann.   J  1   12  oil  immersion,  oc.  4. 

Rabbit  80a  dead  28  days  after  intracerebral  infection.  Mid-brain,  oxj-philic  degenera- 
tion of  the  nuclear  chromatin  forming  a  homogeneous  "block'.  C  =  degenerated 
nucleus;  X=nucleolus;  S  =  satellite  cell. 

Fig.  5.  Staining,  Mann.   1,  12  oil  immersion,  oc.  I. 

ilacacus  rheJus  No.  1.  Spinal  ganglion  of  the  lumbar  region.  Corpuscles  of  type 
Joest-Degen  in  the  nucleus  of  the  ganglion  cells.  C  =  intranuclear  corpuscle  surrounded 
by  a  halo ;  N  =  nucleolus. 


I'rinlMi  under  the  authority  of  His  Majesty's  Station kby  Officb 
by  .lohn  Johnson,  at  the  I'nivpmity  Pre*),  Oxford. 


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^libij  <!Iouncil 
MEDICAL   RESEARCH    COUNCIL 

(Formerlij  Medical  Research  Committee,  National  Health  lusuranee.) 


LIST  OF  PUBLICATIONS 

September,  1928. 

The  following  publications  relating  to  the  work  of  the  Medical  Research  Council  can 
be  purchased  directly  from  H.M.  Stationery  Oflice,  at  the  following  addresses  : 
Adastral  House,  Klngsway,  London,  W.C.  2  ;  York  Street,  Manchester  ;  1  St.  Andrew's 
Crescent,  Cardiff;  120  George  Street,  Edinburgh;  15  Donegal  Square  West,  Belfast; 
or  through  any  bookseller.* 

In  addition,  numerous  memoirs  upon  work  aided  by  the  Medical  Research  Council  have 
appeared  in  Scientific  Journals  :   particulars  of  these  may  be  seen  in  the  Annual  Reports. 

ANNUAL  REPORTS 

Medical  Research  Committee,  Xos.  1-5,  1914-15  to  1918-19. 
Medical  Research  Council,  1919-20  to  1920-7. 

(Price  of  each  report  from  1920-1  to  1925-6,  3i'.  6d.,  post  free  3s.  8d. ;    192(i-7   [Cmd. 
3013],  oS.,  post  free  oS.  2(/.) 

SPECIAL  REPORTS,  &c. 
Alcohol  : 

No.  31.    Alcohol — Its  Absorption  into  and  Disappearance  from  the  Blood  under  different 

conditions.     By  E.  Mellanby.     [1919.]  Out  of  print. 

So.  34.    The  Influence  of  Alcohol  on  Manual  Work  and  Neuro-museular  Co-ordination. 

By  H.  M.  Vernon.    [1919.]    Price  2s.,  post  free  2s.  Id. 
No.  50.    The  Effects  of  .Mcohol  and  some  other  Drugs  during  Normal  and  Fatigued 

Conditions.    By  W.  McDougall  and  May  Smith.    [1920.]    Price  Is.,  post  free  Is.  Irf. 
(Book).     .-Mcohol:     its  .\ction  on   the   Human   Organism.    Second  Edition.    [1924.] 

Price  Is.  paper  covers.  Is.  Gd.  cloth  bound. 
Anaerobic  Bacteria  :  sec  Wound  Infections. 
Animals,  Diseases  of : 

No.  121.     Borna  Disease  and  Enzootic  Encephalo-Myclitis  of  Sheep  and  Cattle.     By 

S.  Nicolau  and  I.  A.  Galloway.     [1928.] 
Bacteriology  (Miscellaneous)  : 

No.  35.    The  Reaction  of  Culture  Media,  by  S.  R.  Douglas,  J.  W.  H.  Eyre,  P.  P.  Laidlaw, 

and  C.  G.  L.  Wolf.     Second  Edition,  re\Tsed  by  P.  P.  Laidlaw,  [1927.]     Price  Gd., 

post  free  7d. 
No.  49.    On  the  Destruction  of  Bacteria  in  Milk  by  Electricity.    By  J.  M.  Beattic  and 

F.  C.  Lew  is.    [1920.]    Price  9d.,  post  free  lOd. 
No.  51.    The  Laboratory  Diagnosis  of. \cute  Intestinal  Infcctions,iucluding  the  Principles 

and   Practice  of  the  .Agglutination   Tests.     By  the  Conunittce   upon   Pathological 

Methods.    [1920.]    Price  4s.  Gd.,  post  free  4s.  8rf. 
No.  64.   Catalogueof  the  National  Collection  of  Type  Cultures.   Second  Edition.    [1925.] 

Price  2s.,  post  free  2s.  Id. 
Blood  Physiology  : 

No.  72.    The  .\cid-base  Equilibrium  of  the  Blood.     By  the  Haemoglobin  Committee. 

[1923.]    Price  2s.,  post  free  2s.  Id. 
Sec  also  Shock,  Surgical. 

*  For  overseas  agencies,  see  p.  viii. 


Special  Reports — contmued. 

Caxicer  : 

No.  99.  .Vn  Investigation  into  the  Statistics  of  Cancer  in  Different  Trades  and  Profes- 
sions.   By  Matthew  Young  and  \V.  T.  Russell.    [1926.]    Price  Is.  Gd.,  post  free  Is.  7d. 

See  also  Radiuji. 
Cerebro-spinal  Fever  : 

No.  2.  Report  of  the  Special  Advisory  Cominittee  upon  Bacteriological  Studies  of 
Cerebro-spinal  Fever  during  the  Epidemic  of  1915.    [191G.]  Out  of  print. 

No.  3.  Bacteriological  Studies  in  the  Pathologj-  and  Preventive  Control  of  Cerebro-spinal 
Fever  among  the  Forces  during  1915  and  1916.  By  M.  H.  Gordon,  Martin  Flack, 
P.  Vi'.  Bassctt-Smith,  and  T.  G.  M.  Hinc  and  W.  ,J.  Tulloch.    [1917.]      Out  of  print. 

No.  17.  (I.)  A  Report  upon  the  Seasonal  Outbreak  of  Cerebro-spinal  Fever  in  the  Navy 
at  Portsmouth,  1916-17.  By  Paul  Fildes  and  S.  I,.  Baker.  (II.)  The  Treatment  of 
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No.  50.  Cerebro-spinal  Fever.  Studies  in  the  Bacteriology-,  Preventive  Control,  and 
Specific  Treatment  of  Cerebro-spinal  Fever  among  the  Military  Forces,  1915-19.  By 
M.  H.  Gordon  and  others.    [1920.]    Price  4s.,  post  free  -ki.  lid. 

Chemotherapy  :  see  Streptococcai.  Ixfections. 
Child  Life  (Antenatal  and  Postnatal  Investigations)  : 

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Brend,  L.  Findlay,  and  J.  Brownlee.    [1917.]    Price  Is.  Cd.,  post  free  Is.  7Jd. 
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Browne.     [1924.]     Price  2s.  Od.,  post  free  2s.  7Sd. 
No.  101.    Poverty,  Nutrition,  and  Growth  :    Studies  of  Child  Life  in  Cities  and  Rural 

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Price  10s. 
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No.  114.    Social  Conditions  and  Acute  Rhcuniutisni.     [1027.]    Price  2s.  Od.,  |>ost  free 

2s.  8d. 
No.  117.    The  Toxaemias  of  Pregnancy  :  A  Clinical  and  Biochemical  Study.     By  J.  N. 

Cniiekshank.  ,1.  Hewitt,  and  K.  L.  Couper.     [1927.]     Price  4x.,  post  five  -Is.  2d. 
No.  118.     The  Cause  of  Foetal  IXalli  in  144  Cases.     By  A.  C.  Palmer.     [1928.]      Price 

3».,  post  free  :{«.  2d. 
See  also  Nutrition  ;    Rickets. 
Dental  Disease  : 

No.  70.    The  Struclure  of  Teeth  in  relation  to  Dental  Disease.     By  .1.  Ilnward  Mununrry. 

[1922.)     Price  2v.,  post  free  2s.  Id. 
No.  97.    'I'he  Incidence  of  Dental  Disease  in  Cliildren.    By  the  Comniittee  fur  llir  Investi- 
gation of  Denial  Disease.     ( 192.>.|     Price  Is.  Od.,  post  free  Is.  Tjd. 
Diphtheria  : 

No.   115.    The  Prevention  of  Diphlheria.      By  .1.  (Jraliani   Forbes.     [1927. |     I'rite  2s.. 

p<i»t  free  2s.  1  jd. 
(Book).    Diphtheria  ;  its  Buclcriology,  Pathology,  and  Immunology.     By  the  Hactcrio- 

logicul  Conunittce.    [1023.]    Price  12s.  Od.,  jmst  free  13s.  3d. 
.S'>>  also  Epiuemioi.u(;y  (No.  75). 


Special  Reports — continued. 

Dysentery  : 

Reports  upon  Investigations  in  the  United  Kingdom  of  Dysentery  Cases  received  from  the 

Eastern  .Mediterranean  : — 
No.  4.    I.  Amoebic  Dysentery  and  the  Protozoological  Investigation  of  Cases  and  Carriers. 

By  Clifford  Dobell."    [1917.]  Out  of  print. 

No.  ;■>.    II.  Report  upon  878  Cases  of  Bacillary  Enteritis.    By  L.  Rajchman  and  G.  T. 

Western.    [1917.]  Out  oj  print. 

No.  G.    III.  Report  upon  recovered  Cases  of  Intestinal  Disease  in  the  Royal  Naval 

Hospital,  Ilaslar,  1915-lG.    By  Paul  Fildes  and  others.     IV.  Report  upon  combined 

Clinical  and  Bacteriological  Studies  of  Dysentery  Cases  from  the  Mediterranean.     By 

S.  R.  Douglas  and  L.  Colebrook.     [1917.]     Price  4s.  Orf..  post  free  in.  7id. 
No.  7.    V.  Report  upon  2,360  Enteritis  '  Convalescents '  received  at  Liverpool  from 

various  Expeditionary  Forces.     By  E.  GljTin  and  others.     [191S.]     Price  6s.,  post 

free  6s.  '2d. 

No.  15.    .\  Study  of  1,300  Convalescent  Cases  of  Dysentery  from  Home  Hospitals  :  with 

special  reference  to  the  Incidence  and  Treatment  of  Amoebic  Dysentery  Carriers.    By 

Clifford  Dobell,  H.  S.  Gettings,  Margaret  W.  Jepps,  and  .1.  B.  Stephens.    [1918.]    Price 

Is.  3d.,  post  free  Is.  4d. 
No.  29.    .\  Contribution  to  the  Study  of  Chronicity  in  Dysenter\'  Carriers.     By  W. 

Fletcher  and  Doris  L.  Mackinnon.    [1919.]    Price  9rf..  post  free  lOrf. 
No.  30.    An  Investigation  of  the  Flexner-Y  Group  of  Dysentery  Bacilli.     By  S.  H. 

Gettings.    [1919.]    Price  Is.,  post  free  Is.  Irf. 
No.  40.    Studies  of  Bacillary  Dysenterj'  occurring  in  the  British  Forces  in  Macedonia. 

By  L.  S.  Dudgeon  and  others.    [1919.]    Price  3s.,  post  free  3s.  IW. 
No.  42.    .\  Study  of  the  Serological  Races  of  the  Flexner  Group  of  Dysentery  Bacilli. 

By  Sir  F.  W.  Andrcwes  and  .\.  C.  Inman.    [1919.]    Price  2s.,  post  free  2s.  lid. 
See  also  Food  Poisoning. 

Encephalitis  : 

No.  108.  The  Sheffield  Outbreak  of  Encephalitis  in  1924.  [1926.]  Price  Is.  9d.,  post 
free  Is.  lOW. 

Enteric  Infections  : 

No.  9.    .A  Report  upon  the  Use  of  .Atropine  as  a  Diagnostic  .\gent  in  T\-phoid  Infections. 

By  H.  F.  Marris.    [1917.]    Price  Is.,  post  free  Is.  Id. 
No.  48.    .A  Report  on  the  probable  Proportion  of  Enteric  Infections  among  Undiagnosed 

Febrile  Cases  invalided  from  the  Western  Front  since  October  1916.    By  W.  W.  C. 

Topley,  S.  G.  Platts,  and  C.  G.  Imrie.    [1920.]    Price  3s.,  post  free  3s.  lid. 
See  also  Bactekiology  ;    Food  Poisoning. 

Epidemiology  : 

No.  75.    The  Schick  Test,  Diphtheria  and  Scarlet  Fever.    By  S.  F.  Dudley.  [1923.]  Price 

Is.,  post  free  Is.  lid. 
No.  111.    The  Spread  of  Droplet  Infection  in  Semi-isolated  Communities.     By  S.  F. 

Dudley.    [1926.]    Price  Is.  6d.,  post  free  Is.  7|d. 
No.  120.     An  Inquiry  into  the  Relationship  between  Housing  Conditions  and  the  In 

cidence  and  Fatality  of  Jleasles.    By  J.  L.  Halliday.    [1928.]    Price  Is.,  post  free  Is.  Id. 
See  also  S.m.41.l-pox  ;    Tuberculosis  ;    etc. 

Flying,  Medical  Problems  of  : 

Reports  of  the  .\ir  Medical  Investigation  Committee  : — 

No.  28.  The  Sense  of  Balance  and  Stability  in  the  Air.  By  Henrj- Head.  [1919.]  Price 
9d.,  post  free  lOd.    (Included  in  No.  53.) 

No.  37.  The  Effects  of  Diminished  Tension  of  Oxygen,  with  especial  reference  to  the 
.•\eti\ity  of  the  Adrenal  Glands.  By  C.  H.  Kellaway.  Tlie  Ear  in  relation  to  certain 
Disabilities  in  Flying.    By  S.  Scott.    [1919.]      Price  Is.   post  free  Is.  Id. 


Special  Reports — continued. 

No.  53.  The  Medical  Problems  of  Flying  (including  reports  on  oxygen  want,  selection 
of  candidates  for  Dying,  sense  of  balance,  and  flying  strain).  [IIVJO.]  Price  6.s\, 
post  free  6s.  -id. 

No.  81.    Tlio  Application  of  tlie  Air  Force  Physical  EITiciency  Tests  to  .Men  and  Wcmifii. 

l!y  I,.  IJ.  (  rij.ps.    [192i.]    Price  Is.  Gd.,  post  free  Is.  7Jrf. 
Food  Poisoning  : 

No.  24.    A  Report  on  the  Investigation  of  an  Epidemic  caused  by  Bacillus  aertrj'cke. 

By  H.  Marrian  Pcrr>-  and  II.  L.  Tidy.    [1919.]    Price  9d.,  post  free  lOrf. 
No.  91.    An  Investigation  of  the  Salmonella  Group,  with  Special  llefcrence  to  Food 

Poisoning.    By  W.  G.  Savage  and  P.  Bruce  White.    [1925.]    Price  3s.  ad.,  post  free 

3s.  Sd. 
No.  92.    Food  Poisoning  :    a  Study  of  100  Recent  Outbreaks.     By  \V.  G.  Savage  and 

P.  Bruce  White.     [1925.]     Price  2s.  Gd.,  post  free  2x.  8rf. 
No.  103.    Further  Studies  of  the  Salmonella  Group.    By  P.  Bruce  While.    [192(!.]    Price 

5s.,  post  free  5s.  2Jrf. 
Haemoglobin  :  see  Blood. 
Heart  Disease  : 

No.  8.    Report  upon  Soldiers  returnc<l  .as  Cases  of  '  Disordered  .Vction  of  the  Heart  ' 

(n..\.H.),  or  Valvular  Disease  of  the  Heart.     By  Sir  Thomas  Lewis.     [1917.]     Price 

Is.,  post  free  Is.  Id. 
Industrial  Fatigue  : 

(The  .\nnual  Reports  of  the  Industrial  F.atiguc  Research  Board,  and  special  reports  on 

particular  subjects, are  published  for  the  Council  in  separate  series.    The  subjects  dealt 

with  include   accident   causation,  rest   pauses,  spells   of  work,  movement   study, 

vocational  selection,  and  problems  of  particular  industries.     .\  list  can  be  supplied 

on  application  to  the  Secretarj'  of  the  Board,  15  York  Buildings,  Adclphi.  W.C.  2.) 
Influenza  : 

No.  30.    Studiesof  Influenza  in  Hospitals  of  the  British  .Armies  in  France,  1918.    [1919.] 

Price  3s.  Gd.,  post  free  3s.  8d. 
No.  C3.    Studies  in  the  Actiologj-  of  Kpidemic  Inlluenza.     By  .1.  Mcintosh.     [1922.) 

Price  2s.  Gd.,  post  free  2s.  7d. 
Jaundice : 

.Nn.   113.    SpirochaetalJaiuulice.     By  G.  Buchanan.    (1927.)    Price  4.s-.,  ikisI  free  Is.  2d. 

Malaria  :  see  Quinine. 

Maternal  Mortality:  xrc  Ciiii.n  I-ifk  and  Sriiii'rooui'Ai.  iNKrcxKiNs. 

Measles:   <(<  l^nni-Mioi.or.v  (No  120.) 

Miners'  Dietaries  :  see  Nutrition. 

Miners'  Diseases,  etc.  : 

No.  89.  ReiHirt  on  Miners'  '  Beat  Knee  *,  '  Beat  Hand  ',  and  '  Beat  Elbow  '.  By  K.  I,. 
Collis  and  T.  L.  Llewellyn.    [192t.]    Price  Is.  Od.,  post  free  Is.  7d. 

Miners'  Nystagmus  :  sec  Vision. 
.Sic  also  .Iainmu  I    (No.  113). 
Nephritis  : 

No.   13.    Albuminuria  and  War  Nephritis  among  British  Troops  in   Friim  c.      Hy   II. 
.\laclx-an.    [1019.]    Price  2s.  fid.,  post  free  2s.  8d. 
Nerve  Injuries  : 

HipiirLs  of  the  Commitlec  upon  Injuries  to  the  Nervous  System: — 

•No.  51.    The  Diagnosis  and  Treatment  iif  Peripheral  Nerve  Injuries.    (1920.)    Price  2s., 

[lost  free  2».  1  Jd. 
No.  88.    Injuries  of  the  Sjiinal  Cord  and  Cauda  lupiina.     |102t  )     Price  Is.  Od.,  post 
free  Is.  7Jd. 


Special  Reports— continued. 

Nutrition  : 

\o.  1:1.    An  Enquiry  into  the  Composition  of  Dietaries,  with  special  reference  to  the 

Dietaries  of  Munition  Workers.    Hy  Viscount  Dunluee  and  Major  Greenwood.    [1918.] 

I'riee  9(/.,  post  free  lOrf. 
No.  :i8.    neport  on  tlie  Present  State  of  Knowledge  of  Accessory  Food  Factors  (Vitamins). 

By  a  Committee  appointed  jointly  by  the  Lister  Institute  and  Medical  Research  Council. 

Second  Edition.    [1924.]    Price  is.  r,fl.,  post  free  4.9.  8^rf. 
No.  87.    Report  on  the  Nutrition  of  Miners  and  their  Families.    By  the  Committee  upon 

Quantitative  Problems  in  Human  Nutrition.    [1924.]    Price  l,t.  3rf.,  post  free  l.f.  4,(1. 
No.  10.').    Diets  for  Boys  during;  the  School  A^e.    By  II.  C.  Corry  Mann.    [1920.]    Price 

2.9.  Gd..  post  free  2.9.  7id. 
See  also  Child  Life  ;    Rickets. 

Pituitary  Extract  ;  sec  STAND.\nDS. 

Pneumonia  : 

No.  79.  Bacteriological  and  Clinical  Observations  on  Pneumonia  and  Empyemata,  with 
special  reference  to  the  Pneumoeoceus  and  to  Serum  Treatment.  By  E.  E.  Glynn  and 
Lettice  Digby.    [192,3.]    Price  5.9.,  post  free  5.9.  3d. 

Pneumothorax,  Artificial  :  sec  Tuberculosis. 
Print,  Legibility  of  :  sec  Vision. 

Protozoan  Infections  : 

No.  59.  A  Report  on  the  Occurrence  of  Intestinal  Protozoa  in  the  inhabitants  of  Britain. 
By  Clifford  Dobcll.    [1921.]    Price  2.9.,  po.st  free  2.9.  IJrf. 

Quinine  : 

No.  90.  Clinical  Comp.ari.sons  of  Quinine  and  Quinidine.  By  the  Committee  upon 
Cinchona  Derivatives  and  Malaria.    [1925.]    Price  1.9.,  post  free  1.9.  Id. 

Radium  : 

No.  02.  Medical  Uses  of  Radium  :  Studies  of  the  Effects  of  Gamma  Rays  from  a  large 
Quantity  of  Radium.    By  various  authors.    [1922.]    Price  5s.,  post  free  5s.  3rf. 

No.  90.  Medical  Uses  of  Radium  :  Summary  of  Reports  from  Research  Centres  for  192;?. 
[1924.]    Price  Is.,  po.st  free  1.9.  Id. 

No.  102.    Medical  Uses  of  Radium  :    Summarj'  of  Reports  from  Research  Centres  for 

1924.  [1920.]    Price  Is.  Gd.,  post  free  Is.  7d. 

No.  112.    Medical  Uses  of  Radium  :     Summary  of  Reports    rom  Research  Centres  for 

1925.  [1920.]    Price  Is.  3rf.,  post  free  Is.  4rf. 

No.  lie.  Medical  Uses  of  Radium :  Summ.iry  of  Reports  from  Research  Centres  for 
1920.     [1927.]     Price  is.  3rf.,  post  free  is.  4d. 

Rheumatism  :  see  Child  Life  (No.  114). 
Rickets  : 

No.  20.  A  Study  of  Social  and  Economic  Factors  in  the  Causation  of  Rickets,  with  an 
Introductory  Historical  Survey.     By  L.  Findl.ay  and  Margaret  Ferguson.     [1918.] 

Out  of  print. 
No.  01.    Experimental  Rickets.    By  E.  Mellanby.    [1921.]    Price  4s.,  post  free  4s.  2rf. 
No.  08.    Rickets  :    the  Relative  Importance  of  ICnvironmcnt  and  Diet  as  Factors  in 

Causation.    By  H.  Corry  Mann.    [1922.]    Price  2s.  Gd.,  post  free  2s.  7Jrf. 
No.  71.    The  Aetiology  and  Pathology  of  Rickets  from  an  experimental  point  of  view.  By 

V.  Korenchevsky.    [1922.]    Price  4s.,  post  free  4s.  3d. 
No.  77.    Studiesof  Rickets  in  Vienna,  1919-22.    [1923.]    Price  7s.  Od.,  post  free  7,9.  lOirf. 
No.  93.    Experimental  Rickets  :    The  Effect  of  Cereals  and  their  Interaction  with  other 
factors  of  Diet  and  Environment  in  producing  Rickets.     By  E.  Mellanby.     [1925.J 
Price  3s.,  post  free  3s.  8d. 


5pecial  Reports— fOH///iH<-(/. 
Salvarsan  :  see  Venereal  Diseases  ami  .STiti-.rTococcAi.  Inkfctioxs. 
Shock,  Svirgical  : 

Kcports  of  the  C'cuiiniiUec  on  Sur<;io;il  Shock  and  Allied  Condition!; : — 
No.  25.    Wound-Shook  .and  Haeniorrhajjo.     (lill!).]     Trice  U.,  post  free -l-s.  5J(J. 
No.  20.    Trannial ic  Toxaemia  as  a  Factor  in  Shock.    [1919.]     Price  Is.,  post  free  Is.  !</. 
No.  27.    Klood   \'oluinc   Changes   in   Wound-Shock  and    I'riniary    Haemorrhage.     By 
N.M.Keith.    [1919.]    Price  <•(/.,  post  free  lOrf. 

Small-pox  : 

No.  98.    Studies  of  the  Viruses  of  Vaccinia  and  Variola.     l?y  M.  II.  Cordon.     [1925.] 

Price  ."is.  Gd.,  post  free  3s.  8J</. 
No.  100.    Small-pox  and  Climate  in  India  :    Forecasting  of  Kpidemics.    Hy  Sir  Leonard 

Rogers.    [1920.)    Price  2s.,  post  free  2s.  lirf. 

Standards,  Biological  : 

No.  09.    Pituitary  Extracts.    Hy  J.  11.  Burn  and  H.  H.  Dale.    [1922.]    Price  Is.  Grf..  post 

free  Is.  7d. 
See  also  Venereal  Disr,\ses  (No.  44). 

Statistics   (Miscellaneous). 

No.  IG.  A  Report  on  the  Causes  of  Wastage  of  Labour  in  .Munition  Factories.  By  Major 
Greenwood.    [1918.]    Price  Is.  Cd.,  post  free  Is.  7d. 

No.  00.  The  Use  of  Death-rates  as  a  Meivsure  of  Hygienic  Conditions.  By  John  Brownlee. 
[1922.]    Price  3s.,  post  free  3s.  1  kit. 

No.  95.  Internal  Migration  and  its  EfTeels  upon  the  Death-Rates  :  with  Special  Refer- 
ence to  the  County  of  Essex.    By  A.  B.  Hill.    [1925.]    Price  3s.  G</.,  post  frt>e  3s.  Sd. 

Streptococcal  Infections : 

No.  119.  A  Study  of  Some  Organic  Arsenical  Compounds  with  a  view  to  their  Use  in 
certain  Streptococcal  Infections.  ByL.  Colcbrook.  [192S.]  Price  Is.  .1//..posf  five  Is.  k/. 

T.N.T.  Poisoning  : 

No.  11.    The  Causation  and  Prevention  of  Tri-nitro-tolucne  (T.N.T.)  Poisoning.     By 

Benjamin  Moore.    [1917.]    Price  Is.,  post  free  Is.  IJd. 
.No.  58.    T.N.T.  Poisoning  and  the  Fate  of  T.N.T.  in  the  .-\nimal  Body.     Ity  W.  J. 

O'Donovan  and  others.    [1921.]    Price  3s.,  post  free  3s.  1  Id. 

Tuberculosis  : 

No.  1.    First   Report  of  the  Special  Investigation  Committee  upon  the  Incidence  of 

Phthisis  in  relation  to  Occupations. — The  Boot  and  Shoe  Trade.    [1915.]    Price  3d., 

post  free  3Jd. 
No.  18.    An  Investigation  into  the  Epidemiology  of  Phthisis  Pulmonalis  in  Great  Britain 

and  Ireland.     Parts  I  and  II.     By  John  Brownlee.    [1918.]     Price  Is.  3d.,  post  free 

Is.  4Jd. 
No.  22.    .An  Inquiry  into  the  Prevalence  and  Aetiology  of  Tuberculosis  among  Industrial 

Workers,  with  special  reference  to  Female  .Munition  Workers.    By  Major  Greenwood  and 

A.  I'",.  Tebb.     (191!>.|     Price  Is.  «d.,  post  free  Is.  7d. 
No.  33.    Pulmonary  Tuberculosis  :    Mortality  after  Sanatorium  Treatment.    By  Noel  D. 

Bardswell  and  J.  II.  U.  Thompson.     [1019.]     Price  2.«.,  post  free  2s.  2d. 
No.  4U.    An  Investigation  into  the  Epidemiology  of  Phthisis  in  (ireat  Britain  and  Ireland. 

Part  HI.     By  John  Brownlee.     [1920.)    Price  2s.  «d.,  post  free  2s-.  7jd. 
No.  07.     Report  on  .Artiticial  Pneumothorax.     [1922.)     By  L.  S.  T.  Burrcll  and  .\.  S. 

MacNalty.     Price  2v.  Od.,  post  free  2s.  Hd. 
No.  70.    Tuberculosis  in  Insured  Persons  accepted  forTreutmcnt  by  the  City  of  llrndford 

Health  Committee.    By  H.  Vnllow.    (1023.)    Price  Od.,  post  free  7d, 


Special  Reports — coniiinicd. 

No.  83.    Tuberculosis  of  the  Lan,nx.    By  Sir  St.  Clair  Tlionisoii.    [1924.]    Price  2s.  0<i., 

post  free  2s.  8d. 
No.  8.5.  An  Inquiry  into  the  After-Histories  of  Patients  treated  at  the  Brompton  Hospital 

Sanatorium  at  Frimley,  during  the  years  1905-14.     By  Sir  P.  H.-S.  Hartley,  R.  C. 

Wingfield,  and  J.  H.  K.  Thompson.    [1924.]    Price  Is.  dd.,  post  free  Is.  7d. 
No.  94.    Tuberculin  Tests  in  Cattle,  with  special  reference  to  the  Intradermal  Test.    By 

the  Tuberculin  Committee.    [1925.]    Price  3s.,  post  free  3s.  .Sd. 

Venereal  Diseases  : 

No.  14.   The  Wassermann  Test.     By  the  Committee  upon  Pathological  Methods.     Xea: 

Edition.     [1921.]     Price  Is.,  post  free  Is.  Id. 
No.  19.    The  Laboratory  Diagnosis  of  Gonococcal  Infections.    Methods  for  the  Detection 

of  Spironenia  pallidum.     By  the  Bacteriological  Committee.     Xew  Edition.     [1923.] 

Price  Is.  6d.,  post  free  Is.  7id. 
No.  21.  The  Diagnostic   Value   of  the   Wassermann   Test.    By  the   Committee  upon 

Pathological  Methods.    [1918.]    Price  Is.,  post  free  Is.  Id. 
No.  23.    An  Analysis  of  the  Results  of  Wassermann  Reactions  in  1,435  Cases  of  Sj'philis 

or  Suspected  Syphilis.    By  Paul  Fildes  and  R.  J.  G.  Parnell.    [1919.]    Price  2s.,  post 

free  2.^.  Id. 
No.  41.    (I.)  An  Investigation  into  the  Ultimate  Results  of  the  Treatment  of  Syphilis  with 

Arsenical  Compounds.    By  Paul  Fildes  and  R.  J.  G.  Parnell.     (II.)  A  Clinical  Study 

of  the  To.xic  Reactions  which  follow  the  Intravenous  Administration  of  '914'.     By 

R.  J.  G.  Parnell  and  Paul  Fildes.    [1919.]    Price  2s.,  post  free  2s.  Id. 
No.  44.    Reports  of  the  Special  Committee  upon  the  Manufacture,  Biological  Testing,  and 

Clinical  Administration  of  Salvarsan  and  of  its  Substitutes.   I.    [1919.]    Price  Is.,  post 

free  Is.  Id. 
No.  45.    Unsuspected  Involvement  of  the  Central  Nervous  System  in  Syphilis.    By  Paul 

Fildes,  R.  J.  G.  Parnell,  and  H.  B.  Maitland.    [1920.]    Price  Is.,  post  free  Is.  Id. 
No.  47.    The  Accuracy  of  Wassermann  Tests,  applied  before  and  after  death,  estimated  by 

Necropsies.     I.  The  Wassermann  Test  applied  before  death.     By  H.  M.  Turnbull. 

[1920.]    Price  2s.  (id.,  post  free  2s.  7id. 
No.  55.    (I.)  Results  of  the  Examination  of  Tissues  from  Eight  Cases  of  Death  following 

Injections   of  Salvarsan.    By  H.  M.  Turnbull.     (II.)    The    Influence    of  Salvarsan 

Treatment  on  the  Development  and  Persistence  of  Immunity,  as  indicated  by  Measure- 
ments of  .Agglutinins.     By  E.   W.  .Ainlcy   Walker.     [1920.]     Price   3s.,   post  free 

3s.  lid. 
No.  66.    Toxic  Effects  following  the  Eniplo>Tnent  of  Arsenobenzol  Preparations.   By  the 

Salvarsan  Committee.    [1922.]    Price  2s.,  post  free  2s.  IJd. 
No.  78.    The  Serum  Diagnosis  of  Syphilis  :     The  Wassermann  and  Sigma  Reactions 

compared.    [1923.]    Price  5s.  6d.,  post  free  5s.  9d. 
No.  107.    The  Effect  of  Treatment  on  the  Wassermann  Reactions  of  Syphilitic  Patients. 

By  E.  E.  Glynn,  R.  E.  Roberts,  and  P.  M.  Bigland.    [1920.]    Price  3s.  6d.,  post  free 

3s.  8d. 

Ventilation,  etc.  : 
No.  32.    The  Science  of  Ventilation  and  Open-air  Treatment.    Part  I.    By  Leonard  Hill. 

[1919.]    Price  10s.,  post  free  10s.  o^d. 
No.  52.    The  Science  of  Ventilation  and  Open-air  Treatment.    Part  II.    By  Leonard  Hill. 

[1920.]    Price  6s.,  post  free  Cs.  4^d. 
No.  73.    The  Kata-thcrmomctcr  in  Studies  of  Body  Heat  and  Elficiency.    By  Leonard 

Hill  and  others.    [1923.]    Price  5s.,  post  free  5s.  2jd. 
No.  100.    Methods  of  Investigating  Ventilation  and  its  Effects.    By  II.  M.  Vernon  and 

others.    [1920.]    Price  2s.,  post  free  2s.  IJd. 

Vision  : 

No.  65.    First  Report  of  the  .Miners' Nystagmus  Committee.    [1922.]    Price  Is.  6d.,  post 
free  Is.  7ld. 


3pecial   Reports — continued. 

No.  80.    Second  Report  of  the  Miners'  Nystagmus  Committee.    [19:23.1    Price  9d.,  post 

free  lOd. 
No.  104.    lUuniination  and  Visual  Capacities.    By  R.  J.  Lythgoe.    [1U2G.]    Price  Os.  Oil.. 

post  free  '2s.  7  id. 
No.  110.     The  Legibility  of  Print.     By  R.  L.  Pyke.    [1926.]    Price  is.,  post  free  is.  2d. 

Vitamins  :  see  Nutrition. 

Wasseimann  Test  :  sec  Vi^nereal  Diseases. 

Wound  Infections  : 

No.  12.    The  Classification  and  Study  of  the  .Vnaerobic  Bacteria  of  War  Wounds.     By 

J. -Mcintosh.    [1917.]  Out  of  print. 

No.  39.    Report  on  the  .Anaerobic  Infections  of  Wounds  and  the  Bacteriological  and 

Serological  Problems  arising  therefrom.    By  the  Committee  upon  .Vnaerobic  Bacteria 

and  Infections.    [1919.]    Price  (is.,  post  free  lis.  Sid. 
No.  57.    Studies  in  Wound  Infections.    By  S.  R.  Douglas,  .\.  Fleming,  and  L.  Colebrook. 

[1920.]    Price  45.  Gd.,  post  free  4s.  S^d. 

The  following  books  were  published  under  the  direction  of  the  .Medical  Research  Com- 
mittee and  are  obtainable  from  the  publishers  named  : 

Milk  and  its  Hygienic  Relations.  By  Janet  E.  Lane-Cla\-pon.  PrieeUs.net.  [Longmans. 
Green  &  Co.] 

The  .Amoebae  living  in  Man.  By  ClilTord  Dobell.  Price  7».  6c/.  net.  [Bale,  Sons  & 
Danielsson,  Ltd.] 

The  Intestinal  Protozoa  of  Man.  By  Clifford  Dobell  and  V.  W.  O'Connor.  Price  1  ,".x.  net 
[Bale,  Sons  &  Danielsson,  Ltd.] 


OVERSE.\S  .AGENCIES  OF  II.M.  ST.ATIONERY  OFFICE 

IRISH  FREE  ST.ATE  :    Messrs.  Eason  &  Son,  Ltd.,  40;  41  Lr.  Ot  onnell  Street,  Dublin. 

CAN.AD.A  :  The  Imperial  News  Company,  Ltd.,  235  F'ort  Street,  Winnipeg  :  77  St.  An- 
toinc  Street,  Montreal;  517  Burrard  Street,  Vancouver;  \Vm.  Dawson  Sub.scriplion 
Scr\-icc,  Ltd.,  91-93  Queen  Street  East,  Toronto. 

AISTR.ALI.A  :  .Messrs.  .Albert  &  Son,  Ltd.,  180  Murray  Street,  Perth,  \Vcsterii  .Australia. 
.Messrs.  Gordon  &  Gotcli,  Ltd.,  Blyth  Street,  .Adelaide  ;  Queen  Street,  Brisbane  ;  Little 
Collins  Street,  Melbounie  ;  Barrack  and  Clarence  Streets,  Sydney  ;  Cimiticre  Street, 
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