The Laboratory Diagnosis of Dysentery Occurring in South African

TYD~KRIF VIR ]
G E..'aESIU1NDE .
B.A.
LABORATORY DIAG NOSI S OF DY SENTERY.
The Laboratory Diagnosis of Dysentery
Occurring in South African Troops
in the Middle East.
By
)1. H . FII'LAYSOX , B. S c . , l\ I. B ., CH.B. , D . P
Major, S. d .J/. C., Pathologis t , J/ .E .F.
.H .,
H E primary purpose of t hi s commun icat ion is to outline
the met hod s used in a So uth Af ri can Hos pit al Lab or a tory
T
These
in the Midd le East for the di agnosis of dysent ery.
met hods are base d on D.M.S. , l\!.E. T echni cal Inst ru ctions,
1941, N o. 59, and on methods used by Colo nel J. S. K. Boyd,
H. A.M .C. , and have been ve ry sat isfact ory.
Dysentery in the l\liddle E a st , as in South A fr ica, may be
caused by h elmi nthic, protozoa l, or bact erial infecti on .
(1) H ELmNTHIc DYSEXTERy.
My ex perience of th is cond it ion was limited t o one case whi ch
was di scover ed in the course of a routine exa mina t ion of fift yfive kitchen p ersonn el for dysentery ca r riers . The pat ie nt,' a
nat iv e fr om the T zaneen D istrict of th e T ransvaal, was
inf ect ed with Schis to soma man soni. The large lateral spine
eggs of t he parasite were very num er ou s. a nd were eas ily
detect ed in the mu cus in the stoo l und er th e low-po wer
object ive .
In ra re ca ses
h omatob ium has been known to cause
dysenteric symptoms. Althou gh seve n cases of in fe ct ion of
the gen it o-urinary sy st em with S. lu smat ob iu m were det ect ed .
no. evidence o f dysentery caused by th is organism was obtaine d.
(2) PROTOZOAL DYSE XTERY.
(a ) A mab ic Dyswtery .-Elev en cases of infect ion with E.
h iMolytica w ere enco untered. In all bu t one o f these cases
sim ilar and
t he naked-eye appearance of th e stools wa
chara ct erist ic of amcebic dvsenter v, These stools consist ed of
sem i-solid b ro wn frecal m~ tter \~i t h some bloo d an d bro wn
mucus . Th e st ools were very foul -smell ing. T he one excenti on was a stool from a case of douhle in fecti on with E,
h i.~ t oly t ica and B. d y.<entu i,p Shiga.
This stool consist ed of
gla iry mucu s an d bl ood and could best be likened to "sago
wit h' red -currant j elly " . N o di ffi culty was found identifyi ng
an E. histobttica infection microscopically. F or micros copic
examinat ion . a port ion of mucus fr om th e stool was was hed
in warm saline, and any area conta ining dark ne crot ic patches
ca ref ullv examined un der a cover-glas s sealed wit h va selin e.
Th e absence of any d efinite cellular exudate such as is present
in acut e ba cilla ry dysentery st ools was an import ant fea t ure.
" ' h ilst few polymorphs wer e seen , the nn mher of mononuclears, macropha ges and re d cell s varied in diff erent specimen s. I n a f ew specimens C harcot- Leyd en cryst als were seen.
T he one ch a racteri st ic feat nr e was t he presence of actively
Gene rally . whe n B.
motile arncebre con t a ining red cells.
I. istol!/t ica we re fo un d, they w~re n umerous. Someti mes a
specimen of m ucus f rom a stool disc lose d one amoeba. In such
cases a second or t hi rd sp eci men oft en showed an area cont a ining num erous a ct ive amreba-,
Two important technical points were strictly obser ved :
Firstly, all st ools were brought to th e lab oratory in bed -pans
within half an hou r of the sto ol be ing passed.
econd ly. a
hot tle of sa line was kept co nsta nt ly in t he in cubator. so t hat
t he mucus could be wa hed with warm sa line immediatelv on
•
a rr iva l in the laborat orv .
I n three cases the prpse nce of E . hist oly t iert cys ts was
di a gn o ed. T he e ca se ha d a pr evious hi story of E. h ietobrtica
infect ion. T he examina tio n for cysts wa carr ied out by rubbing
up a small portion of f:Pees in a watchjrlass with a 1 per cent.
solution of iodine and pota ium iodide. A drop of this
suspension was mounted in th e usu al manner and ex am ined
for t he pre sence of cys ts.
I n diagnosing t he pres ence of cys t s no cell over 14 microns
in diameter was con idered . and examina tion with th e
micrometer eyepiece wa found ext re melv use ful.
(b) Flagellate Dy sentery . -T wo ca ses' o f infection with
Giardia Lamblia were observed . I n nei th er case was a n v blood
or mucus noticed in the stool, nor was there any evi de nce of
cell ular exudate. 'I'richomon as hominis an d Chiloma.< ti x mesnili
we re frequently observed in stools a nd on occa ion alo ng wit h
E. h istolytica. N on e of t hese flagell ates cou ld be accuse d of
ca using a true dysenteric co ndit ion.
(3)
[ JULIE
11 1942.
25I
BA CIL UR Y D YSE :---rER Y.
This cond it ion accounted for the maj ority of t he dys entery
st ools ex amined . I n man v case s th e condition wa s of a mild
natu re and a mount ed to ~ " d iarr hrea", or what is k nown in
the Ca pe as .. appelkoos siekte" . The na ked-eye ap peara nce
of th e stoo ls was charact erist ic. I n these mil d cases t he stools
were of la rge volu me , wat ery a nd pale brown in colour . Small
fla kes of mu cus were pr esen t but no macr oscopic blood was
'
evident.
I n the more sev ere cond iti ons th e st ool consist ed of blood
a nd mucus. F ew of our cases sho wed much macroscopic blood.
T he presence of cop iou s blood is usually an ind ica tion of
.higa in fection. Only one case of B. dy sellt eri,,' ltiga infect.ion was encountered .
I n a fe w cases of more sev ere d vsent er v th e stoo ls were
wa tery , of a yellowish-bro wn colour, ' and co ntained flak es o f
mu co-pus and green ish shreds of mu cus.
T he microscopi c appearance of t he sto ols was extremely
chara ct er ist ic. In a high propor t ion of cases an immedi at e
di agnosi s of the condition was ma de fr om the mi croscopi c
examina tio n an d a repor t forwarde d at once to t he ward . I n
t h is way corre ct treatm en t was in stitu t ed at the earl iest
poss ib le stage of th e infection . A flake of mucus was selecte d
fr om t he stoo l and examined in saline un der a cover-glass.
Th e outs tand ing fe atu re o f t he micr oscopic appearance was
t he . cell ul.a r ity of th e field. The number of red cells present
vari ed WIth t he mu cus sele cte d, e.lJ. a clear piece of mu cus
showed few red cells, a blood -sta ine d shred showed man v.
~\ part f ro m th e red cells, at lea st 90 per cent. of the rema inIlIg cell s wer e pus cells. A ma ll n umber of epithelial cells
macropha.g e? an.d eosinophils were also present, but th~
characterist ic pictu re was tha t o f an acut e inflamrnat orv
exu date, and was a typical .. ba cillary ex ndate".
•
In more ad van ced cases t he number of pus cells decrea sed ,
mor e mononu clear cells were een a nd erv th roc vtes becam e
fewer. T his type of exudate wa s known 'as an' .. ind efinit e
exudate " and was o f no d iagn ostic valu e as an indi cat ion of
bacilla ry dys ent ery , in vie w of its r esembl an ce to th e exu da te
tools showin g an .. ind efinite
found in a moebic dy en te ry.
exudate " were . howev er, cult u red , a nd fr om a certa in port ion
dysentery bacilli wer e isolate d.
It is essen t ial th at a bso lute ly fres h stoo l specimens should
be sub mitted for exa mina t ion if a h igh percen tage of isola t ions
are t o be obt ai ned. I nst ruct ions were, th e refore. i. sued t hat
a ll " d ia rrhreic " s tools sho uld be submitted im med iato lv to
th e lahoratory in a bed -pan, and that on no account sliould
more than thi rty minut es ela pse betw een t he passing of the
stool a nd its a rr iva l at the lahoratorv.
T he pr esence of cresolie d isinfectant in t he bed -pans was
found a t one st a ge to inhibit ha ct eri al gro wt h. Th is was
remed ied by th oroug h wash ing o f t he pa ns in hot wat er a nd
sub stituting chlor ide of lime for r-resolic di sin fectant.
Cult ures wer e mad e on )l cConkev's medium . whi ch was
preferred t o litmus-lact ose-tau rocholate a gar. A portion of
mucus was fished from th e stoo l a nd well wash ed in ste r ile
saline to ~e t ri d of excess H. coli . Tt was th en r ubbed on th e
surface of th e plat e. Th e plat inum loop was th en flamed a nd
th e rubhed ar ea touch ed wit h th e loop . with which a second
a rea o f th e plat e was st roked. Frequ ent ly t he loop was aga in
t1 arned a nd th e second st rok ed a rea touched with the loop. a nd
from thi s inocula t ion a th ird area st roked . I t was found t hat
with . thi s techn iqu e one ha lf o f a 3 ' ·inch ~ l c C onk ev plate
could be used for each stool, although it was p re fera ble t o
use one plate for each stool. T he need for washing mucus
be!ore plat e in oculat ion .ca nnot be ove r-e mphasised . U nless
t his p roced ur e wa. ca rr ied ou t . the isolat ion ra te dropped
almos t to zero . Th e plate was incu bat ed overn ight at 37° C.
T he colonie o f d~· sen t.ery ba cill i wer e usually eas ily recogn ise d
as non-la ctose-fe rme nt ing, small, tran slucen t. del icat e colon ies
with a regular out line. Occasion a llv woolI.-' irregula r colonies
of fJ: dys e nt PT i(~ , onne were observed. 6n some pla te t he
colon ies wer e so n umer ou tha t it was po sihle to carrv out a
I n the majo rity of cases s electNI
di rect ag glutin.at ion te t.
colonies wer e picked off an d inocul a t ed on t o a econ d McConkev
plate. using a met hod suggeste d bv Capt. l\la nifold , T.M. . '
Ir~ th is method a. l\l cConkey plate is divid ed up in to 36
sectio n hy cross-ruling t he gla ss wit h a grea se pencil. On e
st rip con ta ini ng six sect ions is used for each sub -cult ure and
one of six colon ies from each original )lcConkev plat e is
inoc ulated on eac h se cti on. T hu s ix colo nies f ro~n each of
252
J UL Y
L AB OR AT OR Y DIAGNOSIS OF D Y SE N T E R Y .
11, 1942.]
six stools can be inoculated on one .McConkey plate. The
plate is in cubated over nig ht and non -lact ose-fermenting colonies
can be clearly di stingu ished. Direct agglutination tests using
gro u p sera ca n be car ried out and the type of org anism re adi ly
Iden tifi ed . Co nfir ma to ry bioche mica l rea cti ons usin g lactose,
glu cos e and ma rmite p ep t one wat er can th en be don e, using
inocula tions f ro m th e plate colon ies.
T he d irect a~glu ti na t ion te st s wer e car ried out by mak i ng
a thick suspensIOn of the organ isms in saline. Five drops of
this suspens ion w er e pla ced On a glass sli de. One drop of
Shi ga , Schm itz, Sonne, Flexner 1 and Flexn er 2 po ly va lent
anti-sera r esp ecti vely wer e ad de d and mix ed wit h th e d ro ps
of su spe nsion. The slide was then rock ed for a few mi nu t es.
P ositive re sul ts wh ich wer e ea sily see n naked -ey e d evelop ed
in a bo ut five minutes. Th e mixtures whi ch did not ag glutinat e
served as cont ro ls .
The "Flexner " poly valent se ra used we re pre pared agains t
the f ollowing strains :Fl exner I -B. dysenteria: .. . ... .. .
Fl exner 1
2
Flexner 2--B. dysellt er ill ... ... ...
3
Flexner 4
5
6
Boyd
1
Occasionally st rains were isolat ed whi ch d id not agglutinat e
with any o f the type sera, although they con formed in
morphology and biochemical reaction wi th organisms of th e
dysentery gro up . Su ch strains wer e further mvestigated bv
inoculating t ubes of la ct ose and sacc haros e pepton e wat er,
sea ling with paraffin wax a nd in cubating f or three week s.
Most stra ins fermented la ct ose before the end of this peri od.
One strain investi gat ed in this manner pr oved to be B .
d ys pnt eri,c So nn e.
Th e following t abl e summarises th e results of a n in vest.igation of 125 cases of dysentery by t he method s outline d
ab ove :TA RL E.
Number of suspect ed cases in vesti ga ted
125
Number of stools examined .. . ... ... ...
134
Nu mber of cases in w hich .. bacill ary exuda te" wa s
p resent ... ... ... ... . .. .. . .. . ... ... ... ... .. . ... 38
N umber of cases in whi ch " indefinite ex ud ate " was
prese nt
... .. . ... .. . ... ... ... ... ... ... ...
57
N umber of ca ses in whi ch no ex udate was present
30
Number of cases infected wi th H. d ysellt erio S h i[la
1
4
N umbe r of cas es infected wi t h B. dysellt eri,,' Sc h mit z
N umb er of cas es inf ect ed wi t h B . dysclIteri,,' S onn e
2
N umber of ca ses infecte d wi th B . dyscllterilC Ft ez ner 42
Total n umb er of d ysent er y stra ins isolat ed ... ... ...
49
P ercen t age of cases showing ex udate fr om wh ich
53 %
dy sentery st rain s were isolated ... ... ... ... ...
N u mber o f cases showing "bacillary exudate " from
whi ch dysentery st ra in s wer e isolated ... ... 32 or 84 %
N u mber of cas es sho wing " ind efini te exudate" f rom
0
wh ich dysentery stra in s were isolated
17 or 30 ~~
[
S.A.
MEDI CAL
JOURNAL.
directly at the bedside. Two strains of B . dyswt erice Pl ex ner
wer e isolated from scrapings left in glycerol-saline and one
strain of the same organism was recovered from a plate made
at t he bedsid e.
DISCU S SIOX.
I
T he method s used in the exa minat ion of 134 stools from
125 pat ients sus pected of dy sentery have been outl ined ab ov e.
It is cla imed t hat t hese methods ha ve proved very sa tisfact ory
in view of t he result s ob taine d.
The cr it er ia used in th e
diagn osis of a meebic dy sentery, .via., (a) t he pr esen ce of act ively
mot ile amoeb.e, a nd (b) the presen ce o f erythrocytes in t he se
amoebte, led t o t he di scovery of elev en cases of active a rnoebic
dysen t ery or 9 per cen t. o f the total number of ca ses
investigated. T hi s hi gh proport ion of amcebic dysent ery is
expla ine d by t he fact that the maj ority of th e patients ga; ' e a
previous history of amoeb ic dysent ery cont racted in the Un ion
o r in th e E a st Afri can Campaign .
Only three p atients were found ex creting E. h ist olytica cy sts
in the freces. All th ese patients presented evidence of pr evious
E. h ist olytica infection . It is very doubtful if arncebi c cy st s
f ound in the stoo ls of pa tients who have given no previous
history of E. hi st olytica infect ion ca n be regard ed as pathogenic. Many protozoologists are of the opinion that different
r aces of E. !<ist olytica exist, some of which are non -pathogenic
to man.
Thus ' Brumpt (quoted by Mansou-Ba hr , 1940)
recognises a .. physiological species " mor pho logi cally in di stingu ishable f rom pathogenic E. h ist olytica.
•
The majority of cases of dysentery wer e ba cillary, and
organisms of th e Fl exner group formed the highest proportion
of isolat ions . By ca ref ully washing flakes of mu cus bef or e
in oculating pl ates of media, a high percentage of isol ations
was o bta ine d in cases sh owing a •. ba cillary exudate " . This
isolat ion rate com pa red well with that reported by Anderson
and Cr ui cksha nk (1941). These work ers, investigating an outbreak of F lexner dy sentery and usi ng a specially selective
medium for Flexners ba cillus, reported an isolation rate of
97 per cent. fr om the stoo ls of 35 patients exa mined during
t he secon d to t he f ourth day of t he diseas e, and 83 per cent.
isolation f ro m the stools of 18 patients examined during t he
fif t h t o th e seventh d ay of t he di sease. As t he ma j or-ity of
ou r pat ients a r rive d in h osp it al fro m th e th ird t o the se ven th
day of t he di sea se, our isola ti on ra t e of 86 per cent. f rom
38 pat ien ts sho wing bac illary ex uda te may be consid ered
satisfactor y.
T he isolation of B. dysent eriw Fl ex u er f rom th e stoo ls of tw o
members of the kitchen p er sonnel in the cour se of a routine
examinat ion was of consid era ble i nt erest.
Both patients
exhibited stoo ls showing an .. in definite exudat e " . N either
a ppeared ill or appeared to consid er t he ir stools abnormal.
T hese cases wer e probably suffering fro m chronic d vsentery
a nd t hey were obvious ly not suita ble fo r employment' in the
hosp ita l kit chens. Sporadi c cases of dy sente ry ha d occurred
at inte r vals amongst th e hosp ita l sta ff. Al t hough it was not
possible to trace an)' di re ct relationsh ip between t hese ca r riers
a nd such ca ses as d id occur, it was n oted that f ewer ca ses o f
d ysent ery occur red. amo ngst the hospital staff after t he two
carrier s had been Isola t ed and cu red .
T he case of Sc h ist osoma mal/.w ni infect ion detected in a
South Af rica n na t iv e in th e course of rout ine examinatio n for
ca r riers ra ise s an in teresting problem in public hea lth. This
native ca me fr om th e T zaneen d istri ct of th e T ra nsv aa l, and
a ppa rent ly ". man soni infect ion of natives is not un comm on in
this d ist rict. J ust as ". h rematobiu m: infection has gradually
spread down th e East Coa st of the Union , so may we expect
S . ma ns on i infect ion t o sp read if a su itable snail host be comes
infect ed wit h th is helmin th . I t would appear worth wh ile,
wh ilst this infect ion is comparati vely loca lised , as it is at
present, to t ake strenuous meas ures in an attempt to sta mp
out S. manson i in fect ion in th e Un ion.
It will be not ed that althou gh no spe cial select ive medium
was used , t he isolation rate fr om bacill a r v ex uda t es wa s ver v
satisfactory,
' "
Ca rriers. -A rou t in e exa minat ion of kitchen per son nel was
ca r r ied out in an attem pt to detect enteric an d dv senterv
ca r r ie rs. T h ree stoo ls from each member of t he 'h 05pit il
kit ch en st a ff wer e examin ed. Altoget her fifty-fi ve members of
the st aff wer e investi gat ed. T he maj ority of th e stools were
form ed or semi-formed, an d wer e, ther ef or e usele ss for
ex amina ti on for th e pr esence of d ysente ry or ga r;isms . A fe w
" l oo~ e " stools wer e fu r th er invest iga te d by the methods
de scr ibed , a nd fr om two of th ese B . dysellt erirc Fl ex ner was
U) DIAR Y.
isolat ed . N ei th er of t hese ca r r iers. one a X ative and th e
(1) One hu nd red and t wen ty -five cases o f dysent ery wer e
ot her an Italia n P . O.W., re por te d ill at any time or appe ared
in vesti ga te d in a South A fr ica n q en era I H osp it al in th e M id dle
t o consider th eir s tools t o be in an v wa y abnorma l.
E ast : 38 ca ses s howed a .. bac illary exuda te " di agnostic of
Chro llic J)J/sl'lIt uy. - In t wel ve cases o f chronic dvsent er v o n
wh om sigmoidoscop ic exami nat ions wer e carr ied out th e bQwe l
ba cillary dysent er y, 57 cases sh owed a n .. ind efin ite e xudat e ",
whils t 30 ca ses showed no exudat e.
app earan ce suggest ed a chro nic bacillary i nfect ion . Scr a pings
of th e bowel wa ll w ere placed in a sma ll bot t le containing 30
(2) E leven of t h e ea es exam ine d . or 8. 8 per cent . , were
fonnd to be inf ect ed wit h 1-:. h isf olyt ica.
p er cent. gyce rine in nor ma l sa li ne ad j uste d wi th phosphate
(3) T h ree cases were fon nd to be passi ng E . h i.~ toly t ica cy sts.
bu ffer to Ph. 8, an d sent to the la bor ator v. McConkev pla t es
were inoculat ed imm ed iat ely on ar r iva l of "t he spe ci men at t he
Th ese cases all sho wed evi de nce of previous E. h i.~ tol!ltica
lab or atory . I n some cases th e :\IcConkey pl ate s we re inocu lat ed in f ect ion.
S.A.
TYDSKRlF VIRJ
G ENEES K U ND E .
(4) T hir t y -t wo of the 38 cases sho wi nc .. bacilla r v ex uda te ..
•
y ielded dysentery bacilli on culture . 0
(5) E. dys enl er ia: F l ex n er was iso la te d in t he majorit y of
cases . B . dys enler ;;I' :;1,if/a, S clUII;t: a nd S ou ne were ' a 'so
encountere d.
(6) A routine exa mina tiou of 55 suspec ted ca ri-iers viel de d
two cult u r es of B . d y s ellt er ill l Iexner, an d iu ou e case S.
mansoni ova were ident ified .
(7) B. d y senl er ;le Fl ex n er was isolat ed fro m t hree of twelve
sigmoid oscope spec ime ns sub mitted.
A CKX OWLED G~IEXTS .
I ha ve to thank Br igad ier A. J . Orenste in, D . ~I.S . , D. D. F. .
fO,r permission to publish this . pa per, ~n d al so S J Sgts . Lomba r d ,
\\ ellsted and T albot for t heir technical as sis ta nce.
BI BLIOGRAPHY.
Anderson,
D. E.
tr ., aml C r ui d 's han l' , R.
(1941) : Il rit islc
.11 edical J ournal , 497.
.11anso n .B ah T P . H . (1940) : T ro p ical Diseases, pag e 859.
D.M .S., M.E.l (1941) : T'ech n ical ! IIS/ TUC/ ioIlS, .\"0. 59.
The Antigenic Structure of the Bacteria
Incriminated in Four Unrelated Outbreaks of Food Infection.
By
~1.
W . H ENNING, )I.R. C . V.S.. D.Sr.. . UNDERSTEI'OORT.
A
STU DY of th e ant igenic str ucture of four species o f
Sa!m on ~lla .con c~ rned I!I outb rea ks of f ood- i~f ecti on (so called food -p oisoning ] m t he I'ra ns vnal, was made III colla bora t ion with Dr. J a mes Gear of ~ he Scuth Africa n Ir st itute of
Medi cal R esearch .
Deta ils of th e d iffer ent ou t brea ks a re gln~l bv Ge a r R oux
a nd Be van in the p r eced ing paper in th is .ssue, 'r he te /h n io uc
described hy H e nning (1939) was employed,
•
Previous out breaks of Sul moneltn f~o,l - infection ill Sout h
A fri ca wer e descr ibed by Greenfield an d .Iud d (1936 ), H enning
a nd G ree nficld (1937 ). H enni ng (1938) . Bn cha nan an d Bevan
(1009) an d H en ning , Rh od es and Gordon-Johnstone (1941).
O UTBREAK I OF GEAR. Roux AND BEVAN (CULTU RE 1003). A
non-lactose f erm e nt ing G ra m-negati ve or ga nism isolated fro m
the fn-ces, th e vom it. ~nd th e blood of the affe ct ed pat ien ts
re vea led . on a pr elim in a ry st ud y , all t he cha ract ers of a
Sal".lOn ella , a nd was found to fall in G roup C of t he K a u ffrnn n n White Schema. B y mean s of aggluti nat ion t est s it wa s observed
that cu l t u r~ ~003 was ~Iiphasi c an d tha t, .i t ~c c ur r e<! in th e a -fi
ph as e va r rat ron of K a uffm a nn a nd ) d tslll (1930 ).
It was
agg lu tinate d by se ra cont ain ing facto rs P'" en an d enx, b ut
whe n a num ber of si ngle 18-homs -old colonies were t ested some
of the colonies wer e more read ily floccul ut ed b v eh sera while
ot hers wer e agglutin ated better wit h sera couta ining ei ther
fa cto rs en or en.x.
F or fur t her st udy pure fI/-': an d eh sera "Yere therefore pre.
pared by respect ively absor bing ab ort ns-rqui serum wi th eas t bou rn e specific (cont a ining fact ors eh) a nd r a st bourne sp ecific
serum with abo rt us-equi (contai ning fa ct or s wx ). It was not
co n ven ~e n t to obta in a pu re ell serum and the pure PlIX ser um
was SUIta b le for purp oses o f the test. But it was noticed t ha t
phase d issociation occur red so irregularly that it was verv
di fficult to obtai n an antigen that wa s pr esent in either t he a or p
phase in t he pure sta te. By e mpl oyin & cot ts (1934 ) t echn iqu e
for the ac celeration of phase di ssociat ion imp rov ed bv W asse n
(1935 ) and Brune r a nd E d wa rd s (1939. a and b) t h isxlifficu ltv
was .overc ome and it becam e possible to obt ai n pure ant ig ens
of eit her the ao r f3 ph ase. T wo se r res o f sloppy aga r tubes,
of which th e one cont a ined aborlll s· eqlli se r um ( factor elIx )
a nd the ot he r wB / bourll e spe ci fic seru m (f act ors eh l, we re
inoc ula t ed with cult ure 1003. I n each series t he t ra ns fe rs wer e
mad e at int ervals van' in g f rom 6 to 18 hou rs. After six to
eight tran sf er s it was' possibl e to obtain a pure a (eh ) p ha se
fro m th e sloppy aga r enriched with a IJOrtu" -equ i serum a nd a
p UI ~ (enx) phase from the sloppy ag ar cont a ining ea.•t bmlT lI c
serum. From t hese phases it was possible t o ob tain antigens
wbich react ed on ly with pure sera of the sa me phase a nd not
253
[J ULIE 11 1942.
FO OD INFECTION .
wi th pure se ra of t he oppos ite phase -i .e . a -p hase a nt igens
rea cted only wit h pu re eastb ourn e specific se r um from whi ch
the com mon fa ctor wi th a bort us-eq ui (e" x ) was removed by
ex h a u~tion wit h abort us -equi an d t he
fi-p has e ant igens were
ugg lut.in a ted only by pure ahort us-equi serum fr o m which t he
overlapping f act or (ell ) with eastb our ne was re m oved bv abso rp ti on with eas t bou rn e s pecific.
.
After th e prepa ration of pure a and fJ phase ant igen s ag glu t ina t iou an d ab or pt ion t est s wer e performe d : . • U " al//;gel/ : Cu lt ure 1003 rem ove d all the " 0 .. , gglut ini ns
fro m sera li ke choleru -suis whi ch conta iu I uctors \ '1 an d VI I.
while chole rie-sui.• com pletely e xh a ust ed th e .. 0 " aggtut in ins
from 1003 se r um. The "
ant ig ens of cult ure 1003 an d
chol era -suis m ust, t herefor e, be regarded as iden ti cal.
.. 11 " an t.uje ns : Or ganisms like eust bou rn e (specific phase)
wh ich co nt a in fact ors eh nearly com plet ely e xhausted t he a
-phase (eh ) from 1003 ser um without mat eri ally a ltering th e {3
-phase (fa cto rs ell or ell x ), whereas a bact erium like gl os/ru p
(s pecific phase ) contai ning Iactor s ell but no t ch r em oved nearl y
a ll the fJ· phase ag gl uti ni ns f'rom 1003 se ru m.
T hese te sts ind ica t ed t h at cu lt ure 1003 was closelv related
to S. h ruen d er u p , T he recip rocal ab sorption t ests t ha t were
now perform ed wit h b rue n d eru p a nd cult ure 1003 con fir med the se
observa t ion s. Cult u re 1003 remov ed a ll t he .. U " . a (facto rs
ell ) a ndfJ (t act crs cn ) phase ..ggru r iurus fro rn brurn tt er u p serum ,
while braetul eru p co mpletely ex hau st ed H.03 serum. It was
the ref ore e viden t t hat cult u re 1003 and bra e,II l trllp were identica l, conta ini ng t he same a ntigen ic com ponent s , vi z. , VI , V IIeh : en.
o ':
OUTBREAK 2 OF GEAR, R o ux ASD BEVAN (C ULTURE 10(2). An
organis m o f th e S almon ella group was isolated f ro m the urine
a nd freces of a n umb er of t h e pa tients.
P relim ina r y t ests sh owed t hat cult ure 10:12 fa ll s in Group C
of th e Ka u ffma nn-W hit e Schema .
W hen it wa s ca r efully
exam ined by mean s of aggl utinati on a nd absor ption test s its
ant igeni c st ruct ure was found to re sem ble t hat of cult ure 1003
and braend eru p. Cult ure 1002 was, t heref ore, also given the
a ntigeni c assign ment of : VI . VII - eh : ell.
As in the case of cult ure 1003 , a - fJ phase d iss ociation could
not he brough t a bo ut sat isf actor ily withou t th e employme nt of
Scotts (1934) t echn iqu e, mod ified by W assen (1935) and B ru ner
an d Edwa rd s (1939, a an d b )_ T he bioch emi ca l reaction s of
cult ures 1002 an d 1003. h oweve r, re veal ed min or di fferen ces
.
(T a ble I ).
OUTBREAK 3 OF GEAR, Ro ux ANI> BEVAN (CULTU RES 1000 AND
1001). I d en t ical cult ures (1000 and 1001) were isolated from
the vomit an d stools of a numb er of pati ents a nd fr om th e
i::cr imin a t ed me: t. By 1" 1'1 m ina : y t estin g t hese wer e found
10 \;1' S ul m onella » faIlinl( in G,'oup J) of Ka uffma nn -\Vhit e
Sch ernn. By per-form ing agglutinat ion ab sorpt ion t ests cult ure
1001 was foun d tu rem ov e a ll t he .. 0 " ag gl utinin s fro m
as well as
sera . wh er eas eith er
or
e n te rit id is exhau -t ed th e so ma t ic agg lut inins f rom 1001 serum.
Bot h du/,/in and en te r it id is se ra aggluti r-at ed t he" 1I .. a nt i,
ge n o f 1001 to full t it re, hut cult u re 1001 f ailed to exhaust
t he .. 11 .. agg lut inins from du hlin se r um ; it mere lv reduced
t he titre f ro m 12,800 to 6,4 00. C ult ure 1001, how ev er , cornplet ely a bso r bed a.ll th e " 11 .. as well as the " 0 .. a gglut i nins
from r n t n i / id i.' serum (t it re " 0 .. = 800 and " 11 .. = 12.800) .
W hen 1001 se r um (t itle .. 0 .. = 800. .. H .. = 6.400 ) wa s absor bed hy esve rit id i» and d u blin se pa ra tely, en te r it idis re moved
a ll th e flagell ar as well as th e so mat ic agglut in in s. whe rea s
dub/ in only a bs orbed th e so ma t ic agglutini ns , leav ing t he t it re
of t he fla gellar ag gl uti ni ns pract ic nlly unaltered.
It was th er efore clear that cul t ure 1001 was id ent ical wit h
e n t erit id is co nta ining t he sa me an t iuen ic compon ents , viz. ,
lX- gm .
•
e
n
t
e
rit
u
l is
d
u
b
l
i
n
d
u
b
l
i
n
OUTBREA K 4 OF GBR. Ro ux AN n BEVAN (CULTURE 1016). A
pr-elim in a ry t ud y of th e o rganism isola ted f ro m th e excret a
o f a n umhe r of patien t s a nd a l 0 f ro m scr aps of ra w m in ce ·
meat in crilll inat l'd sh ow ed tha t t he\' were id ent ical Sal m ollell ll8
occ uni ng in G roup B of t he Ka u·ffma nu· W h ite Schem a. The
o rg a ni illS we re a p parent ly rel at ed to
/ y plt i.m ur ium .
Agglut iuat ion altd absor pti on t est. sho wed t hat cult ure 1016,
a lt houg h co m plet ely ex ba us ti ng the .. 0 ,. agglu t in in s fr om

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