,the' a , to clinical medicine

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,THE' DISCOVERY OF A SPECIFIC COMPLEMENT.'FIXATION iTESTFOR BILHARZIASIS AND ITS PRACTICAL APPLICA'l'ION" , , , TO CLINICAL MEDICINE. ',' "

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,~ ) By MiJ;'R N; HAMILTON 'F AIR:YE"l', O.RK AU8t~aliar/A~;"y' Mpdical Corps. '., ,','

, . Pathologist, 14tH AUfti'ali(J;~ General Hpspital; Lecture,r '/;nP0~a8itology, Egyptian University.,

f ,' ....

CONTENTS. r:~IN''r'RoDuCTION. II.~FACTORS,LEADING ut: TO THE PRESENT'INVESTIGA~IoN.

, :iII.-O~S~RVATIONS ~N TECHNIQUE AnvociTED., '

IV.-CoNCERNING T':!IE 'SPECIFtcITY OF ,TIns COMPL~ME~T-FrkATI6N REACTION:',',

y.-A~AL~ilIS ~F RJiJI,!ULT~,~.O~TAINEDI~ THE, ExaMINATION OF }lERA o~ 311 HOSPITAL

PA,TIE:NTS' US~NG 'THE SA,LINE EXTRACT AS ANiiGE~.' : , "

V:I.~AN,ALYSIS. OF' RE-s'uDT~ OBll'AiNEl( I~, ,THE" EriMIiNAT~Oi.. OF THE, ,SERA OF 322 CASES', ,,'USING, ,THE'ALCOHOLIC EXTR*,CT A'S A,NTIGEN., " '. " " " '\"

'; 'VII.-,+THE'IM~mNITY' RESPONSE' INExPERIM~NT~i:.;LY" IN:t.')IlCTED MC)NKEYS,; , • ,'(q;) In Morikeysr~covering\f,om tbd)is~ase..' "'" j, ,

, (b) In Monkeys dyirigfrom the Disease. , Vln.-RELATIONS~IP' BETW~EN THE GRADE OF ;EOSINOI''a:ILIA AND TIlE COMPLEMENT FIXATION'

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" ,REACTION IN MAN. ;' ( ,', ",,', '~ , ",' " "';" ' IX, ~ PRACTI~AL V ALPlil OF T~IS 'COMl'~EMENTFIXATioN RlilACTION:' ,

. . ('a) As~' :rv.r~·ap.s of Dia~nosis~~ ,,' , , .. " ' I" , • . ~,.. ,

", (b) As a Ther'apeuticlndex,to tbe~ffeqts:ofaD,Y given,prug on the Bilharzia Worm: ' X;--,CONCLUSIONS. ' ,", -, /' , " "

1.-:-IN'fJ;tODUCTION,. , •

A CAREFdL study of the p~thological effe-cts ~fthe 'Pa:rasitic metazoa, ':very especially~f bilharzia, on the' tissues of the host,as well as the enstiing , ; ,local.Inflammatory reaption and the .celb{lar-hu~oral, resp?nse j inevita1;>ly,'," : ;,,' ,

,. leadsto,tlle conclusion (n0ctat.pres~nt frilIy:appreciatied),tbat ;tbese~niinal' para~ites and their ova exert, their' deleterious .,iIlfiuence, not so much by

,.t4eir mechanical irritation, as,l;>y the secretion or excretion of some soluble' " .toxin.' . '/;".,' ,

, ,', In <this' : re~pe~t: a,s i~manf others, the parasitic- metaz;areserri bIe' 1>rotozoal and bacterial invaders. A cellular~hum6ral:Jesp(lllse on the'part : .qfthe hostis'pommon tq all three groups otJ?arasite~. , rn.,bacterial

',L,

. infections polymorphonuclear leiico<?ytes and mononuclear leucocvtes"are , . ,.es~ecially ,produced ,in ,excess: "The polymorphonucleal' 'lencocytes urespond '

( .to the chemiotact~c injluenceexerted by the toxins -9L. the' pyogenic 'group -, of bacteria,and according toOpie. (1) they contain a;. Umcopr6tease ferment

." .a(}ting in acid or neutral medium:" iilstich ale'u?o~~tosisanexce,ssor, 'neutrophile' myelocytes, can bese,~n in th"e bone marrow';, T~e lymphocyteEi" tre' produced in exces,sjn typhoid and, tubercular -irifection!3 a~d thejr

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contain 'a lympho-protease £erment ~ctjng in a medium wliich_ is /'slightly acid: , " -, I , ' c

,In. protozoal infections~,;maJaria; trypanosomi~sis,· ,Leishmaniasis,.etc.,. ' .there is .an, excess of large 'mononuclear cells in the peripheral' b:lood, ~hiyhprobaply~rigin~te .in the, endothelia;! and pulp cellspf the spleen. ,", . . Metazoal infections generaJly create a response ofeosinophite cells. Eosinophilemyelocytes increase in· the hone marrow, as maybe, seen in :monkeys experimentally infected' with; .bilharziasIs·; ; in, consequence differential- arid ,total ,c6untssbowaIJ, eosinophile leucocytosis. in the., . 'peripheral blood: , / '.' . ' . '.',' , " .. '. '.' . .. ,.

. Similarly recent serological).irlve~tigatiorisrevealea' that an identical! . hU'mora;l re&Ponse,fo~all . groups ofparaErites-,:"bacterial, .p:rotbzoal '1tnd~.; metazoa~-;-:~xi\'lt,s" I'J?:, a nUlll'ber" of.' bac.terial in,f~pti(p:lsantibod~es· of" th.e;Dature of' opsonin, agglutit~:ins):arid ,bac'teriolysin~ may.,be ~demonstrat~d, 1 ,j,

Irt pro~ozoalinfeCtions,~s'pecific eomplement'flxatjon,reactions have been I' des,cribed. .Noguch1[2] demonstrated' b()th antibody and agglutinin,in theseraofsypliiliticsby u'singa specific antigen ma.defronieulture~' ·Trepbnpmata.paUida,: . McG:6wan" [3] demonstr~ted positive complemen.t. 'fixatioIlreactions.in the, sera.of sheepiinfe.cte~with'sarcosporidi~;, .!D0re· rec,entl,y Thomson [4} has reported a positive complete fixation in malaria.

',~erologica;l' investigations ip .helminthio infeo(ions have similarly not beeu'unfruitfuL' ' ". '/- '. . ,'.'. " , ,,:' ('

In' ~estode: infeCtiolls-"positi've 'complemerit',fixation 'teats, h(!,vebeen ; ". - ,', '., .' " " "", j, • •

. oescdbed for tli.e TClJ1ii4 echinococcus by: Gliedini [5l W'einberg and, , Paryu [6]~:Kl"eutkr [7J;'a{Hl by Meye, C8J for T.sagin~tq,. ' " ' , < Fleig and Lisbonne '[9], and laterWelc:h and Chapman [1(,)] ,'described thE\ precipitin reaction in '. hydatid disea~e,and " Langer, [11 J and· ,Le Dantec f3Jled' to:denlOnstrate thisp~~lloihenon'in,othertamia,infeCtions; , .. , " In \'n.~ma.tode infections ,Strobel, 'showeq that'" p~sitive·. complement, . fixat·ionreaetions 'Yere, given by thes.era.ofpatients infected with,

',Trichinella spiral~s:", ' " ," " ,," >,' , .•

, " In,thetrematode i.nfections no .iilllchreaction has Deen sp farde8cr~bed, jmt in the foll~willg pages. I wish to repbrt· the discovery,' 6f ~., spe'cific

.... complemeIltfixatio'n reaction in.bilh·ariiasiiil. . , , , \' . /

The frequency' with which 'La,wton,' and, late~' the wl1itev, noted,: urticar,ia.,~ fever; .and' other sympfoms;~ in,. the early stages' of . bilharziasis, ambngstAuspl'alian' troops, strongly ,suggested, tbepresenc~ of some toxin,. the product of'tbe 'maturingoriof the adul£wormj;. ,Seeing,th,at tbe initial " sympt~}r:ns 'su~~iaed after a ::variable 'period and tha.tthepatienf completely , ,r~covereduiltilth~ sig,ns ~~d ~~mpioms of localized. bilhar:iia~is, SUP((t- I

vened, ir seemed to be probable aria priori grounds that recQvery should ',be • ' . aceotJ;rpariied by't~ef9rmatiori of. some specific imITlprie b<:>dy in the blood. '

. A,specific a.n~igeilwas prepar~dfrom ,the Hver~ of illfected snails of th~ ,.\0'

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N.HamiltohF~irley 451

:Species Planorbis boissyi, the intermediate host .of Biuiarzia! ma~$6ni,and, by,employing themethod6f fixation of co~pkmentfirst desm:ibed by," Bo.rdet. and Gengop,antibody was' aemonst~ated, in ~4e sera~, of patients infected,witheitnerB.,mansoni .0~B. mcemai9bia. ,-~ ,

, I~June,igl'i~. this discovery wa$ reported to the militar:tauthorities and since I then I have very extensively utiliied .the Ipethodof diagnosis of bilharzift~is'inAustialiJn"Btitis:4 and Egy'ptiJ1uhospitals ; with most' ,gx:atifyingresults. ' "".. ,,:. ' " .

,III,.20Bs]JRV ~TIONS QNTECHNIQUE.

, Afttigens,;~Two diffe~entan~igBn~ rverelltilizeCl' durin~ the inve~tiga-' tion,~)lle asaline, extract, the other an alcoholicotie. ,hI the great majority'. 9£ cases the, antigens were prepared,fro'm the infec~edlivers ofsnailE! of ~ the"species' p. boissyi ,(~nt~rmediary host: of;S. mansoni). Eql1~llY8atis- ; factory' arytigens were also made from infected sna,ils of the species Bullinus/ . ,',dyb'ows~i'(B.,h~11lat:obia). '. " , " ", '

'/ 'Antigeps prepared from; either 'species ofsriailsyield~d'positive ',' completement fixation reactions I for . both kinds 'oJ infecticm' in. man' . (B. ha3matobia / or B.,ntansoni); sqowJ~g how cf9~elybiblogicanyalliedare c

,the~e~two sr>~cies.,Therpreparatidn of ,.these antigens was asfoIl9w8:~ , /' \a) Salil~e Extract;~4 number Ofinfe~ted livers w~r~ mac,erated in'~, sqlution composed Qf 0'85 per ,cent saline and 0'5 per cent phellot {one lIver t6011e cubjcgentimet~e solution). This .mixt.ure W!1S 'shali:en' I

thoroughly on an' electric shaker" for,twenty~ minutesan4' 'incubated for:_ twel1ty~four hou~s at 37°C., to permit of extraction., ,<' ' , r'

It was, then filtered and the fresh ,filtrate used as' antigen. .' '." " ., (~)' Alcoholic" Extra~t.-LThis" ;as' found to be the'most satisflwtory,

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I , -antigen.~ A number of, in£ectedsnaiJ~' lIvers Were macerated .in a quantity I of absplute alcohpl, shaken~ thOroughly on, apele9triG'~haker, e~tnlctedfor

. twenty-fol,lr ho1i~s at 37°C.,; and filtered. • The filtrate was evaporated to _ .dry~ess at 45° C. by means of 'a Sprengles exhaul;it pti91P' The~esi~:\le . '

was dried, we1ghed,andshaken vigorously into ~olutionwithO'85 per~ cent saline and 0;5, per cent phenol (0~Q5 gramme of residu~ .. to t;wenty'

'/

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icubi~ ~enti~etres8olution). ;1 '. .,', ", i ,'. . ". ". I' " : " " i,.,:~" \ " { "~Tp~, ,antl.:completilenpary. do~e .was·.,thep .. ;a~cu~a~ely\ estlm\ate~ ,and .. 1~Ot/l I ,>" ~'.

moretlta1L one-thirrJ of th~* arrlrOunt, used' in the test., , ' ' , ' , , Theautigen was tested alscdor anyhffiIDolytictendency.Lately;as ,an

\,: - -alternatiye; I have adopted the}:limpler pj"oced.u:re, ofdilutingthe,eoncl3n-

,tratedalco,hoHc ~xtract with, saliI;le(O:'85 per, cent) and not actually. , " • \ • , ,. I , ~ ~ I ,'\ • • • I / I '

,eyaporatmg It to dryp.ess. (T~~,results of t,hlS. modIficatIOn ha\7e,.~p 'far been. satisfactory. Fresh alcoholic extractf'!,S'l1ouldbe em,ployedas a ',: I;' routine as they are more sensitive, tha~ extraetskept'in the ice~GhestfQr " , several'months., ,<'. '" . ,', -,,' '"" " ',' , . ,., J " " , • ,i'

A fresh s}ispension of three per cent sheep's corpuscle.nvas alwaysm,ade. Thf:), blood was obtained from a sheep especially kept f9r the purpose, aud

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Discovery. of a Spec,ijicOompleJ7l;ent:Fixation Test , " -\,"- " l I" " , ,

'/,' ,;~ft!:lr citration(twq" per cent s6dium citrate in saline)' the'~orp\1scles were' ,repeatedlywashe,dwit9 ,s~iineand 'centrifugalized." T,he ~orl'us()u!ar'

" ~uspensio? wassenlsiti~edbyadding fourM.~.D.s'of p'remplytic ~erp,tpaJ;ld' , ' ~incubatingat; 37° C; fbrone half hour .. This ,n;n,xture,was'then kept (lot,

'SoG.till tequired. ' ' " / , " , , " ,', ' , The blood wa~obtained fr:oin' t4e '-patient witpintU)e~ty-Jour hours 'ot, ' ..

, performing t~etest ,arid,stored in theice-ch,est til.1 reqi1ire~: "The resulting 'serum Was diluted with fottr volumes 6f'lsaline and heated'to 55° G, for' , ,t'Yenty mi~utes to, destroy complement' an( anythe~olabile 'antl.,6om~ 'I

pletilentary hodypr~sent'in, the serum, AnysetuiIl:·exhibiting,anti-coro.-\. ' " 'piementary tendency in the control ,tubes,' or any ~erum inwl:ild~II;licrobic'

,'infection was suspebted,Sw~s, retestedat ~ later date. ," ,,' <'" ',' ',Tli~ r.r;,bbit hCe7lJ:olytic serurp- emp16Yedw,as, prepiiredin this 'laboratOry'

,by 'tbe 'intraveno~s inj~ction 'of 'w~JI-wasbed, ' ~itrated ,', sb,eep',s/, Gb~pu~cles. ",', The titre of the serum used wasneverJess than l'in 2~OOO;, ,.' , .'

I' < "", ~~"! ,'''' I , ' : " ,', >' ,'\' )'" , "

{ , .. ,.Complement was obtaine~ '£ro~, ,a healthy g*ine'~~pig theser~i:n, of 'which had-stood)n contact 'witli ,the clot in, theice-'chest for sixho~rs ~" before use." The, M.'Ii.,D: ofcdmpJemen~w~s alii~ys accurately'determi~ed,,,

A rrr;t,ngeinentOj theSy;tem jorAhe' Test;,' " ", ,The techniquelbaveheen.latterlyadopti~gis,as ~ollows:;-,', '

. ,SmallvollI:m~sof ,the' reagents <ar,e measur~d by,Donald;s, dropping 'pipettes. The, ,tot~I' vplumeiri the fimLIstage,ol the, reaction lS /0'5 ,cubic centimette(five volumes), ,and,each 'unit volume equals O'·lcubi,e'ceritimetre.

. . ~. R~ckscontaining ~6urro~s oft~'bes ar~ utilized ,and tl).esy'stem:is put up' in the following order 1,- .,,', " , ' ' ",'

.' ",

,I 1 ' (1) The firl?t row,contains onevolumeeaph, of, antigeni~ serum! anp,saline, a~d' ope:,volmpe dih:itE1dso:a~, t? con~ai~tht~.eM.H.p.s ,ofconiplem~nt. ~'

. (2) The second row'con,tams ,SImIlar volumes oL the abovereagents~ ex,eept that thevolumeof complement. used contains five M.fi;D.s ,','

',' \ ,. "(3}Tb'e th~rd' row, cQntaiIls similar volllm'es of the (reagents, but, the, ' ::vor~me of ,ebll1plen).ent,usedcbu'tains 'sev~n,~.-II.b.s. ,: ' ,', , .,' " "" '

" (4)" ~he fourth ;r,ow co:iit~ins,/onl;y isetull'l,' saline'and, cdml'leme~t' '. "( thr,ee M.~.D.s) arid serves ~6 det.ect the presence of any a!J.ti~compl~U)e:r;itary .

tendency in:th~ seruinu~de,'rexaIiiinat{on. """,,', ," I~" " "Two ,~n~igencontrols, obe conia~riin,g onevciliurieand the other two' ,

, yoiume~,ofantigep arejric1uded~ and also a pooled neg'ativeand apo'Oled . , p,ositivy serum' " ,. ,,', ",.', ' .... ' ' ',,',,' ' , This sy~teni isin'cubated' for one liour at 37°,C:, andth~n one volume ot

. se1-:lsipized sheep's cOJ;,puscles is add'ed" The racks are, repeatedly' shaken, !

.',' ':an,d re,aqipgsregist,ered ,at fi,Iteen minutes ihterva,l.'Firial readi~g8 are made at theend oI one hoUr. ' . , , , '

. ",'Bie~91tsar~redo~ded: asP+,p.++"p+++;aecordiQg as there is 'complete inhibitidhof hflilmotysis int4e:tubesofthefii~t .row only; of the, 'f}.rst and second only,' ?r'dfthe'first, second and third:' '~. ) ','

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'. 'P + +/and P+++ are rega,rded .. as, defin~tely positivE?; :p, + '0n~he ' other hiindonlyas'positiv'e 'in the pres€mce of~lgnsahd, sYmptoms suggestive ofthe'ais~ase., " " . .,.' '..! ,/

, 'Quantitative estimations of 'the amount o{ complement fixed show' that ' an average/pooled positive serunl', collected from cases of early bilharziasis.; fix .sevE?n M.H.D;sofcoinple,fue:rit: oyer,f.m:d;'abQve,that fixed:hy t,heave~age

'pooled '~;egativeserum •. In IUore!clll'onic cases,t~~ excess fixatiOn· ani(:>!l'llts' t'Q>foj1r :M,H~V.sofcompleinent.,,' , . " '. '

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1,. ' I ~ .) :\, ."',:,,, '. \ ":'1' ,. " ,< '\ '. / " .'. tV;~GON,CERNINGTHESPECl~tciTY OF ~IIIS dOl\IPL~MENT:F1:~A[,IO~ ., "/ {,,; . ' , " REACTIO~'~ ,." ,~;,.,. ,

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,', '(IJEx~minati6n of . a micro~cO~iCJ 'seeti6nof an il1fe~ted,sn'ltit s .H~er, , isno:wsit.to becoIUposedlllainlyof' sporocysts and' cercari~distetidirig the . inter~a~ihous~paces and:causjhg atrophyof,thepal'en6hYma.It' .follow's

" that'any ~xtric~' of· sucli' an organ is la;gelY~9bt~ine'dj~om \th~se ~sporocysts, ' . '; ah.d cerc;arire andi}ot from',the liver cells; , ". '. " . ' " I •.

. ,1 " . (2)1\ Antigens prepared from :tiormal. liyer8()£ 'snails (p; .fJOiS8,!/i) 's&o\\" ." .. 'np:teI?Aency wh,atever tp'fix II:\oreeompltmient,withbilh.a#ittlthanw'ith.· '.' ' nopyahsera. ' , .1' .'. "'. " . I.' ..•. . . • ,'. .• ,.'

,.,' '.1 •. , (3) 'Antigens prepared 'from the livers. of snails of the.speeiesB:dYb'owski \ " i infected with an anie<;lcercaria(Gastrodi8cu8:ciegyp~u,8>, shpweanp~jncre;;"B~d'

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. tendency'to fix~complem€mt withhilbatziasera. .' , , ',"" I •• ' (4} :Pr~~,nlged, \nves.tigati()Ilh~s faiied to;~emc:mstrate ;'any con~t~nt ,. .

t~n<;len:cy to pseudo.,.positivE? re~ctiqllina:t)yoneaisea.se otgrc;mpof,(iiseases. ~ I \ ,J,'

either, in,. bacterial;, metazoal; otprotoz6al; ihfectim:~s'.Th~ ,~eraottp:e "'>-. :'follo~ing -:v;tri~ty .oL helminthic'. ~nfecti9ns"T.eyMiwcoi;c1J,s,A8c~ris;'· /lumbrtQoid({siand 'A ldcylostomddupqe'nqli;2 have 'lluif01in;)-ly yielded negativ~ .. :res~lts.' \,,1 _':' ,',' r,. '.:_." ,: 1~'. _ '- "(, ,) I \~j' .- ' i,~ i ~ _,

.'Iu ,theprbtbpoal' diseases,the sera. of syph{li~ics, ,ofr~lapsil1g"fe"~r',,\ '> Icutaneotis Lelshma:oiallis atid ,ma'larif\i, ~have 'all constantly yielae~ negative' ' . :re:sults., "",.1. :',/. .... ",,' / . ',c'" ./ . ,\,,!, ,:', : "" " '

( \ Inv~l3w 9£ ~hese ob'le'rvedfacts, I. regard.. this:complen;lent fixatIon test,/ r ··,a~ 'a trueBordet~G~:ogoi'i reactiou';iJhatis, it. is\(tepen~e'nt;on/anexces~' .'

frxatiollof compl,etnent bya/pe6ifi(Ji1lP.~uneMdy .contained intlle . sera of .bilhar~iai patients ill theprE?sence_Glfits :specifi.c cercarial antigen.', " . (I!

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"V;-ANALYSIS;'OF RESULTSL'():STAI~E~IN'l'HEEix:AMINA'Tt6N ,OF' ,.s~Ri OF 311 HOSPITAL PATIENTS USING THE 'SALINE ~ EiCTRA~T OF AN'TIGEN.'

\' ' 'Ca) In' a 'group of i cases; whose disease"w.asor'uot inor~npa.ntw6 year~' <1utatioti;·thirty~p:ine out 'offifty'-f.Olilt,orseverity-tw() p~r .rc~p.t; yielded

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pasltlvereactI'on~. .' ,... ." '.. . . .... •.. . . .: ..•. \ . . . . .>/' ."

" . .(b) InitnothfrgrDup of' cases 'ormor~ than two years' dtirl1tion~thiftJ-' . , one outdfforty-six,or 67'4pe'r cen~1 yieldedposi~ive tea:~tior{s.,.,· .• ' .

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-454:, '-.' mscovery-oj aSpecifid Obmplcmerd lfixatiori Test .. · ... - \ - ," -' "

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' •. (c) ThirftY:fivecase!';.l of . syphilis yielded Iiegative-reactionsutilizing' ifresply prepared' anti~en_ ;" ~l1'.ga. veistrongIYr>ositlve' -WasseFmann l'eactiol'ls./

, . (d) One hundr:ed and seventy cases suffeii'ng'Jro,ill different infectiqns, , bacterial, protozoal and inetazoal; were all n!=lgative. ,

(e)-I!} 12 . cases _ a' tyndencytopseudo'::positive reaction was • observeg: \16 easefj yielded; a;J? +; -4. ca~es.yielded a P + +; 2 eases yield~daP + ++.

, 'rhese '~era wereex~rnin~d·earty.in ~he,course oft~e investigation. ' .. ram ..... now awarethatthe use oflresh antigen aud -it~ more acc1,lt/1te ~tanaardization

.'. '. wouldcilrt~inlyhave -eliminated this ,tendency jn certaIn ,ca~es; but".as th~ '~lc0h.olicantigen was sOQn after foun~ ,to be more. sensiti-ye and more stable ' .' thanthE!sab~eymiety/thtl fornieri$n,Ow always. used, ., . '.

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:' '. ·.VI.--'""'ANA1YSIS' OF THEREs'ULTSOB'fAiNED IN AN EXAMINATION/OF TIlE • ,\' ,_' I /; ': ' • i ,~ . j , • ,- :,':, " i!_ " . . , >, , '\ " • ',' ,~ • , , -: i \

/SERA6F32~CASES, USING. 'rHEALCOHOLIC EXTRACT AS. ANrlGF,lN. . , - ~ ~ .. ' '. I' _'.' " .... - .. " :: " '. ' .. ' \ ", !-

,In822'c~o~secutive,c~setl i)ivestigated; the, foIlowinK, rE;s'\llts -were, ,o.btained,:~' . . '. " ," ... ,' " . " " '

. (a)~In :8. group of 36, cases ,of hissthantwoyears,'quration, 32, orB8'8 " per :-Cent, yi.elde~ poslHye: i;eactj~<?Iis. , '. '. " '.' . :: "I ' .'. ': '

': • '(btlri a 'group of 97'chrollic 'ClLses(oftwo ye,aiis'durationand over); 72,. >.' ~''<;r74'2p~\,<oe~t,yieldedpositj~e r~sult's. ' ..•... , ,. " " ,'./ /

. . . ,(c) In forty~fourf- eases of syphilis.riegativ~;::resu1is. were registered; ',.1

ipoughaJI gave 'positiveWasserr;napn reactions'.! In~n,e case, . It "P4 .J7e\tction~ wasobtah1ed, h.ut .this wasdge to a~ti-complementary,tendency in the serum 'control tube.. .. ,:, " " '

I' " , ,~. . I..' '. / : 'f y ,,'" , " . ' i

(d) In 150 other sera, whidhittcludedpt'otqzoal, metazoa? dnd bactet'ial

1\1"

. injections;negati1)eresultswere,un,i/ormly 9btq.ined. I'~ oniy.one'e~8ewas a f

'positiver&sult registered in yv hich .ova wer,e Ilot. found iIlthedejecta" but, as, \. only oneniicroscoplC e~am~nation~ft~:made, aIld ihistory,pfbathing ill'an infect,~d 19?ality~asobtained, and as, moreorer; certain intestinal sfmptp~s .

... were present, bilharzi~sisco,uld not he -excluded. Prob{1bly the ca(le was one .... .of'latent infectj6n.'·· .': .' !."... " . .

, (' .'

VII.~1'iI:E I:MMUNITY'RESPO~SEIN -ExPERiMENTALLY' IN~1i:CTED-" ' MONKEYS. ' .

:r~ IPo~keys(Ge;copitheyu~), ~~, a variable,tirneafter heavy infecti~n~ith: '-cerCari~,~omeor all of the following~~ymptoms 'developf E~a'~iatioi;l, .anorexia/:riiusc~larweak:riess, .trernor apd definite rigors.If ,the monkey.

•. is' goi:qgto (lie, tb~se incre~se'jnintensity;'1f recovery is to ensue,' tbeY' ... ' 41miriish.FoUow~ng liecovety~ :jomLlizings~?nptoms: ~upe:r;verie~at a later'

:dat~: .. Iij !3',1!';Clnsoni,iI).f~ctionsdysente.ri~ sy~ptomsnot :~ncoinmo~ly ,11ppear durmg\ the seveI?-thto tenth w~ek;~ m,.infection with. B.'hcemalob'ia. yesi'clJil.oIieswe~enoted frqm. the.tenth'tq. the twelfth..N ot 'infreq,u~~tly a severe secondary .amemia,.. is present in' these4eavy ,infections. Poikilo-, cytosis,anispcyto~is"PolychroliI~tophqia.and (normoblast;' may he ()hserved

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, in, blo~d smeal's; o~ea~iotiaUy megalol)l~Bts m,a~appear.'Both'the hromo-, globin imd·.tqe' red hlbod,corpuscularcont~ntof, the blood are- Plarkedly, • diminished.' ' , ,,'" .

" ,,' (q,) bb8er'Pati(jns.o~)3lo~d Picture, etc.,in1f!onkeys s.pontgneouifly,

/Yecoveririg from the Disease, " ' . " " " ,', "'" '" , ' ,\ "

f ,An ,an~JYBis OI 'Ta}}le'J ,shows tha:t in ten nionkeys recovE:lring from', :initial,toxic effl;lcts ofal!lassive cercarial jnfection;a iemarkahle,c€)1l111ar, " hurpor~l'Tespons'ewas presehtiriEwerj case, "The:leucocytosis',durihgtbe: fifth t,b, the tw~l£th week variedfrotn13,OOO to 34,000 per cubic ,millimetre. Th~ 'eosinophilia v~riedfroJn' 10 'percent to 63'2;p~r;~ent and th(:)average, over this ,"period equalled 28 per <?erit, Ineve!y'case ,a strongly positive complement~xatlOn reaetio,ndeveloped, ~ The b~~emairdw\ showerJ a

'\lDarked:ht,crease irieosinophile myelocyt~s,greati cellular proHf~ratibn ~nd \, increase, in the normoblasticelemen ts., ' " ",' . •

/. - ,'.' I -; " ",

, : T:A:BLE' r.':"''';THE, CELLULAR-HUMORAL~~SPONSE{iN ¥ol"riy s 1iecoverir<gFRO:M:,

',~iL¥AR~IA.SIS.'" ,', ' >\'[ ,

,\

,Species of ,Time,ofComplement',' ",' \ Leucocytes ' ,Nb. ' infection' obserYQtiol) '. fixation'test Eosinophiles* ':pe~'c,mm, AlitopfoY

_'_' _'_'_,_~ __ I_'_' _":_" _,_,,_~_,_'_,,_,_'_, '7-'~--'-:"-;~--~~~~---' _~~ __ ,_,' 22'2 'pet:, cent . ;15,000 Lateral ':spined

dva:itnd wori:I),s 11th ~eek, /Ii'ositive

" "I'

, "1 B. man~oni

21'6 13,000 " ,10'7

l' 17,500 < ~" '" • 63~2 ", 28,000 H! " 24'0, "H' ,14,700 '" " 41'4 " L_ 34,400, ," 35'0 !" , <27-,000 "

" • 30'S .- " ~+

15'5 -'.'-- 13,800 " ," ,'.J' :'-' 1:6'6, ,,-,,." 18,000, Terihinal-spined , ,

ova a:q,d worms

~ /; .. , 5th ,; /)-t', '1--'"

7th;' , l' <'" ' 4 ~\, ,8th " 5 ./ 7th ,~ -" ,'~ ,

,'6 ,':\ '., \8th " 7 c" ,'6th " :S, " 12th

" .,.

" 9\ ,'12th '/ .J .'. , ". -l-'-' 10 B:' ha!1/uitobia, .' ,9th,

"Ill the Grivet moilk~.Y, tpe homologueofthil, eosiriophile in'man is a,iargercellthanthe" polymorphonuclear. hiucocytei and c(jI\tainslarger oxyphilegranules, ',Its,protop'lasm Ju)'wever" 'has ,a greatflr affinity for basic, stai:l;l, 'and therefore, in staining by Leishman's ,rhetl1oliit is

" necessary to wasJ;t' out lOI1ger than :usual with ,distilled water~todembris;trat\l' the oxyphile -; granule~', . "" ,,' . " ,

/t .

. ~b) Obsefv~tionso~ the Bloo4pictuTi} 'QllJ1()n~eys\dying,~arZp' , ,\ . ,in lheDisease. ~ \ . '

While'alive,:tbe'toxie st~ghlata jnthes'emonkeysat.e obvious and .:thesymptome I h:~tvf3 enum~rated.,pr6noun~ed: Microscopic exam~na~iQn ~ho*~ cloudysw.elling of ~he pa:.renchy~a d£'orgaps aJ;ld \p~ripheral infiltra~ tion 6f.theli ver with rbu~nd cells. The eel1al~:i:-hulmora.l response ,is. min'imaJ'as a general 'rule.: Theanimaif'jdie eitherbe~prether~, is ~time for' .,' an imm'uiiityresponseto dev~16p, or,,Jf theysurvivelonger~, the, intensity of . .the. toxrenii~ oftenieauses .aeomplete d~p'ressiion "'of the mechanismo( "30 -;," . .

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""D~$co~ery Of 'a Specific Oompleinitint,Fi,X;(ktib,n,Tes~ " , ! ~ ~, \ . ',' ','.' i I"" "

. 'lmmunizayion as' is\,s~enip.,T~bfl~t'.; ,'It is!;interesti~g'to p.ote.t,hatsev~ra;l , of these nionkeys, died ,before ova c01ilId he, Q.ep9sited in:,their,tiss~e~, .and,as '

a ,direct; resuI,t of the' '~dion,,6f, a ,circulating;. to:x:in~ tl}e product o~"the ,develo~iilg' worms., ' ,

" .", i '., ~ , . , " " ,,' ,'! 'I

TABI:.E II.-~' Tl;iE,CillLLuL&R-nUMoRAtREfjPONSE '1:,( M01:'KEYS dying FRoMBILIiU"l\~~ASIS.'" .. ' ' ' ". ' _; I , ',' !,',", ~_ . . '\, f "" • :, /

/ {,'

" No., ~, Species of , infection

Tihl~ of ' , ,Complement 'obs~'rvation' 'fixation test :.!Eosihophiles

(

l~'~. m,ansoni,,-' - ~:hw~ek 'N~ga~;7 -;~ per:eriT -::-~;~- Lat:~l' :SPi!l~d-, '. ova and worms'

, 12 8th F " 11'9 ,,8,800 ;, I' It'

,13 : ,_ ,'" 14* " " ,

6th";; ;'''' 18'0 ,...:' 18,090 ,,', ,,7th ,,' Positive ,56'2" 25,900\ ' .. ''', 8th ,,' " , . 11'0" , 6,000· ""

.'3pd, .,': Neg~tive ,:-: 0'.6 ,,,2,200 " ,.!

"', 15,," - ,~ . ., 16 B. hCBmatobia •. .l7 18

5th H, ,,1, "', ",2,200 ,2nd' I"~ ,0'8 Cl ".' 3,000

, \ • '" " I '

.·':Chis,hIonkey {No.)~) developeq severe bilharzialdYllentery ~nduli"i~ately ilu~c~~bed, , ..' - , \,.' J ~ ,'! I ~" I, " /

;':' \ ' ,

, Vn{"'f'"RELATIONSHIp.'BETWEEN THE ,GRADE OjJoEOSINOPHILIA AND " THE COMPLElfrENT FIXATION RE'AcTION i:t;'fMA~. - , '

'. ". ' , ~ , \,," .', " ! " I , ". I \ '

In fifty cases ,of bilha~ziasis,coqrbine~' ,observatio:ps were made' on' tl;le ecisinop?~liaand: 'theco~pl~I?~~t . fixa~ipil ' reaction" with tr-e, , object. ~f , • determInmg whether ;myr~latfon8hlp efi:lstedbetweentpe degree, of. cellular-, '

, rea:ction ahd the,humoral response t9 p.arasiti() invasion:, '1:\ .'" '

. 'The observ~ti~ns roay pe recorded as follbWS:-'- '. , "'. ' . , '. ' : ;'(l);Intw~,nty:.-thre~cases o(bil~~;r~i~)TieldiQga peJ!centage.of eosirip(~i.les) o~Jrom l.p~r cent to 10 pe~ ceQt",slxteen ~or~9'5 p'~r cent), Ylelqedpmntlve,. compie!Bent6xatioI} react~on~: T~~ in,tenl'lity 'ofth&sereactions In,this, group; however; were nO,t \lls. greatas in groups (2) aQd(3); 1\.'

': ,(2) Ill" twenty-twocasestbe eosinopbilecotlUfvvasb~twe~n 10 per,cent,' and ~5 per,cent:;(lig~teen ,(01: 86'.7perce,ntryie~ded:.strongpoflitive'dolDple':: mentfixationreactib'ns.~'f;' I'·., I .... ""'fl .', I, "

, ~, " -, . , ' I , . I

"\ \

(~) ]:0 six casesjn which the, eosinophile co.unt wa.s between 25 re! cent, and 40 per qent. aU( o:r lOQ·;'per"cent), ,yielded ~trong 'positiie, ~eac,ti0Ins:' , From .. theseobservations' it ,wiU.-be ~een 'that' tpe higher the g:t:IJ.de of. ,eqsinphUia present"the gre!l>t~r is the pxatiQn' of th~comple~e~t. . Positive ,~, ... , fixl:\>tion reactiQns aienot u:Pcoq1lll,only' bp£ai1}edinbiIharziasis without any

:1,

; I'l,

" ,I , h

eosin0ph~lla heing. d?1noristrated, f . ". ,... •• ,.. , ." ;'.:.,,'

,. , ,'-". '- i I. " -:.:- '"," '. - '- ., ,_ ;' .-,..',,/"' \ - \ , -,

, IX.-,THEPRACTICA:L YA:j:.UE.OF lTIftSQOMPLEMEl{TFnt:ATloN,TEST.

l Th,isre'action has 'a twofoldapplicati<>n;~' .. . II

(a) As a means of diagnqsis:.. '. .,'. I i' '". ff '

,(b)'As a,therapeuticindexto,thep~ssib,H~ effect of anygivendv~g on the par~:ptl>ilharlitia Vl\orrh., v , • (';\." '. ,

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,(a,) As a'me,ansoj ;qiagnosis!~. ( L, '

,',JQ,early b'ilharziasis, that is, priQr to th~o~s.e~ofi99aIh:itiglesiollsin tbe, :b]adderandl:'~ec,tum, 'this,':is the 'only re!!-i me'thoaof" diagnosis. Inlat~r /(" cases~ where \ the: locali,zingsymptoIlls/are top~'cu~e ' and, ovaaredifficlllt , ,; , ,to '~emori~t~~te" (especially in B: man~oni);" ~h,averepE:at~dlY<1iagno~ed 1:>ilh:ar:ziasis by t4e:serologicartest'alone;,s}lbseq~entcconfirn;tation, has

'j .,: •• '~" been Obtained by cy~toscopiceXaD;lii1ationabd,'la~erbyth~ demon,stra'tion ,Of' ,oya in' :!t~e~ejeqtai af~e~ rep,~~ted : examin~tion~ . ,Iti,~'st~blishing a~', 'endemic' indeX' ~dr 'bilha,:rziasis, ,thIS reactipn is:i:nore 'applioable:JoF, the' ,>~xamii1ation?f,a(large:;Ilnmber Ofpeople,tha'n~sthe 'mote; tedioJ,ls;, ',> <'microscopi'ci'examinatiqn oLthe dejec~a. For;the,a.veragecase,however;, ' '"the:'establis1;rm,entof .a· diagriosis./by,:,thedemoiIsj;r~tion, ofdva'in~e:'

i .. dejeGlta must, ~l~ays' ri~main, tbe /irOu.tiu,e / 11!!boratory .llletboa, 'provided " ,alwa:ys its' shOJ:tcomings, i.e:, , the neces~jty qfrepeated exarpil1a~ions, '!1re ; ;'

'- ":recbgni~ed;,;;Tne (iiagpos,ticvalue,of this'90IPpl,eHienti, fixation react~on' cau '\perbapsbestbesh<>wri:byreport8onth~,followiiiggroilEis,qf mbS6l'/' , ,"" '

< .. ' R~port( a).---.,. F;'orthree mopthscases presenting"urinary,symp(qffis s,lCh' ,a~ hremattitia, pain arid. frequency,' were, sent to.rue forexamin~tiorl frem' ':.a num~el' ofBr~ti~h. militaryliospitals lIi~irO,. ,'Ih~n~yt~!e,ecas~f,!was I . '., , supplied. withany:preyiou~inforD?-atiqn ast?the, presericeorotoj;lqvise' of ' , ova i~thedejecta,; ,In, si~teen a positive diagn.o,sis,wasmad.e.on: the blood

fi.r;tdings aloJ!le';,pfteen were ~ r,~por~e.d,'as neg~tfve.:' In : elev~1f of )these n~gative :cases,tpe pause/of "the urinarY', s~tU;P~oms ~as. trac~~to, conditi~ns, ., such as n,ephdtis,reiuil calculus, etc., Iuthe Jonrremaitiing<oncs ova were'

.,', ' '" being i>ass~d,in'the urine, w¥ileat tbe\saliH~time tlieserurine~cti(jri wa~ 'negative. In, ,three out6f the six'tee,l!cllll'es whichyieldedia positive reaction., "

"ova had p.ot" been f0~nd il;l the 'uflJ1e; l;)utO~lopelWade; D;S;O;,A;N[.s.: " ,Co~su1ting Surgeon.,'iconfirmeifthe, serolo'gjcal diagn:?sis ,by6ystoscopy.ahd 'subsequ~tly';9v~:'Wer~ dem&nl>~'rated in,~each, 6~~e.: \ }J:;blls out.qftwehty

~af?es Ofbilharziasis(~. 'h,ccmatobia otB. : 1I1:Cf:ns,()ni) ~ ~ixtee~:o{ eighty per c~nt 'w~re'posit:iv'fly , O,iagIiosedby, ·.this / llle~lipd; , "No!)?seudo~positi ve reactions 'were'rec<?rn~<:l;' .'; ':, , " ' . I :' ",', :. ,";""~,', I

/ / - Report '(b).:-"::I: lVas, reqhestedby ,tue D.:Q.M~S., A.:r;R, to inve&pigate , ,hynieflJn's:of this ieactio~llt~putbreak ofbilharziasisin:pale~tine~rilonisF" , "tPt3 ; MlouThted Yeomanry' Divll3ions,;oTh,ese merLhad. bathedt;. driri~g the' /',i)revrouis;'liluuim:~r,in 'fresh, water caI~~ls in ,th~~aJ:'ol1IAwhere !Builinu~ 'snaIls e~ist in large.,numbeJ:s./· "', ", " . /. "" I, ,', , 0

'1': ,'Syst~ll):~tic :utitiary'examinat~bns\ qf iRe whole unit ,-pad,'beeIim~~le, six' , _ j i. " 'wee~s previous to my vj~tt;and,aU cases.pass~ng:ova, 'with the e~~eptfbn\()(: "

SIX m.ild iIifecdons, were evacu:!1teg ,tqt~e,: base.T~e cOIilplelllelltfixilItion/:: rell.c'tioti wasper(orPled in tpe Anzac ,FIeld LaboratOl'Yisom~',ten:to fifteen

, -,<\

" j milesbe4jrid"tbe' firing line. Inall,~ fifty-six, ca,ses w~/reexamhied;firty of. Jhese were,ri:ot 'passing ova dni theu,rine andw~re,:"pp~rep.tly qUIte healthy'

, ',men; in six, ovawe,~e, bleing pas$ed.:c -~utof, the si:f c~s~sin wl10ni oyal;la,u'

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biscdverY/9P~a:Spebi]iiY;: 0 ()111:pZe'in}/nt ; iixafii~~ , --', ' ' i . i ' ~, , ' . . . , "'. " ' . _~ • '

A58 , I '. '.' I ' " ,- " I , '~'" , , . ,I ,< ',' ,,1, "., ~,,' . i ~\ ",' ,,' ", _ / ,'." _. '

" been. delllonstrated iIl:thej~riiiefh)'e' yielqed p'()~i~iveser()lQgical reactions.". Oftbe'remai:i;ting ,50' cases) :~ga;ve' I>dsitive,re~ction~, f»ud/ withipthe next.· £ortnigb.t ova were fOU'Ildiii theutineOf 56!' them.ip nJo·rtuTl'ately. I ,was' no~aple, owirigto wiJitaty.exigenci~~,tb.follo.wup the F~piainirrg'/ three cases over' '~'. longer:;period,pu.t in ;two,at)rasttb~blqoa piptiu;e showed" an eosi~6philia.~· Undoubtedly )naU 'ovawould have be~n delllon,".

·str~ted,4j1drepelJ;tedexamlll~tjolls beep. possible" ..... ....• .' .••• . . .' .... . Replirt (c).~'l'hir6y-t:h1:'eeeas~8:of:l),rina~y .alldlnt~stinal. hilbarzia~is . wereexiuIlilled~ 'from the Asylum of::theInsane,.Ahpas~ia,t~rougbtM(

',,\coll~£es~of Dr,'W,·'W.Warnoc,k) C;l\f(:f:;.:oftli.~s~ 33case~, 39or'SO.per: • <;ent yieldedpoE1itiveser.()logic111Jea~t.im;H~. ;/ ..... ' ') ...- ..... ;

( Repo,rt(dj.-TqiitY:cases.ot,c4ronic Billia,tziasi.s.:'w~Je ~xamiQed rrom, K~8r-el~Ai.n.y . Hospital. at the requesJ .#Mr.,F.:Col~"¥,~ddeb., F .RC:S.· ,

- Twenty~three ofthesf'l,or 76·o.p~ri e~t.yi~ldedp9sitive resu~ts: In sev~ll Tt· was negative';. It isfre:qllei1tryjID:possible~o(denforistr~~ WOl'!llS at 'the' ':' .au·~dpsyincase~ ofGh~ollic,biJh~J:zlasii3f Owingtd sec~ndary infectious, arid' . the repeated deposition of ova in the walls oIt}le hol1owrViscert1 (bl~dderai;ld" .colon )ifocaZ : lesion13 .can~ persist; aod 'even cause; 'de~th, ~Orllah JonKafter

'tqe priniary . ca~~e, i.e.;·tlleWO!msthelli~eJ\res,/~ave c~a,~ed to;exist .. ~n; (", ;c'ases suehas tbes(e.OIiew6l.l~d'I1aturally expect to' pbtai1;l 'a" ~egative

. ,poIllplementfi'x:atiop,' ~nd indeed ,t~isis/often t~eca~e: '. . . . .' . i .', Tge ,antigen, l1se'd, in 'the repQr'ts(a)ana(b)'~~s the.saline extract ;' in'! .' (c)'bo'th ,sa:line\a:p.daiC6holic extracts were used. . This . propably accpunts for tbe' high ~ercentage "of pqsitive 'reactions o~tained in tilts series. In' (d) it was the alcholic extract alone. ( . .

( , ; fi . ~'

(bY A~aTheripeu'lic.Index tothePossibleEffect;j a given Dr'ug,on '. . . ....... the parentB~lharZia~:W6;l'jn.: . I ( ( '"

. '" '.', t' , \ ,/ :-, , -",' ,"" :'" ',>, , • ,,--4"~" ,'i' ,:', ,-',' J:: :. :' <,o ""- » : .... ' " .. ' ! In .the,past itp.a§ be@ cl1stomary to. adjudgethethera.pt1\ltic val)1eo£ ,; '~ny drtig~inbilharziasis ~c.Qrdil7lg:t.o the following criteria :~ _ . " .

'(1) The .temporaiydisappearanceof vef?i~ftl()rint6'stin[tl.symptoms .. '" .( (2)Th!'l diminu,tion In' 'tlie .. n urn her of,o¥completecessa,.tioi:lof passage~

,()f ova in the dejecta, ortl]e • inability of "sqcho:vato', hatcbout ~ira~iclia," ';Suclidata,ho)Vever~' ~ct. m~rely 'as'an, inde~ to' the. ~e}llporary·ameli6rat~ori~· .•

<·of local Co.hditions; They do, hot implY"that actlre of tlre disease has)· . been! attained./~This information' .tells one ilittl€!' concerning the \fate. of .. \.th~ true 'eaq~alagents,' ·the·pa~en.t:"worms~ . Recent adv~Ilc~ in 'Dui kno~led.ge' of hllharzii1sis"b9wevei;.iwould now..seem to ,afford a scientitic, ras opposed to. an emp~rical. ~a~i.s, for es,timating:.theeffects oCany .giyen. .' (;

Jine oftreatmerit.Thus:~ ....: , ,>' .......:( ..." . I

,'(lr~The/lethal: 'action qf ,,,;doll,s, drugsonwotms,andcercari~c~ri, ~e, ," ,investigated in'vitrQ;; .' •....... ' .. , •. ;'; ' .... ~ '.. ..... , ··.·(2) ... M~l1keys'can."pe readily iri,fect~d • '\Vith! the pa~~site, .andJhe ~ffe'ct. of in§rave'i1ous iriiecticin,ofanydrug,foJ;lndt6 ~xe:tt a s~~lective1eth!il>I'action 'in vi't~o!tested:. .., , ' . ., .

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(3) T1iecomplement~ fi:x'at~bfr \teskp~r'haps bontrolled by', post~morte1p examination, can be fused to !l~termine tlie 'effects, ,of the drug on the· I

I parasites, either in man' or in e~peiimentally infected 'mbpk~ys. .' ' ,.' .1u 1917', {"ti~ate~Lfour. casesw'ith"repeated intl"ctvenOus i:qjectiollf; of,

,~U:sol, 'o/ithiIegative re~ults.as iu,<}i:C;ated by th~f&ctsth.at thecomplerlient fixationre'~~tio.n:. i:eD+3tinydPOs~ti'vea.nd the·~osin()p.hm~ 1;lndiJ:pinisheq.. ' .. ",' , .' Onapttor~gr(),~nds'thlsW~S to b~~xpected, slUce·,I h,ave . .,found that· free c~lorin~~exert~ n,6 spe,~itio1ethal $.ct~on. u'pop.the.cercari~. .'. '. ' , Christopl).erion has recently reported Javourably/ o-n the treatD;lentof

bHharziaslswith tartaremetI9.·. adminjstere(l,by the i1ft~~~~nous ,route. ~ ; Further ebrifirm(.Hbry,ev~dence.of c,9mplete ,cnre as controlled by this

coinplemerit fixation~e~c.tton. ahd:demonstr~tioilo{the specifiuorselective ' . action'ot t:artar~emeticjn. hig4aHutio~S~9U theaQUl't. bilharzia. viorms' ~nd, . eercarirein vttro atesuggest~d ,as b~ing\ of tq'euttnost val\leip. elucidatirig: 'thjs point.; , , . . -:

X.~()~NCLU$IONS(· . _~,,' '<- .,',)_'< ',_ ~/~c

As are~ultof :this re~earch J off~~thef61ibwingconcIusiol).senulierated .' unde,rfive'beadings., .' .' ."...... >', 'l • ••• ••

'.(1)' Bilharzial parasite~ and '. their ova exert their,·qele.teriou~ influence '., . ,Oil the tissues .. oftheii:: . definitive host; .. man;'riJ.aiuly ,by the f?ecretion' .

• i:.or ex~re.t~o1;l. of , t()~lils, Iratbett~a.IF ;by jll)..ef~ , mechanical' action,_ as' has 'hitherto/been. supposed.,' . ..' cl' ". .... .

/(2). The cenular~hun;r()raLrespoll~e, both iH~an and in experimentally.! . f)nfecteam~nkeys\.is ~irp.ilarjn·. pr(nciple }othat-observed i in other '

'., . M1illinthio, prptozoalc&nd bacte'bal infElction~ .. ~In. bilharziasis there is an .. ", eosi~ophile'lE!1l:cocytosisand:::'corre$JP9nding',iricr~ase in the eosinophile , • myelo4ytes in theb6ne,marrow. 'The,hum6raLl'e}j!pons~,:prod uces an

immuhe bodyfr'amedtone1J(tralizetheoil,harzia toxins;.' ...... .... '.,." ':, ", . (3} By the empl()yrp.erlfof.aIt a!cohqlic' extract Ofitbe; 'infected liyersof. snails, (p .. bQissyi)asantige:r;t, apd :by' utUiz.irig • the techn,iqu'e, T<have. ,describ~d! positiv~:cornplement\fix!Lti?"ll' r~action.? ,h!j,ve ,b.ee1:~{obtai~ed in 3,.: .;

high p~rcentage. of c'a,sesAnmanas. well. as'm~?,perlmel1t!LI1y~n£ect.ed ,,' ~onkeys;, In tfe.'latter;:p()sit,iveiea;ctioIi~niaybe. 'established asearlya~ _

. thefifth.weel{ bf the djs,~a"sfand invariably::appear berore~he·tw~lfth;., . that~is, if the', a1;limal. recoV~rs,In rijenwhbseillfectioil was; of notmol'e,,· . than twoyears-atir!Ltion,~8'8~ei: cent. yield,ed:posit,ive ,l'esults; in more

'" ~hr9nic,. casEls 74:2 .Pe,I; c~nt. 1\sa:,gelleral rule;it ni~fbestated,thttt',for ; ',man"the.higher t4~ eo~inopliiiiil,;'the stronger tp:~fixation"react'idna,J1d the' .greater.jsthe,percentag~:of cases yielding it., ." '.' 1'./ • / .~.',.' .... , ..

(4) : This is. the first occasion ,,()uWhich a ,specific antigen has/been \' obtairtEiaf~omthe; invertebrate inteFIlleq.iary; whi(il~ wilI give a 'specific

reaction: wi,th the.blood·'o'ftlre'vertehrate definitive host. .,., '.' ,., \ . ........ . .' '. " . . ,

, ',(5), Thepr~~tical application Qf. thisr~ac~~on'istvVof?1d';-:7~ , . ".,. ~(aY It is of :diagnpstic vl}lp.e / eaf1:r:'jnthe'disease·.before localizing

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",469' ,Discovery 6j},a' Bp·eeifi;C;C0111;ple,ment.' Fixatiqlk ,Test l: ,... .'" "!' __ .' ":0, - ,:.. ... ! ">' /./ " !.'::." .',...\ -'" " ,

SY1l1pf~~a'~av~ d'~VOl()~,~d; \and;~lsoin' later\'c~ses~~' ~ 4ichov~!are. ~carttfl»' '. disttfquted in the ,d,eje:cta,'especia,lly ill B,. :mansQ~i;,}t WQ-qIa' pro-ve of ,the,

utmost' vaJu<1 i~'de~ermiN~ngthe endemi(J':~nde.xof \bilharz~asis in, the Nile Val1ey,tif~ucha' scl1enI€l V\1ere ever, uI}d~rt~ken: ", " I,

. {W I(shbuld pe ~ofthe greate,!?.t'lvalu~ i~~,~stinia;tingth~' effe9t of the" intra.venous .a.d~~n~&tfa~ion'of .. ~nydr1igs,l.lpO~ the. ad~lt~ar:asjt~s.1 Ip., tbis\

,.ie~pect ther~actiop: shquld,s~~na,in ,the, future,ijuithe ~sametel!1tiop.ship J' ,

to, ,bilhar,ziasis -as. ~6es tl:le' Wass~rmann ,test <to syphili!'l, at the .preset),t ,. ,moment, QIilyI w0uldelti'pl1as,ize that, ul)Ii~ethe Jatte,r;.it'lsspecific .in/'

the liter~l mea~irig o(tpe:term!Ther'e i'sn,~?h'e~~'inthic.:di~ea~e w~ich "l;ha!ve'iI).vf!stig!j.t~d which,wi~1?give;I)sel1d0,:pq~itive reltctions ·as'.do certain

allieq Pfoto.zbaLdiSease13 in tpe ~las.8icalW:assermaI?n.test. , \ . r .' .;1,

", .. ' I am:' m:uch indebted. t?, M~jor\p.IL 'Bahr,D.S.O.; R:A:i\:LC..;;and ,.to' F~ofes~or.'F. Cole, ))1adden, rF .R:U. S:, for the stir'fiulating, inter.est ~~a:help / theyha've .'rendereft ~;u.ting' th,e~re~eI1~ i:nves~t~gatioi)i;' . i . ' ',' '. >;

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,i ,REFERJl]NCE,S.; ! ': :/ J ",' {""r , /' , '., ./<,,:,. '. '\ • '! ... ' ,

{I ,OPIE,191Q,\ ,BarveyLe,ctures,p.,192.' ,'t<,,: . 'r "

[2] NOGU(:jij;I, 1(1)tJourn. Amer: Med, Assoll., \:oLIViii, p~ '116.3'.' ,.' 13] McGow:u' ~ 1913. ' Journ;, fdth.anq" Bact;, vol.xviii;p.125:" , . '

;--' :[4J THOl!(SON,,'J. 'G.\,' 1918., ~'9ompleme9-tlpeviatiqu' hiCa.sesof 'Malaria: a. New, Aid' to . . . •.. : / Diagnosi!r;" Brit. Med.' Journ;., DecelIlb~ 7, 1918. •. . ' ... c , . . . . ,

1: 5] (hUPINI,1906a.nd 1907.' I ".Retlerche slit Siero di Sll.ngue, etd.', ': Gazz. do Osp.er/,. e. del blin.:, ' .• ·',No.,153 and Nos. 6 'arid 45. > \,. _",-',' • , ',' / ••.• ',' •

f61 WEINBER.Gand'PAavU" 19Q8.: i!' Re~ctidndeBordet,Gengou 'daus les, ~elmin~hiases;" Campt. rend.; de la So'c~deBio!~'ivpl.lxv,No./28!Oct.db.!lr .17, p. 298. WlilINBERG; 1909, . v

. : ' Ann. Inst., Past., 11)09, vol.:x;:xiii,· p, 472... . ' ' .. ,. ,~f\ {7J RREUTER, 1909. Mii;/wh: .. me{l.Woche,ns:cj:h1;'~, vql:'lvi'; \ ,"

\(&]MEYEffi,1910:. Zeitschr.j.:Im-rnuniJtk, vor.vii,p.~732. j . ' •

.(91 Fi.:miG ana. L!SBON~E, 1\l07~. :Corrip.~. :end. de la;Soc; d6 Biol{ vol. lxii, No'. 23; p. 1198: 110] WELCH; P.A., ,and CHAPj\IAl;'!,~H"S.; 1908. '''TJ;tePr~cipitill Rea:ction ,in Hydatid :Disease," \ 1908 •. Laneet,M[1Y 9,p.lBBS; , . . . /;;" . . .. . . .'

·{ll] LANGEB, 1905. ' .Munch. med. Woch~n8chr~,vol.,lii: p.1665; \,; .;.: I ',' ! If , '. ;

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