Informal meeting on the progress of entomological research sponsored by the Onchocerciasis Control Programme: Lama Kara, 21 and 22 August 1981

aWORLD HEALTH OP..GANIZATION ORGANISATION MONDIALE DE LA SANTE Informal I,ieeting on the i'rogress of Entorrologicol llesearch Sponsored. by tl.re Onchocerciasis Conf,ro1 kogra;m,e Larna I(ara, Togo 21 and 22 August, 19BL -'.t..c t,.- . ' ,:^'S\i." I r1/-.' -'-)' - \'7-/lt i , ,::., i'- i i'l rt: E/i //46(-. \t ,,i $ IROCMDIIIGS - 21 AUGUST 19BI [he rneeting was convenecl br i}r 13. ,'iriiippon at 08hr0. ile wel- comed the participants, enphasizing the infornal nature of tire raeeting and inviting everyone to express his id.eas freely. nr .i. Grunewald reviewerJ. the anlenrl.a/-,rueounced. that therc worrlcl /*d be brealcs for refreshments durin,,; Lhc urornirr."; (tl8irtl0-I2hOU) lurd ir-f Lr.rrtr<lor"r sessions (f5nOO-feh00). IIe thanlicd tirc L:una tlara stal'f for kindl-.y nr.rl-j.n,,1 a.rrangements for the meeting. Technical presentations then began. r. Entomolosical situation in the Lama Lara Sector Itir Ad.jonou, after warmly welcoming everyone, on behalf of the sector, presented. the situation in Togo. Rains a.re no!, heavy, vrith consequent flooding and sudden changes in water levels. Itaily caLches of flies have been in general }ower in I9BI than in lxevious ycaxs, bo bh in bhe llone already treated and in the Extension Stud.y Zone where experirnental treat- rnents were camied out. The most irnpressive results were obtained. in the Extension Study Zone in tire Ata-iparne sub-sector vhere the average daily catch at Djodji during JuIy passed. from 43L in I9BO to 17.I in 198I. Treatment failures on the r'lono itivcr renrained an unsolve<l m;7s Ler.y. I,lr Adjonou thanlied Llte corrsultanLs ot' Ltre reLnvLrsic.rn sLudy team I'or taking the tiroe to share their l',nouledge rvith him. II. Entomological situabion in the iiatitin,?ou Sector l4r Oredraogo reported. that Benin uas also experiencing heavy rains and. floods. The reinvasi-on in n<.rrthern ljerri-n has been a,b lovlcr lcvels at Cbasse and Zougou than previous years, but it seerrs r:nllJiely that this is corre- latetl with the experimental treatriients. Some affluents near Gbasse were treated for the first time this year. i\rrangernents are being mad.e for ground. prospections in Nigeria to investigate possible sources there. I)r (lan'ms mcttLionc-cl in hi:; diricur;r;iotr LtrrrL i. l, wor^rlrl Irr.r1rri.Lc rr:;r.l'rrl l,rr knoru the dates when rivers starbed to flow in lligeria. Some perio<lic treatments may be d.one there. ...f2 a -2- ITI. Reinrrasi on and Effects Xxper imental Treatments Dr Garm.s presented. a brief review of the reinvasion situation in northern Togo and Benin, fron LSTT to 1981. In 1981 the experimental treatrnents were intcndecl to treat all potential breeding sites of S. squanosum in the a.rea. In general, checlcs of larval breeding sites ind.icated that the control was excellent in the Asrkawkaw, hlawa, Kou6l and Kpaza river systems in both To6o and- Ghana. Eowever, checks of small headwater strearos in mountainous terrain could. not be made and some breeding may have persisted. there. In the l,lono and and Ani6 systens, the tributaries and. the Ani6 vrere well-controlled. There were, howener, persistent treatment faih:res on the I,trono, parbicu- larly in the lorer reaches near Atchined.ji. At the level of biting catches, the reinvad.ed sites had. ver;; ]esr numbers of flies for the most part. fhe pea.lcs characteristic of reinva- sions were absent and a few nullipares were always present indicating some 1oca1 breed.ing. The cytospecies comlns:i-tion of reinvading flies stayed- the sane as in prerrious yea.rs. However, it was not clear whether this lack of flies was due only to the treatments or al-so to natural conclitions. rn the zone of experimental treatments, catches were very low on the Asukar.rkaw, \[ava, Kpaza and Koue systems. OnIy pale wing-tufted flies re- roai-ned. whilst earlier catches included. rnany d.ark wing-tufted. flies. Catches were h:igher on the }iono. Little time had been available to study the reinvasion in Benin. Ilowever, the presence of high nurnbers of flies in northern Beni-n vrhen the Terou River was dry means that this river is not the sole source of the reinvasion. Suggestions for future work 1. Dr Garms expressed the opinion that many of the residual f.l.ies re- invading the Larna Kara sector a.re S. squarnosum breeding in the mountainous area. on the Tog,o-Ghana frontier. IIe suggested. that additional catching points bc established. west of PagaTa to monitor this popuration. Dr phirippon agreed. on this suggestion. ...f, '1- 2. Dr Cheke will fol1ow the Septenber peak of reinvasionl deter- nining the species composition of biting flies and. prospectin6 1rcssibIe source areaso 'Ibeatment faih.res ort Lhe liorro ltitrerrv. A. Dr Kr:rtak presented. results of susceptibility tests carried out at the end of the treatment period. IIae LC l0 and LC !l were within the normal ra.nge. A high percentage of moribuncl larvae was noted. B. Dr I,ieredith noted. that the larrrae tested. in thc 'breatcd arca were all S. darnrosum s. str. and. that the surviving fenale larvae at the highest concentration of ternephos carried. a high proportion of an inversion associated with nale sex d.eternination. 'Iiris in- nersion is however fai-rIy comnon in the i''lono system, althou3h very rare in lvory Coast. Co A general discussion followed. the following'evirloncc was suggested. for taeatment failure due to under-d"osa"qe ra'bherLinn re- sistance :- surviving larva,e were mostly o1d.er and ruid.elir scattercd. in the breed.ing sites some sections of the I.irono and. the tribataries liere success- fully treated susceptibility uas normal there was some d.oubt about the correctness of the dose clue to lack of gar6e readin5s or changes in river level after the gauge reading:" Ilowever, there was also sonc evidence of sornc lcind of :lr:l.c.c L,ion pro- cess for tolera.nce :- - the prolrcrtion of nales in ernerged flies had fa1len to 15,'. - other Sinnrliun species had disappeared. - S. soubrense larvae were only forrncl in a very low pcrcenta4e, although rore had been found before treatments and bhere r.rere still S. soubrense adults biting. .../4 -4- lhe follovring conclusion tras reached :- Atthoggh the failure was probably d.ue to uncler-dosa6er furbhcr ir:ves- tigations are necess€.rf,o ffurther susceptibility tests will be clr,nc in October, along with carefully controlled. trea'bments rrrcnitorecl in iptter systems by I'ir Ocran. v. cytotaxonomy resul'bs Dr i{ered.ith presented. a reporb on the d.istribution of the cytospecies in Togo and. Benin and. the mar:y d.ifferences on the intraspecific level betr'reen the Eastern and. i'/estern Zones of the llogramme. In general, So Sguanosun is restricted to the mountainous region on the Togo-Ghana frontier and tire u-pper parts of the Ani6 and l'iono Rivers. S. d.arurosurn s.str. and S. sirbanrum are wid.ely distributed. i.n the areat the latter being more northerly. S. soubrense and. S. sanctipauli are widely distributed. in the southt but S. sarrctipaul-i extend.s further north than in the Viest. Sone of the peculiarities of populations in the i4astern Zone can be sunmarized. as follows :- some populatj.ons of these heterozygotes have adult fem:rlcs with pale wing tufts some popula tions of S soubrense S. i;anctlp?Uli , and. "so/sa" heterozygotes have many adult rnales vrith a r''bhree spot'r scutal pabtern which is quite rare in the ',r'est. This pattern is associated. with heterozygosity of the 115/65 inversion. some male larvae have been found. with chromosome a.rrangerncnts com- bining S. sirbanum and. S. d.amnosum s.str. characters. '-Che propor i;ion in the sa.mple is such higher than woul-d be normally expected.. no peculiar chronosome arrangements are associated r.rith pale vring- h:ft (Eastern) and dark wing- tr:ft forms of S. squanosum the presence of an inversion associabed. with male sex-debcrrninaLion on chromosome I is rare in thc::iast but cor.lmon in ttrc llest. ...f, -5- Dr llered.ith also presented some i-deas about morphological separa- tion of three forms of S. squamosum larvae which seem to be allopatr:ic in the west and in mounta-lnous areas. The latter two forns carr be s:'rn- patric in the east hor^rever. The S. souamosum larvae fo'und in the i,Iestern Zone tend. to be cha- racterized by tal} tubercles ancl broad- spatulate scabs. 'fhey can only be confused. with S. yahens_e. A second. type of larvae has the spaLulatc scabs, but nay not have taII tubercles. A third'bype, frorn 3cnin, lacks scales and has snrall trbercles. -L,lle relationships of these larval forns to ad.ult wing h:ft forms is unl:novrn. A Seneral d.iscussion of larval morphologl; followed.. The point ruas made that larvae of all cytospecies found in large rivers .bend. to lave smarrer h:bercles. fn smarl, swift rivers, the trrbercres benci bo bc taller. Uven S. d.anrrosum sostr. uray have hrbcrcles in these cases. nr Garms mentioned. that qJ sum from small mountain strcuns bcrrd Lo have many dark 1rcst-cranial hairs. \II. Possible enzymatic stud.ies Dr }ieredith proposed. three possible enztrrmatic stud.ies :- Io Stud.ies of esterase acti vity in lrorr&11 arrd. rer;.i l;'blr.nL tj. sorrbr.t-'ru;r, and o Lller oy bosltec.les. .l t trra.y be lxrss.LtrLe Lo clevr; Lcrp ii li-Ll.'rl c staining test for the rcsistant strain since resistance to organo- phosphate insecticides lil:e temephos often involves increasecl este- rase activity. Even if simiile coloration bechniciues d.o not r+orlc, stand.ard. electrophoresis may elucidate the enzJrme systems involved. in the resistance. 2o Stud.ies of the pate and darl: forms of S. squan.osuril. IIi gher trehalase activities in the pare fo::m may indicate it is a rnigrant. 5. A peculiar enzJnne pattern in S. damnosum s.str, sirbanum in northern Benin could serve as a label for reinvasion stud.ies. After discussion, Id6. 1 was 6:iven higher prior-i by. Arrangements were made for collectj-ons of material in Ivory Coast in liogid ni'brogen. Dr }ieredith agreed to begin prel.ininary tests in Ouagad.ougou io perfect the technique very soon. .o.f6 6The meeting was ad.jour::.ed for lrrnch at I2H00 and reconvened. by Dr Grunewald at 15H00. 1fII. I'torpholo6l1cel .idcntification of adults _of the' -:.:_-9:gl_,o-:lf-* -contpJ r'x Dr Garms emphasized that rto isoenzJrme sLudi-cs or rerur.i r)i,; r..'(,rl'- vrrr$ done this year, and. all adult iden';i-l'ications wcre done norg.riroloi;ically. ile further reviewed past work in this area and. concl-ud.ed. that none of the existing l'eys ttere suitable, especially in the i-iastern Zone and for use by non-experts. liurnerical and. mensuration characters a.re closeiy related. to overall size of the fl-ies, strch that such charac.bers nmst always be used. as a ratio to .i;horax or wing lerrgbh. In blre r:ias'1, vcl:y snrall "forestt' flies and very lar5e trsavalnal.rt' flies have bcen fcund, contrary to the trnormal'r case ln 'bi:e ';iest. Coloreltion charact-rs carr ile quite useful, but problerns irave been fotrncl in the East such as sou orense i. sanctipauli fenales vlith pale vling tuJts and. S. da-nnosum s.str. trith d.ark po st-cranial hairs and 5. s(luluilosum with all palc pos t-cr;utirr1 hair:s. IIe btren presenbed a ncw rnt.'Llrclcl 1'r.lr.' irl<-'rr Lif LcaLron it;rscd <.rrr l,lrr. r"lr"L-io of bhe thorax and amtenna lengths. Uhen 'l,hese rabios ai.re clc Lerrrfned I'or saaples of at least l0 fli.es, the results can be expressed. as d:stributions (histogrars) or as regression lincs. fn either case, puxe polmlations of S. d.a;:mosum/S. sirbanumr .S._€9S@Hr and. S sanctinauli can be readily separated., the ratio d.ecreasing in the sarrrc or(lc.r. i.iixed populations are ind.icatcd by as;a;rcbrical cristribuLions or ch:ur6cc in the slope of the regression lines. In biris case, further senaration may be attempted. on the basis of antenna segment cornpression of colors of wing tufts, lnst-cranial hairs and anal vej-ns. Dr Garms d.emonstrated. that the method could be used in ctifferential transnrission studies w-ith an exarnpJ-c,of a rnixod Il]orcstt'r'urd "sav:Lrurirl ''' population. t'1ies wittr h-iiJI nunl)(rrs of irrlecbivc ti,.rva../l'l.y vlerc i<lcnl,i- fied as Iforesttr cytospecj.es, (Iovr thorax/antenna rartio) c<.,nsisLerrb vriLh existing krowledge. .o./7 -7 - VIII. Dissection of preserved. S . danrmosurn adults Dr Garms continued. his presentation to describe a modified. method. of {issection incorporating measurenents. The routine is as follows :- A. The abd.omen of the anaesthetized f1y is open a bit of ovary pulled out to determined. parous rate. B. Parous and nulliparous flies are trmt in separate tubes of 80!6 alcohol. C. fhe preserved. flies shoulcl be kept in a cool, darlc place. At any tiroe up to years later, the flies can be neasured arrd tireir morphological featr:res recorded. 'rrleis is best done totatly submerged in alcohol. D. The fly is then placed. in a drop of 2fi1 acetic acid. After rernotring the head and d.isplacing it slightly, the thora:c is macerated.. In the meantime, the acetic acid. clears the macerated. thorax tissues and the earlier sbage Onchoccrca larnae c:ur be easily seen. The ad.vanta8es of this method. are that bhe measuremen'bs are rmrclt easier to d.o in alcohol than vrith tire fresh fIy and the 0nclrocerca larvae axe easy to see artd cannot change their position in the flyrs body, or leave the bod.y d.uring d-issection. The only disadvantage. is the lrolr;i.t,,Ie loss of larvae frc,n Lhe ab/omc.n r.rhen examining the ovaries. 'Ihis is rare and. furtherrnore could be over- come by killing the flies and @!ggg larvae sirultaneously by freezing. In a brief d.iscussion, the c-uestion of the trarisr,rission capacity of resistant S. so}brense near the northern limit o-f its normal ra.11ge was raised.. I'{r Zerbo pointed. out the sb.rdies uere und.er way at Grechan on this subject and Dr Grrtnewal.d inforrnc'd Llre partici.p;rnbs r,f t.ht. rnccL.i.n,,,. that IRIU; Boua.lie would co-cperate in tl'iese stud.ies in Ivory Coast uncier a contractual a6reement with the ihogranme. ...f8 -B- IX. Flieht perfornance of Sirm.rliurn sDecies Dr Cooter surunarized. his worl: $rith S-. ornatum in ilngland. (alread.y presented to OCP in a written report). He d.escribed. the equipment and method and presented. some of the data. The charac'beristics of Ii. ornatutn can be presented. briefly as follows :- 1o Flight speed is constanb for aII ages of flies arrcl fcecling rcgimes 2o Flight distance and. duration increase greatly with sugar feeding (versus water only). .BIoocI or blood. plus sugar d.o not -ncrease distance and cluration as nuch as sugax alone. the longest flights ',rere about five hours (equivalent bo about { lcm). 1. Females fly better than nales 4. Distance and duration cluring the first fJ-ight are about the sane no matter hovr old the fly uhen it first flies 5. Distance and duration d.ecrease with repeatcd flit$ts. 'lhe priorities for wolk vli L,h li. da.rrrrrosunr wcrc Llrerr <Iill<:ur :l;c11. 'l'lrt: effects of the fo1lowin6 facLors on I'ligtrt perforrrr:.urcc nced Lo lre cv;rluarLcd :- I. AgC 2. Blood a:rd./or sugax feed.ing 1. Size 4" Parasite burd.en 5. C, tospecies Af ter sone cliscussion, rl- b tri,ls decided i;haL cornplcLe invosLi;abioos should. be carried. out r.rithoclarkttij. s sum at 3oti IraIIsr Ghana r+ith the possibility of comparative stuclies withrrpalet'S. squamosum at Isatsadu Fal Is, Ghana or So sirbanum at Yabo, Upper Voltao The comparative studies would only be done with flies of optimum age and" feedinli reg:inrc. X. Itearing :rnd. ColonizaLion of 5. d:r.mnosum s.l. Dr Raybould began the presentation with a review of the devclopment of the successful larval rearing system used. in the OCP. 'Ihis systen vras developed for simplicity and. portability and used. mostly for rearing material for tarconontc studies. ...f9 -9- iie continued to d.escribe progress in other aspects of col<.rniza- tion. fttensive tests on b1ood.-feed.ing have been conpleted. at A.i:osorabo in collaboration with I'ir Hans-Peter ilj'rtz, from the Institute of Tropical i'ledicine of the University of 'ftrbingen, Federal Republic of Gerrnany. Several alternatives to feecling on hrrmans a.re now avaj.lable. l.t is vc::y liiiely that mating strains of S. soubrensc rzill con tirrur: Lo lrc f'ountI i.rr southcrn'Jlogo. A few fertile egijs were also once collccbecl frc.rrrr egg batches d.e^oosited. by lab. energed. Lj. 4anmosum s.str. Ar_S.Lflanurn females which were not however seen to nate. Fenales reared. fron these eggs un_ fortunately did not feed.. Dr Simmons continued. r.rith a d.escription of the rearin6 slrstem used. at the Un-iversity of llassachusetts for s. d.ecorum. ,rhis colony nas maintained. for 18 generations. During the lB generations, recluction of adult size and fecundity due to crowd.ing and. under-nutrition of the larvae was clearly documented. Dr Simr'pns then d.escribed Lris successful effort to colonize S. soubrense sanctipauli startin6; from e6,1s provided b.y Dr Lia.ylrcrr16. A new larval rearing sys tem provid.r'-ng hi6;her water veloc.i tics had Lo tre builL, but bhe blood feeding ancl rn;r'tin5 Lcclurj_c1 .ut:s used 1'<lr lj. dcc oru.tll l'rere successful with S. d.annosum. The system of inducing ovitrlosition on corks floating in hrrbutent vater produced a higher pr,,portic.rn of ferbile eggs than the irunersion technique used. previously in 0CP. Although per- centaSe of fenales ovipositin8r fecund.ity, fertility and. larrral su-rvival were all good, the feurales t/ere rel'ractory to matin5.; ancl tlic insenLlnation rate was less than IO%. Unfortunately, the colony failed in the 6th generation when the males disappeared. This nay have been due to fixation of a sex-Iinked. lethal gene since the colony tras based only on the off-spring of thiree feuBIes. Dr Sirnmons also clescribcd l; i.urlics on s[)L,J:ruaLolllrrlre f.'or.r,r;rl,i<irr iurd sperm transfer in S. decoru:a. 'l'hc 61oal of these s Lrrdies is to develop a systen of artificial insenination. douever, it nay also be possible to find' out whether the flies mate nore than onc., by calculating the arnount of sperrn used for each egg batch. In this context, Dr Garms mentioned. he had seen a paxous f1y with a spermatophore. The roeeting was adjourrred. at I!hOO. +-10- ?.1 .'.uluct 19SI r)rGr.rnewaldconvened.themcetlngatOBhiS.;.np}ytotlla:r,:-"}ie paxticipantsfortheirco-operatio:ra:.r.defficj.entuork.The ;iee.br:eg was officially closed. at OBhJO bui; s::ralI r7oups were fcrned' to chscussspccificarroirg'red.iateplaristh:clui.toucthemorninS. frfter sun-set at lBhJ0 Dr Sinunons Cenonstrabed his tcchnique to iniucc. ;ravid. fe;rales to oviposit. 'l'hc u:aitr tri,1;crs seem to bc a movin5 vral,cr surfa.ce (air bubbles) in an acaari'd-r.r a.ncj a :;irmlabccl tvili;irt' To iii:-iate cviposition gfavid fernales feC thrcc da;ys ago on Dr:-lit'tionst af,*x' rvere plac&.cn corlcs drifting on the a:;itetccl wate: surface cfter roorn li'3hts wcre extini,lrished. and sun-set light tras placccL nt Lhe rlater line' ESgs were Iaid. imned.iately on the corks an4 at tlie r+a;er Iine on'the waII' ,Ihe VCIJ au.thorities artd all the pe-rticipani's wartt to express their gratitr.rd.e to i,tr Adjonou for his very kind a:rd friend.Iy hospitalj-ty, and' to con5patrrlate him for the very renaxkable rraterial crganization of the meetinS' ( l' !, .t 1, i,' l' rl ll i l' I I i i t, I ,l' i'. l,' 'i, I I 'l 'I I it,' :l .t.. ,i: I ! ;rl" il :j ..1 I ,,! I .l '.1 't :' AG ENDA Dtrbomological situation in the sectors cf i,:;Jira llara (Aa,onou) anci :iati',ingpu (Ou6araogo ). i)robleias arrd. succes:; of the treati.re:it cf rivers in the zone of exten- sion stuoies and, their effect on the rcinvasion in l{orthern 'logo and 3cnin (carms). I -Discussionsonresea.rchproposarsa:iclprojectsofDrchei:efrom September to November 198I and- of l)r G;^:::s in 1932 ' Susceptibilitytestirrgof}arrraeintreateclrir.ersintheextension zone particularly in t,ee lrono river (ilr:rtar:, ilercd.:-th). Cytotaxonornical situation (Mered'ith)' Discussions on the research project on isoenzyne iden of the vcctor complex artd' on enzJrne str:oies on lesistartt poprlation in i.jep bember/October 1981' tiflcati S. dar,ur cns oSUm ihol;ress on S. da':;rosum compl a:rd. S. souano sum ex, esPeciallY of(carms). the nrorphological d.iffcren Lia;ion of adults of tr'rothe various forr:s of Si . soultr Reinvasion a3d. fLisht perfor:nalce of .i. i::I jlotll:il conrplex specii:s' Su:r:unary of the utrLu"-on flighi perri* e on fl;'ers in ;he uliJ"a",,i;;;; id""."il-*a-ai"""ssion; on tle shrc-ies pra,ned' r'r:-th S. dactrosu4 complex species in Ako=onj:o' progress in establistr.in' a colony of E. c;rmosug in U.S'A' (Sit'rnons) and in Akosonbo (RaYbou1d). li I l' I 1a Dr B. Philippon Dr J. Gnrnewald. i,lr G. Zerbo Dr C. Gboho D,lc J.N. ilaybould, i,Ir J. Ouedraogo l.ir C. Adjonou Dr D. Kurtak Dn R. Garms \)-t'trs li. tiernef lJr S.8.0. I,iered-ith Dr R.A. Cheke \ -l tDr R. Cooter t nr K. Simmons tIS[ OF PAII.ECIPi.].tTS Chlef VCU, OCP Research Coordinatorr OCP Chief of Entornolo8ical Erraluation, VCUr OCP Assistant Chief of Lhtonological Evaluation for Eastextt'Zone, VCUr OCP Reeearch Officer, VCUr OCP Sector Chief, Natitin6ou, VCUr OCP Acting Sector Ctrief , Lan,ta iiara, VCU, OCP lbainlng Officer, VCLI, OCP (Recoraer) Consultants, thopenins titut, Hamburgt Federal ltepublic of Cermany Consultant, Liverpool, United. ICingd.om Consultantsr COPR, London, United. Kingd.on Conoultant, University of i'Iassachusetts, U.S.A.