Informal consultation on the development and evaluation of simulium larvicides 16-18 March 1982: VBC meeting room, ILO building

r E:j.{tr vBC/OCP/rCl 82.3/wP + J WORLD HEALTH ORGANIZATION ORGANTSATION MONDIALE DE LA SANTE lI DIVISION OF VECTOR BIOLOGY AI{D CONTROL ONCHOCERCIASIS CONTROL PROGRAMME IN THE VOLTA RIVER BASIN Informal Consultation on the DevelopmenE and Evaluation of Simulium larvicides 16-18 March L982 VBC Meeting Room, ILO Buildine A REVIEW OF THE LITERATI'RE CONCERNING TIIE FEEDING BEHAVIOUR AND GENERAL ECOLOGY OF SIMULII]M LARVAE, ND OF THE DEVELOPMENT, TESTING AND MONITORING oF THE USE OF LARVICIDES, WITH SPECIAL REFERENCE TO THE CONTROL OF SIIflILIID{ DAIO{oSIIM u. DR J. F. WALSH I,IIIO CONSIILTANT VBC Couunen'ts For the information of the Informal Consultation on the DevelopmenE and Evaluation of Simulium larvicides for onchocerciasis, Dr J.F. lJa1sh, I,JIIO Consultant, has been requested to ""ff""t "nd analyse all available publications and reports on the biology and control ofblackflies with special emphasis on S. damAosum s.1. vector of onchocerciasis in tr{est Africa. The Consultant has also been invited to provide his own independant appraisal of the situation as well as his recormnandations for further research. Ihe report of Dr J.F. Walsh is transmitted for the consideration of the "Infonnal Consul. tation on the Development and Evatuation of Simulium larvicides for onchocerciasis". t Tt. Division of Vector Biology and Control has reviewed the Consultantrs report and pre- sents the following comment.s as regards Ehe conclusions and reconunendations. I t t I a( Conclus ions and recommendations SECTION 2 1. VBC agrees with the recoumendation of the consultant regarding the need for additional information concerning the feeding process of blackfly larvae. VBC feels that this work should be carried out on the target species S. damnosum s.1. at all larval instars. From VBCfs point of view, it appears that some gaps remain in the com- plete understanding of these aspects, which are felt to be of utmost importane for the development of suitable larvicide formulations. 2. VBC ful1y agrees on the need for such a study. However, the grouP is requested Eo give guidance on Ehe ways and means to achieve this study. SECTION 3 3. VBC supports this recornrnendation. SECTION 4 4. Some of the physical properties of the EC formulations have been studied in depth (density, suspensibility, etc.) and constitute specifications for this type of formulation. However, iE has to be ,..ognirr"d that specifications established for Simulium control do not automatically guarantee their biological effectiveness Iherefore, iE. is considered that the role of additional parameters be inves tigated. In Ehis connection, it should be advisable to study in more detail the behaviour of EC formulations at very 1ow dosage in water, along igg Progres- sion downstream. This study should include research on physical transformation of the formulation in relation with physical-cheinical characEeristics of \"rater (turbidity, temperature, pH, resistivity, electrostatic load, etc.). , a 2 I AlEhough the spectrr-un of particule sizes ingested by s. damnosum s'1' larvae has been studied, additional research appears to be necessary so as to refine the specifications for particulate larvicides so that guidance can be provided to the manufacturers. 5. VBC supports the idea of conducting some physico-chemical studies with particular emphasis on colloidal larvicide formulations. 6 Studies reconnnended in point 4 should be extended to Part'iculate formutations. T.ThecourmentsmadebytheconsulEanEapPlytothefirstmicro. capsules tested, for which results have been published so far' Ilowever' it should be underlined that Progress has been made in the development or micro- capsules. It. has to be remembered that 2 types of microcapsules have been developed : i)Microcapsuteswhicharedigestedinthemid-guEofEhetarget. ii) llicrocapsules which progressively release the insectieide in the water as well as in the mid-gut ' The firsE type of capsules would be preferabte since Ehe release of acEive ingredieng occurs only inside the larvae' Ilowever, the development of the second type is more advanced. VBC feels that research in Ehis field should receive a high priority. In fact, good microcapsules could include active ingredient which might be very toxic for environnent, e.z. pyrethroids' In addition, it has been shown that. microcapsules are not ingested by fish' Therefore, rhis rype of formulation might improve the selectivity of the acEive ingredient. t "" .i ..: 8. 9. 10. r[BC supports entirely the consultantsr views on these points, wiEh regards to the screening of candidate larvicides. It should be entirely carried out on target species (e.g. S. damnosum s.1.) and its asso- ciated non-target organisms. Mini-gutters and float,ing-cages should be used for that purpose. Field triats should be conducted as recommended by the consul tant . 11. Although temephos appears to be the best insecticide for the time being, its use will depend on the susceptibility of vector species other than S. soubrense (e.g. S. damnosum, S. sirbanum, S . squamosum) and/or on the development of new insecticides which could possibly be used in rotation with temephos. Some reservation are put on the use of chlorphoxim not. only due to its environmental impact but also on the basis of its cross-resist,ance with temephos. 12. Methoxychlor could be considered subject to Ehe following : i) Improvement of its formulation as this insecticide has not given good results against S. damnosuu s.1. so far. ii) Susceptibility of S. damnosum s.1. to O.Cs. iii) Monitoring of its impact on the non-targeE fauna when used aE weekly intervals for a long period of time. 13. A number of carbamates have been offered by the industry and should be tested on S. damnosum s.1. larvae and if on the associated non-target fauna. ,,* 3 A( r I \. L4. See comments p. 7 promising for Eheir impact / A I SECTION 5 15. 16. L7.18. \IBC fully agrees with these points of section 5. However, it should be underlined that at the t,ime of writing his report, the consultant was not aware of the developmenE of new formulaEions Erore con- centrat,ed in active ingredient and applicable without pre-mixing with water' SECTION 5 19. In his report, the consultant did noE distinguish clearly between the juvenile hormone homologues and the chitin irihibitors' It is fully agreed that a low priority should be given to the first type of IGRs due to their narrow window of action. However' a ProPer evaluation of the second type of IGRs is highlY recournended. Research in the industry is at Present orientated Eowards development of these EYPes of Pesticide- SECTION 7 20. Recommendation of the consultant with regards to safe use of blackfly larvicides is fully justified' Alrhough it is unlikely that Ereated water would Present any hazard for riverine people and animals due to the very 1ow dosages used, WC supPorts the consultant's concerns with regard t,o the risk for the population resulting from unguarded stocks of larvicides and to the safe-handling, by Programme personnel, of concentrated formulations. In additionr hazards might come from accidental spraying of riverine people' Until now, an accuEe oral LDro for rats above Ehan or equal to 500 mg/kg was recoEmended for blackfly Iarvicides. However, from the human safeEy poinE of view, material with an oral LD5O to rats expressbd in terms of formulaEion higher than 1 000 rng/kg and dermal tr5O higher than 2 000 mg/kg 4 [:i, I should be accepted. In other words, chemicals of relatively high manrnalian toxicity may be used provided that their concentration in the formutation applied is adjusted accordingly. SECTION B 2L. It should be stressed that a susceptib il i ty test is not designed for the screeni of insecticides. The method has been developed only to compare susceptibility 1evels of different blackfly populations to the same insecticide. Therefore, the test should include : i) The assessment of the basic susceptibility of untreated popu- lations. ii) The comparison of the susceptibility of an unknor'm population to these basic data. This second part can be carried out by using dia- gnostic doses which require only a few larvae (maximum 100). On these basis, WC recomurends that the I'Iulouchet test" for determining the suscep tibilities of Simulium damnosum s.1. Larvae to insecticides be strictly adhered to as originally described and as approved by the Expert Committee on resistance. In addition, it has been found that sixth instars were sometimes diffi- cult to separate from seven instars which in themselves show variat.ions in susceptibility according to their degree of development. The "Mouchet testrr applies to all chemical insecticides except IGRs. It includes chlorinated hydrocarbons which may require higher insecticide concentrations (see i) above). VBC agrees with the consultant that a different method should be designed for B.t. H-14. According to informations recent.ly received in VBC, IRTO (Bouak6) is considering the development of such a suscePtibility Eest. Ivloreover, in view of their potential inEerest, VBC stresses Ehe need for developing a susceptibility tesE to IGRS (see 19. above). t 6SECTION 9 22 to 30. VBC agrees with the consultantts statements in section 9 on the necessity to continue the monitoring on Ehe invertebrate fauna. As regards fish, VBC takes note of the conclusion of the ecological Pannel reported in the 1981 OCPrs annual report (page 22, paragraph 96) whicl indicates "the absence of significant effects on the fishtt ) CORRIGENDU}I To Dr J.F. Walshrs Report : "A review of the literature concerning the feeding behaviour and general ecology of Simulium larvae, and of the development, testing and monitoring of the use of larvicides, with special reference to the control of Simulium damnosum". Part to be corrected Conclusion and recommendations . Item 1, line 4 : Instead of "changett, please read t'chargett. Item 1, line 7 "Rapid" has to be inserted before'buccess'.'. Item 5, line 2 : "Larger" has to be inserted liefore "parEiculate". Item 8 line 4 A conrna has to be inserted after "larvicide". Item 8 line 5 : A conrna has to be inserted after "activity". Item 16, line 2 InsEead of "noted", please read t'promotedtt. ,A review of the literature concerning the feeding behaviour and general ecology of Simuli-um larvae, and of the development, testing and monitoring of the use of larvicides, with special reference to the control of Simulium damnosum by J.F. I,IALSH, WHO CONSULTANT I 3CONTENTS 1. INTRODUCTION 2. SIMULIIIM LARVAL BIOLOGY 2.L Biological background 2.2 Stream energetics 2.3 Feeding behaviour 2.3.L The food of blackfly larvae 2.3.2 Feeding mechanisms of blackfly larvae 2.3.3 Partieulare feeding 2.3.4 Feeding rate 2.3.5 Factors affecting the feeding rate 2.3.6 Food assimilation 2.4 Non-feeding aspecEs of blackfly larval biology PHYSICO-CHEMICAL CONDITONS OF LARVAL HABITATS 3.1 Current velocity 3.2 Water chemistry 3.3 Water temperature 3.4 Turbidity CONVENTIONAL LARVICIDES 4.L Introduction 4.2 Emulsifiable concentrate formulations 4.2.L Specific gravity 4.2.2 Adsorption properties 4.2.3 Miscibility of formularion solvents 4.3 Colloidal formulations 4.4 Particulate formulations 4.4.L Introduction 4.4.2 Selectivity of particulate formularions 4 t 2CONTENTS (conr'd) 4.4.3 Limitations of temperate zone tests 4.4.4 West African trials of particulate formulations 4 .4.5 Wettable powders 4.4.6 Microencapoulated formulations 5 Testing procedures for potential blackfly larvicides 4.5.1 Laboratory methods 4.5.2 Field methods 4.5.2.L Trough and gutter tests 4.5.2.2 Stream trials 4.5.2.2.1 Effects of differences in formularion 4.5.2.2.2. Effects of stream conditions 4.5.2.2.3 Dosage rates and application methods 4.5.2.2.4 Differ:nces - the sensitivity of target species 4.5 .3 5. Bacillus thuringiensis serotype H-14 4 5.1 5,2 Introduct ion Laboratory studies 5.2.1 Mode of action 5.2.2 Effects of water quality 5.2.3 Effects of temperature 5.2.4 Specific and instar differences in sensitivity 5.2.5 Duration of exposure and concentration 5.2.6 Formulations 7 Need for a standardized bioassay test using Simulium larvae 5.3 Field studies 5.2 5.3.1 The operational situation 36 7 INTRODUCTION (contrd) 5.3.2 Cost implications 5.4 Key reference Insect growth regulators Degree of hazard involved in handling and applying larvicides and to people and animals drinking treated water The determination of 1arva1 suseeptibility 8.1 Introduction 8.2 The "Mouchet test" 8.2.I Choice of larvae for the "Mouchet test" 8.2.2 Limitations and modifications of the "Mouchet test" 8.3 Observations arising from susceptibility tests on African species 8.4 Effect of aeration during exposure period . larvicides on non-target fauna 8 9 The impact 9.1 General of Simulium cons iderat ions 9.2 Laboratory studies 9.3 Field trials in Africa, the "DDT-era" 9.4 Field trials in North America and Europe 9.5 The selection of temephos for the OCP 9.6 Monitoring the environmental effects of the OCP 9 . 6. I Introduct ion 9.6.2 Methods 9.6.3 Implementation 9.6.4 Results 9.6.5 The impact of alternatives to temephos currently in use with the OCP 4INTRODUCT]ON (cont'd) 9. 6.5. 1 Chlorphoxim 9.6.5.2 Bacillus thuringiensis serotype H.14 6.6 Key publications 6.7 Annex 9 9 10. 11. t2. Conc lus ions and re cornrnendat ions Acknowledgements Reference list I. Introduction Onchocerciasis is a disease believed to afflict 3O million people. In West Africa alone it has been estimated that 200,000 people are blind as a result of this disease and that in the absence of any kind of intervention a further 14,500 people would become blind each year. (Walsh in prep. ) In all areas where the parasite-host relationships have been worked Simulium has been found to act as the vector and theout a species of definitive host is man. There appears to be no significant alternative vertebrate host. The disease could be brought under control by breaking the man- parasite-fly relationship. In trrlest Africa, at least, one of the main aims of control is to a1low settlement of uninhabited but often fertile river valleys; accordingly, breaking the link by the geographical separation of man from the habitats of Ehe fly is not an acceptable solution. Unfortunately, to date there is no satisfactory chemotherapeutic method available for large-scale application in the field. Given the time seale for development of such agents we can assume that onchocerciasis control will have to rely on other means for at least the next B to 10 years. the vector. Since 1974 aWe are then left with control of scale prograflme has been mounted aga inst Simulium damnosum s,1 in West Africa. Although it is perhaps as yet too early for final conclusions this prograrrne has certainly been judged worthwhile by the affected counEries and there are plans to extend control over much wider areas ("senegambia"l and Nigeria for exarnple) . Providing the present the Sponsoring Agenciesprograrune is considered successful by and the Donor community, there is no reason to think that such "'t '"Senegambia Project. 0nchocerciasis control in Guinea, Guinea Bisson,MaIi Senegal and Sierra Leone. VBC 8L/2 - 1981. 2schemes cannot be fostered. However, there are eoncerns of the OCP (as at present constituted). These arise from about the future two technical sources. One is the problem of long range migration by flies which may have previously taken blood meals and become infective. At present about 20 per cent of the OCP area work by Sowah et a1. (pers. com.) is affected by this problem, but recent suggests that the population 1ikeIy to be subjected to unacceptably high transmission rates is confined t.o a much smaller area. The second concern stems from the fact that since its inception the OCP has been reliant on one larvicide, temephos (eU.t"R), indeed initially on one formulation from one supplier (presently two formulations from two suppliers are available). Until very recently the only operationally tested alternative was 6hlorphoxim. This insecticide has a much more toxic effect on the aquatic invertebrates found associated with S. damnosum s.1. (nejoux et al. 1981) and its use is deplored by many envirorunentalists. It is feared that a wide-scale and repeated use of chlorphoxim might cause drastic alterations in the composition and abundance of the aquatic fauna, including fish and other taxonomic groups (Ecological Group,19B1). The formulation in use in 1981 did not, in any case, prove to be very saEisfactory against S . damnosum s .1 . In March 1980 the S. soubrense/S. sanctipauli species pair of the S. damnosr:m complex developed resistance to Eemephos in Southern Ivory Coast (Cuillet et al. 198o c). These resistant populations have spread from the original centre of resistance at Tiassale on the lower Bandama into the adjacent river basins to LIest (Sassandra and East (Como6) despite the fact that this forest cytospecies pair is not considered to be such an impressive migrant as are those of the savanna (Walsh et al. 1981 b). 3Resistant S. soubrense have bred, unchallenged by insecticide, in the same breeding sites as S. damnosum/S. sirbanum without the transfer of the resistance moiety for several generations and there is no reason to believe that the two species pairs are not reproductively isolated. In October 198t cross-resistance to chlorphoxim \^Ias rePorted from the original outbreak area, on the lower Bandama and lower Nzi, and soon spread to the lower Como6. It resistant to other O.P. that thes'e populations are also but not to chlorinated hydrocarbons and is probable compounds, carbamates (Jamnback, per. com. ). This means that control of S.soubrense, possibly the main vector of onchocerciasis in the forest and transitional vegetation zones (Qui.116v6r6 et al. 1977, Philippon 1977) of Ivory Coast cannot be achieved unless Bacillus thuringiensis H.14 is available in large quantities and lives up to its promise in sma1l scale trials. Opinions differ as to the overall technical importance of this problem. It may be that (1) S. soubrense will prove capable of replaeing S. damnosum s.s. and S. sirbanum throughout their distribution area and will act as an efficient vector of hurnan onchocerciasis. (2) S. soubrense will populate only those areas in the savanna zone where it was occasionally found before resistance developed, and will act as an efficient vector there. (3) Resistant S. soubrense acting as vectors will be confined to areas where non-resistant populations had previously been important. So far resistant S. soubrense have only been found breeding in areas from which the cyto species had previously been recorded, though now in greater nurnbers. However, man-biting behaviour does appear to decline in the Guinea Savanna zorte. 4Thus, the current situation is somewhere between position 2 and 3. The question of vector capacity is being studied further by IRTO at present. The question also arises of resistance developing in the vastly more i*portant S. sirbanum and S. damnosum s.s. populations of the savanna zones. Given the exceptional migratory ability of these vectors this s,eems unlikely to happen (Comins T977, Georghiou & Taylor L977a and c) but should the area under control be greatly extended, as proposed in the Senegambia Report (WHO 1981), the risks will increase substantially. Owing to the exceptional flight ability of S. damnosum s.1. its sma1l size, and relatively low density, the habits of the adult are still imperfectly understood. In particular, little precise information is available about adult resting habits. Thus an attack on the exoSenous S. damnosum complex by means of a large scale adul t i ciding the totalcampaign 750, ooo interes t cannot provide an alternative to larviciding in two thirds were subjected to some measure firs t 1,lrr2 area at present being treated though it may be of considerable in special situations. Fortunately, the breeding sites of S. damnosum s.I. altho ugh many and widespread are not inntunerable. A recent study indicated that during the wet season there were aboua . 23,000 lqn of flowing rivers and medium-sized streams in the original OCP area of. 654,000 kn2. of these of larviciding during the years of conErol operations, the proportion or 3 years of activity (walsh 1981). Thisfalling to 50 per cent after 2 effectively meant that in the 1980 wet season 11,400 km of flowing water \ras treated on at least one occasion, 4r500 lstr being treated at the height of the dry season. Given no problems of resistance there is no reason why these distances cannot be further reduced in the next Ewo or three years. 22 5 LARVAL BIOLOGY 1 Biological background As Hynes (1970a) has stated the rheophile insect fauna is its the Simuliidae comprise a highly uniform family which.is probably present time (White L978). It is "perhaps the most striking feature of world-wide uniformity. " succes s fu1special ized, and relatively in a state of active evolution at the thus possible to carried out draw useful general conclusions from the extensive studies In additon on stream faunas of the temperate zone, especially in N. America, and with caution, apply them to the much less intensively studied areas where the vectors of onchocerciasis are found. An excellent general account of the biology of blackflies is provided by Crosskey (1973), while Le Berre (1966) deals specifically with S. damnosum s.1. 2.2 Stream energetics It is only in the last decade that the energetic relationships of sEream faunas have begun to be studied in detail. Even in 197O a review on the ecology of stream insects could legitimately be concluded by the statement that the role of allochthonous organic material in the nutrition of aquatic insects was almost investigated (Hynes 1970a). Recent studies by Hynes et aI. (f974), Lush & Hynes (1974), Paugy (1980),Goulding (1980) and reviews by Anderson & Sedell (1979), Cuurnins & Klug (1979) and Wallace & Merritt (1980) show that this is no longer the case. Cuunmins, in particular, G973, L974) has pioneered the concept of the functional feeding group. In his scheme Simuliurn larvae, along with net spinning caddis flies such as Cheumatopsyche (a ve ry important group in Inlest African riffles (Petr L97O), Tricorythid and Oligoneuriellid mayflies, and midge larvae, are classed as filtering collectors. Such animals feed on the fine particulate organie matter, FPOM, (less than lnrn to more than 50 microns) -6- and the ultra-fine particulate organic matter, IJPOM, (less than 50 microns to more than 0.5 microns) fractions of the detritus in transport. This non-living organic matter, in stream environments, probably does not occur without associated micro-organisms. By convention organic matter smaller than 0.5 microns in diameter is considered as dissolved (DOM). Much of this particulate organic matter is allochthonous (e.g. of terrestrial origin (Lush & Hynes 1974)) and the Simuliids thus play a vital role in converting this potential food source and making it availabl-e, in their own tissues, to a variety of predatory animals (Service & Elouard 1980) 2.3 Feeding behaviour It should be noted that Minshall (1978) considers that the importance of allochthonous organic material has been over emphasized. 2.3.L Food of blackf 1y larvae The food of blackfly larvae has been reported in a number of papers. It ranges from bacteria, through diatoms, to attached algae (epiphyton) and insects, including their own kind. In addition to detritus, diatoms feature prominently in the lists from many studies (Puri L925, Pacaud 1942, Grenier L949, Zahar 1951). Both Puri (L925) and Pacaud (1942) reared larvae of S. aureum on algal cultures in the laboratory. However, Pacaud (7942) was very conscious of the variety of material in larval guts and rightly concluded that S. ornatum and S. costatun were detritivores , while Grenier (1949) and Zahar (1951) pointed out the variations in proportions of diatoms to detritus in the gut contents of larvae according to locality and season. Although Fukushima et al. (1976) demonstrated a great variety of sizes of diatoms, from Cymbella tumida to Achnanthes lanceolata in larval guts they concluded that blackfly larvae were indiscriminate feeders. A conclusion also reached by Kurtak (L979) who found that while diatoms made up as much as 50 per cent of the gut content at times, proportions 7in the gut were generally similar to proportions of particulate matter in the water. Nearly sixty years ago, Petersen (1924) suggested that blackfly larvae might feed on particles as small as bacteria. This was confirmed when Fredeen (I960, L964) showed that three Nearctic species could be reared on a laboratory culture of Bacillus subtilis. He suggested that feeding on bacteria may well contribute greatly to the outbreaks of S. arcticum in the Saskatchewan river. Hynes (197ob) also considered that many filter feeders in running \,,rater probably fed to a great extent on bacteria. However, Baker & Bradnam (1976) concluded that at normal field densities the bacterial component in streams hras insufficient to support blackfly 1arva1 growth. It should be borne in mind that not all simuliid larvae feed by filtration, some have reduced mouthparts, and are obliged to feed by browsing, while other although capable of filter-feeding may occasionally browse, e.B. Burton ( L973) records Simuliurn hargreavesi larvae feeding on the fiLaments of the alga Oedegonir:m in Norther Ghana. It may be significant that these species inhabits unusually clear sEreams in the OCP area. Such streams tend to have more macrophytes and provide greater opportunities for attached algal growth. 2.3.2 Feedins mechanisms of blackf ly larvae Most. species of simuliid larvae feed by filtering particles from the surrounding passing current, a fact which has been appreciated for over 15O years (Verdat L823 - in l^lallace & Merritt 1980; Planchon 1844 - in Colbo & Wotton 1981; Cameron 1922; PeEersen L924; Puri 1925). The structure and function of the larval mouthparts were very well described by Grenier (1949). Recently, further elegant work has been carried orrt, some with the aid of Scanning Electron Microscopy (Chance 1970a; Craig 1974, L977; Davies 1974; Matsuo 1979). Undoubtedly the most important contribution -B- in this field is the work of Rose & Craig (1980). Although Kurrak (1973) had earlier suggested that a sticky substance may coat the cephalic fans, Rose & Craig have, for the first time, demonstrated that freshwater organisms produce a muco-secretion. Simulium larvae produce such a secretion which coats the cephalic fan rays from two large glands in the labrun. This provides a means whereby fine particles, 0.091-30 microns, are captured even though the smallest space between the abducted cephalic fan rays is about 35 microns in the species they studied. Rubenstein & Koehl (L977) have considered the mechanisms of filter feeding from a theoretical viewpoint. They point out that particles may be captured by direct interception, inertial impaction, gravitational deposition, diffusion of motile particle deposition, and electrostatic aEtraction. Further for any given particle size and velocity, only one or two of the filtration mechanisms listed will play a significant role in particle capture. (Given the type of habitat chosen by most simuliid larvae, and by all members of the S. damnosum complex, it is exEremely unlikely that gravitational deposition and diffusion of motile particle deposition play any role in their nutrition - Jzu). They also point out that particle capture and filter resistance, in turn, are lower for fibres randomly arranged in planes parallel to the direction of flow and lowest for fibres arranged parallel to the flow. Simulium larvae can be considered as stationary filters, as the cephalic fan rays are constanEly cleaned, and clogging is avoided. One means by which an animal can readily alter its filEer is to change the rate at which it cleans its filtering apparatus. I^Ihen Odagmia ornata was presented with the diatom Nitzschia actinastoides as food there was an increase in the length, or The paper byin the number, of pauses for cleaning (Schroder 1980a). Rubenstein & Koehl should be read in the original. I 9Following their lead, Ross & Craig (1980) have concluded that direct interception is the predominant mode of feeding in blackfly larvae, with inertial impaction playing a secondary role. They point out that some animals in marine environments, such as Brittle Stars, which preferentially filter positively charged particles may use electro-static attraction as a feeding mechanism. They also give some information on the physico-chemical nature of the muco-secretion and as simuliids readily capture negatively charged bacteria (Fredeen 1964) conclude that they may not use electro- static attraction. They postulate that other freshwater insects such as Hexagenia limbata (a burrowing mayfly) in their feeding mechanisms. P. Guillet (pers. com.) confirms larvae produce a muco-secretion which trichia. ParticulaLe material adheres will also prove to use muco-secretion that, as anticipated, S. damnosum s.1. coats the cephalic fan rays and micro- only to the internal surfaces of the rays than and to the microtrichia. These latter face into the current, rather acting to decrease the gap between the lame1lae, an observation which light of remarks on filter resistance by Rubenstein important field observations by Guillet should possible and f urther work stimulated. - JFI,I). colleagues (GuilIet & Escaffre a & b) clearly indicates that it seems certain that it sole criterion may be explicable in the & Koehl (1977). (These be published as soon as The work of Guillet and 1981 a & b, Guillet 1981 1980a, Guillet et a1 a1 though is not the part icle size is of great importance, determining ingestion. Formulations of Bacillus thuringiensis serotype H-14, containing similar mean particle sizes to one another are ingested at markedly different rates (Cui1let et al 1981a) while studies with micro-encapsulated formulations of conventional pesticides have not. demonstrated any systematic influences on effectiveness of differenE coat thickness, density, size, etc. 10 (Cui1let 198lb), indicating that at least another physico-chemical property is involved in determining rates of ingestion. (Rubenstein & Koehl L977 state for example, that non-spherical particles will be more readily ingested). Further, Guillet (1981b) found that the addition of as little as I per cent of a dispersal agent to a formulation can result in a ten-fold reduction in ingestion rate of Actellic M2O. 2.3.3 Particulate feeding Pacaud (L942) showed that the main particle s ize in the guts of S. aureum larvae was L5-25 microns. This type of study was not followed up until t{ilIiams et al (1961) measured the size of particulate matter in the guts of several species of Palaearctic blackfly larvae living together in Welsh mountain streams. They found that the sizes of ingested particles, which were roughly 11-15.5 microns x 6.5 - 8.O microns, were uniform across the four species and the different larval instars collected at, the same time and place. Freshwater filtering collectors capture and ingest essentially the entire size range of FPOM and UPOM from transport (suspension) (Cr-unmins & Klug I979). Simuliid larvae appear to select a substrattun where current flow is rapid and the boundary layer is thinnest, thus avoiding the problem of insufficient mixing of particulate material. They ingest, irrespective of quality, particles of food ranging in size from 0.091 microns (co1loidal) (ttlotton 7976) to 350 microns (Wa11ace & Merritt 1980), with the majority of studies reporting sizes less than 10O microns. In those studies in which the gut contents \^rere compared with the POM in the surrounding current there is little evidence that there is some selection of particle si.ze by different species and instars (Chance L97O; Kurtak L97B; Dinkel 1975; - 1r - Wenk & Dinkel 1981; Schrdder 1981a; There is also some evidence that Cnephia ornithophilia contained a preponderance of particles around the 5 micron diameter mark. A fact which may be correlated with it having unusually long and widely spaced microtrichia on the primary cephalic fan rays (pers. com. B. Thompson). Nevertheless feeding, even with respect to particle size, appeared basically non-selective, given that particles of matter of smaller diameter than the distance between the primary rays of the cephalic fan were less effectively captured (Kurtak 1978). Blackflies are considered by some authors to take larger particles than those ingested by other filter feeders. However, the impressive work of Wotton (1976, L977, I978a & b, L979) does not support this idea. He has shom that late instar larvae of Simulium nitiCifrons ingested latex microspheres (polystyrene) of 0.091 microns diameter (e.g. col1oidal) when placed in a suspension of less than O.02 ,per cent by volume of such microspheres in distilled water (Wotton 1976). In natural conditions Metacnephia tredecimatum was capable of filtering particles in proportion to their content in the surrounding water, even where over 80 per cent of the particles were less than 2 microns in diameter (Wotton 1978b). The fact that bacteria are ingested also provides evidence for the routine ingestion of very small particles (Fredeen L964; Burton et al . 1973; Malone & Nolan 1978). Burton et al. have shovm that aerobic bacteria are concentrated in the larva1 guts of S. damnosum s.1. from Ghana while Malone & Nolone (1978) p rovide smaller evidence for Prosimulium mixtum from Newfoundland and Snoddy & Chipley tI e United States. (I97L) from S. underhilli in the -12- 2.3.4 Rate of feeding The collector strategy may be t.o pass a large amount of comparatively 1ow quality food through the gut rapidly (Curmnins & Klug L979). There is evidence that gut passage times may be longer in shredders and scrappers than in filterers. Gut passage times of thosedetritus collectors so far studied have been remarkably short. For the Arnphipod HyalelLg gZtSSg it was measured as 30 minutes (Hargrave 1970), for tubificid worms, in optimun conditions, L5-2O minutes (Poddubnaya 196l in Ladle et aI. 1972) for the gasEropod Potamopyrgus jenkinsi about t hour (Heywood & Edwards L962) for the trichopteran Hydropsych occidentalis 1.54-2.63 hours (Mccullough et aI. 1979). Studies on gut passage/transit times in Simulium larvae have generally relied on feeding the test organisms with some sort of marker material such as powdered charcoal, french chalk, a fluorescent dye, sephadex beads or some other inert, non-nutrient material. The rate of feeding by Simulium larvae has been investigated by a large number of vorkers in recent years (Chance L97Oa; Kurtak L979; Ladle et al , L972; Mulla & Lacey 1976; Wotton 1978 a & b; Elouard & Elsen L977; Elsen & Hebrard 1979; Elsen 1980; Wotton 1978a & b; Schr8der Lg79, 1981b; P. Guillet pers. com; B. Thompson pers. com.). A11 workers are generally agreed that the gut passage time is very short ranging from about 20 minutes Eo 2 hours. Elsen (1980) and P. Guillet (pers. com.) report very fast gut filling times by very small larvae of the S. damnosum complex. Wotton (1978b) makes the point thaE his marker material (french chatk) formed only 10 per cent of the material of the marked band, and thus should not have distorted the feeding process in any way. This is important because Poddubnaya (1961, in Monakov L972) showed that in the tubificid worm Limnodrilus hoffmeisteri, which consumes 2 ,l 13 - to 8 times its weight 1n food per day, Ehe defaecation rate varied with differences in the nitrogen and carbon as with temperature. It seems possible times recorded for simuliids may be, at content of the sediments, as well that some of the very fast gut passage least in part, the result of fast ingestion to compensate for a low, or non-existent, nutrient content of the marker material. Nevertheless, there is no doubt that much of the ingested material is retained for only a very It is interesting to note that a dam outfall which feeds on very smal1 particles provides short sPecles one of period in the gut. M. tredecimatum the slowest gut transit times so far reported for blackfly larvae (Wotton 1978b). In lacustrine outfalls Carlsson eE al . (1977) reported that certain species not found elsewhere in the river system, crowded together. This was considered to be a result of the availability of very sma11 particles of food. In this situation one of the most nutritious foods consists of faeces of conspecifics. These faeces consist of material that of partly reduced large in Bacillus thuringiensis H-l4friable particles, the sort formulations was found to be most readily ingested and most toxic in gut passage fast times time is of recorded by instar larvae laboratory tests (cuiltet et a1 1981b). The considerable practical importance. The very Elsen & Hebrard (1980) who report that first take only (5 min, p€r. com.)2 minutes for a complete digestive transit and P. Guillet would seem to allow 1ittle time for digestion of ingested material. There is in fact evidence that some types of insecticide formulations such as some micro-encapsulated materials and B. thuringiensis H 14 formulations in which the toxic crystals are embedded in a cement may pass through the 1arva1 gut without the toxic material being released in time to exert its biological effect (Gui1let et a1. 1981a & b). ltI -14- 2.3.5 Factors affecting the feeding rate In North American studies the lack of a diel feeding rhythm has been reported for several Simuliuun species. They appear to ingest round the clock, Lhough Schroder (1980b) reported that O. ornata showed a slightly reduced ingestion rate in the dark. Given the consEant availability of the transport, and its generally low nutrient content, this is hardly surprising despite the fact that many other blackfly activities show distinct diel cycles. The following refer to studies in trIest Africa (oviposition - Walsh & Sowah unpublished; pupation - Elsen 7979: eclosion - Disney 1969; Thompson et al. L972; Elsen 1979: drift-Elouard & Leveque L977). Ingestion rates vary with temperature and \^rater velocity and availability and nutrient quality of particulate food material. These three factors have been well considered for O. ornata by Schrdder (19BOa) who points out that filtering activity increases with increased rate of flow and rising temperatures even in the absence of any particulate matter in the water. There is an inherent fanning rhythm (niggs 1981) but cephalic fans need urinimun current velocity to stay open (Harrod 1965). He concluded from this that the larvae had no control over the loading of the cephalic fans. However, as already mentioned in the same paper he points out that differences in food can give rise to variations in the amount of time spent cleaning the cephalic fans. Lacey & Mulla (f979b) provide similar information for S.vittatum. Ladle et al (L972) in contrast to the previous workers found no increase in feeding rates with increasing temperatures, at ^o and Zloc, though between 50 (the minimun checked) and 8oCI eas t betr^reen there was no feeding activity in S. ornatum (= O. ornata). Elsen & Hebrard (1979) working with the S. damnosum complex found fastest feeding rate -15- ') occured at 25oC and a particle density of 22O particles/sec/mm'. Savanna cytospecies appeared to be faster feeders than forest cytospecies. Several authors report young larvae feeding faster than older larvae (l'tulIa & Lacey 1976 for S. tescoEun). Elsen 1980 for S. damnosum complex, confirmed by p. Guillet in discussion B. Thompson (in litt.) working with Newfoundland species points out that at.'higher -tenperatures digestion rate increases more rapidly than does the ingestion rate. With smaller larvae absorbent surface to body volume ratios doubtless al1ow faster digestion. Another factor affecting the feeding rate is 1arval parasitism. Wotton (1978b) has shown that Ehe larvae of M. tredecimattrn which are parasitized by mermithid worms feed more slow1y than unparasit.ized larvae and Carlsson (1974) noted that S. damnosurn larvae parasitized with microsporidea reacEed more slowly to the passage of temephos larvicide, which he concluded was acting as a stomach poison. In a series of papers Elsen & Hebrard (L979) and Elsen (Lg7g, 198?a and b) have looked at a variety of factors influencing the feeding behaviour of the S. damnosum complex. A number of investigations such as the fine differences between the feeding behaviour of male and female larvae (Elsen et al. 1978) seem of academic rather than practical interest but the series as a whole should be consulted by anyone concerned with the chemical control of the S. damnosum complex. Chance (1970) reported that larvae may ingest more food than required which then passes through the gut unchanged. In addition larvae may starve with full guts in conditions where there is excessive inorganic matter, such as in streams carrying much glacial silt. This in itself reflects the automaticity of filter feeding. Carlsson (t967) gives an example of this in mentioning that while digging and blasting operat.ions are in progress the organic drift dovmstream frequently conEains large amounts of sharp- edged silt particles that are harmful to small animals feeding passively. 16 At Kainji Dam, N. Nigeria, breeding of S. damnosum s.1. ceased below a river channel into which large amounts of blasting dust settled (Llalsh unpublished observations). Gaugler & Molloy (f980) demonstrated experimentally that S. vittatum feeding behaviour could be inhibited when otherwise ingestible particles were offered at concentrations of 5O mg/l or more. This was considered to be a consequence of rapid gut fi1ling. Feeding inhibition appeared to occur more rapidly when the particles involved were of nutritive va1ue, but was nevertheless exhibited when the particles were charcoal, chalk and clay. 2.3.6 Food ass imilation Assimilation efficiency of simuliid larvae seems low even by comparison with other detritivores. Much particulate matter apparantly passes through the g unaffected by the processes of digestion. (Colbo & Wotton 1981) ' (This has considerable implications for control by means of micro-encapsulated larvicides and B. thuringiensis H-14 crystals presented in an inert cement, as well as for future attempts at biocontrol with pathogens. - JFW). Sma1l larvae have extremely short gut retention times, as little as 2 minutes is required for S. damnosum s.1. to fill gut according to Elsen (1980)' Sma1l larvae can undoubtedly digest relatively more rapidly than larger larvae, as already indicated, but it is difficult to see how there nutritional . requirements are met at such speeds. Further fundamental research is needed. It would also be illr-uninating to know precisely how the larvae of such species of S. noleri feed successfully and obtain adequate nutrients when located on the very 1ip of a lake outfall. For most species filtering efficiency declines at very high water velocities. Owing to the fundamental characteristics of filters and stream lines (Rubenstein & KoehI 1977) it is difficult Eo feel confidence in studies carried out in jars without through-f lowing current. I L7 Fundamental work on the capture and ingestion of particles would possibly lead to progress with micro-encapsulated materials and a bet.ter understanding of how to best present Such fundamental work could best be in the developed world. Any filter adequate. Schroder (1981a & b) and Kurtak the toxic crystals of B. thuringiensis H-14 carried out in a highly equipped Iaboratory feeding species of Simulium larvae should be (L979) give information on the time t,aken to digest certain types of diatoms buE little, or nothing, is known of the exact digestive processes in Simulium larvae though et a1. (1978) and Undeen (t979) have conrnented onLacey In S. the gut pH. itis ?verecundum S. vittatr.un and Cnephia orni thophil_a at the anterior of the mid gut declining to neutral at the posterior end of mid gut. This may have some effecE on ingested material. The possibility also arises that some nutrient is derived from gut symbiotic flcra and fauna It would be instructive to have fundamental studies on the digestive processes ir blackfly larvae (much more is known about digestion in the adult f1y). In particular, the peritrophic membrane, a secretion membrane, which acts as a food container, f,ay play a role in regulation of gut transit time of the food It is, however, most probable that the speed of ' the progress of food through the gut depends largely on the quantity of particulate matter suitable for ingestion which is available to the larvae from "upstream" (R. Le Berre in discussion). Also, how the very rapid enzyme digestion and absorption is achieved needs further consideration. It may be that first instar larvae, at leasE, can gain food by direct absortpion of organic maEerials in solution, though this seems unlikely as there is experimental evidence that larvae do not survive in a parLicle free (including free from colloidal particles) nutrient solution. It is knor^rn that Simulium ornatipes absorbed zinc coins from \- 18 - 0.1 mg/l solution, and that this involved uptake into the internal organs, in addition to 24 per cent of zinc which was bound to the cuticle probably by phenolic groups (Carter & Nicholas 1978). The review by tr'lallace and Merritt (1980) on "Filter feeding ecology of aquatic insects", is the key reference for this section. 2.4 Non-feeding aspects of Simulium larval biolo gv Work on the larva1 biology of S. damnosum s.1. populations has so far been very limited. Grenier & f'6raud (1960) and Elouard (1978), rhe latter author working with a mixture of S. damnosum s.s. and S. sirbanum, have demonstrated the existence of seven larval instars. Kurtak (1980b) working mainly with S. soubrense, demonstrated the difficulties of readily characterising the later instars. Within the family instar number varies from six to nine depending on species (Colbo & Wotton 1981). Ross & Merritt (1978) have suggested that for P. urixttrn/fuscum, the instar number can be either six or seven depending on the duration of larval development. The same authors have suggested that Cnephia dacotensis cannot be separated into instars by the use of morphological criteria. (For the importance of correctly designating instars see section on Susceptibility Tests). Larval development averages about 7-12 days in the OCP area, varying with water temperatures and doubtless availability of food. The Programmets seven-day aerial operations cycle is based on this and extending the cycle to 9-10 days has resulted in failures of control. Similar failures were noted in the FED campaign (n. Le Berre pers. com. ) and. aE Kainj i Dam (Walsh unpublished) when treatment cycles \^rere extended to 10 days. Recently in a northerly locality S6chan (1980) has shown that S. sirbanum is capable of completing its larval development in 4 days. The only experimental study on S. damnosum s.1. is that of Elsen (1979). In the laboratory he recorded complete larval development ranging from 7-9 days at 3OoC -L9- to 16-18 days at 24oC, with good nutrition. With poor nutrition development times could be greatly extended. Development of each of the first 4 insEars took 24 hours or less, the 5th instar 24-48 hours and the 6th and 7th instars 48-72 hours. Mokry (1976) in an interesting laboratory study of S. verecundr:m, @ species with 5 instars) reported the 3rd to 5th instars taking considerably less time to develop than the 1st,2nd and 6th instars. As is the case with other simuliids (t"tokry L975) the lst instars of S. damnosum s.1. on hatching drift attached to a silken thread spun by themselves, until they find a suitable substrate from which to feed. They remain in this position, even pupating there unless water levels alter markedly or larvicide is applied (Walstr, unpublished). A general impression is gained of animals inhabiting the surface layers of the water though the only study on dep th distribution of S. damnosum s.1. (Elsen 7977) found a majority of larvae attached below a depth of 40 cm, with settlement and pupation possible at a depth of 3m. E1sen did not report which instars, sampled. In a depth distribution study on S. penobscotensis Granett (L979) found a substantial number of later instars at depths up to lm (the maximum depth sampled) but indicated that the early instars were not sampled to any extent. In general current speed is the key factor and this will usually be greater at the surface. As Simuliun larvae are strongly positively phototactic (Scheren L962) and deeply fixed larvae which finds itself in unsuitable feeding conditions will tend to rise towards the surface (Colbo & Wotton 1981). (The finding of some larvae at greater depths than usually considered for Simulium larvae does not alter the need for larvicidal formulations Eo have a specific gravity of slightly less than one. It can be anticipaEed that deeply attached larvae would only occur in places of high trubulence where mixture of formulation I t20 \,rith river water would be adequate Eo spread larvicide throughout the whole volr:rne of water. Results obtained in the OCP so far indicate that despite using insecticide formulations with specific gravity below 1, all larvae are kiIled, ineluding those living at great depths. 3. PHYSICO-CHEMICAL CONDITONS OF LARVAL IIABITATS 3.1 Current velocity The basic need of Simulium larvae is for flowing water. Probably velocity of the current is the key factor in determining the micro-distribution of most species of larvae in a stream. phillipson (1956) showed that S. ornatum is not affected by Iow oxygen concentrations but responds to current speed. Later Harrod (1965) reported that this species could not keep its cephalic fans open at current speeds of less than 19 cm/sec, with most specimens opening their fans at current speeds of 22-28 cm/sec. Thus, although in food rich water larvae may occur in unusually low current speeds, for most filter feeding species there is a lower limit below which they will not thrive (Grenier 1949). Possibly species such as S. adersi, which have been taken in near stagnant waters (Gibbins t934), and those from a Rotenone treated stream (see below) fed by browsing. The fact that Simulium larvae of some species are only found in fast flowing water cannot be taken as proof that they are confined to such sites by abiotic factors. Cook & Moore (1969) relate an interesting case of a californian stream treated with Rotenone to eliminate coarse fish populations prior to restocking. I^Iithin Ewo weeks of this drastic action young Simuliurn larvae (species not identified) had virtually taken over all the available attachment space in moving !r'ater and a few were found in pools from which they had previously been absent. This suggests that their normal confinement to riffles was, at least in part, the result of predatory pressure. t-2t- Members of the S. damnosum complex have been recorded from 1arva1 sites where the general current velocity ranged from 0.4 m/sec to 2.4O m/sec (Grilnewald 198I). In East Africa the Kisiwani and Sanje forms of the S. damnosum complex were found at fairly high current velocities buL could be reared in the laboratory at substantially lower velociEies. In contrast the Kibwesi form, which was never found at velocities below 7.72 m/sec in the field, could not be reared in the laboratory at flow rates less than 1 .4om/sec (Grlinewal d L976) . 3.2 Water chemistry In studies on faunas of limited areas of N. America Lewis & Bennett (i975), Ross & Merritt (1978) and Merritt et a1. (1978) could find no correlation between water chemistry and species distribution. There was a broad correlation between the distribution of some cytotypes of the S.venustum/verecundum complex in New York State and physico-chemieal factors (Gordon & Cupp 1980). In all these studies temperature appeared to be the most important physical factor. Lewis & Bennett (1975) concluded that distri- bution was probably influenced mostly by oviposition behaviour. Thus S. verecundrrn oviposite on vege tation while S. venustrsn rarely does so (nothjels in Colbo & Porter 1979). Rhtun (1971) showed that different polararctic species had more or less marked differences as regards oviposition behaviour and selection of oviposition which could not in many cases be influenced by water chemistry. In Africa Grlinewald (in a series of papers especially L976) and Qui116v6r6 et al. (1976 & 1977) have attempted to relat.e the chemical characterisEics of streams to the distri- bution of the various members of the S. damnosum complex. Both amassed considerable data on 20 chemical components of flowing water and are agreed in considering pH of importance in determining cyto-species distribution- Grrlnewald & GrrJnewald (f 978) in laboratory rearing experiments showed 22- that the Kibwesi and Kisiwani forms were sensitive to pH and condutivity conditions and failed.in water near neutrality and conductivity near 7OS, conditions in which the Sanje form occured naturally. this elegant demonstration the author remains unconvinced S. damnosum complex, in the OCP area, have distributions determined by such and rivers fromfactors. The complex occurs in a wide variety of streams within a few lcn of the Atlantic ocean to the edge of the ancient Saharan dune systems. Individual cytospecies zones and to some extent in different are found in different vegetation sized rivers, and it seems likely that it is the general habitat and climatic conditions which determine the behaviour of the gravid female). There is no evidence to suggest that larvae of certain cytospecies cannot flourish in streams where others do so. Out of 187 sites listed by Vajime & Qui116v6r6 (1978) 24 had 3 or 4 cytospecies, and 5 cytospecies have been recorded from several sites (Fiasorgbor, Meredith, unpublished OCP reports), despite the fact that in most cases only very sma1I ntmbers of larvae have acEually been cytotaxonomically identified. Further the 6 main LIest African cytospecies can be raised in aerated tap water provided they are suitably fed. Given the wide variety of water conditions found in the OCP area and elsewhere in l,Iest Africa (useful additional data in Rai 1974, Petr, 1970, and Carlsson 1968) it is difficult to see how the development of an insecticide formulation can be tailored to match a particular set of conditions, though formulators should certainly have to hand the data provided in Grilnewald. (1976) and Qui116v6r6 et a1. (1976 & 1977) and Grijnewald's valuable review paper (f981). In two respects the streams of West Africa, which harbour the vectors of onchocerciasis are markedly different from Ehose in North America in (oesp ite that the members of the _23_ which so much of the detailed research on blackflies has been carried out and in which many larvicide studies have been made. These are water temperatures and the amount of suspended solids. 3.3 Water t eratures Qui116v6r6 et al . (1977) report a temperature range from 22oC to 33oC from rivers of Ehe Ivory Coast. Generally lower temperatures occur during the wet season. Grtlnewald (1976) gives 22oC to 31.1oC for cytospecies which occur in the OCP area. Other temperature data are given by Crisp 1956, carlsson (1968), Hynes (Lg75), trrlalsh (1973) while Grtinewald (1931) sruunarizes the situation for damnostnn complex siEes throughout Africa. A detailed analysis by ocran et a1. (1981) provides an extremely valuable guide to \,rater temperatures at 3O sites in the Western half of the OCP area. They record a range from mean monthly minima of 17.soC in December at three sites in December to mean monthly maxima of 32oC at two sites in May. They show conclusively that temperature is most closely related to the size of the rivers, with smaller rivers cooler. oShaded rivers are usualLy 3-4 cooler than open ones of similar size in the same latitudinal zones. Their data should be available to and borne in mind by all concerned to develop and test Simulium larvicides for use in West Africa. 3.4 Turbidity Most West African streams and rivers carry very considerable loads of suspended solids aE the beginning of the I^7et season. However, the only data available appears to be that of Secchi disc readings made by the aquatic monitoring teams (in Fairhurst et al . t979) and by Cheke (1981). An example for the Leraba River at the fronEier bridge is given which shows the regular seasonal pattern (from Fairhurst). For many rivers Secci disc -24- readings are consistently below 20 cm for considerable periods of tiure (Walstr 1978). Indeed the naming of some of the rivers in the OCP area e.g. White, Red and Black Volta reflects the varying amounts and types of suspended solids being transported. For the Niger River, in Nigeria the wet season flood characterized by a high silt load is called the "White Flood" whereas the lesser, but nevertheless substantial dry season flood which peaks in February-March and is the result of the previous years rain, in Guinea, percolat.ing through the Inundation Zone is designated the "Black I'lood". This flood carries 1ittle silt. It is known from the work of Fredeen and colleagues (Berck 1953; (1980) f'redeen et al 1953a & b; Fredeen et al L975; Fredeen L962) that both DDT and Methoxychlor adsort on to suspend solids (clay, silt and very fine sand) in transport. in river water. Following Fredeen et al. 1953 Noel-Buxton (1956) experimented with mixing a 1O per cent solution of DDT in kerosene with hard soap and clay for 3 hours at 85-95oC, at a ratio of I part of DDT, 1O parts of soap and 100 parts of clay. This material was applied at a rate of 0.03 mg/l for 23 minutes to the Kamba river, a relatively small, clear, tributary of the Black Volta in Ghana. IE was completely effective for 39 lcn. Using the same material in the Black VoIta, during a clear \,rater period larvae lsere completely controlled for a distance of at least 112 lan. Fredeen Q962) reported tests made in in 1954 in which DDT and Heptachlor variously formulatedAlbert.a, Canada with and without the addition of benEonite granules or celite (a diatomaceous earth) were applied to irrigation canals in which Simulium larvae bred. Best results were obtained with 2.5 per cent Heptachlor in celite. This was effective at O.1mg/l for 15 minutes formulaEions and emulsions of both DDT and Heptachlor were clearly most over a distance of 37 tcn. 0i1 effective in turbid r^/ater. Blackfly controllers in l{est Africa 25- (n. le Berre pers. com. and J.F. Walsh unpublished) have noted the greater effectiveness of both DDT and temephos in turbid \nlaters. Miles & Mount (1976) showed that temephos is rapidly adsorbed on to the particles of clay from dilute suspensions of the pesticide in water. As Miles (1978) points out the 1ow solubility of temephos (0.O29 mg/l in water, Bovnnan et al. 1958) and its high affinity for surfaces may explain its effectiveness against Simulir:m larvae. Guillet (1978) has clearly demonstrated the importance of suspended solids to the efficacy of temephos in gutter (cages flottantes) tests. fn turbid water (wet season) the effective dose in such tests (Procida AbaEe 200 EC) is 0.4ng/l/lO minutes wheareas in the dry season (relatively clear water) the effective dose rate is O.Amg/I/fO minutes. The effectiveness of OMS-1821 (permethrin ) was equally clearly shown, in the same paper, to be greater in Eurbid waters. Given the importance of this topic hard data are very sparse. It would be valuable if in all field trials of larvicides in West Africa, and elsewhere for that matter, a measurement of the amount of suspended solids be made by sedimentation and filtration techniques, and that visual observations e.g. Secchi disc readings and colour comparisons be noted (in discussion with Drs Grllnewald and Kurtak). In exceptional cases the nature and size of naturally occuring particles should be examined. The collecting of monthly assessments of quantity of suspended solids for a few key rivers in the OCP area should be considered to provide background information for the insecticide evaluators and formulators. -26- o @ o\o + f,- O LJoOtE .o u LL <t)rFz C)r c) U) Cf OO ut-a t!-F (n H c) .r{ dp tuL c) Fi C) bJ4 '-l t- -.1 (g .o aul{ G) F] =OgN= Fa O n- O\oO@OI O C) r\ JS iC iJ.,,-]> J -27- o @ O\o t F- -,O trJ C]O{-I C)all (n IFzOE z C) U\ C] (J(J LLJa n- tlF a S{ o .-{ ts C) o cc c pa o {J o 14 OHN= F o ON oot aO\oO@O? o : 35IC IJ3]S , a I I I I I I -28- I ,o o o\o +f- C)t!ootI Ou t.l- @rFzoI C)(n O C) c) trJa @ lrJF U) }-{ 0) .?1 tr () os CS l-{ CU o l.{ o hJ c (rJg cU =ogN= F , O o$' a o\oO@OIO: ():-)/\J f,sic t.).,ti: 29 F0 BRIDGf , RjD VOL?A i?IV:it fi-,,o tj.L-"/.4t-5r1 (tt-t ) IRANSPARENCY oF WATER SECCHI DISC REAOINGS II?6*ri t?5 '76 .77 tr .-5.2 25 2L 23 z2 2l 20 l9 t7 t6 r5 r3 r2 ito tr .x ".ttrt I I I I I I I I I I I I I I I I x x trx\\ '\ .'\ t a I f \ \/ x \x \ \X- '9 I 6 ;) July August Scprember Octobcr Novcmber DeccmberJunr 44 30 CONVENTIONAL LARVICIDES 1 Introduction The overriding importance of formulation for blackfly larvicides is very clearly demonstrated by the fact that several temephos E.C. formulations vrere rnact rve in field tests against S. damnosum while one r^ras outs tanding (l^rHO 1973) and has been the mainstay of the OCp to date. In temperate zones laboratory tests (see Jamnback & Means 1968 for references) emulsions proved generally inferior to other formulations as blackfly larvicides. This is probably owing to the fact that in laboratory or small scale "gutter" tests larvae do not feed normally. Gjullin et aI. (1949) in pioneer work recorded that a much higher dosage of larvicides was required in troughs than in streams to obtain equivalent mortality. This is well shown by Guillet (1978) and Guillet & Escaffre (1979c) who solved the discrePancy by the judicious use of suspended solids and for the first time provided a sma1l scale experimental system which gave results comparable to those obtained in large-sca1e river trials. Owing to the nature, and limitations, of laboratory studies a number of workers in the temperate zone favoured the use of suspensions or wettable powders (effectivety particulate formulations) on the grounds that these are likely to be ingested only by filter feeders and will thus be selective (e.g. Travis & Wilton 1965; Travis & Schuchman 1968). Travis er al. (1967) and Travis & Schuchman (1968) reported that a methoxychlor suspension worked better than emulsions and wetEable powders per second, while temephos suspensions were also more effective than temephos emulsions. Although oil solutions are favoured for aerial application by some workers (Jamnback & Means 1968; Jamnback & r'rempong-Boadu 1966) such formulations require highly specific materials and are 1ike1y to be extremely costly. In fact, emulsifiable concentrates have been much the most widely _31 _ used formulations in the field. They have several practical advantages, in that by dilution with water they can be applied for a wide range of agricultural and other uses and are generally much cheaper to produce than other types of formulations. 4.2 Emulsifiable concentrate formulations The vast bulk of all blackfly larviciding has been carried out using emulsion formulations of temephos, DDT and methoxychlor (Brown 1962; Chance 1970b; Jamnback 1973; WHO 1973). Such formulations have proved to have excellent carry in the large turbid rivers of Canada and West Africa, DDT in the Saskatchewan River (Fredeen et al. 1953b) and the Niger River (Walsh 1970a), methoxychlor in the Athabasca river (Haufe 1980). In West Africa, most rivers are fairly turbid, especially in the rainy season (see section ii) temephos at the low dosage of o.o5 rrrg/l/10 mins. has a carry of over 40 km in medium sized rivers, and has pr.-rved outstanding throughout the OcP area (Le Berre er al. L978; walsh er al. 1981). 4.2.L Specif ic gravity The best results have been obtained with E.C. formulations with a density inferior to that of water, in order to ensure a satisfactory dispersal and carry, with most of the insecticide staying in the upper layers . of the \,rater in non-turbulent conditions. Although blackfly larvae may occur in deep water (Elsen 1977) this probably only happens in conditions of relatively fast flow where the insecticide is adequately mixed with the whole volume of \,rater. Certainly formulations of slightly lower density than water have proved the best in field trials. Surprisingly, Qu6lennec 0962) appears to be the only experimenter to have compared two similar formulations, -32- except as regards their specific gravity, under similar field conditions. He compared 30 per cent DDT emulsions of S.G. 0.968 and 0.995 at 1 mg/1/30 mins in a small stream. The less dense formulation was effective to 28 km while the latter carried only 19 lcn. Ovazza & Valade (1963), using the 0.958 spec. grav. formulation at O.1 mg/l/30 min on a large r^7et season river, obtained complete control over a distance of at least 42 kfi. The temephos formulation used so successfully in the OCP has a density not Sreater than 0.980 g/ml. Thus the requirement laid down in WHO (I979a) concerning density of formulations for use as blackfly larvicides seems fu1ly justified, as do the other specifications. 4 .2.2 Adsorp tion properties Another property of larvicides, and in particular emulsions, which has a major bearing on their effectiveness seems to be that of adsorption on to suspended solids in river l^Iater. It may also explain much of their selectiveness. Fredeen et al (1953a) and Berck (1953) proved that DDT adsorbed on to silt and later Fredeen et a1. (f975) provided evidence that methoxychlor acted in a similar way. Miles & Mount (1976) have shovm, in the laboratory, that temephos has a great affinity for clay particles s uspended dealt with in water. (The question of turbidity in hlest African rivers is in section 3.4 of this report). Although the chemisEry of silt is generally 1ittle understood (Golterman 1975) some attempt should be made to further study the question of adsorption of larvicides on the naturally occuring particulate maEter in West African rivers, and the phenomenon in general. The physico-chemical relationships beCween particles and insecticide is clearly of greaE importance. Freeden et al. (1975) demonstrated that suspended solids from water samples taken 15 to 30 minutes after passage of an insecticide wave contained 40 to 47 per cent of the methoxychlor emulsion from the sample. 33 Ilerna et al. (L972) showed that the majority of methoxychlor in a sample of lake water containing plankton was in the particulate fraction, while Charnetski et al. (1980) showed that at 1.9 km below a dosing point on the Athabasca river, Canada, methoxychlor was sti1l in the water phase in its emulsion. By 16.9 krn below the dosing point, and at subsequent sampling points, the methoxychlor was mainly associated with the suspended solids in transport. Bailey & White (1964) reviewed the question on adsorption and desorption of organic pesticides by soil col1oids. Wang et al. (L972) found that Rhodamine B greatly enhanced the adsorption of parathion onto clay particles in water. I,rlallace et al. (1973a) used Rhodamine B as a marker with methoxychlor oil emulsion and found that the dye could provide valuable information of dispersal and earry of insecticide. They did not, however, report on the absorption of insecticide on to particles, contrary to the implication in Wallace and Hynes (1981). The question of adsorption enhancing properties of certain materials should be considered by larvicide formulators . Some requirement for a standard of adsorption should be included in the specifications for emulsion concentrates for Simulium conrrol (WHO 1979a). 4.2.3 Miscibility of formulation solvents Kurtak (1981) has raised the question of the relatively poor carry of chlorphoxim formulation used in 1981 ty the OCp. In discussion with Dr F. Peterson of Uniroyal Chemicals, Bethamy, Connecticut, iE was concluded that the two solvents used in the Bayer formulation, cyclohexanol and isophorone, are both miscible in water at the concentrations used in the OCP. Thus, a good emulsion is not achieved, as can readily be seen from the transparency of chlorphoxium/water mixtures. Non-miscible solvents such as xylene or toluene might result in a bett.er emulsion anri hence better carry. 34 A satisfactory emulsion is basically a function of the hydrophilic lipophilic balance (uln) of the non-ionic surfacta.nts. An HLB of 1o means a molecule with about equal hydrophilic and lipophilic ends' To obtain a good emulsion the HLB rating should be between 8 and 18. Whether this would prove to be the case as regards blackfly larvicides should be tes ted. 4.3 Colloidal formulations The possibility that co1loidal formulations might prove satisfactory blackfly larvicides has been strengthened by the increasing knowledge of the feeding efficiency of Simulium larvae. It is now known that these may feed on relatively small particles (Elsen, 1979b1 Wotton, 1978b; Dinkel , Lg75; Wenk & Dinkel, 1981). Wotton (1976) showed, exPerimentally, that blackfly larvae can feed efficiently on true colloids (particle size 0.091 microns) as can filter feeding mosquito larvae (Dadd,1975). In the field (Qu61ennec, Lg76) tested Didicol 6R reputed to be a stable, almost col1oidal, suspension of solid pesticide (602 DDT). The particles have a mean equivalent diametre (Stokes) of one micron, and thus actually faI1 into the UPOM (ultra fine particulate matter) range (Cummins and Klug, L979). Although this forrrulation, which aPpears to be the finest particulate forrnulation hitherto tested, hlas not as effective as a DDT emulsion (in fact at a dose of 60mg/1/6Onrm it did not eliminate all larvae from a breeding site) it was capable of crossing very quiet streLches of water. In fact the effectiveness of this formulation tended to increase with distance from the dosing point suggesting that a long period of passage, during which the larvae fed, was necessary for a toxic dose ro be imbibed. True colloidal materials might be worthy of further s tudy. -35- 4.4 Particulate formulations 4.4.1 Introduction Following Ehe appreciation, firsE by Fredeen and his colleagues (Fredeen et a1., 1953a and b; Berch, 1953) that DDT adsorbed on to suspended solids in the SaskaEchewan river, many workers (Noel-Buxton, L956; Fredeen, 1962; Fredeen et a1., L975; Guillet, L978; Guillet & Escaffre, L979c) tested emulsifiable concentrates in conjunction with suspended solids. (See also secrion on rurbidity 3.L.2.ii and emulsions above.) That turbidity greatly influences the effectiveness of larvicides, especially their carry, had been noted by Le Berre, Qu6lennec and Guillet pers. com.; Philippon et al., 1973; Walsh unpublished; in West Africa. 4.4.2 Selectivity of particulate formulations The potentially selective effect of larvicide presented in such a way was noted by Fredeen et aI.(1953b) and arising from this Hynes (1950b) made a plea for the use of particulate formulations to reduce damage to the non-filter feeding components of stream faunas. Following this suggestion of Hynes (1960b) Williams et al. (i961) determined the particle size of food ingested by the Simulium larvae of a Welsh mountain stream. Later Kershaw et a1. (1968) treated several Welsh streams with OOT in a water dispersable powder form, with particle size range of from 4-15 microns, similar to thaE ingested by Simulium larvae (Williams et al., 1961). Dosage rates varied from 0.04 nglL/30 min. to 0.4 ng/L1120 min. aE very low stream discharge rates. The minimum -36- dose was not completely effective against simuliid larvae but a dose of 0.2 ng/l/30 min. eliminated all larvae for 140 m below the dosing point. Non-target invertebrates were unaffected by this. At 0.4 nglL/120 min. Simulium larvae disappeared for 1 krn but this dosage also removed Baetis naiads and caused Ganrnarus to move downsEream. Nelson "r,u ,"".1il-, carried out experiments in canada using particulate formulations of methoxychlor and temephos. They used hlater suspensions of particulate larvicides, the methoxychlor having particles mostly in the size range B to 15 microns with L27" over 40 microns. The specific gravity of Ehe formulation was about 1.5. Of the wettable temephos (eUateR) powder 637" by weight was in tne 2.5 to 20 microns particle size range, with 202 less than 2.5 microns and L7Z greater than 20 microns. Dosage rat,es r.rere usually 0.L ng/Lll5 min. They compared their results with those obtained by Wallace et al. (1973) and lrlallace & Hynes (1975) who used liquid formulations in similar circumstances. They showed clearly that in such conditions the particulate larvicides were less harmful to some of the important stream insects than were the corresponding liquid formulations. -37- However, though no information is given regarding stream discharge the maximum distance of control referred to in the summary of results is 960 m. Onr trial (out of 5) with methoxychlor at O.L ngll-/lS min. was irteffecEive against simuliid larvae, whereas in the following year, the same stretch of stream was successfully treated with 0.16 rrrg/ll 15 min' In another stream temephos was effective at 175 m below Ehe dosing point (dosage rare O.l ngllltO min. but not at 450 m. (There were 5 EemePhos trials.) Nelson and WesE (1978) also mention that in a preliminary experiment an aerial application of particulate at 1.5 Lb/a.i./flieht mile did not provide saEisfactory control. Frost (1980) also carried out trials with particulate temephos against Newfoundland blackfly larvae. In May, with temperaEures of 2.5-6.5oC, 0.1 and 0.3 nel:-.l20 min. doses had little or no effect on the targe QO-24oC) t Prosimulium fuscum and the non-target fauna. In August treatment of 0.3 *e/tl+S min and 0.1 *e/tlZO min.. were equally effective in killing S. tuberosum and S. venusturn larvae while leaving much of the non-target fauna intact. There is no information about the carry of these treatments. -38- 4.4.3 Limitations of temperate zone tests These interesting results, viewed against would have been the fact that the unsatisfactory to that the use of the controller of onchocerciasis vectors. particulates poses the Most. test.s in temperat.e areas have been carried out in It is clear than those habitually by those populated by as regards Ehe non-target fauna, must be degree of Simulium tarval control obtained populated by the S problem of carry streams which are damnosum complex some other onchocerciasis vectors). As formulationsparticulate much smaller (though Fredeen improved canals, not (L952) pointed out "although the use of the larvicidal effect in clear water in the same resutts should not be expected certain irrrgatron in all clear-water streams. Unlike irrigation canals these generally show ponding extent. A pond, however small, would not only dilute would also a1low some settling out of the particles". 4.4.4 West African trials of particulate formulations to a greater or lesser the larvicide but In the search for larvicides for the OCP a number of Water Dispersible Powder formulations, in addition to Didicol 6 already referred to, have been tested, as have microencapsulated formulations, of the "slow release" and "digestible" capsule Eype. Most recently a variety of different formulations of Bacillus thuringiensis serotype H-14, which are all classed as particulates, have been assayed and in some cases tested in the fie1d. -39- 4.4.5 Wettable powder formulation The earlier phase of larvicide testing included river trials of OMS 708 (Mobam) r^rater dispersable powder which was ineffective, probably because the insecticide rapidly settled out (Qu6lennec, L971-). Dimethrin (OMS 187) as a W.D.P. carried badly, perhaps due to sedimentation (Qu6lennec, 1976a). Sevin (OMS 29) W.D.P. had some merit, but to carry through sluggish water it has to be used at fairly high concentrations (over L nel1l3} min. ) Thus it is unlikely to be cost effective (Qu6lennec et al., L967; Le Berre et al., L976a). Permethrin (Ort fgZf) 257. wettable powder proved to be very effective against S. damnosum larvae in the "cages flottantes" tests, especially when in the presence of suspended solids. It was in facE ten times more toxic than wettable powder formulations of temephos and chlorphoxim (Gui11et et a1., L979a). Precisely why the presence of suspended solids should have this effect with a larvicide already in a particulate form is not clear. It may be that the rate of larval ingestion was stimulated by the concentration of particulate matter (see section on larval feeding behaviour 3.1.1) and that a toxic dose was ingested. In the presence solely of Eoxic particulate matter feeding behaviour may have ceased, owing to paralysis, or some other inhibitory factor, with the larvae subsequently recovering (J.F.W.). -40- 4.4.6 Micro-encapsulatqd formulatigns A wide range of tests with micro-encapsulated materials have been carried out by P. Guillet and his colleagues in recent years (see especially Guillet et al., 1979a; 1981a). Guillet (1981b) sununarised the work on digestible microcapsules (Actel1ic I'{20), He concluded that encapsulation has not brought the hoped for advantages but rather had resulted in technical difficulties. Standardization of production appears to have been a difficulty. Large batches of materiat intended for river tests almost invariably were degraded or had different characteristics to the smaller quantities provided for laboratory tests. It must be concluded that the elegant idea of micro-encapsulation is extremely unlikely to be achieved for blackfly larvicides in the quantities required for West African vector conErol prograrunes. It also seems likely that any such formulation would be prohibitively expens ive. As regards primiphos methyl (Actellic) slight variarions in form of the capsu radically effected the ingestion by S. damnosum larvae. This would appear to be a physico-chemical question and not one of particle size. It would be valuable for biophysicisEs and biochemists to make use of these materials in their studies of larval ingestion (Gui1let, 198lb). -4L- The high speed of ingestion and ingestion of food materials by S. damnosum s.1. larvae also poses a problem as regards micro-encapsulated formulations. P. Guillet et al. 1981a) found that some formulations of Actellic M20 passed through the larvaI gut so rapidly that the toxic material was not released. This problem has also been encountered in a formulation where B. thuringiensis H-I4 Eoxic crystals where imbedded in an inert cement (see section on B. thuringiensis below). Given this high speed of ingestion, and consequent rapid guE transit timer atrY particulate formulation in which the toxin is not inunediately available for absorption across the gut wall is likely to be inefficient. This makes successful encapsulation a very sophisticated matter. -42- 4.5 Testing procedures for potential blackfl y larvicides Larvicide testing procedures against Simulidae have been evolved over the last 35 years. Effectively they can be divided into jar tests, trough tests (in the laboratory and besides or in the stream) and fu11 scale river tests. Gjullin et al. (1949, 1950) were out,standing pioneers in developing testing methods, as were Travis and colleagues (a11 especially Travis 1956 and 1968; Travis & Guttman L966 - who tested B.t.; Travis & Wilton L9653 Travis & Schuehman 1968; Travis er al 1967; Wilton & Travis 1965) and Jamnback and his colleagues (especially Jamnback 1962, 1964, L969; Jamnback & Frempong-Boadu 7966; Jamnback & Means 1968; Frempong-Boadu L966). In Africa, Qu6lennec carried out a long series of river trials between 196O and I97O . (See also WHO (1973) for literature on the choice of temephos for the OCP). 4.5.1 Laboratory methods A number of workers used jars to make bioassays of blackfly larvicides (Jamnback L962) but the use of troughs which could give some semblance of the field situation rrere generally preferred. Jamnback (1964) changed to trough testing after finding that fenthion rated better that DDT in jar tests but was far less effective in streams (Travis 1966). Travis & Wilton 0965) also considered larvae did not react favourably in the unnatural environment of jars and the same authors (1968) listed some of the probl€msr of using simulated stream testing. (See also comments in section 9.2 on non-target organisms ) . Using troughs in laboratories, or at the stream side, enabled workers to closely control the various parameters which could be expected to influence the outcome of tests (see the Travis and Jamnback paper cited above) . Nevertheless, tests using troughs did not suggest that emulsions would be as effective as oil solutions or wettable powders (Jamnback & Frempong-Boadu 19661, Jamnback & Means 1968; Travis et a1 L967), nor that temephos would prove to be an excellent larvicide. It was the last in a ranking list of sixteen promising a 43 materials tested in acetone sol,ution (Jamnback & Frempong-Boadu 1966). 4.5.2 Field methods The advantages of natural habitat tests are obvious when the test organisms have such specific habitat requirements as have Simulium larvae. Also, for onchocerciasis vector conErol the distance downsLream of the application point to which a larvicide is fu1ly effective ("the carryrr) is fundamental factor in determining its usefulness. However, the contraints a on the experimenter are severe and no two tests In addition large quantities of larvicide rnay be test when one is concerned with the control of a rivers such as S. damnosum s.1. are strictly comparable. needed to conduct a single species breeding in large 4,5.2.1 Trough or gutter tests To solve these problems, Fredeen (1969) devised a system for carrying out a series of snall-scale tests in a very large river. However, his technique which could be applied to an innnensely wide but shallow river (the St. Lawrence, Canada) is not suitable for the deep riversof West Africa. Guillet and Escaffre experimented with gutters placed in the river and eventually devised a minigutter test" The apparatus is readily portable and allows eight replicate tests to be carried out at the same time (Escaffre pers. com.). By using the larvicide in conjunction with mud, which presrurably stimulates normal feeding behaviour, results very similar to those in ful1 scale river tests are obtained (Gui11et L978; Guillet & Escaffre L979c). This is a significant piece of progress. 4.5.2.2 Stream trials 5.2.2.1 Effects of differences in formulation In l.rlest Africa a long series of field trials \^rere carried out during the 1960's and 1970's. These indicated that emulsifiable concentrates' with -44- density below l.O were Ehe formulations most likely to perform well over long distances. A11 particulate materials so far tested have tended to have poor carry, except those produced by absorbing on the naturally occuring suspended solids (Noel Buxton 1956). Ihose particulates with the smallest size of particles, such as Didicol 6 (Qu6lennec 1976b) and Sandoz Tecknar 4O2 (Cui1let et al. L979; Lacey 1980) are the ones with the most promising long distance effect in West African conditions. Most of the emulsifiable concentrates which proved ineffective in the field did not in fact form satisfactory stable emulsions (Escaffre et al. L976). This appears to be a problem with chlorphoxim (Kurtak 1981) and with the methoxychlor formulations hitherto tesEed (Le Berre et al. 1976; Qui116rEr6 et aI. 1976). Jamnback et a1 (1970) had a fairly high level of success, when spraying swaths of methoxychlor into the White Volta River in Ghana. Though in fact, using the same techniques and type of formulation of DDT it did not produce markedly better results. However, some E.C. formulations, which were ineffective in the field, did produce satisfactory emulsions. Thus, Escaffre et aL O976) tested three formulations of temephos from American Cyanamid against the Procida,' , standard in both the laboratory and the field. One of the Cyanamid formulations (Lot 72) did not emulsify but lots 73 and 74 appeared very similar. Only lot 73 was effective in the field, and comparable with the Procida standard. Miles & Cooper (1974) studied the physical properties of these four temephos formulations. The three Qyanamid formulations generally performed in a similar manner, and were not very different frol, the Procida standard except that the standard was considerably less vo1atile. Again, the discrepancy between lots 13 and 74 in the field was not expl icable. 45 It is possible that this lack of success might be attributable to a lack of adsorptive action resulting from the particular choice of emulsifier. The work of Wang et al. (L972) suggests that it might be worthwhile to try to provide an adsorption enhancing material in larvicide formulat ions . 4.5.2.2.2 Effects of stream conditions In West Africa chemical conditions as such do not appear to have a major influence on the effectiveness of larvicides. The two matters of overriding importance are stream discharge and turbidity. In general with increased discharge most larvicides travel farther, and are much more effective. It is standard practise in the OCP to use temephos at O.lmg/l/lO min in sma1l streams but O.Olng/tllQrun. on larger, especially turbid rivers (Walsh et al. 1981). The great importance of turbidity has already been covered (sections 3.4 and 4.2.2). Temperature also influences all chemical reactions and in this regard it is necessary to bear in mind Ehe marked differences between stream temperatures in North America and Africa. Temephos shows a positive correlation between temperature and effectiveness and has not always been effective when used at 1ow temperatures. Back et al. (L979) working in Quebec found it effective when applied at 0.3 mg/1/20 min at temperatures above l5oc but not at 8o - 12oC. In contrast, chlorinated hydrocarbons tend to have an inverse temperature - effectiveness relationship and this may explain, to some extent, vrhy methoxychor has been the preferred larvicide in North America but hitherto unsatisfactory in West Africa. In general who are carrying to give details the hydrological data provided by workers from all regions out stream tests are meagre. More concern should be shown of stream width, flow rate, discharge rate, turbidity temperatures etc., as well as very precise information on the 46 "effective carry" of the larvicide. 4.5.2.2.3 Dosage rates and application methods Hoeking et al. (1949) proposed that for blackfly larvicides concentration and time = constant mortality. Many subsequent \.rorkers (for example Travis et al. L967) have proved that this is not correct. This is because blackfly larvicides, in the fie1d, have been effective as stomach poisons, and not as contact poisons. As a result there is usually a minimum exposure time below which the larvicide, even at high dosages, fails to kill blackfly larvae. Those insecticides which are effective at high concentrations for very short periods have tended to kil1 a broad spectrun of non-target invertebrates and have accordingly been rejected for use in large scale operations. Application techniques have been designed to solve this problem of the need for a contact time of several minutes, despite the larvae inhabiting a constantly changing mediun. Where treatments are applied very close to the larvae breeding site, a slow release of insecticide over a period of 20 to 30 rnm may be desirable. This was how ground controllers of African onchocerciasis vectors applied their larvicides. Very fast applications! a consequence of aerial treatments, usually do not take effect, until.!he insecticide has become longitudinally dispersed in the ' river and has passed a point for a considerable time. In the aerial application of larvicide in the OCP the aim is normally to achieve overlap of doses, relying on the carrying pov/er of rivers. The main technique has thus been the "Vide Vite" fast release of E.C. (see WHO/VBC series Nos. 76.614 to 620 for details). For the very different conditions of attempting to control S. ochraceum. an inhabitant of tiny streams in mountainous country of Central America, a solid block impregnated with temephos fo res t des igned 1e81).Eo be released at 1 mg/I/60 n,in, has been developed (Nakamara et a1. 47- Application techniquesmust be developed for each special circumstance. It may be that a fundamentally different aerial application technique of t.he Tecknar 4O2 f'ormulation would obviate, or at least reduce, the amount of water in which it must be pre-mixed. This, however, seems unlikely. 4.5.2.2.4 Ditf.erences in the sensitivity of target species Inevitably different blackfly species are going to vary in inherent as well as acquired susceptibility to larvicides. It is likely that S. tridens is less sensitive than S. damnosum s.1. to both DDT (Wa1sh L970, and temephos (Kurtak et al. 1980) . There appears to be specific differences to Bacillus thuringiensis H-14 (Lace y 1980; Mo11oy et a1. 1981). This is hardly surprising and given slight differences in the configuration of the cephalic fans of different species any larvicide ingested in particulate form may have a slightly differential effect, though this will usually be minor. However, it is clearly necessary at some stage to test a larvicide against the intended target species, rather than to rely entirely on results extrapolated from elsewhere. -48- 5. BACILLUS TITURINGIENSIS SEROTYPE H-14 5.1 Introduction Since its isolation (Goldberg and Margalit, 1977) Bacillus thuringiensis var. israelensis or B. thuringiensis H-14 (de Barjac, 1978) has proved to be a highly active agent against a variety of mosquito and blackfly larvae, and has consequentty excited great interest (a cursory literature search unearthed nearly 60 papers with some relevance to Ehis section of the report) . This is in marked contrast to earlier strains of B. thur1nq]-ens1s. which although at times showing some toxic activity against mosquito larvae proved of no interest when 13 of them were bioassayed against late instar larvae of S. vittatum (Lacey and Mulla,1977a). Of great benefit is the apP arent selective effect of B. thuringiensis H-14 against certain groups of Diptera. This is presumably mainly a matter of physiological sensitivity aided by the fact that as a particulate material iE exerts its toxic effect only on ingestion. 5.2 Laboratory studies 5.2.L Mode of action This appears to be the destruction of the mid-gut 'epithelium. Lacey et al. (1978) have suggested that a high gut pH, which is known for boLh mosquiEos (Dadd, 1975) and the blackflies S. vittatum (Lacey et aI. , 1978) , and S. vittatum S. verecundum and Cnephia orqithoohilia (Undeet 1979), dissolves the parasporal crystal thus releasing its toxin. Mortality may occur within 15 minutes or aft.er many hours (Lacey and Federici, 1979) and may not necessarily be accompanied by detachment. -49- 5.2.2 Effects of water quality When the primary powder was used against mosquitos there was an 8 to 9 fold decrease in activity when tap water containing chlorine was used (SinEgre et al., 1980). However, both Sindgre et al. (1980) and Ignoffo et al. (1981b) give no reason for thinking that the pH of blackfly breeding sites is likely to influence the ef.ficiency of the toxin. Of possible significance is the finding by Ignoffo et al. (1981b) that B. thuringiensis israelensis was much more active aga inst Aedes aegypti in distilled water than in pond water. The important factor appeared to be sediment. About half of the B.t. israelensis activity was found by a sediment concentration of 2 per cent (particle size of 53 per cent of the sediment exceeded 500 microns, and orr1-y 24 per cent was of less than 100 microns). Dr P. Guillet (pers. com.) sEates that normal turbidity does not effecg the acEion of B.t. israelensis however, it is possible Eo get feeding inhibition (of S. damnosum s.1.) by using flour or barium sulphate There is some evidence that feeding inhibition occurs sooner with organic than inorganic materials. Gaugler and Mo11oy (1980) found that a variety of inorganic and organic materials caused feeding inhibition in S. vittatum larvae once the concenEration reached 50 ngl1-. It is important that the possibility of B.t. .israelensis bonding on to naturally occurring suspended solids in West African rivers be rigorously investigated. It may be that the bonding on the UPOM fraction would increase the efficacy and range of B.t. israelensis However, should much material be bonded on to large particles carry would be reduced and ingestion un1ike1y. -50- 5.2.3 Effects of temperature Bacillus thuringiensis israelensis activity is proportioned to temperature at least up to 30oC according to P. Guillet (pers. com.) working with S. damnosum s.1., a finding confirmed for S. vittatum, at least uP to 24oC, by l,Iolloy et a1. (1981) . 5.2.4 Spe cific and instar differences in sensitivity As with conventional chemical larvicides there is some evidence for specific difference in sensitivity . Nevertheless B.t. israelensis is highly toxic Eo at least two species of Palae arctic blackflies (I{eiser and Vankova,1979), six species of Nearctic blackflies (Undeen and Nagel, 1978), one Neotropical species, the vector S. ochraceum (Undeen et al.,1981), as well as the Afro-trop ical S. damnosum s.1., S cerviconutum S. unicornutum and S. alcocki group (Gui1let et Escaffre, L979 a and b), and S. hargreavesi (Guillet et al., 1981b). Guillet et al. (1981b) show how earlier instars of S. damnosum s.1. are markedly more sensitive to B.E. israelensis than are older instars, as is the case with conventional larvicides. Molloy et al. (1981) rep orted similar results with S. vittatum. 5.2.5 Duration of exposure and concentration Frommer et al. (1980) investigated the effects of exposure time using Abbott powder (6406-L25) against S. vittatum larvae. They found that 1arva1 morEaliEy increased with exposure time to 60 min but above that there was little change in tC50 and LC90 levels. However, Guillet et a1. (1981b) stress the importance of formulation type in determining this relationship (see below). -51 - 5.2.6 Formulations As Burges (198la) points out rrpathogens pose formulation problems of keeping them suspended in the correct position in the water. This must be solved before their fu1l potential can be realized". He instances the case of Goldberg and colleagues who encaP sulated B.t. israelensis in jojoba oi1 without reducing insecticidal activity after ingestion by mosquiros. This gives the bacterial material both added buoyancy and a hydrophilic surface which holds the material just below the water surface. Correct formulation of blackfly larvicides is an even more tricky problem than that posed by larvicides for mosquito control. Nevertheless, valuable lessons may be learnt from the experience of formulating other D. thuringiensis strains (Couch and Ignoffo, 19S1) and from conventional larvicide studies. Unfortunately, as Guillet et al. (1981b) points out, the problems posed by the utilization of serotypes 1 and 3 against lePidoPtera have been studied for 20 years without being satisfactorily resolved. Bacillus thuringiensis israelensis is already available as primary powder, wettable powder, aqueous suspension concentrate, granule and emulsifiable concenErate formulations. Their advant.ages and disadvant.ages have been reviewed by Couch and Ross (1980) (this PaPer not seen). -52- Molloy et a1. (1980) have compared the toxic effects of Abbott and Roger Bellon primary powders and of Roger Be1lon prinary powders, wetable powders, and aqueous suspensions aga inst S. vittatum. Ro per Be11on (Rl53-78) primary powder proved much more active than the Abbott formulation though both contained equal cel1 concentrations. In the comparison of the aqueous suspension proved to have low overall toxicity while an emulsion gave erratic and unanalyzible results. When judged on the basis of weight of active ingredient there was no significant difference between the primary and wettable powder formulat.ions. These results are quite similar to rhose previously obtained by Guillet and Escaffre (1979b) using these formulations aga inst S. damnosum s.1. larvae. Guillet et al. (1981a and b) further pursued their studies on Ehe acEivity of B. thuringiensis H-14 formulations against S. damnosum s.1. by investigating the way in which the toxin was ingesLed and in bioassaying 11 differenE formulations. In this work they have clearly demonstrated that the formulations having larger parEicles r,rere the most efficacious. They separated the Abbott wettable powder formulation (ABG 6108) into 4 fractions by particle size, ( 40 microns, 50 to 63 microns, 80 to 100 microns and over 120 microns. The most efficacious fraction was that containing the 80 to 100 micron particles, excepE for the first to third instar larvae which suffered most mortality with the 50 to 63 micron particles. A similar type of result was obtained by fractionating the B-oger Be11on B.t. H-14 673 powde 53 However, Ehe optimum size of particle may vary from powder to powder and depends on other physical properties of the particles such as density and friability. This last property is very important and explains why the most efficacious formulation tested in the laboratory, so far, has been the Roger Bellon R153/78 primary powder. It is essential to read the reports of Guillet and his colleagues concerning this work in fulI. Mo1[oy and Jamnback (198lb) also studied particle size and fornulation efficiency a1ainst S. vittatr:m. Efficacy was positively correlated with particle size. Sandoz TechnarR aqueous suspension concentrate was much less effective thau pither the Biochem Products BactimosR or TPS-78. However, in contre the settling rate of the toxic components of Technar was significantly slower; its much smaller particles stayed in suspension long after the other two formulations had settled. 5.2.7 Need for a standard ized bioassay test using Simulium larvae As B. thuringiensis II-14 becomes ever more widely tested, and indeed used operationally, there is a need to ensure that materials for testing conform to Ehe producerrs specifications (Frommer et al., 1980b; Guillet et al., 1981b). It is also necessary to have a bioassay test which can be used to provide pre-control susceptibility data, especially as regards popu lations of S. damnosum s.1. in l,Iest Africa. -54 Already several teams of workers have been involved in bioassay work using Simulium larvae (Frourner et al.r 1980a; Gaugler et al., 1980; Guillet et al. 1981b; Hembree et al. , 1981; Lacey et al. , 1981; Undeen and Nagel, 1978). It is thus desirable thaE, as far as possible, testing procedures be standardized, a point emphasized by Guillet (pers. com.) and Lacey et al. (unpublished). This would seem to eliminate the method of GauBler et a1. (1980) which, though elegant' seems inappropriate to many laboratories, while the simple and compact apParatus devised by Hembree et al. (1980), though suitable for S, vittattmr, seems unlikely to yield such good results with the much tess amenable larvae of the S. damnosum complex It has to be accepted that given the wide variety of Simulitrm species used in these tests Ehere may be inevitable differences between t.est procedures as regards flow rate (current speed), temperatures and feeding regime. Given that the Colbo and Thompson (1978) rearing system has been used successfully in bioassays with S. damnosum s.1. (Guillet et al. 198lb) as well as with Nearctic species (Undeen and Nage1. L977) the testing procedure devised by Guillet et a1. (1981b) may prove the most satisfactory overall. I,IHO should attempt to facilitate the adoption of a standard bioassay using Simulium larvae. -55- 5.3 Field studies Several field checks have been carried out aga inst S. damnosum s.1. in l^Iest Africa and the Sandoz Technar 402 formulaEion is being used currently on a semi-operational basis. This is inevitably a compromise but it is the only formulation that can be readily applied by helicopter and is likely to carry a substantial distance. In field trials the Roger Bel1on primary powder, when hand applied, was very effective at short range (Guillet and Escaffre 1979a) and a wettable powder formulation also showed promise (Guillet and Escaffre 1979b). Lacey (1980) reported on a large river tria.l in Ivory Coast of Sandoz 402 Technar. 0n advice from P. Guillet treatment was carried out at 1.5 mg/t/tO min. The formulation was diluted with 80 per cent water. At a discharge of 457 *37"""., temperature 26.8oC, and pH 7.2 the 770 L of diluted formulation was added Eo the river from boats. Complete control of S. damnosum s.1. to 19 lsn was achieved with very few survivors up to 28 km. Other Simulium species occurred for 4 km suggesting lower sensitivity. There were apparently no adverse effects on Ephemeroptera and chironomidae. Davies and Zerbo (1981) report a dry season experiment against temephos resistanE S. soubrense population on the Leraba river. At a discharge of a only 0.80 m'/sec. treatmenE was made at 1.6 mg/t/tO min. on the advice of Dr Guillet. The 402 Technar formulation was mixed with 60 per cent by volume of water before being applied by hand. Some treatments were also made by helicopter, the dosage time being 5 seconds. Results were rated as good with carry equal to that of Procida AbateR 200 EC. -56- Cheke (1981) during October and November in Togo carried out a series of field trials to test the carry of t,emephos, chlorphoxim, Technar 402 and Solvay. Altogether, 4 tests of each of the conventional larvicides were made, with 10 of Technar and 5 of Solvay, It was clear that the l,l - lt( Solvay formulation of B.t.t{-was totally unsatisfactory. Given at a dosage -\rate of 6.4 ng/L/10 min. the maximum carry was 2.4 kn at a discharge of 6 mr/sec. Chlorphoxim at an operational dose of 0.025 ngll-/lO mins. gave uuarkedly poorer carry than temephos, dosed at 0.1 mg/1/10 min. and appeared ro be roughly equal to the cirry of Technar 402 dosed at 1.5nglLll-l mins. ' and premixed with 20 per ce+t by volume of river water. Two trials with A'I near Technar ar 1.5 ngllr/lO mins. failed completely. ItE*, H. Agoua, ' ;. *1*rt"a and It. ocran (pers. com.) also report total failure with Technar when used without pre-mixing with !'rater. i' 5 .3.1 The operational situation 1 I I As yet it is not clear whether Technar 402 will carry as well as temephos in the wet season and in turbid rivers, though it seems unlikely to do so \n the small clear streams of Southern Togo. AE Ehe dosage rates required 0.9 1 of Technar is used p.. 13 of discharge, as against 0.3 1 for temephos. To achieve success the Technar ' must be premixed with at least 20 per cent by volume of water' Thus the amount to be dosed is 3.6 times that required for temephos treatments, even if only the same number of treatments are required with each product' It is. thus an urgenE mat,ter that the quantity of active material per litre of formulat.ion is increased aE least 3 fold, always assuming that chis will result in a 3 fold increase in efficacy. Further experimentation \\ -57- with application equipment may enable the amount of water which must be mixed with the formulation before application to be reduced. This should reduce logistic costs. 5.3.2 Cost implications At present cost of insecticide accounts for 8.52 of the OCP budget. As one litre of temephos costs 2.2 times as much as one litre of Technar, but at least three times as much Technar is needed, a change from temephos to Technar would mean a 50 per cent increase insecticide costs. Given the almost certainly poorer carry of Technar, there will be a somewhat greater cost increase both directly, through the use of more larvicides and indirectly through the need for more flying hours. 5.4 Key reference Gaugler and Finney (1981) have provided a valuable review, unfortunately as yet unpublished, of B. Ehuringiensis H-14 as a blackfly larvicide. -58- 6. INSECT GROWTH REGI]LATORS These so -cal1ed "third-generation pesticides"do not seem 1ike1y to be good agents for the control of Simulium damnosum s.1. on a large-scale though borh merhoprene (AlrosiaR) (oMS-l697) and diflubenzuron DimilirrR(ot'ls-tgoa) have shown some promise in preventing normal pupation by larvae and normal adult emergence when larvae were exposed for periods of 24 hours or more. They have been tested against a ntmber of N. American blackfly species (Cuming & McKague 1973: McKague & Wood 1974; Dove & McKague 1975; McKague et al. 1978; McKague & Pridmore L979; Lacey & Mu1la L977, L978a and b, I979a; Thompson & Adams 1979). Emergence of adult S. verecundum and S. decorum was reduced by 57 to 94 per cent following exposure of late instar larvae to O.l mg/llht Altosid. Slow release of Altosid, over a 48 hour period, from formulated blocks, giving a concentration of 0.002 to 0.006 mg/t in a smal1 stream suppressed adult blackfly emergence for 23 days 18 metres downstream' and for 17 days 43 metres downstream. Later, laboratory tests showed that Altosid caused pupal rather than larval mortality in S. venustum/ verecundum and pulse dosing proved to be more effective .nrn "orIII-,r, dosing (Thompson & Adams L979). Susceptibility increased with larval . age at time of dosing, and pupae were not affected. This specificity poses a severe limitation on the use of Altosid especially on fast developing species, such as many of the tropical vectors, unless their very frequent larval meglting causes them to be more susceptible generally, than are N. American species. This should be tested. Diflubenzuron Dimilin has been shown to c'a,use mortality in several species of blackflies in laboratory and field tests. Dosage rates of 0.1 to 0.2 ng/L/Lb ususally gives 8O-1OO per cent mortality. However, there are specific i 59 differences in susceptibility, with S. tescorum apparantly highly susceptible Second and third instarand S. vittatum less so (Lacey & Mu1la 1978a). larvae of S. vittattrn proved to be much more susceptible than penultimate instar larvae and there was a greater mortality with increasing water temperatures. S. vittattun eggs exposed for 24-48 hours failed to hatch unless they were already 72 hours old at the time of treatment (Lacey & Mu1la L97l and 1978b). Pupae were unaffected. Although both methoprene and difluben uron have relatively selective effects it is 1ikely that some valuable non-target invertebrates would be destroyed in treated streams, as occurs in sEatic water systems (Norland & Mulla L975; A1i & Mulla 1978). Difficulties in using these substances operationally against blackfly larvae are likely as a result of the long contact time required for high mortality. Such contact times are rarely achieved in dry season OCP aerial treatments for example. Further, Ehere is as yet 1ittle information concerning the carry of these materials, though the superiority of diflubenzuron wp formulations over emulsions (Lacey & Mulla 1979) suggests that this may not be greaE. Against this mortality form diflubenzuron shows a clear positive correlation with temperature, at least up to 24oC, and it may be even more active at normal water temperatures in West Africa. Another major problem would appear to be that even diflubenzuron was most effective on only part of the aquatic stages (2 and 3 instar larvae) whereas conventional larvicides are very efficient against all but the prepharate pupae. i7 60 THE DEGREE OF HAZARD INVOLVED IN HANDLING AND APPLYING LARVICIDES TO PEOPLE AND ANIMALS DRINKING TREATED WATERS In the OCP, as at present For this operated most of the insecticide is applied by insect icide aircraft. task there are over lOO durnps countries of the of scattered throughout the seven rogramme area. Most of these durnps are unguarded and unprotected from htunan interference in any \"ray. Larvicide has been stolen from at least 10 of these dumps at one time or another. The main reason for theft of larvicide is probably to use as fish poison. In such cases Ehe drtuns are frequently slashed open with a panga which is likeIy to lead to splashing of the thief. Another reason for theft is to use the drums, which will often be used as drinking !,/ater containers, after a perfunctionary washing out. The pi1ot, in the course of a treatment cyc1e, will refill his insecticide tank at numerous dumps which may be a considerable distance from the river. Temperatures are high and protective clothing, except gloves in some cases, are not worn. Should the pilot get splashed he may have no opportunity of washing except for the limited amount that could be achieved if he used his emergency drinking water supply. During treatments, especially in the dry season when the cockpit temperatures may be 45oC, insecticide is sometimes placed precisely during hovering flight or in lines as the helicopter tacks back and forth. Personal experience confirms that such operations spray may be wafted into the cokpit through open windows. A smal1 amount of the larvicide is applied from the ground using Hudson sprayers or by simply pouring directly from drums. This work, which is likely to increase, is carried out by sub-sector Personnel, whose primary role is fly catching and searching for larvae. They have no training in insecticide application techniques and in potential hazard. 61 - The targets of this larviciding activity are rivers and streams which for many conrnunities provide the sole available water supply for drinking, cooking and washing, as well as for watering stock; at least in the dry season (up to 7 months). These rivers are usually treated in the dry season at the rate of O.I rrlg/L110 mins. in the case of temephos. Where the larvicide is applied more s1owly' at the treatment point the concentration may briefly approach 6O ppm. In the Anglophone countries of West Africa, insecticide is referred to as "medicine" in pidgin English, as are pharmaceutical products. Every- where among subsistance farming and fishing communities, which are largely illiterate, there is an almost total lack of appreciation of the potential hazards of pesticides. Given these operational "facts of 1ife" it is clear that any compound chosen for use in the suppression of blackfly larvae in the tropics must have a 1ow level of toxicity to mammals. For the compounds initially considered as potential candidate larvicides for the OCP the Oral LD'O hras a minimum of 5OO rirC/kE and the dermal toxicity was very low (Wt10 1973, Annex III-3). Jamnback & Frempong-Boadu (1966) also gives a valuable list of oral and dermal LD5O for 17 compounds which they considered to show some promis as blackfly larvicides. There is no doubt that these standards must be strictly adhered to in the future. 8. DETERMINATION OF LARVAL SUSCEPTIBILITY 8.1 fntroduction The aim of a susceptibility test should be solely to provide an assessment of sensitivity to an insecticide in standard conditions, or as Quel6nnec (1976) prt ir "the purpose . is to obtain a relative estimate of the sensitivity ....". Essentially all jar tests are -62- susceptibility tests and provide 1ittle other valuable information though they have frequently been tried in screening procedures. In the jar the insecticide is as it may actually work without a current of at as a contact poison, and not by ingestionacting in the stream. Harrod (1965) has shor^m that least 19cm/sec. S. ornatlrm var. nitidifrons cannot keep its cephalic fans open. It can be assr:med that the S. damnosum complex, v*rich is typically found in fast water, will respond in a similar way, and not feed in still water. Apart from important work on other species, (Gjullin et al. 19504 Jamnback 1962; wilton & Travis 1965; Jamnback & west 1970) there have been a number of reporEs of studies on S. damnosum (t'tuirhead-Thomson 1957; Ovazza & Valade 1963; Qu6lennec & Vervent L97O; Walsh L973; Raybould L975; Thompson 1975). Qu6lennec (1976) reviewed the situation. Qu6lennec & Vervent (1970) were successful in obtaining satisfactory regression lines by exposing larvae to low concentraEions of insectide for 24 hours without aeraEion. Such long exposures are also likely to resulE in the loss of insecticide by codistillation (Bovnnan et al. 1959; Weidhaas et a1. 1960). Further, long holding periods require control over the ambient temperature, which in the OCP area may exceed 4OoC at times. This means a permanent power supply is necessary if tests are to be carried our. Walsh (1973) & Thompson (L975) tried to avoid this problem by carrying out tests in the field. Exposures were made for 30 minute periods, early in the mornings with ice used to regulate Ehe temperatures of the test solutions which were placed in water baths. After exposure the larvae v/ere returned to the breeding river in cloth bags for the holding period. Both workers found that dead larvae tended to disintegrate, though Thompson, who counted out exact number of larvae, at the beginning of the test \n7as able to report saEisfacEory LD'O levels. A holding period with larvae enclosed in a _63_ b"g, in natural conditions \^ras also used by Travis & Itlilton (1965). Laboratory studies have varied with regards to the methods of retrieving larvae from natural substrates, exposure times, holding times (though in mosL cases exposure times were 30 minutes and holding tiroe 24 hours as recommended in WHO 1970), use of river or distilled water aeration of test and holding solutions, solvent for technical insecEicide (acetone or eEhanol) and of course temperatures. Qu6lennec & Vervent (1970) avoided excessive manipulation and questions of aeration. However, at the very 1ow doses required by their technique there is a greater risk of experimental error. Very good summaries of this earlier work are given in Qu6lennec (1976) and Mouchet et al, (\977). As Qu,6lennec (L976) states "it seems pointless to attempt to reproduce the conditions found in the natural larval breeding places The only essential is that the controls should survive during the test period". 8.2 The "Mcuchet test" Mouchet et aI (L977) provided a satisfactory test for West African conditiot which has been accepted by the Expert Cornrnittee (WHO). They have pointed out that testsmust be reproducible, simple and executable in the OCP area (or any other control area). Thus a sysEem which does not need mains for electricity is desirable. Further distilled r^rater must be used, as any river water any case is 1 ikely use of riverto be contaminated by the control operaEions, and in r^rater imparts an unquantif iable variability between the test of lulouchet et al . consists of the exposure of larvae for 3 hours to distilled water and insecticide tests. 4-5rh in a 250 m1 Es s ent ial 1y ins t ar container at 2O-2LoC. Mortality is recorded at the end of the exposure period. The distilled water is aerated before the test, but not during the exposure period. Moribund larvae are counted as dead. 64 The sixth and seventh instar larvae are discounted as they have been noted as much more tolerant. Tests with a six hour exposure provided no advantage and temperatures of 25oC led to abnormal results. This method has now been used in very many tests by IRTO and OCP staff and has proved very satisfactory. 8.2.1 Choice of larvae for "Mouchet" test While preparing a detailed protocol for use by the OCP technicians, Kurtak (1980a & b) restudied the question of larval age selection and also gathered some data on variation due to observer error. As regards Ehe first question, he confirmed that older larvae were indeed about 10 times more tolerant than younger ones, as suggested by Mouchet et al . (1977) for S. damnosum complex, and originally for Alaskan blackfly larvae by Gjullin et a1. (1950). This change seems to come very rapidly between the 5th and 6th instars. Third instar larvae are also significantly less tolerant than 4th and 5th instar larvae, and should onty be used when no other larvae are available. He concluded, however, that both "young" (4th and 5th instar) and "o1d" (6ttr and 7th instar) larvae could be used in tests, provided that comparisons of LC'O were made within the same age group. This being done in the OCP. Kurtak further discussed the various methods proposed for separating the instars and found that they were somewhat ambiguous and difficult to apply consistently and that the different methods could lead to significantly different tC5O,". He proposed some simpler criteria, which although not giving a perfect separation, could be applied consistently by large numbers of observers. However, Ehe determination of larval instar varies from species to clear that species including within the S. damnosum complex, and it is some species cannot be separated into instars by the use of 65 C. (Cnephia dacotensis (Ross & Merritt 1978);morphological criteria, e C. ornithophilia - (colbo biology). Accord ing in larva1 age, of at least two s us cept ib i1 ity thuringiens is times in the LC of50 8.2.2 Limitations and modifications Although the "Mouchet test" has proved its worth, especially in screening for resistance to the OP compounds, tem.ephos and Chlorphoxim, a 24 hout holding period may be advisable for chlorinated hydrocarbons. Both DDT and Methoxychlor produce many moribund larvae, someLimes more larvae are moribund than dead in short tests. This can cause mis-scoring and give rise to false suspicions of resistance. With a 24 hour holding period this problem is resolved. Any such modified holding period should follow the standard 3 hour exposure time of the "Mouchet test" rather than reverting to the half hour exposure time reconruended in the earlier literature. Thus riverside 3 hour test data could be extrapolated as could the mass of OCP data (wtto 1981b). Another urgent need is for a test which can provide a measurement of for particulate insecticides, and especially for Bacillus & Wotton 1981). (See also section on larval to Kurtakrs data, when observer error is combined with variation the detection limit of the susceptibility test is a change the population. H-14. A Eest based on the rearing technique of Colbo & by Guillet & Escaffre (fggOa)Thompson (1978) as modified for bioassay could form the basis for a standardised Eest. 8.3 Observations arisine from susceptibility tests on African simulicides Certain interesting factors have and during field control operations. to a large extent on their tendency to arisen during susceptibility testing Ease of manipulation of larvae depends attach in still water. S. damnosum s.1. : -66- S. neavei and S. hargreavesi are noticeablydifferent in this respect (Raybould & Clark L974, Raybould 1975, Walsh 1973). Both Qu6lennec & Vervent (1970) and Walsh (1973) found that S. hargreavesi was more amenable to testing and provided much better regression lines than did S. damnosrm s.1. It may be that the different speeds of development of these larvae affect their testability . Thus hargreavesi appears to dwell in clearer water than S. damnosurn in many areas and may grow, and moult more sIowly. It would be valuable to know whether proximity to uroulting time affected the susceptibility. Kurtak et al. (1980) carried out tests on S. griseicolle which is known to survive where all S. damnosum are kitled by DDT (trrlalsh l97Oa, 1970b) and temephos (many observers in OCP area). Tests suggested thaE S. griseicolle was much more tolerant than S. damnosum but in the absence of pre-control data whether this had been acquired since the onset of control was not known. However, dosage mortality curves for exposed pop ulations of S. riseicolle and did not become horizontal at higher dosage levels. Data in Walsh (197Ob) shows that griseicolle in the Niger river above Kainji, was more tolerant than damnosr.rn after three years during which only 11 DDT treatments had been made. This suggests that tolerance may have been the pre-existing condition. The question of tolerance in griseicolle should be checked, as it should in adersi and schoutedeni. Apparant tolerance in the wild could of course arise from ingestion of a different. range of particle sizes, if the larvicide was acting as a stomach poison, and could be a result of behavioural adaptation as well as physiological tolerance per second. Only the latter condition is determinable by susceptibility testing. 67 8.4 Effect of aeration during exposure period Jamnback & West (1970) pointed out the interesting fact that LC 1eve1s50 a factor of ten between tests carried out with aeration exposure period, and without. In both situations the larvae were confined in organdy bags. It is 1 ikely feeding that. provision of aeration during the Eest period affects larval differed by during the fan activity believed to (Harrod 1965). As the insecticide form a co11oida1 suspension rather (Bor^rman et aI . 1959) larvae in the aerated waterr may have been poisoned by ingestion as well as contact. That mosquito and simuliid larvae can feed on colloids is established (Dadd L975; Wotton L976). The precise reasons for the results of Jamnback & tlest should be investigated. Esterase activity Recent work has indicated that the S. sanctipauli/soubrense cytospecres pair in Southern Ivory Coast e>rhibited a broad spectrum of esterase activity in 1978 before the introduction of routine larviciding. A similar broad spectrum of esterase activity was also exhibited by Ehe presumably non- resistant (uncontrolled) S. soubrense population from Soubr6 on the lower Sassandra. Temephos resistant populations of this cytospecies pair, hore.r"r, ' showed a narrowing of esterase activity to one end of the spectn:m. In contrasE Togo populations of soubrense have a narrower spectrum of esterase activity concentrated at the opposite end of the spectrum to thaE exhibited by resistant populations in Ivory Coast, (S. Meredith pers. com.) It may be that the Eastern population of S. soubrense are intrinsically less 1ikely to become resistant than were those in Ivory Coast. Similar studies on S. griseicolle populations might be instructive. no aeration, no cephalic in question, DDT, is than a true solution 99 -58- IMPACT OF SII{ULIW LARVICIDES ON NON-TARCET E.^IINA I General considerati-nng Simulium control operations have two the other indirect. basic types of environmental impact, one direct and The direct impact is that which occurs to the environment from the use of insecticides and other agents to reduce Simulirm populations and the indirect impact is that which arises from changes in the distribution and activity of human populations following successful application of control measures. The former is both more obvious and more rapid in its manifestations. Nevertheless, indirect impact must be borne in mind as the redistribution of human populations is one of the basic aims of Onchocerciasis control activities, at least when these are being carried out on a large scale. The introduction of insecticide into running water systems can have fairly drastic effects on the aquatic fauna and result in marked deteriorations in environmental quality. As Hynes (1959) sEated "the biological effects of poisonous pollution are primarily simple elimination of individual species for varying distances below the outfall. If conditions are suitable, the absence of surviving species may become unusually abundant because of some biotic factor which normally controls them". This control Simulium larvae, of the fauna or flora. However, in most Simulium control operations the larvicide will be introduced at many po:-nts on numerous occasions, every week for up to statement applies equally to Lhe effects of of pesticides into flowing water in order to "trash fish" or any other unwanted elements rivers, and thus unless care pollution. There are numerous examples , is taken, may exceed of the changes which have the accidental introduction of agricultural, (196Oa) in a the deliberate introduction 20 years in many of the OcP those occasioned bv accidental and more taken place following especially, forestry pesticides into rivers. Hynes classic exposition of the -69- effects of pollution in general gives some valuable data on these matters, especially in relation to chlorinated hydrocarbon compounds. Muirhead- Thomson (1971) gives extensive consideration to the subject and provides a particularly valuable source of information regarding the laboratory testing of the impact of larvicides. However, his account is unduly negative in its discussion of DDT and contains 1ittle information regarding the two Simulium larvicides most frequently used at the present time (temephos and methoxychlor). 9.2 Laboratory studies These are valuable in that detailed observation on individuals of one or a few species testing has been Muirh ead-Thoms on the subject Fales et a1. can be made under rigidly carried out most elegantly controlled conditions. Such ].na long series 1981 a, b). of studies by He also reviewed aspect are Sanderson & (1973, 1977, 1978a, b, c, 1979, in detail (1971). Other important works on this 1968; Fredeen 1972; Jensen & Gaufin 1964a, b; Cope 1966, 1968; Yasuno et a1 . 1978. A11 such studies are of limited applicability and as trrlallace and Hynes (1981) state "approaches used in laboratory testing make predictions of couununity dynamics of benthos in running waters difficult, if not impossible". Nevertheless, laboratory . toxicological tests do determine specific toxicant effects and accurately compare precise responses to specific chemicals and their various formulations. After all one of the first steps in choosing a blackfly larvicide is generally a laboratory test which shows a particular material to be toxic to a variety of mosquito larvae. I-7 0- The Erouble is that differences in technique make such studies extremely hard Jto "orp.... Differences in aeration, temperature etc., are inevitable if the organisms to be tested are to be comfortable, and not subject to excessive control mortality. Thus, Sanders & Cope (1958) and Jensen & Gaufin (L964a & b) both studied the effects of pesticides on naiads of Nearctic species of stoneflies (Plecoptera). However, the former authors did not use aeration in their tests while the latter did so. Despite such problems, general trends can be detected in the results. Many authors indicated that smaller specimens hrere clearly more sensitive than larger ones, of the same species. The effects of temperature varied according to the class of compound . Thus, toxicity of all chlorinated hydrocarbons to Trichoptera larvae varied inversely with temperature, while organophosphorous compounds varied directly (Fredeen 1972). These temperature findings are supported by the work of Yasuno et al. (1978) on Crustacea and to some extent by Sanders & Cope (1966) on Cladocera. Jensen & Gaufin (1964 a, b) found that one of their species of stonefly was consistently more tolerant to the wide range of pesticides tested. However, this is not always the case even among congeners. Sanders & Cope (1966) showed that three species of Daphnia and one of Simocephalus generally exhibited the same order of sensitivity to each of the test insecticides, but Daphnia magna, alone, was much more tolerant Eo endrin than were the other three species, a result confirmed by independant groups of workers. In general OPs were more toxic to Cladocerans than were chlorinated hydrocarbons. Muirhead-Thomson (1973) found that fenthion was as toxic to Odonata naiads as it was to Baetis naiads and Simulium larvae, 3 strong Ipointer against using this effective at Odonata naiads to kill Simuliurn and Baetis. Temephos is also relatively innocuous control dosages (JFt{). In laboratory tests that deltamethrin was 2O times more letha1 -7 L- material against blackflies, while DDT was not at concenErations forty times those required to Odonata at normal Simulirmr Muirhead-Thomson (1981a) found to Simulium larvae than was temephos and 4 times more than ihlorphoxim in one hour exposure tests. The same author (1979) showed that the predatory caddis larvae Hydropsyche and Rhyacophila were more tolerant Ehan Simulium larvae to both temephos and chlorpyriphos methyl. However, the difference was insufficient to be safe. rn tests on six non-target organisms the tolerance levels to chlorphoxim and deltamethrin were elucidated by Muirhea<i-Thomson (1981b). These tests showed that deltamethrin was highly lethal to all the non-target organlsms excep t Ephermerella, (which is relatively tolerant of some other classes of insecticide-JFW). All the non-target organisms were more tolerant of chlorphoxim than were Simulium larvae, though this was not marked in Baetis. Muirhead-Thomson goes on to argue that such laboratory testing in the OCP area' with appropriate non-target organisms, would aid the interpretation of the field data provided by Dejoux and his colleagues. . Further valuable information on the impact on non-target organisms is available from studies concerned with the use of mosquito and Mulla larvicides, see for example the reviews of Mulla et al. (1979) & Mian (1981). Apart from tests carried important developments in the larvicides has been the use of 1 aboratory the effects of potential Simulitun out in the testing of the trough one of the most test located at the riverside. -72- l Using such Eechniques, the ORSTOM hydrobiologists have been able to ohrain a relatively rapid and reliable assessment of likely impact of a larvicide on the non-target fauna, without the need to undertake a full scale river test. The technique 9.3 Field trials of is described in Dejoux (L976) and Troubat (1981). larvicides in Afriea the "DDT era" Early African attempts at insecticidal control of blackflies used DDT and dosages which were generally rather high. Garrriram & McMahon (Lg47) discussed the elimination of S. neavei from the Kodera focus. They used a locaI1y made up emulsion of DDT at a rate of between 2 and 5 ^g/ 1/30 min. (ft should be noted that by the mid-sixties, controllers were having good success against S. damnosum using dosage rates of between 0.1 and O.O5 ng/ll3o min., even less, on rivers as large as the Niger and Congo). They gave a graphic account of the effects of treatment and it seems worthwhile to quote a little from this to give an idea what such high dosages of DDT could do, but alsb to record the ephemeral nature of the damage. Garnham & McMahon (L947) state "the first confirmation of Ehe efficacy of DDT treatmenE was an agitated report from the local 1ittle 1ater, however, inhabitants that fish were dying they care and asked when the next take place. They found it such an easy way of getting No controller could write along those lines now, the eradication of S. damnostnn and indeed, the use of of insecticide would be prohibitively expensive. In the case under consideration treatments r^/ere in large numbers. A application hras to fish". even if he was reporting such large quantities carried out over of 4 months, about once a intensive searching showed one mudfish. However, tv/o fortnight. One month after the cessation of the presence of months later the some snails, Ieeches, crabs a period treatments, and larvae of Ephemeroptera, Odonata, -73- i Hydrometridae (bugs), Corixidae, Gyrinidae, Culicidae were already present in some abundance. Unfortunately, the ecology of African rivers and known before the advent of the OCP and Beadle (1974) and Simulium alcocki streams TroS not well in his definitive work a lesser extent (Mormyrus victoriae which initially fed "The inland waters of t.ropical Africa" deals almost entirely with 1akes. Thus, only one or two investigations had been made into the influence of pesticides on African flowing waters. In one of these studies, Corbet (1958a, b)-took advantage of the periodic temporary closing of the sluices of the Owen Fal1s Dam at Jinja (Uganda) on the Victoria Nile to make collections below the Dam during the 1ow water periods. Fortunately he had the opportunity to make his collections before and after the use of DDT to control S. damnosum. Before treatment, he found that the invertebrate fauna v/as very dense with five genera of Ephemerontera and seven genera of Trichoptera present as well as 20 species of mollusc. Amongst the most abundant were a filter feeding mayfly Tricorythus tinctus and a caddisfly Phanostoma senegalense. The most obvious effect of treatment with O.44 mglt DDT was the disappearance of all mayflies and caddisflies. Other groups survived well and there appeared to be no diminution of molluscs, leeches or crabs. However, heavy recolonisation . occurred within six weeks. Several fish which before dosing fed predominantly on the lithophitic larvae of mayflies and caddisflies switched to a wide range of plant and animal food, some of terrestrial origin (C1ariallabes petricola Gnathonemus longibarbis and to kannune but others such as Mastacembelus almost exclusively on lithophilic mayfly naiads became very scarce, either , -7 4- through moving auray or starvation. As the invertebrate fauna recovered the fish reverted to their original feeding habits. Nevertheless, Corbet (1958a) predicted that with regular be greatly modified and many Hynes & IJilliams (1962) treatments the character of the river would of DDT on the invertebrate fauna of and fish species would be eliminated. the effects of a single application the Manafwa River (Mount Elgon, Uganda). after a single treatment of DDT at ins e ct s tudied Collections were made 29 and 31 days 1 mg/1/30min. The mayflies, stoneflies and caddisflies had all been ""Jiously affected as shown by the absence of larger larvae. However, on nunbers of animals, the only major change was the increase in Simuliurn larvae. This is to be expected following a single treatment which leaves eggs unharmed and large populations of adults in the vicinity, and which may have destroyed the predators regulating the Simulium population. Walsh (1970a) and Walsh & Mellink (1970) made some observations during the control of Simuliun at Kainji, N. Nigeria. After eight years of intermittent rrearmenr of the Niger River (nOt at O.O5 to 0.033 rr.g/l/lo min.) collections showed that a very rich fauna still existed, inclrlding the stonefly Neop.erla spic complex and many species of mayflies and caddisflies. Seven species of mollusc had been collected before control operations started, after six years of control nine species were collected. Invertebrate collections were ' made on the perennial Kontagora River after six years of continuous regular acriviEies (DDT at 0.1 to 0.5 mg/1/30 min), Fish,crabs and molluscs were fairly numerous and Coleoptera and Odonata larvae were found. After six months interrupEion of control (through one dry season) five species of Simuliun were re-established as were chironomids and mayflies. No caddisflies or stoneflies were taken. I a-7 5- Post & Garms (1966) studied the effects of DDT and Baytex (fentkrion) on West African fish after observing a fish kiI1 on the Niger in Guinea following treatment with O.l ng/l/ 30 min. DDT. They found that the solvent was highly toxic. Baytex was the more acutely toxic material to fish but at concentrations used in Simulium control the safer material. They also reported, as have many other workers, remarkable variations in tolerance of different fish species. In their study Platypoecilus maculatus was particularly tolerant. Apart from the work of the ORSTOM scientisEs which led to the decision to us temephos in the OCP, which is dealt with later and the papers of Blanc and i colleagues (1956a, b,d; 1958), these appear to be virtually the only earlier references to the effects of the deliberate introduction of insecticide for Simulir-m control in Africa on the non-target fauna. However, as pointed out by Hynes (1970a) perhaps the most striking feature of the rheophilic insect fauna is its world-wide uniformity. This fact enables us to utilise the results of workers in other regions to get some idea of the likely effects of insecticide usage. 9.4 Field trials in North America and Europe Much of our knowledge concerning the impact of insecticides in flowing water comes from a consideration of the effects of extensive forest aerial spraying against Spruee budworm and Gypsy moth in Canada and the USA. Elson & Kerswil1- 1966; Hastings et a1. 196l; Hitchcock, L965a; Ide 1957 & 1967; Cope 1961 & 1966; Flannagan L973). In most cases, these studies concern the heavy use of DDT, at the rate of f.rom 2Lbl acre gradually reduced, partty because of the objections of sports fishermen, to about 0.5lb/acre. -7 6- Hynes (1961) gives an account of the effects of an accidental spillage of a sheep-dip conEaining BHC into a small Simulium stream in UK, Collections were made one month after the spi1lage. Firstly there was a marked development of the flora in the "treated" reaches of the stream, particularly tlg4optrcEg and S-pirogyra. A11 trout Salmo trutta numerous above the treated stretch, had been eliminated. The crustacean Gammarus, mayflies, caddisflies, stoneflies were eliminated as were many beetles, most Diptera and hydracarine mites. Tricladida appeared unaffect.ed, and Oligochaete worms, molluscs and chirono.mid midges had greatly increased. Cook and l"loore (1969) reported on the use of Rotenone to control "rough fish" in a Californian River (i.e. they used the rotenone with the deliberate intention of wiping out the fish fauna so that it could be replaced with more "valuable" species). After the fish kil1 they found that the invertebrate fauna of riffles and pools increased greaLly. Simuliidae and chironomid midges were among the groups Eo show this population explosion, as did Ephemeroptera and Trichoptera to a lesser extenE. Simulium larvae were even found in pools, suggesting that predation rather than physical facEors had previously confined some species, at least, to Ehe rapids areas. The treatment was carried out in September and by }fareh invertebrate population 1evels were similar to those in untreated streams. -77 - Coming to the effects of blackfly control progranrnes, Jamnback and Earby (L962), working in the Adirondacks, compared streams which had never been treated and those which had been regularly treated with DDT for from five to ten years. The arthropod fauna appeared to have been reduced by a third. Trichoptera, EphemeroPtera and PlecoPtera were the worst hit groups. Biomass per sq. m. had been reduced from 0.378 to 0.247 ga. Fredeen (1974, 1975) reported on the treatment of the Saskatchewan River with single applications of meEhoxychlor at rates of 0.L8,0.24 and 0.6 rigl1-l1,,5 mins. He concluded that at these rates Simulium species could be effectively controlled without. serious environmental damage' Careful monitoring showed that fish were unharmed. Of the non-target invertebraEes the Plecoptera were the most sensitive equalling the target Simulium species in this respect, with, in decreasing order of susceptibility, Ephemeroptera, Trichoptera and Chironomidae. An important finding was that insect larvae located within the bottom deposits were not affected. Residues in water, sand and invertebrate and fish tissue were negligible and short lived. Burdick et al. (1974>, like Jamnback and Earby, worked in the Adirondacks. They found that there was a slight reduction in the diversity of the arthropod fauna of riffles when both DDT and methoxychlor were used, but recovery was very fast. After treatnent the average daily standing crop was reduced by 407" for DDT and by 2LZ f.ot methoxychlor. -78- Flannagan et al. (1979, 1980) have studied the effects of methoxychlor on the Athabasca River. Treatment was at 0.3 ^glt/tS mins. for 15 minutes and caused catastrophic drift for 400 km of this large glacial river (mean fLow 572 *3/".".). Long-term changes to the Ephemeroptera' Trichoptera and Plecoptera occurred. Some of these organisms were more sensitive than the target Simulium. Methoxychlor levels well above ambient water levels were recorded in natural populations of invertebrates, with different levels being recorded in caged clams and crayfish. The authors therefore did not recournend the use of animals in cages, or the use of methoxychlor. The critical view of methoxychlor as a blackfly larvicide, taken by Flannagan and his colleagues was not supported by other investigators of the Athabasca blackfly control study. The reports of this exceptionally detailed work (Haufe, 1980; Haufe and Croome (eds.), 1980) should be read in full by all concerned with the larvicidal control of blackflies. Wallace and Hynes (1975) and Wallace eE at. (1976) also studied the effects of methoxychlor in laboratory and field experiments. They reported acute catastrophic drift taking plaee for about 150 min. which represenEed a reduction in standing crop of the stream. The insecticide ki11 was very non-selective and Plecoptera, in particular, were severely affected. Nevertheless Surber samples indicated that species in the drift, including the target Simulium, survived in the bottom deposits. MosL importantly they found considerable amounts of methoxychlor residues in stream vegetation eight weeks after treatment. -7 9- Moving on to temephos, the insecticide chosen by WHO for its anti-S imul ium damnosum campaign in WesE Africa, there was very little Dre-control data. Wallace et al. (1973) working in Canada used artificial substrates, Surber samplers and drift nets to check the effects of temephos Abate} Dursban and uethoxychlor. A11 Ehree compounds produced catastrophic drift, but Surber samples showed that no invertebrate species had been eliminated. Among the non-target species killed were predaceous Plecoptera and Trichoptera, including Hydropsychidae, a grouP knor,rn to be important in trIest African rivers (Petr, l97O), some species of which are probably significant predators of S. damnosum s.1. (Burton and McRae, 19721 Service and Lyle, 1975; Service and Elouard, 1980). It was not possible to make detailed comparisons beLween these insecticides as conditions varied between tests which inevitably had to be carried out on different streams or at different times. Nevertheless from reading these papers we learn that several different types of insecticide can be formulated and applied so as to give a satisfactory ki1I of Simulium larvae. In all cases a wide variety of non-target organisms are caused to detach and are probably killed. However, blackfly larvae are particularly sensitive so that with carefully conErolled treatment rates it is possible to reduce Ehis ki11 of non-target organisms substantially. Unfortunately among the -80- species which appear to be especially sensitive to larvicide are the Plecoptera and the Hydropsychidae (Trichoptera) which, as already mentioned, can be important predators of S. damnosr:m. Fortunately most organisms which are located within the bottom deposits escape the effects of treatmentr So that streams quickly recover from the occasional treatments required to control pest species in temPerate zones which form the basis for most of the above sEudies. 9.5 The selection of temephos for the OCP This subject has been covered definitively in Annex III-3 of the PAG Mission Report (WHO, L973). With regard to the testing on non-target organisms, carried out in West Africa on behalf of WIIO, the key references are Dejoux and Troubat (L973, L974, 1975), Lauzanne (1973) and Lauzanne and Dejoux (1973). 9.6 Monitoring of the Onchocerciasis Control Prograuune 9.6.1 Introduction From the beginning, wHo treated the envirorunental- asPects of the prograr0ne very seriously as is indicated by ghg faet that the first of the advisory corrnittees to be established was the Ecological Panel on which the four agencies, FAO, UNDP, WHO and the world Bank which are responsible for the running of OCP, are each represented by a Technical Advisor, .usua11y someone from outside the Agencies. In addition, uNEP are represented on this cos'mittee. This panel met as early as January T974. An ad hoc Sroup of hydrobiologists met in May t9 74 and eventually produced an aquatic monitoring protocol. -81- 9.6.2 Methods The monitoring protocol laid down a standard set of environmental measurements and standard sampling methods to be carried out at regular specified intervals at certain representative sites. Initially it was decided that at each site to be monitored invertebrate animals would be sampled by means of Eckmann grabs, Surber samplers and Drift nets. The samples to be taken at set times of day and night at monthly intervals. In addition some simple data related to the condition of the river was also to be collected (e.g. width, depth and speed of current, temperature, pH and turbidity). Sites would also be visited at three monthly intervals when fish populations would be sampled using batteries of gill nets and the catches weighed and measured, and the gonads examined (Lev6que et a1., L977). 9.6.3 Implementation Unfortunately owing to the lack of knowledge of tropical rivers in general and of those in West Africa in particular together with the lack of skilled personnel localIy, except in Ghana, it took sorne littIe time for the monitoring activities to sEarE, and in parEs of Ivory Coast and Ghana the Simulium control operation (Phase I) had already started. However, monitoring eventually got under way at nine sites where both invertebrate and fish monitoring were being undertaken, with two additional sites for fish only and two for inverEebrates only. Three sites in Ghana -82- were chosen situaEed on large rivers near their confluences with the Volta Lake. Other sites in Ivory Coast and Upper Volta covered typical S. damnosum breeding rivers, one of which was a temporary stream. 0n seven of the rivers monitoring began 4 to 36 months before Simulium control activities. Experience soon showed that the Eckmann grab was generally very inefficient with the type of river bottom found in the West African savanna zorre and in order to study the bottom fauna reliance was placed on the Surber samples and the use of artificial substrates. Nevertheless Ehe programre relies most heavily on the taking of drift net samples to determine the rate of daytime and nighttime drift. Such samples can be taken at any water 1eve1 and are the only ones possible with rivers in flood (providing there is a suitable bridge from which to suspend the nets). A11 the data collected is entered on standardised forms and stored in the W[10 computer. Copies of the computer tapes have been transferred to Salford University where a detailed analysis of the data is being undertaken at present. Some idea of the complexity of this data can be gathered from the fact that the invertebrate form includes sixty taxa, whilst no less than 272 fish taxa have been reported. -83- This biological monitoring progranme is aimed at identifying major ecological change in the biology of the iivers of the OCP area (Lev6que eE al., L977). To achieve this the sites were carefully chosen and included two rivers which were noE expected to be EreaEed for several years. The collection techniques are simple and sLandardised. Each group of field workers uses exactly the same type of sampling equipment, the field teams meet annually Eo discuss the field programne and Eo compare t.axonomic findings and to suggest revisions of the monitoring protocol. This latter is a major problem of working in Africa where the 1arva1 stages of fresh water insects can only very rarely be identified to species. Most of the taxa on the inverEebrate reporting form are, in fact, families. Inevitably there is a fair level of imprecision and it is difficult, if not impossible, to deal in concepts such as diversity indices. Added to these problems is the fact that savanna rivers vary hydrologically from season to season and from year to year to an extent not encountered in temperate zones where most of the basic study of running water faunas has been undertaken. It is clear that smal1 changes caused by use of insecticide might well be masked by the natural annual variations which occur and have hitherto not been sLudied. From the OCP we have four years pre-control data from one river giving a unique amount of information from a tropical river. -84- As the OCP has developed so has the monitoring programme which is now being carried out in five countries at about 20 different sites. Most of the work has been carried out by the ORST0M team based in Bouak6, Ivory Coast, which in addition to the routine Progranrne have conducted a targe number of research investigations revolutionizing our knowledge of the running water faunas of West Africa. 9.6.4 Results The ORSTOI"I studies have been published in a series of about 35 reports from the ORSTOM Hydrobiological Laboratory, Bouak6. In addition annual reports to the OCP are prepared by all monitoring teams and numerous papers have been published in international journals. See, for example, Dejoux (1975, L978), Dejoux and Elouard (L977), Dejoux et a1. (1979), Elouard and Lev6que (1977), Paugy (1980), Samman and Pugh Thomas (1978a,b, 1979), and Troubat (1981). Usually examination of the rivers treated by OCP reveals a healthy looking aquatic fauna with a wide range of species present and often abundant. On numerous occasions it has been found that S. damnosum has been eliminated by treatmenEs while even some other species of blackflies are still present (S. adersi, S. tridens, S. schoutedeni). This is important because in riffles blackflies may be a significant proportion of the total biomass. The ideal control would in fact be to eliminate S. damnosum s.1. and have its niche occupied by say S. erlersi, this sometimes occurs, but only temporarily. Unfortunately fish catches -85- in West African savanna rivers have been falling during the lifetime of OCP, but as these reductions are siurilar in treated and untreated river basins this presumably cannot be the fault of the temephos treatmenEs. They are much more 1ike1y to be Ehe resulE of a series of unfavourable hydrological seasons or of heavier fishing pressure. It is valuable to have some monitoring outside the control area in order tohave sueh comparative data available should disputes arise, and for a more satisfactory interpretation of the daEa from controlled sites. The monitoring teams themselves have carried ouE many studies whilst undertaking the routine work. Dejoux and Etouard (1977) studied the action of a single Eemephos treatmenE during control operations by taking a long series of drift net samples each hour for a period before and after the passage of the insecticide. Like the north American workers previously quoEed they found that the treatment with temephos caused a catastrophic drift involving a wide range of organisms, including the mayflies Baetidae and Caenidae, the caddis flies Macronema and Orthotrichia and the chironomid midges 0rthocladiinae along with the Siuruliidae. It should be noted that Plectoptera do not usually form a numerically important part of the fauna of African streams (Table I), though they feature greatly in the literature on temperate running r^/ater studies. Another point is that organisms such as the llydropsychidae tend not to appear in the drift even though they succumb to the insecticide. Dejoux and his colleagues have gone on to show that the first treatment of a river with temephos may well result in the destruction of up to 60 or 707" of the fauna, but with subsequent treatmenEs the level of catastrophic drift is greatly reduced. -86- Sasunan and Pugh Thomas (1978a) studied the effects of temephos treatment by means of Surber sampling. They noted significant reductions of the mayflies Baetidae and Leptophlebiidae but no reduction in the important Caenidae. Orthocladiinae were also severly affected but, somewhat surprisingly, none of the Trichoptera were. The same authors (1978b) studied the effects of Eemephos treatment on the zooplankton microcrustacea, in particular Thermocyclops hyalinus, in laboratory and field. In the field they found no effect despite the fact that at the time of treatment temephos concentrations were much above the 24 hour LC50 levels for this animal. This is explained on behavioural grounds. In later studies (Sauunan and Pugh Thomas, L979) treatment with temephos was shovm to cause a short period of catastrophic drift of Cladocera, Copepoda and Chaoboridae which started within one hour of the passage of the insecticide. However, drift was normal and of the same magnitude as pre-control drift 24 hours later. Walsh (t979) carried out a monitoring programne on a river which was seasonally without flow and indeed without standing water for up to thTo months each dry season. Sanpling was carried out in 1975 (pre-control) and the subsequent control years. The health of a stream is usually monitored by calculation of an index figure expressed by the formula: Mean Night Index of Drift Mean Day Index of Drift -87- The results for the tr^7o end of season uonths sampled each year (October and November) are as follows: Year Index Z of pre-control figuqe t97 5 34 .L 100 t976 13.5 39.5 1977 14.0 41,2 t97I 22.L 64 .9 Surber samples for November in each year show no reduction in numbers of animals during the control period. In fact the lowest figure is that recorded in 1975, the pre-control year. Thus it is clear that there is no evidence for any serious effect of the control activity. L975 t976 1977 l97B ) No of animals/m- 7,999 8,737 13,858 9,688 A further confirmation of this state of affairs is the fact that no tax on routine recorded in 1975 was lost in subsequent years. However, a detailed examination of the data reveals that the proportions of different groups of organisms present had changed markedly, and that the Chironominae had greaEly increased in relative abundance. 1975 L976 L977 1978 Chironominae as Z of total fauna 20 48 76 7t This group is generally taken to indicate a pollution problem. Results of the other field teams have proved generally similar to those of Walsh. At a meeting held in September 1979 attended by represenEatives of the field tearns and by outside exPerts including Hynes and Fredeen it \^ras agreed that the exploratory analysis of the data carried out using the Salford University computer confirmed: (a) the general impression of the hydrobiologists thaE treatment had had remarkably litt1e effect on the biota; treatments with an emulsifiable concentrate B9 gross changes, it is nevertheless clear that six years of regular (weekly) of temephos has left remarkablY at least as regards thelittle sign of environmental impoverishment, invertebrate fauna. As regards fish the time scale is as yet too short to be that no deleterious effects have occurlred. As the ORSTOMabsolutely certarn Annual Report for as well as abiotic 198O states "natural variations from one year to another, factors, can have a considerable effect on a river totally masking the perfectly by of Eutropius effects of a pesticide if these are slight. This is illustrated the ichthyological results where the sudden massive return mentalis (i.e. to the Comoe in December L979) could not have been suspected even six months earlier". It is perhaps of interest to draw attention to the finding regarding the amount of flowing waters actually treated by the OCP \,rithin its area. It has been estimated that in the first year of control in any one area about 66 per cent of the total available flowing \^/ater (at least as regards river length) was treated in both wet and dry seasons. After about three years of routine control the proportion had dropped to about 50 per cent (Walsh 1981). Thus a very considerable body of flowing \,nater is rarely or never treated. This includes important headwater stretches of major rivers such as the Black and White Voltas, and a nr-unber of widely scattered rocky streams which though suitable for Simulium hargreavesi are rarelY colonized by S. damnosum. These untreated stretches form extremely valuable refugia for non-target organisms. 9.6.5 The impact of currently used alternatives to temephos 9.6.5.1 Chlorphoxim Recently Dejoux and his colleagues have turned their attention to the impact of chlorphoxim in Southern Ivory Coast. This larvicide had been studied -90- earlier by statzner (1g77) with inconclusive results. Dejoux & Troubat (Lg76), Gibon & Troubat (1980), Dejoux et al. (1981) and oRSTOI'I (1981) alI clearly indicate that chlorphoxim has a much more devasEating effect on the invertebraEe fauna. There does not aPpear to be any direct effect of the fish but the danger lies in a reduction in available fish food (oRSToI"l 1981). ft is probable that west African fish can show a considerable degree of plasticity in this respect as was shovrn by those of the victoria Nile (corbet 1958a) . In any case some species, such as Alestes nurse rely on terestrial, allochthonous insects for a considerable part of their sustenance in the normal course of events (Paugy 1980) ' 9.6.5.2 Bacillus thuring iensis serotyPe H-14 Preliminary tests on aquatic organisms living in mosquito breeding habitats indicated that this serotype was uninocuous to a wide range of inverrebrate species (wt{o 1979). Colbo and undeen (1980) evaluated B.t. H-14 against the fauna of a smalI Newfoundland stream, although the simuliid larvae suffered a mortality of more than 93 per cent no significant decreases \^7ere recorded for the major grouPS of non-target insects (EPhemeroptera t Trichoptera, Ple tera). Molloy & Jamnback (1981a) treated a smal1 stream in New york State and achieved an 89 per cent reductioh in Blackfly larvae, at the same time and Ephemeroptera' Trichoptera, Plecoptera, Chironomide and Flumidae populations were increasing' then no adverse effect on any of these non-target populations was evident' Dejoux (1979) tested Roger Bellon primary powder k-153-78, the most effective formulation of B.t. H-14 in laboraEory studies in troughs and in a small river in the Ivory Coast. He concluded that the compound was virtually innocuous to the non-target fauna in that term' while showing high toxicitY to S. damnosum s.1. - 91 - q Gibon et al. (1980) studied the effects of an experimental treatment of the Maralon6, a substantial river in the Ivory Coast. A Sandozliquid formulation was used, being applied rapidly from a boat after dilution with 60 per cent water to provide a dose of 1.6 mg/1/10 min. Several trough tests were also carried out. Only the simuliidae were noticeably affected by the treatment. Overall the night drift doubled, from its usual on the night following treatment. In a trough Eest at a dose of 3.2 mg/t/10 min there was an important detachment of all types of Ephemeroptera and Chironomidae. However, the chemato psyche (Trichoptera) were usually unaffected at this concentrat ion. There is now little doubt that B. thuringiensis sero type H-14 when applied to the breeding sites of S. damnosum s.1., cause even less damage to the non-target i,nvertebrtsE fauna than does temephos. However, there is a need for long-term studies to confirm these very positive preliminary findings. 9. 6.6 Key publicatiorrs Apart from the work of the ORSTOM team, other key publications with respect to the impact of Simuliun larvicides on non-target organisms are Muirhead-Thomson (197L); Garner & Bailey (Lg75); Haufe & Croome (1980) and Wallace & Hynes (1981). -92- Tuble I F.eglon No. ot Co] 1 ect lons - Inverr,etlrare Groups Classtf':ed bl Or'der of' Btonrass f'rom JO At'rtcrn Hlvers (by I'. Walsh, 1973) a WesL Africa OCP area tJesL Africa alIW;st Ar'rIca not, OCP are East Africa 20I4UIO )zI12I)2I)Iriank bY Blomass 1 ) 4 i I 1 I I o 2 o l 2 4 4,i I o o o o 2 IOI ) 4 1 o I I I ) ) 0 2 0 0 I I 4 I 0 o 0 o I 2 o 0 I o o I I I 2 o o o I I c. ) o 0 0 o o I 0 4 2 o ) I o o 2 ) 4 o 0 o o I 2 6l o ) 0 I o o o Simultidae 'frichoPLera^ [.:phemero Pr.era Chlronomidae flecoPtera ColeoPtera Odonata OEher I TABLE I! S.ITE 7 BAGOIJE NIGHT DRIFT TA.YON r'latar\a s.E P Baetidae before, a f t,er 95.15 28.?O 61 .08 9.37 o.316 Hydropsychidae before a fter 34.54 4.L2 22.55 2.67 o.204 Philopo tomidae before a f t,er 9. 16 4.48 4.96 t.37 o.394 0.128 o.97 5 o. 711 O Eher Simul iidae be for e aft.er 5.2s o.25 3. OO o. r8 Chironominae be fore a fter 9 .57 9. 39 3.81 5.20 0r Ehocladinae be for e afcer 6.L7 7 .L5 2.24 2.23 92 9.6.7 ANNEX I Appendix to the Report of the Informal Working Group in the Onchocerciasis Control Programme in the Volta River West Africa. (OCP/WG/81.1) on Temephos Resistance Basin Area of A Note on Criteria for Jud 1n the Environmental Acceptability of Insecticides for the Control of Simulium Damnosum The choice of temephos as the best compound darnnosum was made ten Years ago. to be used for the control of Simuliuu One of the most important criteria considered in its selection was its relatively 1ow roxicity for Ehe non-t.arget aquatic fauna. AfEer several years of application, it has been proved that temephos can be considered as an environmentally "safettinsecticide. This implies that: the acute effects of acceptablel temephos on the invertebrate fauna are ecologically temephos reduces the density of invertebrates at the breeding sites in proportions which have never 1ed to a disequilibrium under normal conditions of application; there is no ezidence of the disappearance of species; no fish nortality or any other notable change in fish poputations has been recorded after temephos applications; i accumulation in the food chain seens to be low. For all Ehese reasons, the use of that compound is, from an ecological point of view, without major risks for the aquatic ecosystem. It is felt that the crireria referred ro above should be adopted for the selection gf alternative compounds for Simulium control.II a It - 93 - ANNEX 2 Report of the Second Sess ion of the Ecological Group, Ouaqadougou 25 - ?9 Ma1 1981 Criteria for tl-re environreirtal- acceptabilit.r of insecticidcsioi tl:e con'b::o] liiluliun darurosrrm 29. The i.loricii:g Groui-l on Temephos Resrsta:ce clre',.,' Lrp a list of criteria for jud.girr.S tlie environmental accepte,bili[' of insecticidesfor th,e control of Sisrulium Ja.:nncsrua. ;':.fter trany years of stud;,- a;1<l in'bensive scieirtific C-iscu,-ssionr the ecclo5f istsr hi,c-Eobiolo,3ists and. enviroitr:eirtal tox.lcol-o,.;ists coir:rected. '*ith the Progranmc have reachecL the ccncensus ths.t, from an ecolo3;ical point of vier.r, the rrse of teinol:hos is uithout iiajor risks fo:: tbe aolu:,ric ecosysteni. Given ti"e ;rcsent use4;'o a.:+ tenephos fornulation, it i:as been conclud.ed that: (r) ti:e j-n-necliate effects of tcrrrephos on tne invertebrate fauna a::e ecologically accepts.ble; acute iaortality nay be consi.- d.eral-ile amorg thc inverteb::ate artlrropods but there is afair pro:orticn of sr:rvivors a?ong practically all ta:rono:lic gnDuls t (z) tenepi'rcs red.uccs the d.e:rsit;,' cf irvertebrates at the breed.ing sites ir proportions '"Aicir have never led. to a d.j.sequilibriur:r uncler norinal co:Citio.rs of application; l=\\) ) -ihere is lj.ttle evidei.,ce of d.rsappea-raitce of invertebrate species , (+) iro -erch nortality or arry other notable cha.nge in fish ponul-a'bions iras been recorded. af ter te_lephos api:licr.tions; (l) accumulation in. the food. chain seens to l-'re low. 10. Tire 5colo,:1 ical llroup aclapted. these consid.era.tions into criteria alp1i-cab1e to t:e selectj-on of a.Iter:.ative ccri)lou-n{s for Sir-nulir.rn corrtrolr' as f ollor+s; r1) Tnc ac:,-ro effects of a cand.icate pcsticiJe, i:l the fon:nu- I:r-ti cn and d-,rse ra.te as approrirr-3-te for. r bs use a6ainst *=,"1!= should not red.uce fl:e nrr:rlters of invertebrate saecies to a l-eveL at lrhich ti:eir s,,rrvival ir a giveirloc:li-t;r r,,ouf c'l- -he enci-aru3erecL. (z) Tl:e nesticitle si:.oul-d. nct g:.-re r-isc to tr:.c regicnai loss of ::-rf i-rrvertebratc s'-.ecies; tlle tenpor3ry searional l_oca1 d.isanlca-r.'a:rcc of : s.,-tes ci Si::u.ii,;;r one i:rvertebrate s-)ecies at t-:e -ureed,in6: u:a1- havc to i:c e"cccutcd,. .t) '-T:e pesti-ci,Le shou,-rL rot cal-.se a 1c.1;-rerr,r ,i i.e. extcnd,i^1T l;ei-oncl t-.e re:<t sc,-.son) i:ba.l a::ce': .-1r :1o.-:1al coed itions of airi:1ic:ti,:n, c.5. i:ia::l:erl s:rif ts i-r -c,_e .i:e1a.l j.ve aou:ic1a.rce cf siccieo s_:ou1C nc-t occur. ?he ',rs., :i t'.e les-;rcid,e s:icul* :rave nei ,.:ter arrJ olrecti.i:tr31 :-: f:-s:: "or r.:.,;,, ef'fecl on;-re I:f:-cycl-e of fisir. ,'rl\ ANNEX 3 -94- (i) Cor-rpo,.,.l:r)slil:e1y to accumulate ir: tirc food. rreb should be avoided. (5) In the proccss of selectirg pesticj-d.es for !jrlg].i.un control in an a,::ca fulI accourit shoulct be tallen of hr"man activities r,rhich either by thei,.rselves or in conbi:iation v;ith the vector con'Lrol cperations might canr,se adversc effects cn the environ- na_:* .. rv-r v a 3l-. The last criterion Lras been addecl bec.--.use v$:ious other forms of Iand.-use or cf sani-ta4' control are 1il:e1;- to rlevelop si:lultaneously clr-rrii:€,; tl.:e next fe,.,r d.ecad.es in tlie regions urcler consid.erl'"tioa. flrus, for instance, the release of chemicals, or ot1:er acti',rities, n:ay hamper the iecoverT- cf rive:-'ine po-pulations r'rhich nalr l12.vs been reducecl by Si:ruli'Lr::l cont,:oI operations . -95- 11. ACKNOWLEDGEMENTS At ORSTOM Bouak6, I am especially grateful to Dr P. Guillet for spending considerable time discussing the problems of larvicide testing. Mr J. Troubat provided documentation from the hydrobiology laboratory. Mr & Mrs J. Assetine, also of ORSTOM, provided hospitality in Abidjan. Messrs H. Agoua, M.Ocran and M. Ouedraogo of the OCP staff, Bouak6, briefed me with up-to-date information on the resistance problem in the Ivory Coast and of the control activities using B. thuringiensis H-14. At the OCP HQ, I am especially grateful to Dr D. Kurtak for advice about susceptibility testing, larval feeding behaviour and formulation chemistry. Dr H. Jaurnback answered a wide variety of question concerning larvicides and their testing. Dr J. Grunewald and Mr. G. Zerbo helped in discussions and Drs S. Meredith and R.A. Cheke told me about their work with consultants. Dr R.W. Crosskey, British Museum (Uat. Hist.) loaned several rare offprints and provided photocopies. Dr B. Thompson, Environmental Protection Service, Alberta, most generously sent me some results from his, as yet unpresented, thesis. Dr D. Mol1oy, New York State Science Service, provided unpublished manus cripts . Dr R. Wolton, Goldsmithfs College, London, Clarified my ideas about larval feeding behaviour. Professor D.H. Molyneux, Salford University encouraged me in the project and provided facilities at Salford University. 96- Ms E. Lee, Salford University kindly provided time-series graphs of Secchi Disc readings. Finally, Dr R. Le Berre, VBC provided sound editorial advice and Mme A. Frongia typed the report from a very untidy manuscript with the help of Miss A. Cathie. 97 CONCLUSIONS AND RECOMMENDATIONS Section 2 t. Larval feeding biology and the mechanics of the feeding process are now fairly well understood. IE would be of interest to have further details regarding the possible ability of larvae to select particles within a particular size range. Ihe possibility that particle change effects capture and/or ingestion, should be considered. There seems to be no need for such fundamental studies to be carried out on any particular species. This approach is, however, unlikely search for new larvicides. 2. Given the possibility that the to provide an avenue to success in the distribution of S. soubrense is limited by competition with S. damnosum s. str and S. sirbanum it mighr be instructive to have experimental studies of these species living together. Section 3 3. Basic knowledge 1arvaI habitats seems of the physico-chemical conditions of S. damnosum s.1. adequate. However, attention should be paid to all aspects of the turbidity of vector breeding streams at different seasons. Section 4 4. The physical properties of E.C. formulations are knovrn in general terms. The spectrum of particle sizes ingested by S. damnosr:m s.1. larvae is knovrn and provides a rational basis for the choice of particulate materials. Adsorption properties of formulations might be given more emphasis with a view to adding adsorption enhancing materials to E.C. formulations. 5. Simuliurn larvae are knor,m to be capable of ingesting col1oida1 materials in the absence of particulate matter. Acetone solutions of technical DDT were probably presented as colloids and were among the most effective formulations tested in the laboratory. The question of operationally -98- applying colloidaI larvicides should be investigated. 6. Particulate formulations appear to have severe operational limitations unless they are of very sma1l particle size. In contrast there is proof that the exceptional efficacy and carry of DDT and methoxychlor, in large Canadian rivers, owed much to adsorption on to naturally occuring partieles. It is probable that temephos, as used in the OCP, also acts in this way. 7. Micro-encapsulated formulations offer a theoretically eleganE means of delivering insecticide to filter-feeding larvae. However, in practise the technology of production seems to be over-sophisticated and uncertain. It is felt that efforts to develop such formulations should receive 1ow priority. 8. Larvicide screening procedures, against both target and non-target organisms, using jar tests without a through-flow system, seem of very marginal value. Trough tests are preferable. Although an indication of the parameters which influence the activity of any particular candidate larvicide and its likely specErum of activity can be achieved using the fauna of any region, screening against the operational target species and its associated non-target organisms is most important. Extrapolation from other species seems to be fraught with difficulty. 9. The minigutter (and to some extent the large gutter) test, in which larvae feed normally when provided with suspended solids, offers a relatively reliable and rapid method of screening larvicides, when used in an area of operational control. 10. Sgream trials are necessary to check on application techniques, on carry, and on the results obtained in artificial conditions. Experimenters need Eo pay particular attention to recording the hydrological conditions in which the trials take place in order to enable valid comparisons between tests to be made. -99 11. Despite the resistance which has developed in some S . soubrense populations, it seems likely that temephos will remain the main larvicide for the OCP. Where resistance develops the use of ehlorphoxim appears to offer 1ittle benefit. Given its serious environment.al impact, at least in the dry season, it should not be used at that time. L2. Alternative larvicides should be urgently sought among the chlorinated hydrocarbons and carbamaLes. Although not so far proved adequate for West African conditions methoxychlor must be considered the most promising contender. It has the required safety to vertebrates and lack of persistence in the environmental is ts environment. It has proved generally acceptable to in North America, though its impact on the non-target It is alsoinvertebrate fauna in West Africa remains to be ascertained. cheaper than most alternatives. If it cannot be put into suitable formulation for use in West Africa, some of the other non-persistent analogues of DDT shouXd be tried. 13. There seems less likelihood of finding an environmentally acceptable and reasonably priced carbamate. L4. There seems 1ittle likelihood that any of the synthetic pyrethroids could be used operationally on a large-sca1e r.rithout substantial damage to the riverine fauna. 1 100 Section 5 15. B. thuringiensis serotype H-14 shows great promise. At present an inadequate percentage of increased at least three-fo1d, preferably the operational formulation contains active ingredient. This needs to be more, without a corresponding increase in cost per litre of formulation. 16. A standard bioassay test against Simulium larvae should be agreed to and noted by WHO. The modification of the Colbo/Thompson rearing technique employed by Guillet seems a promising method. L7. Consideration should be given to application techniques with a view Eo reducing the need to pre-rnix with water. 18. Reconsideration of the formulation might also lead to a reduction in the need to add r^rater. Section 6 19. Insect growth regulators are most unlikely to become operational low priority in thisaga ins t regard. Sect ion S. damnosum s. 1. They should be given very 20. The totally unguarded nature of most of the OCPs stocks of larvicide, the lack of sophistication in the locaI community, and the use of treated rivers as domestic water supplies must constantly be borne in mind during Ehe search for new larvicides. Section 8 2L. Thanks to the "Mouchet test" the situation regarding susceptibility testing of conventional chemical larvicides can now be considered as generally s at is factory . 1arva1 instars onchocercias is FurEher consideration of the methods of determining the in differenE members of the S. damnosum complex, and of other vectors, is required. The question of "holding time" during needs further study. 7 tests with chlorinated hydrocarbons also { I101 Entirely different procedures are required (see recournendation 16) . Section 9 for B. thuringiensis serotype H-14. 22. Results from the OCP indicate that, at least as far as the non-target invertebrate populations are concerned, 6 years of regular (often weekly) applications of temephos, at rates of about O.L/ngl1/10 min. have caused no serious impoverishment of the environment (Annex). More time needs to elapse before data concerning fish populations can be accurately assessed. 23. It seems that the moniEoring protocol is adequate to provide evidence of any gross changes in the fauna. 24. Without more taxonomic data on the West African river fauna, analytical techniques such as species diversity indices cannot be used Eo detect small changes. In this regard the forthcoming production of a well illustrated key to the fauna prepared by Dejoux and his colleagues is extremely opportune. It is to be hoped that this document can receive quality reproduction, befitting the fine art-work, and be widely distributed to interested workers. Possibly OCP/WHO could help in this. 25. The position regarding the effects of ehlorphoxim on the non-target ' invertebrate fauna appears to be unsatisfactory (see Recommendation II). The views of the Ecological group are given in the Annex 26. B. thuringiensis serotype H-14 is likely to be basically innocuous but adequate data remains to be collected. 27. Monitoring, according to the standard protocol the life of the OGP. Monitoring should continue for should continue throughout at least one year on any speed river which ceases to be treated. This may provide information on the of recovery of depleted populations. 28. The present was achieved only investigation by equally necessary conditions. of potential sma11-s cale LO2 satisfactory position regarding the monitoring protocol because there qlas a considerable voltrme of research experienced hydrobiologis ts . for any large scale control This type of imput would be 29. Although laboratory programre established in different studies can provide some information on the selectivity and on their likely mode of action, the only satisfactory to be that using the "cages flottantes" method must be carried out using larvicides test appears devised by Dejoux and his colleagues. Such tests non-target species normally found associated with 30. The WHO and the OCP should continue to take the Ecological Group (Report of 2nd session) to the selection of alternative compounds for the Annex. operational target species. advice of internationally the the respected ecologists and hydrobiologists. In this regard the recommendations of the Working Group on f,emephos Resistance and those of concerning the criteria applicable Simuliun control are given in /-1- Ali, A. and Mulla, M.S. (1978) Impact of the insecr growrh regulator Diflubenzuron on invertebrates in a residential recreational lake. Arch. Environ. ConEam. Toxicol. 7. t 483-491- Back, C., Lanouette, J.G., Aubin, A. G979) Preliminary tests on the use of temephos for the control of blackflies (Diptera : Simuliidae) in Northern Quebee. Mosquito News, 39 z 762-767. Bailey, G.w. and white, J.F. (1964) Review of adsorption and desorprion of organic pesticides by soil co1loids, with implications concerning pesticide bio-activity . J. Agr. food Chem. , L2 z 324-332. 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