The cytotaxonomy of Simulium sanctipauli and Simulium soubrense (Diptera: Simuliidae)

It is noted that the chromosomal inversion 2L-7, which has been used in the past to separate S. Sanetipauli from S. soubrense, occurs as an intraspecific polymorphism and hence cannot be considered diagnostic, although in some populations 2L-7 can still strogly indicate the presence of two species. However, two newly recognised inversions, 1L-A and 2L-A, can be used in combination to identify S. sanctipauli, S. soubrense and a new species S. soubrense B. The absence of the relevant heterozygotes for these two new inversion confirms the separate specific status of S. sanctipauli from S. soubrense from S. soubrense B as well as providing a reliable means of larval identification. The misuse of 2L-7 as a species diagnostic inversion has undoubtedly led to past misidentifications of S. sanctipauli and S. soubrense, and it is possible, for example, that only S. sanctipauli is resistant to organophosphate insecticides in Ivory Coast and not S. soubrense. Beffa form appears to be a distinctive geographic race of S. soubrense, but forme konkouré remains as yet unassigned. A cytotaxonomic key for the identification of members of the S. sanctipauli sub-complex is presented.     

Temephos-resistant larvae of Simulium sanctipauli associated with a distinctive new chromosome inversion in untreated rivers of south-western Ghana

Larvae of the Simulium damnosum Theobald complex (Diptera: Simuliidae) were sampled in June 1996 from two sites in south-west Ghana where larviciding has not been applied: Sutri Rapids on the Tano river (05 degrees 23 minutes N 02 degrees 38 minures W) and Sekyere-Heman on the Pra river (05 degrees 11 minutes N 01 degrees 35 minutes W). All specimens were identified as Simulium sanctipauli Vajime & Dunbar sensu stricto (Diptera: Simuliidae). Bioassays with temephos (organophosphorus larvicide employed by the Onchocerciasis Programme for systematic treatment of most rivers across West Africa since the 1970s) showed about five-fold resistance in the Tano population (LC95 2.37-3.14 mg/L) and slight tolerance to temephos in the Pra population (LC95 0.67-0.76 mg/L), vs. the diagnostic concentration of 0.625 mg/L. Larval salivary polytene chromosomes of S. sanctipauli showed fixed inversions 1S-24/24, standard IIL-6 and a new inversion IL/36 polymorphism at Sutri on the Tano. These karyotype characteristics differ from those of temephos-resistant S. sanctipauli in rivers of C te d'Ivoire and other sites on the Tano in Ghana. Thus, temephos resistance in S. sanctipauli at Sutri is associated with distinct chromosomal configurations, showing that immigration was unlikely. This resistance could have been locally selected by exposure of S. sanctipauli larval populations to agrochemicals run-off from cocoa, coffee and oil plantations flanking the rivers.     

Establishment and spread of Gyranusoidea tebygi noyes and Anagyrus mangicola noyes (hymenoptera: encyrtidae), two biological control agents released against the mango mealybug Rastrococcus invadens williams (homoptera: pseudococcidae) in Africa

Two specific endophagous parasitoids Gyranusoidea tebygi and Anagyrus mangicola, of Indian origin, were mass‐reared at the International Institute of Tropical Agriculture in Cotonou and released against the mango mealybug Rastrococcus invadens, in collaboration with national biological control programmes. G. tebygi was released in the following countries: Benin, Gabon, Ghana, Nigeria, Sierra Leone and Zaire. In Togo, it had been released earlier and studied during another project. This parasitoid is now established in all areas infested by the mango mealybug. In addition, it established itself without previous release in Congo and Côte d'Ivoire. A. mangicola has been released in Benin, Gabon and Sierra Leone since 1991, and by mid‐1993 was recovered from a few sites. It seems locally established in southern Benin.     

Elimination of the Djodji form of the blackfly Simulium sanctipauli sensu stricto as a result of larviciding by the WHO Onchocerciasis Control Programme in West Africa

Cytotaxonomic identifications of larvae of members of the Simulium damnosum Theobald (Diptera: Simuliidae) complex collected in forest zones of southeast Ghana and southwest Togo between 1977 and 1996 showed that the Djodji form of Simulium sanctipauli Vajime & Dunbar, a vector of onchocerciasis, was eliminated in 1988 by larvicide operations conducted by the World Health Organization (WHO) Onchocerciasis Control Programme (OCP) in West Africa. No members of the form were identified amongst 997 larvae collected up to 8 years after systematic control operations began in February 1988. The results are discussed in relation to estimates of the numbers of samples required to certify elimination and the possibility that other members of the S. damnosum complex were also eliminated by the OCP.     

Assessment of rDNA IGS as a molecular marker in the Simulium damnosum complex

For five cytospecies of the Simulium damnosum Theobald complex of blackflies (Diptera: Simuliidae) from West Africa, both ends of the intergenic spacer region (IGS) of the rDNA have been sequenced with the aim of developing specific molecular markers. No specific differences in these two regions were detected between Simulium sanctipauli V. & D., Simulium sirbanum V. & D., Simulium soubrense V. & D., Simulium squamosum Enderlein and Simulium yahense V. & D., except in the number of A subrepeats at the 5' end of the IGS (two in S. squamosum and four or five in the others) and in position 310 of the 3' end (a C in S. squamosum and a G in the others). However, genetic distances within and between species overlapped. These DNA sequences had no strong phylogenetic signal, and the trees obtained were mostly unresolved. Although most sequences from S. squamosum clustered together, a few of them were more similar to those in other cytospecies. These results could be explained either by hybridization with genetic introgression or by ancestral polymorphism and recent speciation.     

DNA probes for the identification of members of the Simulium damnosum complex (Diptera: Simuliidae)

Genomic libraries in plasmid have been constructed from various sibling species of blackflies of the Simulium damnosum complex from West Africa. Three cloned repetitive sequences, which show variation in copy number between sibling species, have been isolated. These clones can be used as probes for the dot-blot identification of larvae, pupae or adults into the three main West African subcomplexes, i.e. damnosum, squamosum and sanctipauli subcomplexes. The sequences also show some intraspecific variation in copy number.     

Completion of the sequence of the nuclear ribosomal DNA subunit of Simulium sanctipauli,with descriptions of the 18S, 28S genes and the IGS

We describe the IGS-ETS, 18S and 28S ribosomal gene sequences of Simulium sanctipauli Vajime & Dunbar, a member of the S. damnosum Theobald (Diptera: Simuliidae) complex of blackflies (Diptera: Simuliidae). These regions, together with the ITS-1, ITS-2 and 5.8S rDNA presented elsewhere (accession number U36206), constitute the composite sequence of the entire rDNA unit, making S. sanctipauli the second dipteran species of medical importance for which the entire rDNA has been sequenced. Despite the lack of sequence identity, the IGS of S. sanctipauli showed some structural similarities to other Diptera, i.e. the mosquito Aedes albopictus Skuse (Culicidae), the fruitfly Drosophila melanogaster Meigen (Drosophilidae) and the tsetse Glossina (Glossinidae). Two blocks of tandemly repeated subunits were present in the IGS of S. sanctipauli and, unlike other species of Diptera, they contained no duplications of promoter-like sequences. However, two promoter-like sequences were identified in the unique DNA stretches of the IGS by their sequence similarity to the promoter of Aedes aegypti L. (Diptera: Culicidae). The observed sequence variation can be explained, as in the case of Drosophila spp., by the occurrence of slippage-like and point mutation processes, with unequal crossing-over homogenizing (to a certain extent) the region throughout the gene family and blackfly population. The 18S and 28S rDNA genes show more intraspecific variability within the expansion segments than in the core regions. This is also the case in the interspecific comparison of these genes from S. sanctipauli with those of Simulium vittatum, Ae. albopictus and D. melanogaster. This pattern is typical of many eukaryotes and likely to be the result of a more relaxed functional selection in the expansion segments than on the core regions. The A + T content of the S. sanctipauli genes is high and similar to those of other Diptera. This could be the result of a change in the mutation pressure towards AT in the Diptera lineage.     

Progress in controlling the reinvasion of windborne vectors into the western area of the Onchocerciasis Control Programme in West Africa

Since vector control began in 1975, waves of Simulium sirbanum and S. damnosum s.str., the principal vectors of severe blinding onchocerciasis in the West African savannas, have reinvaded treated rivers inside the original boundaries of the Onchocerciasis Control Programme in West Africa. Larviciding of potential source breeding sites has shown that these 'savanna' species are capable of travelling and carrying Onchocerca infection for at least 500 km northeastwards with the monsoon winds in the early rainy season. Vector control has, therefore, been extended progressively westwards. In 1984 the Programme embarked on a major western extension into Guinea, Sierra Leone, western Mali, Senegal and Guinea-Bissau. The transmission resulting from the reinvasion of northern C?te d'Ivoire and Burkina Faso has been reduced by over 95%, but eastern Mali has proved more difficult to protect because of sources in both Guinea and Sierra Leone. Rivers in Sierra Leone were treated for the first time in 1989 and biting and transmission rates in Sierra Leone and Guinea fell by over 90%. Because of treatment problems in some complex rapids and mountainous areas, flies still reinvaded Mali, though biting rates were approximately 70% lower than those recorded before anti-reinvasion treatments started. It was concluded that transmission in eastern Mali has now been reduced to the levels required to control onchocerciasis.     

The Hydrochemical and Physical Characteristics of the Breeding Sites of the Simulium neavei Roubaud Group and their Associated Crabs in the Eastern Usambara Mountains in Tanzania

Investigations on the hydrochemical and physical parameters of the breeding sites of Simulium woodi (an onchocerciasis vector). and Simulium nyasalandicum, and on their aquatic stages and associated crabs, were carried out for 13 months in the Zigi (Sigi). River system. The conductivity, pH and concentration of the main ions and silicate were all higher in the breeding sites of S. woodi than in those of S. nyasalandicum. S. woodi was restricted to small heavily shaded streams and did not enter rivers and large streams which contained higher concentrations of ammonium and nitrite. S. nyasalandicum was most numerous in rivers and large streams, but invaded small heavily shaded streams during the rainy season when conductivity and pH values were reduced.     

Development of Onchocerca volvulus microfilariae injected into Simulium species from Cameroon

Microfilariae (mff) of the savanna and forest strains of Onchocerca volvulus (Leuckart) were injected intrathoracically into adult females of Simulium damnosum Theobald sensu stricto, S.sirbanum Vajime & Dunbar, S.squamosum Enderlein and S.mengense Vajime & Dunbar. Nine days post infection (pi) 27-29% of the savanna mff and 31-38% of the forest strain had developed to third-stage larvae (L3), irrespective of the fly species, size or injection dose (5, 10 or 15 mff). Savanna flies supported the development of forest O.volvulus better than forest flies, in contrast to the results after per os infections. Therefore, in these four species of the S.damnosum complex from Cameroon, the peritrophic membrane is considered to be the main factor limiting the success rate of microfilarial development following the ingestion of blood infections, while the fly's haemolymph and intracellular environment play minor roles.     

Inverse density dependence of parity rates in the onchocerciasis vector Simulium damnosum s.l.

A correlation between parity rates and an index of adult numbers of Simulium damnosum s.l. (Diptera: Simuliidae) indicates an association, but does not prove causality or show the direction of any causal relationship. The question of whether adult numbers affect parity rates or vice versa is reminiscent of the age‐old query of which of the chicken and the egg came first. A method for resolving such issues based on analyses of pairs of time series was proposed by Granger in 1969. When Granger's method was applied to monthly numbers of adult female S. damnosum s.l. caught attempting to bite humans at Asubende, Ghana, and their parity rates, a significant relationship (P = 0.005) emerged, clearly showing that parity rates were dependent on adult numbers. Implications of this inverse density dependence and the results of analyses of other similar time series are presented and discussed.     

尼日利亚西南部奥孙河沿线黑蝇Simulium damnosum Theobald complex的形态分类研究(双翅目:蚋科)

黑蝇Simulium damnosum sensu lato是由多个姊妹种组成的复合体,这些种在生态学和盘尾丝虫病的传播方面各不相同.本文对奥孙河沿线的尼日利亚西南部森林区域的黑蝇S.damnosum s.l.复合体的组成以及成虫的形态学特征进行了研究.本研究所用的黑蝇S.damnosum s.l.成虫标本来源于奥孙河沿线的3个区,分别是Osun Eleja,Osun Ogbere和Osun Budepo.标本采集通过人体诱捕的方式,采集时间从2008年2月至2009年6月上午7:00到下午6:00,每两周采集1次.通过观察成虫的翅毛簇(wing tufts)和其他形态特征,对其进行分类研究.结果表明:存在同域分布的森林种和稀树草原种.在奥孙河沿线3个区内,森林种为优势种,占总捕获量的99.18%,而稀树草原种仅占0.82%;森林种和稀树草原种在多度上存在显著差异(P<0.05).所捕获的所有稀树草原种的翅毛簇均为灰白色,而捕获的森林种的翅毛簇颜色存在显著差异(P<0.05).为了更清楚地揭示该地区黑蝇S.damnosum s.l.的物种组成,建议进行更深入的研究.     

Anthropophilic blackflies (Diptera: Simuliidae) and onchocerciasis transmission in southwest Ethiopia

Abstract. From May 1984 to May 1985, the transmission of onchocerciasis by blackflies (Dipteria: Simuliidae) was studied around Gilgel Ghibe and Gojeb rivers in southwest Ethiopia. The predominant anthropophilic species of Simulium at both localities were S.(Edwardsellum) damnosum sensu law and S.(Lewisellum) ethiopiense. Some S.(Anasolen) dentulosum also landed on human bait at Gilgel Ghibe river.
Dissections of S. damnosum from Gilgel Ghibe and Gojeb revealed mean parous rates of 92% (n = 18,291) and 84% (n = 9530), respectively. S. ethiopiense from Gilgel Ghibe and Gojeb showed mean parous rates of 53% (n = 322) and 93% (n = 14), respectively. Of the parous S. damnosum , 1.3% at Gilgel Ghibe and 0.5% at Gojeb harboured infective third-stage larvae (L3) of Onchocerca volvulus (or morphologically indistinguishable from it). Unknown filariae of animal origin, indicative of zoophily, were found in 0.3% and 0.7% of Simulium damnosum complex females from Gilgel Ghibe and Gojeb, respectively. S. ethiopiense harboured developing (L,) larvae only, with 7.7% infection rate in both localities. In contrast, S. dentulosum did not harbour any filaria larvae. The annual infective biting rate (AIBR) and transmission potential (ATP) of the S. damnosum complex at Gilgel Ghibe river were 858 and 5478, respectively. The AIBR and ATP of S. damnosum s.I. at Gojeb river were 519.5 and 1963, respectively. These results emphasize the predominant role of the S. damnosum complex in the transmission of O. volvulus in southwest Ethiopia     

Estimation of preferred water flow parameters for four species of Simulium (Diptera: Simuliidae) in small clear streams in South Africa

Blackfly larvae typically occur in fast-flowing riffle sections of rivers, with different blackfly species showing preferences for different hydraulic conditions. Very little quantitative data exist on hydraulic conditions linked to the blackfly species occurring in South African streams. Stones-in-current biotopes (i.e. fast riffle flows over cobbles) were sampled from four sites in three small clear streams in the Eastern and Western Cape provinces of South Africa. Mean water column velocities at each sampled stone were measured using a mini current meter, while flow velocities closer to the boundary layer where blackfly larvae occurred were estimated using indirect techniques (standard hemispheres and aerating tablets). Standard hemispheres were also used to calculate more complex hydraulic parameters such as Froude and Reynolds numbers. Four species of Simuliid were sampled in sufficient numbers to show trends in flow velocity preferences. Simulium impukane and S. rutherfoordi both occurred at their highest densities at velocities of 0.3m s^?1, while S. merops preferred velocities of 0.7m s^?1. Simulium nigritarse SL attained the highest densities of all the blackfly species sampled, and its relative abundances were greatest at velocities of 0.8–0.9m s^?1. Within the streams surveyed, all blackfly species occurred in subcritical-turbulent flows — based on a classification using Froude and Reynolds numbers — although two of the species were also found in high densities in supercritical flows where these existed at the sites. Local hydraulics within the stones-in-current biotope are complex, but in the absence of fine-scale equipment for measuring micro-velocities, standard hemispheres are a useful, cost-effective technique for the initial quantification of hydraulic parameters in small, clear streams. Such an approach facilitates further understanding of links between hydraulics and aquatic invertebrates in South African streams.     

Cytotaxonomy of four species in the Simulium perflavum species group (Diptera: Simuliidae) from Brazilian Amazonia

Polytene chromosomes of four members of the Simulium perflavum species group in Brazil are described, and a standard map for the species group is presented. Simulium rorotaense Floch & Abonnenc, S. perflavum Roubaud and an undescribed species (S.‘X’) are chromosomally conservative, each representing a single species with a unique preimaginal habitat. Chromosomal, morphological and ecological evidence indicates that S. maroniense Floch & Abonnenc, previously considered synonymous with S. rorotaense, is a good species. Independent morphological and chromosomal analyses yielded 99.4% agreement in separating larvae of S. rorotaense and S. maroniense. The two species can be distinguished by gill morphology or by a subterminal inversion on the long arm of chromosome III. Simulium maroniense consists of at least four cytotypes, each with different sex chromosomes and autosomal polymorphism profiles and associated with a particular landscape type, altitude, temperature regime or geographical location. Simulium rorotaense and S. maroniense share one unique inversion, as do S. perflavum and S. ‘X’, indicating two pairs of sister species. The anthropogenic S. perflavum probably dispersed into Central Amazonia sometime after the mid-1970s.     

Population relationships of Simulium yahense and Simulium sanctipauli in the Firestone Rubber Plantation at Harbel, Liberia

Comparative genetic differences for the phosphoglucomutase and trehalase loci were surveyed in larval and adult blackfly populations of the onchocerciasis vectors Simulium yahense Vajime and Dunbar, and S.sanctipauli Vajime and Dunbar. Genotype frequencies for each stage and all populations were at Hardy-Weinberg equilibrium, indicating that S.yahense and S.sanctipauli remain genetically distinct. S.yahense populations from three different locations were found to be genotypically comparable. The larval population of S.yahense found in closest proximity to a S.sanctipauli population was found to express the phosphoglucomutase allele 1.33, characteristic of S.sanctipauli, with significantly greater frequency than other larval populations of S.yahense. This may constitute evidence of limited genetic introgression of S.sanctipauli with the S.yahense population.     

A new species of the Simulium damnosum complex from Uganda, and comparative morphology of the tarsal claws in females of this complex

The female, male, pupa and larva of Simulium (Edwardsellum) pandanophilum sp. nov. (Diptera: Simuliidae) from western Uganda are described. Diagnostic features are the shape of the tarsal claws of females and, cytogenetically, new intraspecific inversions on chromosomes I and II. Simulium pandanophilum is assigned to the Simulium damnosum complex. Different tarsal claw shapes of West African members of the S. damnosum complex are also described.     

Blackfly vectors of zoonotic onchocerciasis in Japan

Studies of blackfly vectors of Onchocerca dewittei japonica Uni, Bain & Takaoka (Spirurida: Onchocercidae), a parasite of wild boar implicated in the aetiology of zoonotic onchocerciasis in Japan, and six other zoonotic Onchocerca species of this country are reviewed. Molecular identification of infective larvae found in wild‐caught female blackflies showed that Simulium bidentatum (Shiraki) (Diptera: Simuliidae) is a natural vector of O. dewittei japonica, and also Onchocerca sp. sensu Fukuda et al., another parasite of wild boar. Inoculation experiments demonstrated that Simulium arakawae Matsumura and four other Simulium species are putative vectors. Similarly, S. arakawae, S. bidentatum and Simulium oitanum (Shiraki) are putative vectors of Onchocerca eberhardi Uni & Bain and Onchocerca skrjabini Rukhlyadev, parasites of sika deer. Morphometric studies of infective larvae indicated that Onchocerca lienalis Stiles, a bovine species, is transmitted by S. arakawae, Simulium daisense (Takahasi) and Simulium kyushuense Takaoka, and that Onchocerca sp. sensu Takaoka & Bain, another bovine species, is transmitted by S. arakawae, S. bidentatum, S. daisense and S. oitanum. Prosimulium sp. (Diptera: Simuliidae) and Simulium japonicum Matsumura are suspected vectors of Onchocerca suzukii Yagi, Bain & Shoho and O. skrjabini [Twinnia japonensis Rubtsov (Diptera: Simuliidae) may also transmit the latter], parasites of Japanese serow, following detection of the parasites' DNA genes in wild‐caught blackflies.     

Poor genetic differentiation but clear cytoform divergence among cryptic species in Simulium damnosum complex (Diptera: Simuliidae)

A complex cryptic species taxonomy has been developed for the flies that vector Onchocerca volvulus L euckart in A frica. This taxonomy names cytoforms of Simulium damnosum T heobald s.l. based on chromosomal inversion polymorphisms. Researchers have linked variation in cytoforms to variation in geographical distributions and biological traits, but have failed to demonstrate that many of these cytoforms are differentiated genetically or morphologically. This may be explained by recent divergence and incomplete lineage sorting of ancestral polymorphisms. Alternatively, it may be the outcome of ongoing gene flow, with inversions maintaining sets of co‐adapted alleles and selection driving inversion frequency differences across habitats. Here, our goals were to test species delimitations in the S. damnosum complex, and determine if the poor differentiation between these species is better explained by the ancestral polymorphisms or gene‐flow hypotheses. Geometric morphometric analysis and more intensive genetic sampling of populations in N igeria only reinforced the lack of clear divergence among S. damnosum complex cytoforms. By contrast, the first statistical estimates of evolutionary relationships based on chromosomal inversion polymorphism data revealed clear signs of divergence, which lends support to the ancestral polymorphisms hypothesis. But the latter analyses may be biased by non‐neutral evolution of inversion polymorphisms; more rigorous tests of cytoform‐based species hypothesis will require deeper sampling across populations and neutral genetic markers along with explicit modelling of genetic isolation and migration.