Rangewide population genetic structure of the African malaria vector Anopheles funestus

Anopheles funestus is a primary vector of malaria in Africa south of the Sahara. We assessed its rangewide population genetic structure based on samples from 11 countries, using 10 physically mapped microsatellite loci, two per autosome arm and the X (N = 548), and 834 bp of the mitochondrial ND5 gene (N = 470). On the basis of microsatellite allele frequencies, we found three subdivisions: eastern (coastal Tanzania, Malawi, Mozambique and Madagascar), western (Burkina Faso, Mali, Nigeria and western Kenya), and central (Gabon, coastal Angola). A. funestus from the southwest of Uganda had affinities to all three subdivisions. Mitochondrial DNA (mtDNA) corroborated this structure, although mtDNA gene trees showed less resolution. The eastern subdivision had significantly lower diversity, similar to the pattern found in the codistributed malaria vector Anopheles gambiae. This suggests that both species have responded to common geographic and/or climatic constraints. The western division showed signatures of population expansion encompassing Kenya west of the Rift Valley through Burkina Faso and Mali. This pattern also bears similarity to A. gambiae, and may reflect a common response to expanding human populations following the development of agriculture. Due to the presumed recent population expansion, the correlation between genetic and geographic distance was weak. Mitochondrial DNA revealed further cryptic subdivision in A. funestus, not detected in the nuclear genome. Mozambique and Madagascar samples contained two mtDNA lineages, designated clade I and clade II, that were separated by two fixed differences and an average of 2% divergence, which implies that they have evolved independently for approximately 1 million years. Clade I was found in all 11 locations, whereas clade II was sampled only on Madagascar and Mozambique. We suggest that the latter clade may represent mtDNA capture by A. funestus, resulting from historical gene flow either among previously isolated and divergent populations or with a related species.     

Distribution of a knockdown resistance mutation (L1014S) in Anopheles gambiae s.s. and Anopheles arabiensis in western and southern Kenya

In Kenya, insecticide-treated mosquito nets (ITNs) distributed to pregnant women and children under 5 years old through various programs have resulted in a significant reduction in malaria deaths. All of the World Health Organization-recommended insecticides for mosquito nets are pyrethroids, and vector mosquito resistance to these insecticides is one of the major obstacles to an effective malaria control program. Anopheles gambiae s.s. and Anopheles arabiensis are major malaria vectors that are widely distributed in Kenya. Two point mutations in the voltage-gated sodium channel (L1014F and L1014S) are associated with knockdown resistance (kdr) to DDT and pyrethroids in An. gambiae s.s. While the same point mutations have been reported to be rare in An. arabiensis, some evidence of metabolic resistance has been reported in this species. In order to determine the distribution of the point mutation L1014S in An. gambiae s.s. and An. arabiensis in southern and western Kenya, we collected larvae and screened for the mutation by DNA sequencing. We found high allelic and homozygous frequencies of the L1014S mutation in An. gambiae s.s. The L1014S mutation was also widely distributed in An. arabiensis, although the allelic frequency was lower than in An. gambiae s.s. The same intron sequence (length: 57 base) found in both species indicated that the mutation was introgressed by hybridization. The allelic frequency of L1014S was higher in both species in western regions, demonstrating the strong selection pressure imposed by long-lasting insecticide-treated nets (LLITN)/ITN on the An. gambiae s.s. and An. arabiensis populations in those areas. The present contribution of the L1014S mutation to pyrethroid resistance in An. arabiensis may be negligible. However, the homozygous frequency could increase with continuing selection pressure due to expanded LLITN coverage in the future.     

Multimodal pyrethroid resistance in malaria vectors, Anopheles gambiae s.s., Anopheles arabiensis, and Anopheles funestus s.s. in western Kenya

Anopheles gambiae s.s., Anopheles arabiensis, and Anopheles funestus s.s. are the most important species for malaria transmission. Pyrethroid resistance of these vector mosquitoes is one of the main obstacles against effective vector control. The objective of the present study was to monitor the pyrethroid susceptibility in the 3 major malaria vectors in a highly malaria endemic area in western Kenya and to elucidate the mechanisms of pyrethroid resistance in these species. Gembe East and West, Mbita Division, and 4 main western islands in the Suba district of the Nyanza province in western Kenya were used as the study area. Larval and adult collection and bioassay were conducted, as well as the detection of point mutation in the voltage-gated sodium channel (1014L) by using direct DNA sequencing. A high level of pyrethroid resistance caused by the high frequency of point mutations (L1014S) was detected in An. gambiae s.s. In contrast, P450-related pyrethroid resistance seemed to be widespread in both An. arabiensis and An. funestus s.s. Not a single L1014S mutation was detected in these 2 species. A lack of cross-resistance between DDT and permethrin was also found in An. arabiensis and An. funestus s.s., while An. gambiae s.s. was resistant to both insecticides. It is noteworthy that the above species in the same area are found to be resistant to pyrethroids by their unique resistance mechanisms. Furthermore, it is interesting that 2 different resistance mechanisms have developed in the 2 sibling species in the same area individually. The cross resistance between permethrin and DDT in An. gambiae s.s. may be attributed to the high frequency of kdr mutation, which might be selected by the frequent exposure to ITNs. Similarly, the metabolic pyrethroid resistance in An. arabiensis and An. funestus s.s. is thought to develop without strong selection by DDT.     

Dynamics of gene introgression in the African malaria vector Anopheles gambiae

Anopheles gambiae is a major malaria vector in Africa and a popular model species for a variety of ecological, evolutionary, and genetic studies on vector control. Genetic manipulation of mosquito vectorial capacity is a promising new weapon for the control of malaria. However, the release of exotic transgenic mosquitoes will bring in novel alleles in addition to the parasite-inhibiting genes, which may have unknown effects on the local population. Therefore, it is necessary to develop methodologies that can be used to evaluate the spread rate of introduced genes in A. gambiae. In this study, the effects and dynamics of genetic introgression between two geographically distinct A. gambiae populations from western Kenya (Mbita) and eastern Tanzania (Ifakara) were investigated with amplified fragment length polymorphisms (AFLPs) and microsatellite markers. Microsatellites and polymorphic cDNA markers revealed a large genetic differentiation between the two populations (average F(ST) = 0.093, P < 0.001). When the two strains were crossed in random mating between the two populations, significant differences in the rate of genetic introgression were found in the mixed populations. Allele frequencies of 18 AFLP markers (64.3%) for Mbita and of 26 markers (92.9%) for Ifakara varied significantly from F5 to F20. This study provides basic information on how a mosquito release program would alter the genetic makeup of natural populations, which is critical for pilot field testing and ecological risk evaluation of transgenic mosquitoes.     

Population Structure of Anopheles gambiae in Africa

The population structure of Anopheles gambiae in Africa was studied using 11 microsatellite loci in 16 samples from 10 countries. All loci are located outside polymorphic inversions. Heterogeneity among loci was detected and two putative outlier loci were removed from analyses aimed at capturing genome-wide patterns. Two main divisions of the gene pool were separated by high differentiation (F(ST) > 0.1). The northwestern (NW) division included populations from Senegal, Ghana, Nigeria, Cameroon, Gabon, Democratic Republic of Congo (DRC), and western Kenya. The southeastern (SE) division included populations from eastern Kenya, Tanzania, Malawi, and Zambia. Inhospitable environments for A. gambiae along the Rift Valley partly separate these divisions. Reduced genetic diversity in the SE division and results of an analysis based on private alleles support the hypothesis that a recent bottleneck, followed by colonization from the NW populations shaped this structure. In the NW division, populations possessing the M rDNA genotype appeared to form a monophyletic clade. Although genetic distance increased with geographic distance, discontinuities were suggested between certain sets of populations. The absence of heterozygotes between sympatric M and S populations in the DRC and the high differentiation in locus 678 (F(ST)>0.28) contrasted with low differentiation in all other loci (-0.02相似文献   

Flight tone of field‐collected populations of Anopheles gambiae and An. arabiensis (Diptera: Culicidae)

Abstract.Laboratory colonies of the human malaria vectors Anopheles gambiae Giles and An. arabiensis Patton have distinct flight tones. If flight tone similarly distinguishes natural populations of these sympatric sibling species, it may play a role in reproductive isolation of swarms that are otherwise behaviourally identical. To assess the fidelity of flight tone differences in natural populations, flight tone was measured in the F1 progeny of mosquitoes of both species captured in western Kenya. Flight tone distributions of wild An . gambiae and An. arabiensis were similar to their laboratory conspecifics. However, interspecies comparisons of flight tone of wild mosquitoes revealed significantly different but overlapping distributions for both sexes. Furthermore, when the effect of body size on flight tone was determined, there was a positive correlation between wing length and flight tone for both sexes of An. gambiae and An. arabiensis , suggesting that mosquito size is a significant variable affecting flight tone. Although these findings diminish any practical benefit of flight tone as a diagnostic tool in species identification, its potential role in pre‐mating species recognition needs further investigation.     

The association between the phytoplankton, Rhopalosolen species (Chlorophyta; Chlorophyceae), and Anopheles gambiae sensu lato (Diptera: Culicidae) larval abundance in western Kenya

Algae are important food resources of the larvae of the African malaria vectors, Anopheles gambiae Giles and Anopheles arabiensis Patton (Anopheles gambiae sensu lato), and other zooplankton, but empirical evidence remains meager about the agal flora in ephemeral water bodies. The animals present in natural aquatic habitats in western Kenya were sampled from July to November 2002 to study abiotic and biotic environmental factors determining A. gambiae sl larval abundance. The five highest concentrations of third and fourth instars and pupae (hereafter referred to as old-stage larvae) were sampled in conjunction with the unicellular epizoic green algae, Rhopalosolen species (Chlorophyta; Chlorophyceae). Canonical correspondence analysis revealed that the presence of Rhopalosolen species was the most important determinant of the animal assemblage. The density of old-stage A. gambiae sl larvae was positively correlated with the presence of Rhopalosolen species, but the density of first and second instars of A. gambiae sl was not. The water bodies with Rhopalosolen sp. yielded larger mosquitoes in spite of the higher density of larvae. We demonstrated that the productivity of water bodies in terms of the larvae of malaria vectors can differ in magnitude depending on the agal flora. We discuss phytoplankton as a regulator of mosquito larval populations.     

Revisiting the role of introgression vs shared ancestral polymorphisms as key processes shaping genetic diversity in the recently separated sibling species of the Anopheles gambiae complex

The role of interspecific hybridisation in the evolution of pest species is poorly understood. In mosquito disease vectors this is of particular importance due to the evolution of insecticide resistance and the proposed release of transgenic strains that are refractory to the malaria parasite. In this study, we apply population genetic methods in a novel manner to determine whether mitochondrial DNA sequences have introgressed between the closely related African malaria vectors Anopheles gambiae and A. arabiensis. Our results suggest that speciation was geologically recent and ancestral haplotypes at the ND5 locus are retained in both species. In addition, comparing haplotype frequencies in allopatric and sympatric populations, suggest locale specific unidirectional introgression of mitochondria from A. arabiensis into A. gambiae.     

Distribution of the species of the Anopheles gambiae complex and first evidence of Anopheles merus as a malaria vector in Madagascar

BACKGROUND: Members of the Anopheles gambiae complex are amongst the best malaria vectors in the world, but their vectorial capacities vary between species and populations. A large-scale sampling of An. gambiae sensu lato was carried out in various bioclimatic domains of Madagascar. Local abundance of an unexpected member of this complex raised questions regarding its role in malaria transmission. METHODS: Sampling took place at 38 sites and 2,067 females were collected. Species assessment was performed using a PCR targeting a sequence in the IGS of the rDNA. Analysis focused on the relative prevalence of the species per site, bioclimatic domain and altitude. Infectivity of Anopheles merus was assessed using an ELISA to detect the presence of malarial circumsporozoite protein in the head-thorax. RESULTS: Three species were identified: An. gambiae, Anopheles arabiensis and An. merus. The distribution of each species is mainly a function of bioclimatic domains and, to a lesser extent, altitude. An. arabiensis is present in all bioclimatic domains with highest prevalence in sub-humid, dry and sub-arid domains. An. gambiae has its highest prevalence in the humid domain, is in the minority in dry areas, rare in sub-humid and absent in sub-arid domains. An. merus is restricted to the coastal fringe in the south and west; it was in the majority in one southern village. The majority of sites were sympatric for at least two of the species (21/38) and two sites harboured all three species.The role of An. merus as malaria vector was confirmed in the case of two human-biting females, which were ELISA-positive for Plasmodium falciparum. CONCLUSION: Despite the huge environmental (mainly man-made) changes in Madagascar, the distribution of An. gambiae and An. arabiensis appears unchanged for the past 35 years. The distribution of An. merus is wider than was previously known, and its effectiveness as a malaria vector has been shown for the first time; this species is now on the list of Malagasy malaria vectors.     

Linear and spatial organization of polytene chromosomes of the African malaria mosquito Anopheles funestus

Anopheles funestus Giles is one of the major malaria vectors in Africa, but little is known about its genetics. Lack of a cytogenetic map characterized by regions has hindered the progress of genetic research with this important species. This study developed a cytogenetic map of An. funestus using ovarian nurse cell polytene chromosomes. We demonstrate an important application with the cytogenetic map for characterizing various chromosomal inversions for specimens collected from coastal Kenya. The linear and spatial organization of An. funestus polytene chromosomes was compared with the best-studied malaria mosquito, An. gambiae Giles. Comparisons of chromosome morphology between the two species have revealed that the most extensive chromosomal rearrangement occurs in pericentromeric heterochromatin of autosomes. Differences in pericentromeric heterochromatin types correlate with nuclear organization differences between An. funestus and An. gambiae. Attachments of chromosomes to the nuclear envelope strongly depend on the presence of diffusive beta-heterochromatin. Thus, An. funestus and An. gambiae exhibit species-specific characteristics in chromosome-linear and -spatial organizations.     

Gut content identification of larvae of the Anopheles gambiae complex in western Kenya using a barcoding approach

Although larvae feeding and food source are vital to the development, survival and population regulation of African malaria vectors, the prey organisms of Anopheles gambiae larvae in the natural environment have not been well studied. This study used a molecular barcoding approach to investigate the natural diets of Anopheles gambiae s.l. larvae in western Kenya. Gut contents from third- and fourth-instar larvae from natural habitats were dissected and DNA was extracted. The 18S ribosomal DNA gene was amplified, the resulting clones were screened using a restriction fragment length polymorphism method and nonmosquito clones were sequenced. Homology search and phylogenetic analyses were then conducted using the sequences of non-mosquito clones to identify the putative microorganisms ingested. The phylogenetic analyses clustered ingested microorganisms in four clades, including two clades of green algae (Chlorophyta, Chlorophyceae Class, Chlamydomonadales and Chlorococcales families), one fungal clade, and one unknown eukaryote clade. In parallel, using the same approach, an analysis of the biodiversity present in the larval habitats was carried out. This present study demonstrated the feasibility of the barcoding approach to infer the natural diets of Anopheles gambiae larvae. Our analysis suggests that despite the wide range of microorganisms available in natural habitats, mosquito larvae fed on specific groups of algae. The novel tools developed from this study can be used to improve our understanding of the larval ecology of African malaria vectors and to facilitate the development of new mosquito control tools.     

Evidence for late Pleistocene population expansion of the malarial mosquitoes, Anopheles arabiensis and Anopheles gambiae in Nigeria

Anopheles gambiae Giles s.s. and Anopheles arabiensis Patton (Diptera: Culicidae) are major vectors of malaria in Nigeria. We used 1115 bp of the mitochondrial COI gene to assess their population genetic structures based on samples from across Nigeria (n = 199). The mtDNA neighbour-joining tree, based on F(ST) estimates, separated An. gambiae M and S forms, except that samples of An. gambiae M from Calabar clustered with all the An. gambiae S form. Anopheles arabiensis and An. gambiae could be combined into a single star-shaped, parsimonious haplotype network, and shared three haplotypes. Haplotype diversity values were high in An. arabiensis and An. gambiae S, and intermediate in An. gambiae M; all nucleotide diversities were relatively low. Taken together, patterns of haplotype diversity, the star-like genealogy of haplotypes, five of seven significant neutrality tests, and the violation of the isolation-by-distance model indicate population expansion in An. arabiensis and An. gambiae S, but the signal was weak in An. gambiae M. Selection is supported as an important factor shaping genetic structure in An. gambiae in Nigeria. There were two geographical subdivisions in An. arabiensis: one included all southern localities and all but two central localities; the other included all northern and two central localities. Re-analysing an earlier microsatellite dataset of An. arabiensis using a Bayesian method determined that there were two distinctive clusters, northern and southern, that were fairly congruent with the mtDNA subdivisions. There was a trend towards decreasing genetic diversity in An. arabiensis from the northern savannah to the southern rainforest that corroborated previous data from microsatellites and polytene chromosomes.     

Sex-linked differentiation between incipient species of Anopheles gambiae

Emerging species within the primary malaria vector Anopheles gambiae show different ecological preferences and significant prezygotic reproductive isolation. They are defined by fixed sequence differences in X-linked rDNA, but most previous studies have failed to detect large and significant differentiation between these taxa elsewhere in the genome, except at two other loci on the X chromosome near the rDNA locus. Hypothesizing that this pericentromeric region of the X chromosome may be accumulating differences faster than other regions of the genome, we explored the pattern and extent of differentiation between A. gambiae incipient species and a sibling species, A. arabiensis, from Burkina Faso, West Africa, at 17 microsatellite loci spanning the X chromosome. Interspecific differentiation was large and significant across the entire X chromosome. Among A. gambiae incipient species, we found some of the highest levels of differentiation recorded in a large region including eight independent loci near the centromere of the X chromosome. Outside of this region, no significant differentiation was detected. This pattern suggests that selection is playing a role in the emergence of A. gambiae incipient species. This process, associated with efficient exploitation of anthropogenic modifications to the environment, has public health implications as it fosters the spread of malaria transmission both spatially and temporally.     

Influence of soil quality in the larval habitat on development of Anopheles gambiae Giles.

Larval ecology is an important aspect of the population dynamics of anopheline mosquitoes (Diptera: Culicidae), the vectors of malaria. Anopheles larvae live in pools of stagnant water and adult fitness may be correlated with the nutritional conditions under which larvae develop. A study was conducted in Mbita, Western Kenya, to investigate how properties of the soil substrate of Anopheles gambiae breeding pools can influence development of this mosquito species. An. gambiae eggs from an established colony were dispensed into experimental plastic troughs containing soil samples from a range of natural Anopheles larval habitats and filtered Lake Victoria water. The duration of larval development (8-15 days), pupation rate (0-79%), and adult body size (20.28-26.91 mm3) varied among different soil types. The total organic matter (3.61-21.25%), organic carbon (0.63-7.18%), and total nitrogen (0.06-0.58%) levels of the soils were positively correlated with pupation rate and negatively correlated with development time and adult body size.     

Selective Introgression of Paracentric Inversions between Two Sibling Species of the Anopheles Gambiae Complex

The Anopheles gambiae complex includes the major vectors of malaria in sub-Saharan Africa where >80% of all world-wide cases occur. These mosquitoes are characterized by chromosomal inversions associated to the speciation process and to intraspecific ecological and behavioral flexibility. It has been postulated that introgressive hybridization has selectively transferred inversions on the second chromosome between A. gambiae and A. arabiensis, the two most important vectors of malaria. Here we directly test this hypothesis with laboratory experiments in which hybrid populations were established and the fate of chromosomal inversions were followed. Consistent with the hypothesis, ``foreign' X chromosomes were eliminated within two generations, while some ``foreign' second chromosomes persisted for the duration of the experiments and, judging from the excess of heterozygotes, established stable heterotic polymorphisms. Only those second chromosome inversions found naturally in the species could be introgressed.     

Topi, an IS630/Tc1/mariner-type transposable element in the African malaria mosquito, Anopheles gambiae

IS630/Tc1/mariner elements are diverse and widespread within insects. The African malaria mosquito, Anopheles gambiae, contains over 30 families of IS630/Tc1/mariner elements although few have been studied in any detail. To examine the history of Topi elements in An. gambiae populations, Topi elements (n=73) were sampled from five distinct populations of An. gambiae from eastern and western Africa and evaluated with respect to copy number, nucleotide diversity and insertion site-occupancy frequency. Topi 1 and 2 elements were abundant (10-34 per diploid genome) and highly diverse (pi=0.051). Elements from mosquitoes collected in Nigeria were Topi 2 elements and those from mosquitoes collected in Mozambique were Topi 1 elements. Of the 49 Topi transposase open reading frames sequenced none were found to be identical. Intact elements with complete transposase open reading frames were common, although based on insertion site-occupancy frequency data it appeared that genetic drift was the major force acting on these IS630/Tc1/mariner-type elements. Topi 3 elements were not recovered from any of the populations sampled in this study and appear to be rare elements in An. gambiae, possibly due to a recent introduction.     

A comparison of Anopheles gambiae and Plasmodium falciparum genetic structure over space and time

Population genetic structure and subdivision are key factors affecting the evolution of organisms. In this study, we analysed and compared the population genetic structure of the malaria parasite Plasmodium falciparum and its mosquito vector Anopheles gambiae over space and time in the Nianza Province, near Victoria Lake in Kenya. The parasites were collected from mosquitoes caught in six villages separated by up to 68 km in 2002 and 2003. A total of 545 oocysts were dissected from 122 infected mosquitoes and genotyped at seven microsatellite markers. Five hundred and forty-seven mosquitoes, both infected and uninfected, were genotyped at eight microsatellites. For the parasite and the vector, the analysis revealed no (or very little) genetic differentiation among villages. This may be explained by high local population sizes for the parasite and the mosquito. The small level of genetic differentiation observed between populations may explain the speed at which antimalarial drug resistance and insecticide resistance spread into the African continent.     

Mosquito behavioural aspects of vector-human interactions in the Anopheles gambiae complex

The behaviour of two of the most anthropophilic malaria vectors in the world, Anopheles gambiae Giles and An. arabiensis Patton, is revisited with respect to recent studies on their host preferences and the chemical ecology of host-seeking. Issues are discussed in relation to the ways anthropophily may have arisen in the complex, and the opportunities the study of olfaction and host-seeking behaviour offers to malaria control in Africa.     

Feeding and survival of the malaria vector Anopheles gambiae on plants growing in Kenya

The propensity of the malaria vector mosquito Anopheles gambiae Giles (Diptera: Culicidae) to ingest sugars from various plants, and subsequent survival rates, were assessed with laboratory-reared males and females offered eight species of plants commonly cultivated and/or growing wild in western Kenya. In cages (no-choice bioassay), mosquitoes given the opportunity to feed on castorbean (Ricinus communis L.) had the longest survival times (mean and median survival time of 6.99 +/- 0.23 and 5.67 +/- 0.17 days, respectively), comparable to mosquitoes given 6% glucose (mean and median survival time of 8.70 +/- 0.23 and 6.67 +/- 0.33 days, respectively). Survival rates of An. gambiae were low on the other plants, comparable to mosquitoes given only water. Three plants: sweet potato (Ipomoea batatas L.), wild sage (Lantana camara L.) and castorbean provided levels of sugar ingestion by both sexes of An. gambiae detectable using the cold anthrone method, showing a positive correlation between median survival and sugar consumption (Spearman rank correlation coefficient = 0.905, P < 0.0001). Equal numbers of males and females were released in an enclosed semi-field screenhouse system containing a range of local plants, but no host for blood, and allowed to feed ad libitum: 6.7 +/- 0.5% (11/64) of those recaptured were found to contain detectable fructose (all females). Common plants are clearly a viable source of nutrition for adult female An. gambiae, as well as males, and may constitute and important resource for this important malaria vector.