Anopheles arabiensis and An. gambiae chromosomal inversion polymorphism, feeding and resting behaviour in relation to insecticide house-spraying in Tanzania

Abstract. Differential responses of the mosquitoes Anopheles arabiensis and An. gambiae sensu stricto to house-spraying with DDT or lambda-cyhalothrin were evaluated in relation to chromosomal inversion polymorphism, feeding and resting behaviour of these malaria vectors in Tanzania. Blood-fed mosquitoes from pit traps outdoors, exit traps on windows and indoor-resting catches were identified cytogenetically and the chromosomal inversion frequencies compared between samples and species. Their outdoor-resting behaviour was assessed by a mark–release–recapture experiment and by determining the proportion of freshly blood-fed individuals in exit traps. The source of bloodmeals was analysed by an ELISA method. Endophagic females of An. arabiensis were more likely than those of An. gambiae to exit from a house on the night of blood-feeding. Only in one out of three villages was there evidence that chromosomally distinct individuals within a species had different preferences for resting sites. There were indications, but not conclusive evidence, that mosquitoes caught indoors or outdoors had a tendency to return to the same type of resting site. In villages sprayed with either insecticide, the mean age of the vector populations was greatly reduced, compared with those in the unsprayed villages. An. arabiensis females exited from DDT sprayed houses after blood-feeding, whereas with lambda-cyhalothrin those exiting were mostly unfed and there was a decline in the human blood index. The excitorepellency of DDT was perceived as a disadvantage, whereas lambda-cyhalothrin apparently had more impact on malaria transmission by An. arabiensis.     

Host selection by Anopheles arabiensis and An. quadriannulatus feeding on cattle in Zimbabwe

In the Zambezi valley, mosquito females of the Anopheles gambiae Giles complex (Diptera: Culicidae) were collected from a hut containing pairs of cattle distinguishable by known DNA markers. DNA was extracted from the blood-fed mosquito abdomens and primer sets for ungulate and mosquito DNA loci were used to identify the mosquito sibling species and individual host source(s) of their bloodmeals. The 67 mosquitoes comprised a mixture of An. arabiensis Patton (31%) and An. quadriannulatus Theobald (69%). DNA from one or both of the cattle present in the hut was detected in 91% of samples. When the hut contained an adult and a calf, the percentage of bloodmeals from the adult, the calf and adult + calf were 58%, 27% and 15%, respectively; the trend towards meals from the adult host was consistent but not always significant. When the pair of cattle comprised two adults of roughly equal size and age, then mosquitoes generally showed no significant bias towards feeding from one individual. There was no significant difference in the pattern of host selection made by An. arabiensis and An. quadriannulatus but the former had a significantly higher percentage (20%) of mixed meals than An. quadriannulatus (9%). These two members of the An. gambiae complex appear to be less selective in their choice of cattle hosts compared to day-active Diptera such as tsetse and Stomoxys, possibly because the hosts are generally asleep when Anopheles are active and there is therefore less selective pressure to adapt to host defensive behaviour. The slight bias of Anopheles towards older and/or larger cattle may be related to the host's larger surface area.     

Cytogenetics of the Anopheles gambiae complex in Sudan, with special reference to An. arabiensis: relationships with East and West African populations

The species composition of malaria vector mosquitoes belonging to the Anopheles gambiae complex (Diptera: Culicidae) from >40 localities in Sudan, representing most ecological situations, was determined by analysis of ovarian polytene chromosomes. Of 2162 females, 93% were identified as An. arabiensis Patton and 7% were An. gambiae Giles sensu stricto. No hybrids were found between the two species. Anopheles arabiensis occurred in all but two sites, whereas An. gambiae s.s. was effectively limited to the southernmost, more humid localities. For chromosomal paracentric inversions, the degree of polymorphism was low in An. gambiae s.s. (inversions 2La, 2Rb and 2Rd), higher in An. arabiensis (inversions Xe, 2Ra, b, bc, d1, s; 3Ra, d). Anopheles gambiae samples from Sudan were all apparently panmictic, i.e. they did not show restricted gene flow such as observed among West African populations (interpreted as incipient speciation). Chromosomal inversion patterns of An. gambiae in southern Sudan showed characteristics of intergrading Savanna/Forest populations similar to those observed in comparable eco-climatic situations of West Africa. Anopheles arabiensis was polymorphic for inversion systems recorded in West Africa (2Ra, 2Rb, 2Rdl, 3Ra) and for a novel 2Rs polymorphism, overlapping with inversion systems 2Rb and 2Rd1. Samples carrying the 2Rs inversion were mostly from Khashm-el-Girba area in central-eastern Sudan. In the great majority of the samples all polymorphic inversions were found to be in Hardy-Weinberg equilibrium. Sudan populations of An. arabiensis should therefore be considered as generally panmictic. Anopheles arabiensis shows more inversion polymorphism in west than in east African populations. Sudan populations have more evident similarities with those from westwards than those from eastwards of the Great Rift Valley. The possible influence of the Rift on evolution of An. arabiensis is discussed.     

The impact of permethrin-impregnated bednets on malaria vectors of the Kenyan coast

Abstract. The effects of introducing permethrin-impregnated bednets on local populations of the malaria vector mosquitoes Anopheles funestus and the An.gambiae complex was monitored during a randomized controlled trial at Kilifi on the Kenyan coast. Pyrethrum spray collections inside 762 households were conducted between May 1994 and April 1995 after the introduction of bednets in half of the study area. All-night human bait collections were performed in two zones (one control and one intervention) for two nights each month during the same period. PCR identifications of An.gambiae sensu lato showed that proportions of sibling species were An.gambiae sensu stricto > An.merus > An.arabiensis.
Indoor-resting densities of An.gambiae s.l. and the proportion of engorged females decreased significantly in intervention zones as compared to control zones. However, the human blood index and Plasmodium falciparum sporozoite rate remained unaffected. Also vector parous rates were unaltered by the intervention, implying that survival rates of malaria vectors were not affected. The human-biting density of An.gambiae s.l. , the predominant vector, was consistently higher in the intervention zone compared to the control zone, but showed 8% reduction compared to pre-intervention biting rates - versus 94% increase in the control zone.
Bioassay, susceptibility and high-performance liquid chromatography results all indicated that the permethrin content applied to the nets was sufficient to maintain high mortality of susceptible vectors throughout the trial. Increased rates of early outdoor-biting, as opposed to indoor-biting later during the night, were behavioural or vector composition changes associated with this intervention, which would require further monitoring during control programmes employing insecticide-treated bednets.     

Cannibalism and predation among larvae of the Anopheles gambiae complex

Among the aquatic developmental stages of the Anopheles gambiae complex (Diptera: Culicidae), both inter- and intra-specific interactions influence the resulting densities of adult mosquito populations. For three members of the complex, An. arabiensis Patton, An. quadriannulatus (Theobald) and An. gambiae Giles sensu stricto, we investigated some aspects of this competition under laboratory conditions. First-instar larvae were consumed by fourth-instar larvae of the same species (cannibalism) and by fourth-instar larvae of other sibling species (predation). Even when larvae were not consumed, the presence of one fourth-instar larva caused a significant reduction in development rate of first-instar larvae. Possible implications of these effects for population dynamics of these malaria vector mosquitoes are discussed.     

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.     

Identification and expression of odorant-binding proteins of the malaria-carrying mosquitoes Anopheles gambiae and Anopheles arabiensis

Host preference and blood feeding are restricted to female mosquitoes. Olfaction plays a major role in host-seeking behaviour, which is likely to be associated with a subset of mosquito olfactory genes. Proteins involved in olfaction include the odorant receptors (ORs) and the odorant-binding proteins (OBPs). OBPs are thought to function as a carrier within insect antennae for transporting odours to the olfactory receptors. Here we report the annotation of 32 genes encoding putative OBPs in the malaria mosquito Anopheles gambiae and their tissue-specific expression in two mosquito species of the Anopheles complex; a highly anthropophilic species An. gambiae sensu stricto and an opportunistic, but more zoophilic species, An. arabiensis. RT-PCR shows that some of the genes are expressed mainly in head tissue and a subset of these show highest expression in female heads. One of the genes (agCP1588) which has not been identified as an OBP, has a high similarity (40%) to the Drosophila pheromone-binding protein 4 (PBPRP4) and is only expressed in heads of both An. gambiae and An. arabiensis, and at higher levels in female heads. Two genes (agCP3071 and agCP15554) are expressed only in female heads and agC15554 also shows higher expression levels in An. gambiae. The expression profiles of the genes in the two members of the Anopheles complex provides the first step towards further molecular analysis of the mosquito olfactory apparatus.     

Effects of adult body size on fecundity and the pre-gravid rate of Anopheles gambiae females in Tanzania

Abstract. The influence of adult body size on the pre-gravid state and fecundity was studied in Anopheles gambiae Giles females hand-caught inside houses and virgin females collected as pupae in Tanzania. Blood-fed mosquitoes were kept for 2-3 days before dissection and examination for insemination and ovarian condition. Those females which did not develop eggs were classified as pre-gravid. The number of mature eggs in those mosquitoes which became gravid was counted. Virgin females were fed and kept for egg maturation in the laboratory. Wing-length of females was measured as an index of mosquito size. The overall pre-gravid rate in the resting An. gambiae population was found to be 21% and, of these, 66% had been inseminated. In the virgin females the pre-gravid rate was 92.6%. The mean wing-length of wild females which became gravid was significantly larger than those which remained pre-gravid. There was a positive correlation between fecundity and wing-length. Smaller females tended to require two or three bloodmeals to facilitate completion of the first gonotrophic cycle. The critical size permitting oviposition from the first blood-meal was a wing-length of 3 mm.     

The relationship between female body size and survival rate of the malaria vector Anopheles arabiensis in Ethiopia

Abstract. The relationship between female mosquito body size and survival rate was studied in field populations of Anopheles arabiensis in the Awash valley, central Ethiopia. Body size was quantified by measuring the wing-length. Highly significant correlations were found between size, parousness and insemination. It was concluded that larger An.arabiensis females have a higher probability of survival, being inseminated and producing more egg batches than smaller adults. Implications for vectorial capacity and vector competence of mosquitoes are discussed.     

Electroantennogram and behavioural responses of the malaria vector Anopheles gambiae to human-specific sweat components

Afrotropical malaria vectors of the Anopheles gambiae complex (Diptera: Culicidae), particularly An. gambiae sensu stricto, are attracted mainly to human hosts. A major source of human volatile emissions is sweat, from which key human-specific components are the carboxylic acids (E)- and (Z)-3-methyl-2-hexenoic acid and 7-octenoic acid. Electrophysiological studies on the antennae of An. gambiae s.s. showed selective sensitivity to these compounds, with a threshold at 10(-6) g comparable to that of known olfactory stimulants 1-octen-3-ol, p-cresol, isovaleric acid, and lower than threshold sensitivity to L-lactic acid and the synthetic mosquito repellent N,N-diethyltoluamide (DEET). A combination of the acids released at concentrations > 10(-5) g in wind tunnel bioassays significantly reduced the response to CO2, the major attractant released by human hosts, for strains of An. gambiae s.s. originating from East and West Africa. Field trials with odour-baited entry traps (OBETs) in Burkina Faso showed that 7-octenoic acid significantly increased (by 1.7-fold) the catch of females of An. gambiae sensu lato (comprising two sibling species: An. arabiensis Patton and An. gambiae s.s.) in OBETs baited with CO2, whereas combinations of the acids significantly reduced the catch in CO2-baited traps (by 2.1-fold) and in whole human odour-baited traps (by 1.5-fold). The pure (E) and (Z) geometric isomers of 3-methyl-2-hexenoic acid gave comparable results to the (EIZ) isomer mixture. These results provide the first experimental evidence that human-specific compounds affect the behaviour of highly anthropophilic An. gambiae s.l. mosquitoes. The compounds appear to inhibit the upwind flight' response to known long-range attractants, and may serve either to mask' the attractants present or, more probably, to 'arrest' upwind flight when mosquitoes arrive at a host under natural conditions. In the final approach to hosts, vectors are known to reduce their flight speed and increase their turning rate, to avoid overshooting the source. In our experimental apparatus, these changes in flight behaviour would reduce the number of mosquitoes entering the ports of the collection devices.     

Interspecific competition between sibling species larvae of Anopheles arabiensis and An. gambiae

Mosquito larvae of the sibling species Anopheles arabiensis Patton and An. gambiae Giles sensu stricto (Diptera: Culicidae) were investigated for interspecific competition. Single-species and mixed-species populations were reared at 27 degrees C from the first instar to pupation at different densities (100, 200 or 400 larvae/200 cm2 tray) with a constant amount of food, 0.2 mg/larva/day. Pupae obtained from mixed populations were identified to species using PCR. Both species had a 1:1 sex ratio at pupation. Development time to pupation averaged about one day less for An. arabiensis compared to An. gambiae, ranging from 0.93-1.49d for males and from 0.44-0.84 d for females in single populations. In mixed species populations the difference for males ranged from 0.99-1.58d and for females from 0.93-1.62d. Survival rates of An. gambiae s.s. were significantly higher than those of An. arabiensis in both the single-species and mixed-species populations. Mixed-species rearing did not have an effect on the survival of An. gambiae, whereas the mortality rate of An. arabiensis was significantly higher in mixed populations than when only this species was reared at the same densities, suggesting a competitive disadvantage for An. arabiensis in mixed populations. High proportions of larvae (4-35%) were lost during development; these losses could not be accounted for by corpses found in the rearing pans. The possibility of cannibalism was investigated by rearing each species separately in small containers (five per 50 ml), inspected every 6h, but no cannibalism was detected at any stage of development in either species. It was concluded that, under these experimental circumstances, interspecific competition between both species did occur but with a detrimental effect on An. arabiensis only. Relevance of these findings to the ecology of both species in the field is discussed briefly.     

Zoophily of Anopheles arabiensis and An. gambiae in Madagascar demonstrated by odour-baited entry traps

In Madagascar we used odour-baited entry traps (OBETs) for host choice tests of wild female anopheline mosquitoes (Diptera: Culicidae) at representative localities on the East and West sides of the island (villages Fenoarivo and Tsararano, respectively) and at the southern margin of the central plateau (Zazafotsy village, 800 m altitude). No insecticide house-spraying operations have been undertaken at these villages. Odours from a man and a calf of similar mass, concealed in different tents, were drawn by fans into separate OBETs set side by side. Traps were alternated to compensate for position effects, and different pairs of individual baits were employed for successive replicates. Totals of 266 An. funestus Giles sensu stricto and 362 An. gambiae Giles sensu lato were collected in 48 trap nights during March-June 1999. For each mosquito species the 'index of anthropophily' was defined as the proportion of females caught in the human-baited trap. For An. funestus this index was found to be consistently greater than 0.5 (value for random choice between traps/hosts), indicating that this species 'preferred' human to calf odour (index=0.83). Conversely, the index of anthropophily for An. gambiae s.l. indicated they 'chose' calf in preference to human odour (index=0.26). No significant differences of relative preference for calf or man were detected between villages; geographical variance accounted for <8% of the total experimental variance. Molecular identifications of 181 specimens of the An. gambiae complex (approximately 50% of the samples) revealed only An. arabiensis Patton at Tsararano and Zazafotsy, but >97% An. gambiae Giles sensu stricto at Fenoarivo, in accordance with prior knowledge of the differential distributions of these sibling species on the island. Predominant zoophily (i.e. intrinsic 'preference' for cattle odours) by both An. arabiensis and An. gambiae s.s. in Madagascar contrasts with their greater anthropophily in continental Africa.     

Use of a male-specific DNA probe to distinguish female mosquitoes of the Anopheles gambiae species complex

Abstract. A method has been developed to distinguish between mated female individuals of the sibling species Anopheles gambiae Giles sensu stricto and Anopheles arabiensis Patton. The DNA probe pAnal, reported by Gale & Crampton (1987a) to be useful for the specific identification of An.arabiensis males, is here shown to be sufficiently sensitive to deduce the species identity of inseminated females from the identity of the sperm contained within the spermatheca.     

Observations on the Anopheles gambiae complex in the Senegal River Basin, West Africa

1. Three sibling species of mosquitoes of the Anopheles gambiae complex are found in the Senegal River Basin: An. melas Theobald, An. gambiae Giles and An. arabiensis Patton. 2. An. melas is restricted to the river delta and environs where saltwater breeding places are present. 3. An. gambiae and An. arabiensis are sympatric in the study area; An. arabiensis predominates in coastal zones where it breeds also during the dry season; An. gambiae predominates in inland areas where breeding is mostly restricted to the rainy season (July-September). 4. An. arabiensis is chromosomally polymorphic all over the study area, with much variation of inversion frequencies, particularly for the 2Ra arrangement. 5. An. gambiae is characterized by a very high frequency of the 2Rb-2La inversion arrangement which is typical of the Savanna chromosomal form.     

Responses of Anopheles gambiae complex mosquitoes to the use of untreated bednets in The Gambia

Population dynamics of the Anopheles gambiae complex of malaria vector mosquitoes were studied in four small hamlets in The Gambia. Bednets were used to reduce man/vector contact in two of the hamlets. High densities of An. gambiae, sensu lato, were present for only 3-8 weeks during the rainy season, depending on the position of the hamlet within the study area. The proportions of blood-fed mosquitoes caught indoors (83.0%) and existing from houses (11.6%) were lower in hamlets where bednets were used than in hamlets without (96.5% and 33.1% respectively). Fewer of the blood-fed mosquitoes had fed on man in houses where people slept under bednets (68.2%) than in those without (81.5%). However, the average number of infective bites received by children was still greater than one a year in hamlets where bednets were used. Consequently bednets are considered unlikely to be an effective malaria control measure so long as they are untreated with insecticide.     

Responses of adult mosquitoes of two sibling species, Anopheles arabiensis and A. gambiae s.s. (Diptera: Culicidae), to high temperatures

It is well known that amongst the sibling species of the Anopheles gambiae complex, A. arabiensis Patton predominates over A. gambiae sensu stricto Giles in hotter, drier parts of Africa. Here it was investigated whether A. arabiensis is better adapted to higher temperatures than A. gambiae s.s. at the microclimatic level. Bioassays were used to assess behavioural avoidance activity of adult mosquitoes in the presence of increasing temperature. Female mosquitoes were introduced into a holding tube from which they could escape into a cage through a one-way funnel. From a starting temperature of 28 degrees C they were exposed to a 2 degrees C rise in temperature every 30 min until all mosquitoes had escaped or been knocked down. As temperature increased, A. arabiensis left the holding tube at higher temperatures than A. gambiae s.s. (A. arabiensis mean activation temperature = 35.7 degrees C, 95% CIs = 35.4-36.1 degrees C; A. gambiae s.s. = 33.0 degrees C, 32.5-33.5 degrees C). To determine the relative ability of both species to survive at extremely high temperatures, batches of insects were exposed to 40 degrees C for different periods. It took considerably longer to kill 50% of A. arabiensis at 40 degrees C than it did A. gambiae s.s. (112 min vs. 67 min). These data show that adult A. arabiensis are better adapted to hotter conditions than A. gambiae s.s., a characteristic that is reflected in their spatial and temporal distribution in Africa.     

Impact of permethrin-treated bednets on malaria transmission by the Anopheles gambiae complex in The Gambia

Malaria vector mosquitoes belonging to the Anopheles gambiae complex were studied in four hamlets in The Gambia. All inhabitants were given bednets treated either with a placebo (milk) in two hamlets or with the pyrethroid insecticide permethrin (500 mg/m2) in two other hamlets. Malaria transmission occurred mainly during a few weeks of the rainy season, in September and October 1987. The indoor resting densities of mosquitoes in permethrin-treated hamlets were reduced, and we estimated over 90% reduction in biting on man by An. gambiae Giles sensu stricto in these hamlets. No mosquitoes were found under permethrin-treated bednets compared with eighty-one recovered from placebo-treated bednets. Mosquitoes exited more readily from rooms where permethrin-treated bednets were used than from rooms with placebo-treated nets. The annual mean probability that a child would receive an infective bite was estimated to be 0.09 in hamlets with insecticide-treated bednets, compared with 1.9 where placebo-treated bednets were used. Permethrin-treated bednets are therefore recommended as a means of effectively reducing the risk of exposure to malaria transmission, particularly in areas of low seasonal transmission.     

Mark–release–recapture experiments with Anopheles gambiae s.l. in Banambani Village, Mali, to determine population size and structure

Mark–release–recapture experiments with Anopheles gambiae s.l. were performed during the wet seasons of 1993 and 1994 in Banambani, Mali. All recaptured mosquitoes were identified to species by PCR analysis and, when possible, by chromosomal analysis to chromosomal form. Two species of the An. gambiae complex were present: An. gambiae s.s. and An. arabiensis ; their ratio differed greatly from one year to the next. Three chromosomal forms of An. gambiae s.s. were found – Bamako, Savanna and Mopti. The drier 1993 was characterized by a high frequency of An. arabiensis and of the Mopti chromosomal forms of An. gambiae s.s. These trends were consistent with large-scale geographical patterns of abundance along a precipitation gradient. We observed no difference in dispersal between the two species, nor among the chromosomal forms of An. gambiae s.s. Therefore, in this situation at least, it is reasonable to group such data on the An. gambiae complex as a whole for analysis. Population size of An. gambiae s.l. females in the village was estimated to be 9000–11 000 in 1993 and 28 000 in 1994. The corresponding numbers were somewhat higher when independently-derived values of daily survival were used. These were consistent with estimates of effective population size obtained from patterns of gene frequency change.     

Unexpected anthropophagic behaviour in Anopheles quadriannulatus

The strongly anthropophilic behaviour of Anopheles gambiae Giles sensu stricto (Diptera: Culicidae), the most important malaria vector in Africa, has been demonstrated by field and laboratory studies. Other members of the An. gambiae complex express varied degrees of anthropophily. Anopheles quadriannulatus (Theobald) species A and B are more zoophilic members of the complex and hence are considered to be of no medical importance. Olfactometer experiments with An. quadriannulatus species A have demonstrated attraction to both human and cow odour. To extend these olfactometer observations a choice experiment was conducted in an outdoor cage with a human and a calf as baits, using laboratory-reared mosquitoes. Anopheles gambiae s.s. (from Liberia) and two strains of An. quadriannulatus species A (SKUQUA from South Africa, SANGQUA from Zimbabwe), marked with different coloured fluorescent powders for identification purposes, were released simultaneously and given an equal opportunity to feed on either host. The experiment was repeated six times. Bloodmeals were identified using the precipitin technique. Anopheles gambiae s.s. showed highly anthropophagic behaviour, taking 88% of bloodmeals from the human host. In contrast, both strains of An. quadriannulatus fed with equal frequency on the human or the calf; the response to either host was not significantly different. These results confirm the olfactometer findings and demonstrate anthropophagic behaviour not previously recorded in this species. This finding has implications for prospective manipulation of host preference for genetic control purposes.     

Reduced susceptibility to DDT in field populations of Anopheles quadriannulatus and Anopheles arabiensis in Malawi: evidence for larval selection

Abstract.  Bioassays for insecticide resistance in adult mosquitoes were conducted on samples of Anopheles gambiae Giles s.l . (Diptera: Culicidae) species collected as larvae from breeding sites in the lower Shire Valley, Malawi. The results indicate full susceptibility to permethrin, deltamethrin and malathion, but reduced susceptibility to DDT in one sample from Thom (LT₅₀ of 8.39 min for females and 25.09 min for males). Polymerase chain reaction-based species identification of the mosquitoes assayed revealed a mixture of Anopheles arabiensis Patton and Anopheles quadriannulatus (Theobold). The LT₅₀ did not differ significantly between species. Genotyping of the L1014F and L1014S kdr alleles showed all mosquito specimens to be homozygous wild type; thus the reduced susceptibility detected is not attributable to target site insensitivity and instead is likely to be metabolic in nature. Anopheles quadriannulatus is characteristically zoophagic and exophilic. Indeed, of 82 Anopheles collected through knockdown collections within dwellings, only one was An. quadriannulatus and the rest were An. arabiensis . They are unlikely, therefore, to have been exposed to selection pressure arising from insecticide-treated net usage or to DDT indoor residual spraying. Therefore, it is suggested that this example of reduced susceptibility to DDT in An. quadriannulatus reflects selection in the larval stages.