Emergence of behavioural avoidance strategies of malaria vectors in areas of high LLIN coverage in Tanzania.

Despite significant reductions in malaria transmission across Africa since 2000, progress is stalling. This has been attributed to the development of insecticide resistance and behavioural adaptations in malaria vectors. Whilst insecticide resistance has been widely investigated, there is poorer understanding of the emergence, dynamics and impact of mosquito behavioural adaptations. We conducted a longitudinal investigation of malaria vector host choice over 3 years and resting behaviour over 4 years following a mass long-lasting insecticidal nets (LLINs) distribution in Tanzania. By pairing observations of mosquito ecology with environmental monitoring, we quantified longitudinal shifts in host-choice and resting behaviour that are consistent with adaptation to evade LLINs. The density of An. funestus s.l., declined significantly through time. In tandem, An. arabiensis and An. funestus s.l. exhibited an increased rate of outdoor relative to indoor resting; with An. arabiensis reducing the proportion of blood meals taken from humans in favour of cattle. By accounting for environmental variation, this study detected clear evidence of intra-specific shifts in mosquito behaviour that could be obscured in shorter-term or temporally-coarse surveys. This highlights the importance of mosquito behavioural adaptations to vector control, and the value of longer-term behavioural studies.

Investigating associations between biting time in the malaria vector Anopheles arabiensis Patton and single nucleotide polymorphisms in circadian clock genes: support for sub-structure among An. arabiensis in the Kilombero valley of Tanzania.

BACKGROUND: There is growing evidence that the widespread use of Long-Lasting Insecticidal Nets (LLINs) is prompting malaria vectors to shift their biting towards times and places where people are not protected, such as earlier in the evening and/or outdoors. It is uncertain whether these behavioural shifts are due to phenotypic plasticity and/or ecological changes within vector communities that favour more exophilic species, or involve genetic factors within vector species to limit their contact with LLINs. Possibly variation in the time and location of mosquito biting has a genetic basis, but as yet this phenomenon has received little investigation. Here we used a candidate gene approach to investigate whether polymorphisms in selected circadian clock genes could explain variation in the time and location of feeding (indoors versus outside) within a natural population of the major African malaria vector Anopheles arabiensis. METHODS: Host-seeking An. arabiensis were collected from two villages (Lupiro and Sagamaganga) in Tanzania by Human Landing Catch (HLC) technique. Mosquitoes were classified into phenotypes of "early" (7 pm-10 pm) or "late" biting (4 am -7 am), and host-seeking indoors or outdoors. In these samples we genotyped 34 coding SNPs in 8 clock genes (PER, TIM, CLK, CYC, PDP1, VRI, CRY1, and CRY2), and tested for associations between these SNPs and biting phenotypes. SNPs in 8 mitochondrial genes (ATP6, ATP8, COX1, COX2, COX3, ND3, ND5 and CYTB) were also genotyped to test population subdivision within An. arabiensis. RESULTS: The candidate clock genes exhibited polymorphism within An. arabiensis, but it was unrelated to variation in the timing and location of their biting activity. However, there was evidence of strong genetic structure within An. arabiensis populations in association with the TIM, which was unrelated to geographic distance. Substructure within An. arabiensis was also detected using mitochondrial markers. CONCLUSIONS: The variable timing and location of biting in An. arabiensis could not be linked to candidate clock genes that are known to influence behaviour in other Diptera. This finding does not rule out the possibility of a genetic basis to biting behaviour in this malaria vector, but suggests these are complex phenotypes that require more intensive ecological, neuronal and genomic analyses to understand.

Comparative evaluation of the Sticky-Resting-Box-Trap, the standardised resting-bucket-trap and indoor aspiration for sampling malaria vectors.

BACKGROUND: Understanding mosquito resting behaviour is important for the control of vector-borne diseases, but this remains a challenge because of the paucity of efficient sampling tools. We evaluated two novel sampling methods in the field: the Sticky Resting Box (SRB) and the Resting Bucket trap (RBu) to test their efficiency for sampling malaria vectors resting outdoors and inside houses in rural Tanzania. The performance of RBu and SRB was compared outdoors, while indoors SRB were compared with the Back Pack Aspiration method (BP). Trapping was conducted within 4 villages in the Kilombero Valley, Tanzania over 14 nights. On each night, the performance for collecting Anopheles vectors and Culicinae was compared in 4 households by SRB and RBu outdoors and by SRB or fixed-time Back Pack aspirator in 2 of the 4 focal households indoors. FINDINGS: A total of 619 Anopheles gambiae s.l., 224 Anopheles funestus s.l. and 1737 Culicinae mosquitoes were captured. The mean abundance of An. arabiensis and An. funestus s.l. collected with SRB traps inside and outdoors was significantly lower than with BP or RBu. The SRB however, outperformed BP aspiration for collection of Culicinae indoors. CONCLUSIONS: Of the methods trialled indoors (BP and SRB), BP was the most effective, whilst outdoors RBu performed much better than SRB. However, as SRB can passively sample mosquitoes over a week they could provide an alternative to the RBu where daily monitoring is not possible.