Solicited Cough Sound Analysis for Tuberculosis Triage Testing: The CODA TB DREAM Challenge Dataset.

Cough is a common and commonly ignored symptom of lung disease. Cough is often perceived as difficult to quantify, frequently self-limiting, and non-specific. However, cough has a central role in the clinical detection of many lung diseases including tuberculosis (TB), which remains the leading infectious disease killer worldwide. TB screening currently relies on self-reported cough which fails to meet the World Health Organization (WHO) accuracy targets for a TB triage test. Artificial intelligence (AI) models based on cough sound have been developed for several respiratory conditions, with limited work being done in TB. To support the development of an accurate, point-of-care cough-based triage tool for TB, we have compiled a large multi-country database of cough sounds from individuals being evaluated for TB. The dataset includes more than 700,000 cough sounds from 2,143 individuals with detailed demographic, clinical and microbiologic diagnostic information. We aim to empower researchers in the development of cough sound analysis models to improve TB diagnosis, where innovative approaches are critically needed to end this long-standing pandemic.

Evaluation of the DN-Mini (miniaturized double net) trap for sampling host-seeking Anopheles mosquitoes in malaria-endemic villages of southern Tanzania.

BACKGROUND: Surveillance of malaria vectors is crucial for assessing the transmission risk and impact of control measures. Human landing catches (HLC) directly estimate the biting rates but raise ethical concerns due to the exposure of volunteers to mosquito-borne pathogens. A common alternative is the CDC-light trap, which is effective for catching host-seeking mosquitoes indoors but not outdoors. New, exposure-free methods are needed for sampling mosquitoes indoors and outdoors in ways that reflect their natural risk profiles. The aim of this study was therefore to evaluate the efficacy of the miniaturized double net trap (DN-Mini) for sampling host-seeking mosquitoes in south-eastern Tanzania, where malaria transmission is dominated by Anopheles funestus. METHODS: Adult mosquitoes were collected from 222 randomly selected houses across three villages (74 per village) in Ulanga district, south-eastern Tanzania, using the DN-Mini traps, CDC-Light traps, and Prokopack aspirators. First, we compared CDC-light and DN-Mini traps for collecting indoor host-seeking mosquitoes, while Prokopack aspirators were used for indoor-resting mosquitoes. Second, we deployed the DN-Mini and Prokopack aspirators to collect host-seeking and resting mosquitoes indoors and outdoors. Generalized linear mixed models (GLMM) with a negative binomial distribution were used to compare the effectiveness of the traps for catching different mosquito species. RESULTS: The DN-Mini was 1.53 times more efficient in collecting An. funestus indoors (RR = 1.53, 95% CI: 1.190-1.98) compared to the CDC-Light trap. However, for Anopheles arabiensis, the DN-Mini caught only 0.32 times as many mosquitoes indoors as the CDC-Light traps (RR = 0.32, 95% CI: 0.183-0.567). Both An. funestus and An. arabiensis were found to be more abundant indoors than outdoors when collected using the DN-Mini trap. Similarly, the Prokopack aspirator was greater indoors than outdoors for both An. funestus and An. arabiensis. CONCLUSION: The DN-Mini outperformed the CDC-light trap in sampling the dominant malaria vector, An. funestus species, but was less effective in capturing An. arabiensis, and for both vector species, the biting risk was greater indoors than outdoors when measured using the DN-Mini trap. These findings highlight the importance of selecting appropriate trapping methods based on mosquito species and behaviors.

Passive Outdoor Host Seeking Device (POHD): Designing and Evaluation against Outdoor Biting Malaria Vectors.

Odor-baited devices are increasingly needed to compliment long-lasting insecticidal nets (LLINs) and indoor residual spraying (IRS) for control of residual malaria transmission. However, the odor-baited devices developed so far are bulky, dependent on the source of electricity and carbon dioxide (CO2), and they are logistically unsuitable for scaling up in surveillance and control of malaria vectors. We designed a passive and portable outdoor host seeking device (POHD) and preliminarily evaluated suitable components against Anopheles arabiensis that maintains residual malaria transmission. Experiments were conducted using semifield reared An. arabiensis within the semifield system at Ifakara Health Institute (IHI) in southeastern Tanzania. These mosquitoes were exposed to Suna traps® baited with BG lures or source of light and augmented with carbon dioxide (CO2) in view of identifying best attractants necessary to improve attractiveness of designed POHD. Two Suna traps® were hanged at the corner but outside the experimental hut in a diagonal line and rotated between four corners to control for the effect of position and wind direction on mosquito catches. Furthermore, mosquitoes were also exposed to either a bendiocarb-treated or bendiocarb-untreated POHD baited with Mbita blend, Ifakara blend, and worn socks and augmented with warmth (i.e., 1.5 liter bottle of warm water) inside an experimental hut or a screened rectangular box. This study demonstrated that mosquitoes were more strongly attracted to Suna trap® baited with BG lures and CO2 relative to those traps baited with a source of light and CO2. The POHD baited with synthetic blends attracted and killed greater proportion of An. arabiensis compared with POHD baited with worn socks. Efficacy of the POHD was unaffected by source of warmth, and it was reduced by about 50% when the device was tested inside a screened rectangular box relative to closed experimental hut. Overall, this study demonstrates that the POHD baited with synthetic blends (Mbita and Ifakara blends) and bendiocarb can effectively attract and kill outdoor biting malaria vector species. Such POHD baited with synthetic blends may require the source of CO2 to enhance attractiveness to mosquitoes. Further trials are, therefore, ongoing to evaluate attractiveness of improved design of POHD baited with slow-release formulation of synthetic blends and sustainable source of CO2 to malaria vectors under semifield and natural environments.

The Use of Granular Cyclopentanone as Alternative to Artificial Source of Carbon Dioxide in Improved Passive Outdoor Host Seeking Device (POHD).

Reliable sources of CO2 that are relatively cheap, obtainable, and easy to sustain are immediately required for scaling up of odor-baited mosquito surveillance and control devices. Several odor-baited devices are in the pipeline; however, their scale-up against residual malaria transmission, particularly in resource poor areas, is limited by the unavailability of reliable sources of CO2 and reliance on electrical power sources among other factors. We evaluated the use of granular cyclopentanone as an alternative to artificial or yeast fermentation-produced CO2 in passive outdoor host seeking device (POHD). Experiments were conducted against semifield reared An. arabiensis within the semifield system (SFS) at Ifakara Health Institute. Mosquitoes were tested against odor-baited POHDs augmented with yeast fermentation-produced CO2, granular cyclopentanone, attractive blends (Mbita or Ifakara), or their combinations. An insecticide, bendiocarb, was a killing agent used as a proxy for marking the mosquitoes visit the POHDs. Relative attractiveness of different treatment combinations was compared based on the proportion of dead mosquitoes that visited the POHD. The POHD augmented with granules of cyclopentanone alone was attractive to An. arabiensis as much as, or more than, POHDs augmented with yeast fermentation-produced CO2. The POHD baited with CO2 attracted more mosquitoes than those POHDs baited with synthetic blends alone; when these blends are combined with CO2, they attracted more mosquitoes than individual blends. More importantly, such POHDs baited with cyclopentanone attracted far greater proportion of mosquitoes than the POHD baited with either Mbita or Ifakara blend alone. The granular cyclopentanone strongly enhanced/potentiated the attractiveness of POHD baited with Mbita blends against mosquitoes compared to that of POHD baited with Ifakara blend. Moreover, the granular cyclopentanone retained its residual activity against An. arabiensis for up to 2 months after application particularly when used in combination with Mbita blend. In conclusion, this study demonstrates that cyclopentanone granules have the potential to substitute sources of CO2 in outdoor-based surveillance and control devices, thus warranting evaluation of such alternative under realistic field conditions.

Semifield Evaluation of Improved Passive Outdoor Host Seeking Device (POHD) for Outdoor Control of Anopheles arabiensis Mosquitoes.

Despite the considerable progress made so far, the effectiveness and mass application of odour-baited outdoor mosquito control devices in pipelines is limited by several factors. These include the design and size of the devices, optimal placement of attractive blends, and nature of materials into which the blends are impregnated. The primary aim of this study was to manipulate these factors to improve the attractiveness of our recently developed passive outdoor host seeking device (POHD) to outdoor biting Anopheles arabiensis. Specifically, the study aimed to determine optimal placement of odour blends and killing bioactives in POHD for maximum attraction and killing of An. arabiensis and to assess the effects of blend types, formulation, and residual activity on attractiveness of the POHD to An. arabiensis. The POHDs baited with attractive blends, carbon dioxide (CO2), and bendiocarb-treated electrostatic netting were placed either towards the top or bottom openings, and other modifications were exposed to An. arabiensis under the semifield system at Ifakara Health Institute (IHI). Each night, a total of 100 starved female, 3-7-day-old, semifield reared An. arabiensis mosquitoes were released, collected the next morning (alive or dead), counted, and recorded. Live mosquitoes were maintained in the semifield insectary and monitored for 24 hours mortality. Each treatment combination of the POHD was tested in three replicates. Overall, the results indicated that the proportion of mosquitoes attracted to and killed in the POHD varied with position of attractants and killing agent (bendiocarb). The POHD with bottom placed attractants and bendiocarb attracted and killed higher proportion of mosquitoes compared to the POHD with top placed attractants and bendiocarb. The highest mortalities were observed when the POHD was baited with a combination of attractive blends and CO2. Moreover, the residual activity of attractive blends applied inside POHD varied with type and formulation of attractive blend. The POHD packed with Mbita and Ifakara blend in microencapsulated pellets (granules) attracted higher proportion of mosquitoes than that baited with soaked nylon-strip formulation of either blends. Interestingly, POHD baited with Mbita blend in microencapsulated pellets (granules) formulation attracted and killed higher proportion of mosquitoes (>90%) than that baited with Ifakara blend even 9 months after application. Conclusively, the POHD remained effective for a relatively longer period of time when baited with bottom placed synthetic blends and CO2 combination, thus warranting further trials under real life situations.

Ivermectin-treated cattle reduces blood digestion, egg production and survival of a free-living population of Anopheles arabiensis under semi-field condition in south-eastern Tanzania.

BACKGROUND: Anopheles arabiensis feed on cattle and contributes to residual transmission of malaria in areas with high coverage of long-lasting insecticide-treated nets and indoor residual spraying in East Africa. This study aimed to evaluate the effects of ivermectin-treated cattle as a complementary vector control tool against population of An. arabiensis under the semi-field conditions in south-eastern Tanzania. METHODS: The free-living population of An. arabiensis was allowed to forage on untreated or ivermectin-treated cattle in alternating nights within the semi-field system in south-eastern Tanzania. Fresh blood fed mosquitoes were collected in the morning using mouth aspirators and assessed for their blood meal digestion, egg production, and survivorship. The residual activity of ivermectin-treated cattle was also determined by exposing mosquitoes to the same treatments after every 2 days until day 21 post-treatments. These experiments were replicated 3 times using different individual cattle. RESULTS: Overall, the ivermectin-treated cattle reduced blood meal digestion in the stomach of An. arabiensis, and their subsequent egg production and survival over time. The ivermectin-treated cattle halved blood meal digestion in mosquitoes, but reduced their egg production for up to 15 days. The ivermectin-treated cattle reduced the survival, and median survival times (1-3 days) of An. arabiensis than control cattle. The daily mortality rates of mosquitoes fed on ivermectin-treated cattle increased by five-fold relative to controls in the first week, and it gradually declined up to 21 days after treatment. CONCLUSION: This study demonstrates that long-lasting effects of ivermectin-treated cattle on egg production and survival of An. arabiensis may sustainably suppress their vector density, and reduce residual transmission of malaria. This study suggests that ivermectin-treated non-lactating cattle (i.e. calves, heifers and bulls) could be suitable option for large-scale malaria vector control without limiting consumption of milk and meat by communities in rural settings. Furthermore, simulation models are underway to predict the impact of ivermectin-treated cattle alone, or in combination with LLIN/IRS, the frequency of treatment, and their coverage required to significantly suppress population of An. arabiensis and reduce residual transmission of malaria.

Eave tubes for malaria control in Africa: initial development and semi-field evaluations in Tanzania.

BACKGROUND: Presented here are a series of preliminary experiments evaluating "eave tubes"-a technology that combines house screening with a novel method of delivering insecticides for control of malaria mosquitoes. METHODS: Eave tubes were first evaluated with overnight release and recapture of mosquitoes in a screened compartment containing a hut and human sleeper. Recapture numbers were used as a proxy for overnight survival. These trials tested physical characteristics of the eave tubes (height, diameter, angle), and different active ingredients (bendiocarb, LLIN material, fungus). Eave tubes in a hut with closed eaves were also compared to an LLIN protecting a sleeper in a hut with open eaves. Eave tubes were then evaluated in a larger compartment containing a self-replicating mosquito population, vegetation, and multiple houses and cattle sheds. In this "model village", LLINs were introduced first, followed by eave tubes and associated house modifications. RESULTS: Initial testing suggested that tubes placed horizontally and at eave height had the biggest impact on mosquito recapture relative to respective controls. Comparison of active ingredients suggested roughly equivalent effects from bendiocarb, LLIN material, and fungal spores (although speed of kill was slower for fungus). The impact of treated netting on recapture rates ranged from 50 to 70 % reduction relative to controls. In subsequent experiments comparing bendiocarb-treated netting in eave tubes against a standard LLIN, the effect size was smaller but the eave tubes with closed eaves performed at least as well as the LLIN with open eaves. In the model village, introducing LLINs led to an approximate 60 % reduction in larval densities and 85 % reduction in indoor catches of host-seeking mosquitoes relative to pre-intervention values. Installing eave tubes and screening further reduced larval density (93 % relative to pre intervention values) and virtually eliminated indoor host-seeking mosquitoes. When the eave tubes and screening were removed, larval and adult catches recovered to pre-eave tube levels. CONCLUSIONS: These trials suggest that the "eave tube" package can impact overnight survival of host-seeking mosquitoes and can suppress mosquito populations, even in a complex environment. Further testing is now required to evaluate the robustness of these findings and demonstrate impact under field conditions.

Eave tubes for malaria control in Africa: an introduction.

In spite of massive progress in the control of African malaria since the turn of the century, there is a clear and recognized need for additional tools beyond long-lasting insecticide-treated bed nets (LLINs) and indoor residual spraying (IRS) of insecticides, to progress towards elimination. Moreover, widespread and intensifying insecticide resistance requires alternative control agents and delivery systems to enable development of effective insecticide resistance management strategies. This series of articles presents a novel concept for malaria vector control, the 'eave tube', which may fulfil these important criteria. From its conceptualization to laboratory and semi-field testing, to demonstration of potential for implementation, the stepwise development of this new vector control approach is described. These studies suggest eave tubes (which comprise a novel way of delivering insecticides plus screening to make the house more 'mosquito proof') could be a viable, cost-effective, and acceptable control tool for endophilic and endophagic anophelines, and possibly other (nuisance) mosquitoes. The approach could be applicable in a wide variety of housing in sub-Saharan Africa, and possibly beyond, for vectors that use the eave as their primary house entry point. The results presented in these articles were generated during an EU-FP7 funded project, the mosquito contamination device (MCD) project, which ran between 2012 and 2015. This was a collaborative project undertaken by vector biologists, product developers, modellers, materials scientists, and entrepreneurs from five different countries.

Electrostatic coating enhances bioavailability of insecticides and breaks pyrethroid resistance in mosquitoes.

Insecticide resistance poses a significant and increasing threat to the control of malaria and other mosquito-borne diseases. We present a novel method of insecticide application based on netting treated with an electrostatic coating that binds insecticidal particles through polarity. Electrostatic netting can hold small amounts of insecticides effectively and results in enhanced bioavailability upon contact by the insect. Six pyrethroid-resistant Anopheles mosquito strains from across Africa were exposed to similar concentrations of deltamethrin on electrostatic netting or a standard long-lasting deltamethrin-coated bednet (PermaNet 2.0). Standard WHO exposure bioassays showed that electrostatic netting induced significantly higher mortality rates than the PermaNet, thereby effectively breaking mosquito resistance. Electrostatic netting also induced high mortality in resistant mosquito strains when a 15-fold lower dose of deltamethrin was applied and when the exposure time was reduced to only 5 s. Because different types of particles adhere to electrostatic netting, it is also possible to apply nonpyrethroid insecticides. Three insecticide classes were effective against strains of Aedes and Culex mosquitoes, demonstrating that electrostatic netting can be used to deploy a wide range of active insecticides against all major groups of disease-transmitting mosquitoes. Promising applications include the use of electrostatic coating on walls or eave curtains and in trapping/contamination devices. We conclude that application of electrostatically adhered particles boosts the efficacy of WHO-recommended insecticides even against resistant mosquitoes. This innovative technique has potential to support the use of unconventional insecticide classes or combinations thereof, potentially offering a significant step forward in managing insecticide resistance in vector-control operations.

The impact of livestock on the abundance, resting behaviour and sporozoite rate of malaria vectors in southern Tanzania.

BACKGROUND: Increases in the coverage of long-lasting insecticidal nets (LLINs) have significantly reduced the abundance of Anopheles gambiae sensu stricto in several African settings, leaving its more zoophagic sibling species Anopheles arabiensis as the primary vector. This study investigated the impact of livestock ownership at the household level on the ecology and malaria infection rate of vectors in an area of Tanzania where An. arabiensis accounts for most malaria transmission. METHODS: Mosquito vectors were collected resting inside houses, animal sheds and in outdoor resting boxes at households with and without livestock over three years in ten villages of the Kilombero Valley, Tanzania. Additionally, the abundance and sporozoite rate of vectors attempting to bite indoors at these households was assessed as an index of malaria exposure. RESULTS: The mean abundance of An. gambiae s.l. biting indoors was similar at houses with and without livestock. In all years but one, the relative proportion of An. arabiensis within the An. gambiae s.l. species complex was higher at households with livestock. Livestock presence had a significant impact on malaria vector feeding and resting behaviour. Anopheles arabiensis were generally found resting in cattle sheds where livestock were present, and inside houses when absent. Correspondingly, the human blood index of An. arabiensis and An. funestus s.l. was significant reduced at households with livestock, whereas that of An. gambiae s.s. was unaffected. Whilst there was some evidence that sporozoite rates within the indoor-biting An. gambiae s.l population was significantly reduced at households with livestock, the significance of this effect varied depending on how background spatial variation was accounted for. CONCLUSIONS: These results confirm that the presence of cattle at the household level can significantly alter the local species composition, feeding and resting behaviour of malaria vectors. However, the net impact of this livestock-associated variation in mosquito ecology on malaria exposure risk was unclear. Further investigation is required to distinguish whether the apparently lower sporozoite rates observed in An. gambiae s.l. at households with livestock is really a direct effect of cattle presence, or an indirect consequence of reduced risk within areas where livestock keepers choose to live.

The impact of host species and vector control measures on the fitness of African malaria vectors.

Many malaria vector mosquitoes in Africa have an extreme preference for feeding on humans. This specialization allows them to sustain much higher levels of transmission than elsewhere, but there is little understanding of the evolutionary forces that drive this behaviour. In Tanzania, we used a semi-field system to test whether the well-documented preferences of the vectors, Anopheles arabiensis and Anopheles gambiae sensu stricto (s.s.) for cattle and humans, respectively, are predicted by the fitness they obtain from host-seeking on these species relative to other available hosts. Mosquito fitness was contrasted, when humans were fully exposed and when they were protected by a typical bednet. The fitness of both vectors varied between host species. The predicted relationship between host preference and fitness was confirmed in An. arabiensis, but not in An. gambiae s.s., whose fitness was similar on humans and other mammals. Use of typical, imperfect bednets generated only minor reductions in An. gambiae s.s. feeding success and fitness on humans, but was predicted to generate a significant reduction in the lifetime reproductive success of An. arabiensis on humans relative to cows. This supports the hypothesis that such human-protective measures could additionally benefit malaria control by increasing selection for zoophily in vectors.

The fitness of African malaria vectors in the presence and limitation of host behaviour.

BACKGROUND: Host responses are important sources of selection upon the host species range of ectoparasites and phytophagous insects. However little is known about the role of host responses in defining the host species range of malaria vectors. This study aimed to estimate the relative importance of host behaviour to the feeding success and fitness of African malaria vectors, and assess its ability to predict their known host species preferences in nature. METHODS: Paired evaluations of the feeding success and fitness of African vectors Anopheles arabiensis and Anopheles gambiae sensu stricto in the presence and limitation of host behaviour were conducted in a semi-field system (SFS) at Ifakara Health Institute, Tanzania. In one set of trials, mosquitoes were released within the SFS and allowed to forage overnight on a host that was free to exhibit a natural behaviour in response to insect biting. In the other, mosquitoes were allowed to feed directly on from the skin surface of immobile hosts. The feeding success and subsequent fitness of vectors under these conditions were investigated on six host types (humans, calves, chickens, cows, dogs and goats) to assess whether physical movements of preferred host species (cattle for An. arabiensis, humans for An. gambiae s.s.) were less effective at preventing mosquito bites than those of common alternatives. RESULTS: Anopheles arabiensis generally had greater feeding success when applied directly to host skin than when foraging on unrestricted hosts (in five of six host species). However, An. gambiae s.s. obtained blood meals from free and restrained hosts with similar success from most host types (four out of six). Overall, the blood meal size, oviposition rate, fecundity and post-feeding survival of mosquito vectors were significantly higher after feeding on hosts free to exhibit behaviour, than those who were immobilized during feeding trials. CONCLUSIONS: Allowing hosts to move freely during exposure to mosquitoes was associated with moderate reductions in mosquito feeding success, but no detrimental impact to the subsequent fitness of mosquitoes that were able to feed upon them. This suggests that physical defensive behaviours exhibited by common host species including humans do not impose substantial fitness costs on African malaria vectors.

Entomopathogenic fungi, Metarhizium anisopliae and Beauveria bassiana reduce the survival of Xenopsylla brasiliensis larvae (Siphonaptera: Pulicidae).

BACKGROUND: Entomopathogenic fungi, particularly those belonging to the genera Metarhizium and Beauveria have shown great promise as arthropod vector control tools. These agents, however, have not been evaluated against flea vectors of plague. FINDINGS: A 3-h exposure to the fungi coated paper at a concentration of 2 × 108 conidia m-2 infected >90% of flea larvae cadavers in the treatment groups. The infection reduced the survival of larvae that had been exposed to fungus relative to controls. The daily risk of dying was four- and over three-fold greater in larvae exposed to M. anisopliae (HR = 4, p<0.001) and B. bassiana (HR = 3.5, p<0.001) respectively. Both fungi can successfully infect and kill larvae of X. brasiliensis with a pooled median survival time (MST±SE) of 2 ± 0.31 days post-exposure. CONCLUSION: These findings justify further research to investigate the bio-control potential of entomopathogenic fungi against fleas.

Does Cattle Milieu Provide a Potential Point to Target Wild Exophilic Anopheles arabiensis (Diptera: Culicidae) with Entomopathogenic Fungus? A Bioinsecticide Zooprophylaxis Strategy for Vector Control.

Background. Anopheles arabiensis is increasingly dominating malaria transmission in Africa. The exophagy in mosquitoes threatens the effectiveness of indoor vector control strategies. This study aimed to evaluate the effectiveness of fungus against An. arabiensis when applied on cattle and their environments. Methods. Experiments were conducted under semi-field and small-scale field conditions within Kilombero valley. The semi-field reared females of 5-7 days old An. arabiensis were exposed to fungus-treated and untreated calf. Further, wild An. arabiensis were exposed to fungus-treated calves, mud-huts, and their controls. Mosquitoes were recaptured the next morning and proportion fed, infected, and survived were evaluated. Experiments were replicated three times using different individuals of calves. Results. A high proportion of An. arabiensis was fed on calves (>0.90) and become infected (0.94) while resting on fungus-treated mud walls than on other surfaces. However, fungus treatments reduced fecundity and survival of mosquitoes. Conclusion. This study demonstrates for the first time the potential of cattle and their milieu for controlling An. arabiensis. Most of An. arabiensis were fed and infected while resting on fungus-treated mud walls than on other surfaces. Fungus treatments reduced fecundity and survival of mosquitoes. These results suggest deployment of bioinsecticide zooprophylaxis against exophilic An. arabiensis.

Selection of mosquito life-histories: a hidden weapon against malaria?

BACKGROUND: There has recently been a substantial decline in malaria incidence in much of Africa. While the decline can clearly be linked to increasing coverage of mosquito vector control interventions and effective drug treatment in most settings, the ubiquity of reduction raises the possibility that additional ecological and associated evolutionary changes may be reinforcing the effectiveness of current vector control strategies in previously unanticipated ways. PRESENTATION OF HYPOTHESIS: Here it is hypothesized that the increasing coverage of insecticide-treated bed nets and other vector control methods may be driving selection for a shift in mosquito life history that reduces their ability to transmit malaria parasites. Specifically it is hypothesized that by substantially increasing the extrinsic rate of mortality experienced in vector populations, these interventions are creating a fitness incentive for mosquitoes to re-allocate their resources towards greater short-term reproduction at the expense of longer-term survival. As malaria transmission is fundamentally dependent on mosquito survival, a life history shift in this direction would greatly benefit control. TESTING THE HYPOTHESIS: At present, direct evaluation of this hypothesis within natural vector populations presents several logistical and methodological challenges. In the meantime, many insights can be gained from research previously conducted on wild Drosophila populations. Long-term selection experiments on these organisms suggest that increasing extrinsic mortality by a magnitude similar to that anticipated from the up-scaling of vector control measures generated an increase in their intrinsic mortality rate. Although this increase was small, a change of similar magnitude in Anopheles vector populations would be predicted to reduce malaria transmission by 80%. IMPLICATIONS OF HYPOTHESIS: The hypothesis presented here provides a reminder that evolutionary processes induced by interventions against disease vectors may not always act to neutralize intervention effectiveness. In the search for new intervention strategies, consideration should be given to both the potential disadvantages and advantages of evolutionary processes resulting from their implementation, and attempts made to exploit those with greatest potential to enhance control.

Exploiting the behaviour of wild malaria vectors to achieve high infection with fungal biocontrol agents.

BACKGROUND: Control of mosquitoes that transmit malaria has been the mainstay in the fight against the disease, but alternative methods are required in view of emerging insecticide resistance. Entomopathogenic fungi are candidate alternatives, but to date, few trials have translated the use of these agents to field-based evaluations of their actual impact on mosquito survival and malaria risk. Mineral oil-formulations of the entomopathogenic fungi Metarhizium anisopliae and Beauveria bassiana were applied using five different techniques that each exploited the behaviour of malaria mosquitoes when entering, host-seeking or resting in experimental huts in a malaria endemic area of rural Tanzania. RESULTS: Survival of mosquitoes was reduced by 39-57% relative to controls after forcing upward house-entry of mosquitoes through fungus treated baffles attached to the eaves or after application of fungus-treated surfaces around an occupied bed net (bed net strip design). Moreover, 68 to 76% of the treatment mosquitoes showed fungal growth and thus had sufficient contact with fungus treated surfaces. A population dynamic model of malaria-mosquito interactions shows that these infection rates reduce malaria transmission by 75-80% due to the effect of fungal infection on adult mortality alone. The model also demonstrated that even if a high proportion of the mosquitoes exhibits outdoor biting behaviour, malaria transmission was still significantly reduced. CONCLUSIONS: Entomopathogenic fungi strongly affect mosquito survival and have a high predicted impact on malaria transmission. These entomopathogens represent a viable alternative for malaria control, especially if they are used as part of an integrated vector management strategy.

Anopheline and culicine mosquitoes are not repelled by surfaces treated with the entomopathogenic fungi Metarhizium anisopliae and Beauveria bassiana.

BACKGROUND: Entomopathogenic fungi, Metarhizium anisopliae and Beauveria bassiana, are promising bio-pesticides for application against adult malaria mosquito vectors. An understanding of the behavioural responses of mosquitoes towards these fungi is necessary to guide development of fungi beyond the 'proof of concept' stage and to design suitable intervention tools. METHODS: Here we tested whether oil-formulations of the two fungi could be detected and avoided by adult Anopheles gambiae s.s., Anopheles arabiensis and Culex quinquefasciatus. The bioassays used a glass chamber divided into three compartments (each 250 × 250 × 250 mm): release, middle and stimulus compartments. Netting with or without fungus was fitted in front of the stimulus compartment. Mosquitoes were released and the proportion that entered the stimulus compartment was determined and compared between treatments. Treatments were untreated netting (control 1), netting with mineral oil (control 2) and fungal conidia formulated in mineral oil evaluated at three different dosages (2 × 1010, 4 × 1010 and 8 × 1010 conidia m-2). RESULTS: Neither fungal strain was repellent as the mean proportion of mosquitoes collected in the stimulus compartment did not differ between experiments with surfaces treated with and without fungus regardless of the fungal isolate and mosquito species tested. CONCLUSION: Our results indicate that mineral-oil formulations of M. anisopliae and B. bassiana were not repellent against the mosquito species tested. Therefore, both fungi are suitable candidates for the further development of tools that aim to control host-seeking or resting mosquitoes using entomopathogenic fungi.

A new resting trap to sample fungus-infected mosquitoes, and the pathogenicity of Lecanicillium muscarium to culicid adults.

Some soil-dwelling entomopathogenic fungi that are widely used in pest control are also able to reduce the survival of adult mosquito vectors under laboratory conditions. However, there is still little information about the naturally occurring fungal pathogens affecting culicid mosquitoes. As such, we hypothesized that fungi that already kill mosquitoes in realistic domestic environments could be effective against these vectors in human habitations. A simple, inexpensive, handmade, cylindrical kiln-fired clay pot (30 cm height, 24 cm inner diameter, 0.8-1cm wall thickness) was modified into a trapping device for resting adult mosquitoes and to sample fungus-infected moribund and dead individuals. The entomopathogenic fungus Lecanicillium muscarium was isolated from a dead culicid mosquito collected with this trap in southeastern Tanzania. This isolate is the first L. muscarium reported to occur naturally on adult culicids in Tanzania and was found to be pathogenic also to adults of Aedes aegypti, Anopheles arabiensis and Culex quinquefasciatus under laboratory conditions. The trapping device confirmed its efficacy to sample mosquito-specific fungi in domestic locations and that the isolated fungus might have potential for mosquito control.

First report of Metarhizium anisopliae IP 46 pathogenicity in adult Anopheles gambiae s.s. and An. arabiensis (Diptera; Culicidae).

The entomopathogenic fungus Metarhizium anisopliae isolate IP 46, originating from a soil sample collected in 2001 in the Cerrado of Central Brazil, was tested for its ability to reduce the survival of adult male and female Anopheles gambiae s.s. and An. arabiensis mosquitoes. A 6-h exposure to the fungus coated on test paper at a concentration of 3.3 x 106 conidia cm-2 reduced the daily survival of both mosquito species (HR = 3.14, p < 0.001), with higher risk of dying in An. gambiae s.s relative to An. arabiensis (HR = 1.38, p < 0.001). Fungal sporulation was observed in >95% of mosquito cadavers in the treatment groups. The results indicate that M. anisopliae IP 46 has the potential to be a bio-control agent for African malaria vector species, and is a suitable candidate for further research and development.

Ecological and evolutionary determinants of host species choice in mosquito vectors.

Insects exhibit diverse resource-exploitation strategies, including predation, herbivory and parasitism. The ecological and evolutionary factors that influence the resource selection of some insects (e.g. herbivores) have been extensively investigated because of their agricultural importance. By contrast, there has been little investigation of the selective forces that mediate host choice in haematophagous insects, despite their importance as vectors of disease. Here, we review potential determinants of host species choice in mosquitoes, the most important insect vectors of human disease, and discuss whether these could be manipulated to yield new disease-control strategies based on vector behavioural change.