Search

Background: Nchelenge District in northern Zambia suffers from holoendemic malaria transmission despite a decade of yearly indoor residual spraying (IRS) and insecticide-treated net (ITN) distributions. One hypothesis for this lack of impact is that some vectors in the area may forage in the early evening or outdoors. Anopheles gibbinsi specimens were identified in early evening mosquito collections performed in this study area, and further insight was gleaned into this taxon, including characterizing its genetic identity, feeding preferences, and potential role as a malaria vector. Methods: Mosquitoes were collected in July and August 2019 by CDC light traps in Nchelenge District in indoor sitting rooms, outdoor gathering spaces, and animal pens from 16:00–22:00. Host detection by PCR, COI and ITS2 PCR, and circumsporozoite (CSP) ELISA were performed on all samples morphologically identified as An. gibbinsi, and a subset of specimens were selected for COI and ITS2 sequencing. To determine risk factors for increased abundance of An. gibbinsi, a negative binomial generalized linear mixed-effects model was performed with household-level variables of interest. Results: Comparison of COI and ITS2 An. gibbinsi reference sequences to the NCBI database revealed > 99% identity to "Anopheles sp. 6" from Kenya. More than 97% of specimens were morphologically and molecularly consistent with An. gibbinsi. Specimens were primarily collected in animal pen traps (59.2%), followed by traps outdoors near where humans gather (24.3%), and traps set indoors (16.5%). Host DNA detection revealed a high propensity for goats, but 5% of specimens with detected host DNA had fed on humans. No specimens were positive for Plasmodium falciparum sporozoites. Animal pens and inland households > 3 km from Lake Mweru were both associated with increased An. gibbinsi abundance. Conclusions: This is the first report of An. gibbinsi in Nchelenge District, Zambia. This study provided a species identity for unknown "An. sp. 6" in the NCBI database, which has been implicated in malaria transmission in Kenya. Composite data suggest that this species is largely zoophilic and exophilic, but comes into contact with humans and the malaria parasites they carry. This species should continue to be monitored in Zambia and neighbouring countries as a potential malaria vector. [ABSTRACT FROM AUTHOR]

Background: Nchelenge District in northern Zambia suffers from holoendemic malaria transmission despite a decade of yearly indoor residual spraying (IRS) and insecticide-treated net (ITN) distributions. One hypothesis for this lack of impact is that some vectors in the area may forage in the early evening or outdoors. Anopheles gibbinsi specimens were identified in early evening mosquito collections performed in this study area, and further insight was gleaned into this taxon, including characterizing its genetic identity, feeding preferences, and potential role as a malaria vector. Methods: Mosquitoes were collected in July and August 2019 by CDC light traps in Nchelenge District in indoor sitting rooms, outdoor gathering spaces, and animal pens from 16:00–22:00. Host detection by PCR, COI and ITS2 PCR, and circumsporozoite (CSP) ELISA were performed on all samples morphologically identified as An. gibbinsi, and a subset of specimens were selected for COI and ITS2 sequencing. To determine risk factors for increased abundance of An. gibbinsi, a negative binomial generalized linear mixed-effects model was performed with household-level variables of interest. Results: Comparison of COI and ITS2 An. gibbinsi reference sequences to the NCBI database revealed > 99% identity to "Anopheles sp. 6" from Kenya. More than 97% of specimens were morphologically and molecularly consistent with An. gibbinsi. Specimens were primarily collected in animal pen traps (59.2%), followed by traps outdoors near where humans gather (24.3%), and traps set indoors (16.5%). Host DNA detection revealed a high propensity for goats, but 5% of specimens with detected host DNA had fed on humans. No specimens were positive for Plasmodium falciparum sporozoites. Animal pens and inland households > 3 km from Lake Mweru were both associated with increased An. gibbinsi abundance. Conclusions: This is the first report of An. gibbinsi in Nchelenge District, Zambia. This study provided a species identity for unknown "An. sp. 6" in the NCBI database, which has been implicated in malaria transmission in Kenya. Composite data suggest that this species is largely zoophilic and exophilic, but comes into contact with humans and the malaria parasites they carry. This species should continue to be monitored in Zambia and neighbouring countries as a potential malaria vector. [ABSTRACT FROM AUTHOR]

Background: Malaria vector control using insecticide-based approaches has proven to be an effective strategy. However, widespread insecticide resistance among malaria vector populations across sub-Saharan Africa threatens to derail control efforts. This study was conducted in Chikwawa district, an area in rural southern Malawi characterised by persistent malaria transmission and reports of insecticide resistance in the local mosquito population. The aim of the was to characterise the intensity of insecticide resistance within a population of Anopheles funestus sensu lato (s.l.), a major vector of malaria in this district. Methods: Live adult females belonging to the An. funestus group were collected from households by indoor aspiration. The CDC bottle assay was used for phenotypic quantification of resistance to deltamethrin, permethrin and alpha-cypermethrin at 1×, 2.5×, 5× and 10× the recommended diagnostic dose for each of these insecticides. WHO tube assays were used to determine susceptibility to bendiocarb, dichlorodiphenyltrichloroethane (DDT) and pirimiphos-methyl insecticides at diagnostic concentrations. Results: Anopheles funestus s.l. exposed to 10× the recommended diagnostic dose was highly resistant to alpha-cypermethrin (mortality 95.4%); in contrast, mortality was 100% when exposed to both deltamethrin and permethrin at the same dose. Despite showing susceptibility to deltamethrin and permethrin at the 10× concentration, mortality at the 5× concentration was 96.7% and 97.1%, respectively, indicating moderate resistance to these two insecticides. WHO susceptibility assays indicated strong resistance against bendiocarb (mortality 33.8%, n = 93), whereas there was full susceptibility to DDT (mortality 98.9%, n = 103) and pirimiphos-methyl (mortality 100%, n = 103). Conclusions: Strategies for managing resistance to insecticides, particularly against pyrethroids, must be urgently implemented to maintain the effectiveness of insecticide-based vector control interventions in the area. Such strategies include the wide-scale introduction of third-generation synergist insecticide-treated bed nets (ITNs) and next-generation dual active ingredient ITNs. The use of effective non-pyrethroids, such as pirimiphos-methyl, clothianidin and potentially DDT, could provide a window of opportunity for indoor residual spraying across the district. This strategy would support the current Malawi Insecticide Resistance Management Plan which aims at rotating insecticides to minimise selection pressure and slow down the evolution of resistance to approved insecticides. These actions will help to prevent malaria vector control failure and improve progress towards malaria elimination. [ABSTRACT FROM AUTHOR]

Background The SEMOS gold mine in Sadiola, southwestern Mali, has been implementing a malaria vector control programme for 15 y using indoor residual house spraying and sporadic larval control. Periodic screening of the vector populations have been carried out over the years to provide information to the control programme, mainly on vector species present and their insecticide resistance status. The data from five entomological surveys, carried out in 2006, 2011, 2014, 2016 and 2018, are presented. Methods Adult mosquitoes were collected resting on walls inside houses and on verandas. Insecticide susceptibility assays were carried out and mosquitoes subsequently identified by species using molecular assays. Results The major malaria vector mosquitoes, Anopheles gambiae and Anopheles arabiensis were abundant at each sampling period with Anopheles coluzzii and Anopheles funestus being rare or absent. Anopheles rivulorum was identified in 2006 and Anopheles leesoni in 2016. The presence of Anopheles rivulorum -like, identified for the first time in 2018, was not screened for in previous surveys. Insecticide susceptibility bioassays showed resistance in both A. gambiae and A. arabiensis to pyrethroids, carbamates and dichlorodiphenyltrichloroethane over the 12 y. Conclusions This is the first record of A. rivulorum -like west of Côte d'Ivoire. Resistance levels to the three classes of insecticides were variable but appeared to decrease after pyrethroids were discontinued for house spraying. [ABSTRACT FROM AUTHOR]

The association between insect development and temperature is well established. Thermal summation using accumulated degree‐day measures is commonly used. However, the time at which evidence is collected is important in these estimates. The aim of this study was to provide a simulated model of the effect of temperatures on six dipteran species commonly associated with cadavers, from the death scene to the refrigerator, and finally at the time of autopsy. Temperatures measurements were sampled over a 16‐month period from the external environment (external to the mortuary), within the mortuary refrigerator, and within the mortuary autopsy suite. Monte Carlo simulation using accumulated degree‐days (ADD) was used to estimate the variations based on the mean and standard deviation of the temperature measurements. It was found that there was a negative correlation between the base temperature of the fly species (lowest temperature at which the flies will survive) and developmental likelihood. Species with high base temperatures (Chrysomya albiceps, Chrysomya chloropyga, and Musca domestica) were less likely to continue development in refrigerators than species with lower base temperatures (Lucilia sericata and Piophila casei). The findings of this study highlight the importance of recording temperature measurements and the period of refrigeration on PMI estimation especially when continued development occurs in spite of a period of cooling of the insect evidence. [ABSTRACT FROM AUTHOR]

Background: Anopheles funestus (s.s.) is a primary vector of the malaria parasite Plasmodium falciparum in Africa, a human pathogen that causes almost half a million deaths each year. The population structure of An. funestus was examined in samples from Uganda and the southern African countries of Malawi, Mozambique, Zambia and Zimbabwe. Methods: Twelve microsatellites were used to estimate the genetic diversity and differentiation of An. funestus from 13 representative locations across five countries. These were comprised of four sites from Uganda, three from Malawi and two each from Mozambique, Zambia and Zimbabwe. Results: All loci were highly polymorphic across the populations with high allelic richness and heterozygosity. A high genetic diversity was observed with 2–19 alleles per locus and an average number of seven alleles. Overall, expected heterozygosity (He) ranged from 0.65 to 0.79. When samples were pooled three of the 12 microsatellite loci showed Hardy–Weinberg equilibrium. Unsupervised Bayesian clustering analysis of microsatellite data revealed two clusters with An. funestus samples from Mozambique, Uganda and Zambia falling into one group and Malawi and Zimbabwe into another. The overall genetic differentiation between the populations was moderate (FST = 0.116). Pairwise differentiation between the pairs was low but significant. A weak but significant correlation was established between genetic and geographical distance for most populations. Conclusions: High genetic diversity revealed by the loci with low to moderate differentiation, identified two clusters among the An. funestus populations. Further research on the population dynamics of An. funestus in east and southern Africa is essential to understand the implications of this structuring and what effect it may have on the efficient implementation of mosquito vector control strategies. [ABSTRACT FROM AUTHOR]

Background: In 1987, Gillies and Coetzee published a pictorial key for the morphological identification of adult female mosquitoes. Since then, several new species of anopheline mosquitoes have been described. Methods: The 1987 key to adult female mosquitoes was used as the template for the current key. Results: New species described in the literature over the past 32 years have been included. A list of all currently known Afrotropical species is provided. Anopheles stephensi is included for the first time as occurring on the African continent. Conclusions: An updated key for the morphological identification of Afrotropical anopheline species is presented. [ABSTRACT FROM AUTHOR]

To accelerate malaria elimination in areas where core interventions such as insecticide-treated nets (ITNs) are already widely used, it is crucial to consider additional factors associated with persistent transmission. Qualitative data on human behaviours and perceptions regarding malaria risk was triangulated with quantitative data on Anopheles mosquito bites occurring indoors and outdoors in south-eastern Tanzania communities where ITNS are already used but lower level malaria transmission persists. Each night (18:00h-07:00h), trained residents recorded human activities indoors, in peri-domestic outdoor areas, and in communal gatherings. Host-seeking mosquitoes were repeatedly collected indoors and outdoors hourly, using miniaturized exposure-free double net traps (DN-Mini) occupied by volunteers. In-depth interviews were conducted with household representatives to explore perceptions on persistent malaria and its control. Higher proportions of people stayed outdoors than indoors in early-evening and early-morning hours, resulting in higher exposures outdoors than indoors during these times. However, exposure during late-night hours (22:00h–05:00h) occurred mostly indoors. Some of the popular activities that kept people outdoors included cooking, eating, relaxing and playing. All households had at least one bed net, and 83.9% of people had access to ITNs. Average ITN use was 96.3%, preventing most indoor exposure. Participants recorgnized the importance of ITNs but also noted that the nets were not perfect. No complementary interventions were reported being used widely. Most people believed transmission happens after midnight. We conclude that insecticide-treated nets, where properly used, can still prevent most indoor exposures, but significant risk continues unabated before bedtime, outdoors and at communal gatherings. Such exposure is greatest for rural and low-income households. There is therefore an urgent need for complementary interventions, particularly those targeting outdoor-biting and are applicable for all people including the marginalised populations such as migratory farmers and fishermen. Besides, the differences in community understanding of ongoing transmission, and feedback on imperfections of ITNs should be considered when updating malaria-related communication and interventions. [ABSTRACT FROM AUTHOR]

Background: Anopheles funestus mosquitoes currently contribute more than 85% of ongoing malaria transmission events in south-eastern Tanzania, even though they occur in lower densities than other vectors, such as Anopheles arabiensis. Unfortunately, the species ecology is minimally understood, partly because of difficulties in laboratory colonization. This study describes the first observations of An. funestus swarms in Tanzania, possibly heralding new opportunities for control. Method: Using systematic searches by community-based volunteers and expert entomologists, An. funestus swarms were identified in two villages in Ulanga and Kilombero districts in south-eastern Tanzania, starting June 2018. Swarms were characterized by size, height, start- and end-times, presence of copulation and associated environmental features. Samples of male mosquitoes from the swarms were examined for sexual maturity by observing genitalia rotation, species identity using polymerase chain reaction and wing sizes. Results: 581 An. funestus (98.1% males (n = 570) and 1.9% (n = 11) females) and 9 Anopheles gambiae sensu lato (s.l.) males were sampled using sweep nets from the 81 confirmed swarms in two villages (Ikwambi in Kilombero district and Tulizamoyo in Ulanga district). Six copulation events were observed in the swarms. Mean density (95% CL) of An. funestus caught/swarm/village/evening was 6.6 (5.9–7.2) in Tulizamoyo and 10.8 (5.8–15.8) in Ikwambi. 87.7% (n = 71) of the swarms were found in Tulizamoyo, while 12.3% (n = 10) were in Ikwambi. Mean height of swarms was 1.7 m (0.9–2.5 m), while mean duration was 12.9 (7.9–17.9) minutes. The PCR analysis confirmed that 100% of all An. funestus s.l. samples processed were An. funestus sensu stricto. Mean wing length of An. funestus males was 2.47 mm (2.0–2.8 mm), but there was no difference between swarming males and indoor-resting males. Most swarms (95.0%) occurred above bare ground, sometime on front lawns near human dwellings, and repeatedly in the same locations. Conclusion: This study has demonstrated occurrence of An. funestus swarms for the first time in Tanzania. Further investigations could identify new opportunities for improved control of this dominant malaria vector, possibly by targeting the swarms. [ABSTRACT FROM AUTHOR]

Abstract: In spite of widespread insecticide resistance in vector mosquitoes throughout Africa, there is limited evidence that long‐lasting insecticidal bed nets (LLINs) are failing to protect against malaria. Here, we showed that LLIN contact in the course of host‐seeking resulted in higher mortality of resistant Anopheles spp. mosquitoes than predicted from standard laboratory exposures with the same net. We also found that sublethal contact with an LLIN caused a reduction in blood feeding and subsequent host‐seeking success in multiple lines of resistant mosquitoes from the laboratory and the field. Using a transmission model, we showed that when these LLIN‐related lethal and sublethal effects were accrued over mosquito lifetimes, they greatly reduced the impact of resistance on malaria transmission potential under conditions of high net coverage. If coverage falls, the epidemiological impact is far more pronounced. Similarly, if the intensity of resistance intensifies, the loss of malaria control increases nonlinearly. Our findings help explain why insecticide resistance has not yet led to wide‐scale failure of LLINs, but reinforce the call for alternative control tools and informed resistance management strategies. [ABSTRACT FROM AUTHOR]

Background: Anopheles funestus has been recognized as a major malaria vector in Africa for over 100 years, but knowledge on many aspects of the biology of this species is still lacking. Anopheles funestus, as with most other anophelines, mate through swarming. A key event that is crucial for the An. funestus male to mate is genitalia rotation. This involves the 135° to 180° rotation of claspers, which are tipped with claws. This physical change then enables the male to grasp the female during copulation. The aim of this investigation was to molecularly characterize wild An. funestus swarms from Zambia and examine the degree of genitalia rotation within the swarm. Methods: Anopheles funestus swarms were collected from Nchelenge, northern Zambia, during dusk periods in May 2016. All the adults from the swarm were analysed morphologically and identified to species level using a multiplex PCR assay. Anopheles funestus s.s. specimens were molecularly characterized by restriction fragment length polymorphism type and Clade type assays. The different stages of genitalia rotation were examined in the adult males. Results: A total of six swarms were observed during the study period and between 6 and 26 mosquitoes were caught from each swarm. Species analysis revealed that 90% of the males from the swarms were An. funestus s.s. MWtype, with 84% belonging to clade I compared to 14% clade II and 2% failed to amplify. Very few specimens (3.4%) were identified as Anopheles gambiae s.s. Eighty percent of the males from the swarm had complete genitalia rotation. Conclusions: This is the first time that An. funestus swarms have been molecularly identified to species level. Anopheles funestus swarms appear to be species-specific with no evidence of clade-type differentiation within these swarms. The An. funestus swarms consist mainly of males with fully rotated genitalia, which strongly suggests that swarming behaviour is triggered primarily when males have matured. [ABSTRACT FROM AUTHOR]

Background: The correct identification of disease vectors is the first step towards implementing an effective control programme. Traditionally, for malaria control, this was based on the morphological differences observed in the adults and larvae between different mosquito species. However, the discovery of species complexes meant that genetic tools were needed to separate the sibling species and today there are standard molecular techniques that are used to identify the two major malaria vector groups of mosquitoes. On the assumption that species-diagnostic DNA polymerase chain reaction (PCR) assays are highly species-specific, experiments were conducted to investigate what would happen if non-vector species were randomly included in the molecular assays. Methods: Morphological keys for the Afrotropical Anophelinae were used to provide the a priori identifications. All mosquito specimens were then subjected to the standard PCR assays for members of the Anopheles gambiae complex and Anopheles funestus group. Results: One hundred and fifty mosquitoes belonging to 11 morphological species were processed. Three species (Anopheles pretoriensis, Anopheles rufipes and Anopheles rhodesiensis) amplified members of the An. funestus group and four species (An. pretoriensis, An. rufipes, Anopheles listeri and Anopheles squamosus) amplified members of the An. gambiae complex. Conclusions: Morphological identification of mosquitoes prior to PCR assays not only saves time and money in the laboratory, but also ensures that data received by malaria vector control programmes are useful for targeting the major vectors. [ABSTRACT FROM AUTHOR]

Background: The Democratic Republic of the Congo (DRC) is characterized as a holoendemic malaria area with the main vectors being Anopheles funestus and members of the Anopheles gambiae complex. Due to political instability and socio-economic challenges in the region, knowledge of insecticide resistance status and resistance mechanisms in these vectors is limited. Mosquitoes were collected from a mining site in the north-eastern part of the country and, following identification, were subjected to extensive testing for the target-site and biochemical basis of resistance. Quantitative real-time PCR was used to assess a suite of 10 genes frequently involved in pyrethroid and dichlorodiphenyltrichloroethane (DDT) resistance in An. gambiae females and males. In An. funestus, gene expression microarray analysis was carried out on female mosquitoes. Results: In both species, deltamethrin resistance was recorded along with high resistance and suspected resistance to DDT in An. gambiae and An. funestus, respectively. A total of 85% of An. gambiae carried the kdr mutations as either homozygous resistant (RR) (L1014S, L1014F or both) or heterozygous (RS), however only 3% carried the rdl mutant allele (RS) and no ace-1 mutations were recorded. Synergist assays indicated a strong role for P450s in deltamethrin resistance in both species. In An. gambiae, analysis of transcription levels showed that the glutathione-S-transferase, GSTS1-2, produced the highest fold change in expression (7.6-fold in females and 31-fold in males) followed by GSTE2, thioredoxin peroxidase (TPX2), and cytochrome oxidases (CYP6M2 and CYP6P1). All other genes tested produced fold change values below 2. Microarray analysis revealed significant over-transcription of cuticular proteins as well as CYP6M7, CYP6P9a and CYP6P9b in insecticide resistant An. funestus. Conclusions: These data show that high levels of deltamethrin resistance in the main malaria vector species, conferred by enzymatic detoxification, are present in the DRC. [ABSTRACT FROM AUTHOR]

Background: Insecticide resistance is the major emerging challenge facing the malaria vector control programmes in Tanzania. Proper monitoring and detection is of paramount importance guiding the vector control programmes. This paper presents the effect of mosquito aging on insecticide resistance status in Anopheles arabiensis populations in dry and rainy seasons in northern Tanzania. Methods: Anopheles gambiae s.l. larvae were sampled from rice fields in both dry and rainy seasons and reared in the insectary to adults. The emerged females in batches of 2, 3, 5, and 10 days old were exposed to six insecticides (deltamethrin, permethrin, lambda-cyhalothrin, DDT, bendiocarb and pirimiphos-methyl) to see the effects of age on insecticide resistance. Mosquitoes were exposed to insecticides using WHO standard susceptibility test kits. Knockdown was recorded during the 1-h exposure, while mortality and resistance ratio were recorded 24 h later. Mosquito specimens were identified to species level using the polymerase chain reaction (PCR) method. Results: Among the 326 specimens processed by PCR, 323 (99.1%) were identified as Anopheles arabiensis. There was reduced mortality (ranging from 61 to 97.7%) when adults reared from larvae were exposed to all pyrethroids and bendiocarb in both dry and rainy seasons, while they were fully susceptible to DDT and pirimiphos-methyl. There was a significant increase in mortality rate with increase in mosquito's age in both dry and rainy seasons following exposure to pyrethroids (DF = 1, P < 0.05). Mosquitoes showed significantly higher mortality rates in the rainy season than in the dry season after being exposed to pyrethroids (DF = 1, P < 0.05). Higher mortality rates (94.0-99.8%) were observed in all ages and seasons when mosquitoes were exposed to bendiocarb compared with pyrethroids. Pirimiphos-methyl was only tested in the rainy season so no comparison with dry season mosquitoes could be made. Conclusions: Results showed that An. arabiensis were resistant to pyrethroids in both seasons and that the young age groups exhibited higher levels of resistance compared with the older age groups. Mosquitoes were full susceptible to DDT and pirimiphos-methyl irrespective of the season and age. [ABSTRACT FROM AUTHOR]

Background: The extensive use of indoor residual spraying (IRS) and insecticide-treated nets (ITNs) in Africa has contributed to a significant reduction in malaria transmission. Even so, residual malaria transmission persists in many regions, partly driven by mosquitoes that bite people outdoors. In areas where Anopheles gambiae s.s. is a dominant vector, most interventions target the reduction of indoor transmission. The increased use of ITNs/LLINs and IRS has led to the decline of this species. As a result, less dominant vectors such as Anopheles funestus and Anopheles arabiensis, both also originally indoor vectors but are increasingly biting outdoors, contribute more to residual malaria transmission. The study reports the investigated community perceptions on malaria and their implications of this for ongoing outdoor malaria transmission and malaria control efforts. Methods: This was a qualitative study conducted in two rural villages and two peri-urban areas located in Kilombero Valley in south-eastern Tanzania. 40 semi-structured in-depth interviews and 8 focus group discussions were conducted with men and women who had children under the age of five. The Interviews and discussions focused on (1) community knowledge of malaria transmission, and (2) the role of such knowledge on outdoor malaria transmission as a contributing factor to residual malaria transmission. Results: The use of bed nets for malaria prevention has been stressed in a number of campaigns and malaria prevention programmes. Most people interviewed believe that there is outdoor malaria transmission since they use interventions while indoors, but they are unaware of changing mosquito host-seeking behaviour. Participants pointed out that they were frequently bitten by mosquitoes during the evening when outdoors, compared to when they were indoors. Most participants stay outdoors in the early evening to undertake domestic tasks that cannot be conducted indoors. House structure, poor ventilation and warm weather conditions were reported to be the main reasons for staying outdoors during the evening. Participants reported wearing long sleeved clothes, fanning and slapping themselves, using repellents, and burning cow dung and neem tree leaves to chase away mosquitoes. Conclusions: Community understanding of multiple prevention strategies is crucial given changes in mosquito host seeking behaviour and the increased incidence of outdoor biting. The current low use of outdoor control measures is attributed largely to limited awareness of outdoor transmission. Improved community understanding of outdoor malaria transmission is critical: efforts to reduce or eliminate malaria transmission will not be successful if the control of outdoor transmission is not emphasized. [ABSTRACT FROM AUTHOR]

Background: Insecticide use via indoor residual spraying (IRS) or treated nets is the primary method for controlling malaria vector populations. The incidence of insecticide resistance in vector populations is burgeoning globally making resistance management key to the design of effective malaria control and elimination strategies. Vector populations can be assessed for insecticide resistance using a binary (susceptible or resistant) classification based on the use of the standard WHO insecticide susceptibility assay for adult anopheline mosquitoes. However, the recent scaling up of vector control activities has necessitated a revision of the WHO bioassay protocol to include the production of information that not only diagnoses resistance but also gives information on the intensity of expression of resistance phenotypes detected. This revised protocol is expected to inform on the range of resistance phenotypes in a target vector population using discriminating/diagnostic insecticide concentrations (DC) as well as their potential operational significance using 5× DC and 10× DC assays. The aim of this project was to use the revised protocol to assess the intensity of pyrethroid resistance in a range of insecticide resistant Anopheles strains with known resistance mechanisms and for which there is evidence of operational significance in the field setting from which these colonies were derived. Methods: Diagnostic concentration (DC) bioassays followed by 5× DC and 10× DC assays using the pyrethroid insecticides permethrin and deltamethrin were conducted according to the standard WHO bioassay method against pyrethroid resistant laboratory strains of Anopheles funestus, An. arabiensis and An. gambiae. Results: Low to moderate resistance intensities were recorded for the An. arabiensis and An. gambiae strains while moderate to high intensities were recorded for the An. funestus strains. Conclusions: It is evident that resistance intensity assays can add predictive value to the decision making process in vector control settings, although more so in an IRS setting and especially when bench-marked against resistance phenotypes of known operational significance. [ABSTRACT FROM AUTHOR]

Background: It is widely acknowledged that modifications to existing control interventions are required if South Africa is to achieve malaria elimination. Targeting indoor residual spraying (IRS) to areas where cases have been detected is one strategy currently under investigation in northeastern South Africa. This seroprevalence baseline study, nested within a targeted IRS trial, was undertaken to provide insights into malaria transmission dynamics in South Africa and evaluate whether sero-epidemiological practices have the potential to be routinely incorporated into elimination programmes. Methods: Filter-paper blood spots, demographic and household survey data were collected from 2710 randomly selected households in 56 study wards located in the municipalities of Ba-Phalaborwa and Bushbuckridge. Blood spots were assayed for Plasmodium falciparum apical membrane antigen-1 and merozoite surface protein-119 bloodstage antigens using an enzyme linked immunosorbent assay. Seroprevalence data were analysed using a reverse catalytic model to determine malaria seroconversion rates (SCR). Geospatial cluster analysis was used to investigate transmission heterogeneity while random effects logistic regression identified risk factors associated with malaria exposure. Results: The overall SCR across the entire study site was 0.012 (95% CI 0.008-0.017) per year. Contrasting SCRs, corresponding to distinct geographical regions across the study site, ranging from <0.001 (95% CI <0.001-0.005) to 0.022 (95% CI 0.008-0.062) per annum revealed prominent transmission heterogeneity. Geospatial cluster analysis of household seroprevalence and age-adjusted antibody responses detected statistically significant (p < 0.05) spatial clusters of P. falciparum exposure. Formal secondary education was associated with lower malaria exposure in the sampled population (AOR 0.72, 95% CI 0.56-0.95, p = 0.018). Conclusions: Although overall transmission intensity and exposure to malaria was low across both study sites, malaria transmission intensity was highly heterogeneous and associated with low socio-economic status in the region. Findings suggest focal targeting of interventions has the potential to be an appropriate strategy to deploy in South Africa. Furthermore, routinely incorporating sero-epidemiological practices into elimination programmes may prove useful in monitoring malaria transmission intensity in South Africa, and other countries striving for malaria elimination. [ABSTRACT FROM AUTHOR]

Mosquitoes, especially Aedes aegypti, are becoming important models for studying invasion biology. We characterized genetic variation at 12 microsatellite loci in 79 populations of Ae. aegypti from 30 countries in six continents, and used them to infer historical and modern patterns of invasion. Our results support the two subspecies Ae. aegypti formosus and Ae. aegypti aegypti as genetically distinct units. Ae. aegypti aegypti populations outside Africa are derived from ancestral African populations and are monophyletic. The two subspecies co-occur in both East Africa (Kenya) and West Africa (Senegal). In rural/forest settings (Rabai District of Kenya), the two subspecies remain genetically distinct, whereas in urban settings, they introgress freely. Populations outside Africa are highly genetically structured likely due to a combination of recent founder effects, discrete discontinuous habitats and low migration rates. Ancestral populations in sub-Saharan Africa are less genetically structured, as are the populations in Asia. Introduction of Ae. aegypti to the New World coinciding with trans-Atlantic shipping in the 16th to 18th centuries was followed by its introduction to Asia in the late 19th century from the New World or from now extinct populations in the Mediterranean Basin. Aedes mascarensis is a genetically distinct sister species to Ae. aegypti s.l. This study provides a reference database of genetic diversity that can be used to determine the likely origin of new introductions that occur regularly for this invasive species. The genetic uniqueness of many populations and regions has important implications for attempts to control Ae. aegypti, especially for the methods using genetic modification of populations. [ABSTRACT FROM AUTHOR]

Background: Despite large reductions in malaria burden across Zambia, some regions continue to experience extremely high malaria transmission. In Nchelenge District, Luapula Province, northern Zambia, almost half the human population carries parasites. Intervention coverage has increased substantially over the past decade, but comprehensive district-wide entomological studies to guide delivery of vector control measures are lacking. This study describes the bionomics and spatio-temporal patterns of malaria vectors in Nchelenge over a two and a half year period, investigates what household factors are associated with high vector densities and determines why vector control may not have been effective in the past to better guide future control efforts. Methods: Between April 2012 and September 2014, twenty-seven households from across Nchelenge District were randomly selected for monthly light trap collections of mosquitoes. Anopheline mosquitoes were identified morphologically and molecularly to species. Foraging rates were estimated and sporozoite rates were determined by circumsporozoite ELISAs to calculate annual entomological inoculation rates. Blood feeding rates and host preference were determined by PCR. Zero-inflated negative binomial models measured environmental and household factors associated with mosquito abundance at study households such as season, proximity to the lake, and use of vector control measures. Results: The dominant species in Nchelenge was An. funestus (s.s.) with An. gambiae (s.s.) as a secondary vector. Both vectors were found together in large numbers across the district and the combined EIRs of the two vectors exceeded 80 infectious bites per person per annum. An. funestus household densities increased in the dry season whilst An. gambiae surged during the rains. Presence of insecticide treated nets (ITNs) and closed eaves in the houses were found to be associated with fewer numbers of An. gambiae but not An. funestus. There was no association with indoor residual spraying (IRS). Conclusion: In Nchelenge, the co-existence of two highly anthropophagic vectors, present throughout the year, is likely to be driving the high malaria transmission evident in the district. The vectors here have been shown to be highly resistant to pyrethroids used for IRS during the study. Vector control interventions in this area would have to be multifaceted and district-wide for effective control of malaria. [ABSTRACT FROM AUTHOR]

Background: Constant and fluctuating temperatures influence important life-history parameters of malaria vectors which has implications for community organization and the malaria disease burden. The effects of environmental temperature on the hatch rate, survivorship and development rate of Anopheles arabiensis and An. quadriannulatus under conditions of inter- and intra-specific competition are studied. Methods: The eggs and larvae of laboratory established colonies were reared under controlled conditions at one constant (25 °C) and two fluctuating (20-30 °C and 18-35 °C) temperature treatments at a ratio of 1:0 or 1:1 (An. arabiensis: An. quadriannulatus). Monitoring of hatch rate, development rate and survival was done at three intervals, 6 to 8 h apart depending on developmental stage. Parametric ANOVAs were used where assumptions of equal variances and normality were met, and a Welch ANOVA where equal variance was violated (a = 0.05). Results: Temperature significantly influenced the measured life-history traits and importantly, this was evident when these species co-occurred. A constant temperature resulted in a higher hatch rate in single species, larval treatments (P < 0.05). The treatment 18-35 °C generally reduced survivorship except for An. arabiensis in mixed, larval species treatments where it was similar to values reported for 25 °C. Survivorship of both species at 20-30 °C was not significantly impacted and the adult production was high across species treatments. The development rates at 25 °C and 20-30 °C were significantly different between species when reared alone and in mixed species from larvae and from eggs. The effect of temperature was more pronounced at 18-35 °C with An. arabiensis developing faster under both competitive scenarios and An. quadriannulatus slower, notably when in the presence of its competitor (P < 0.05). Conclusions: The influence of temperature treatment on the development rate and survival from egg/larvae to adult differed across species treatments. Fluctuating temperatures incorporating the extremes influence the key life-history parameters measured here with An. arabiensis outcompeting An. quadriannulatus under these conditions. The quantification of the response variables measured here improve our knowledge of the link between temperature and species interactions and provide valuable information for modelling of vector population dynamics. [ABSTRACT FROM AUTHOR]

Background: Insecticide resistance carries the potential to undermine the efficacy of insecticide based malaria vector control strategies. Therefore, there is an urgent need for new insecticidal compounds. Black pepper (dried fruit from the vine, Piper nigrum), used as a food additive and spice, and its principal alkaloid piperine, have previously been shown to have larvicidal properties. The aim of this study was to investigate the larvicidal effects of ground black pepper and piperine against third and fourth instar Anopheles larvae drawn from several laboratory-reared insecticide resistant and susceptible strains of Anopheles arabiensis, An. coluzzii, An. gambiae, An. quadriannulatus and An. funestus. Methods: Larvae were fed with mixtures of standard larval food and either ground black pepper or piperine in different proportions. Mortality was recorded 24 h after black pepper and 48 h after piperine were applied to the larval bowls. Results: Black pepper and piperine mixtures caused high mortality in the An. gambiae complex strains, with black pepper proving significantly more toxic than piperine. The An. funestus strains were substantially less sensitive to black pepper and piperine which may reflect a marked difference in the feeding habits of this species compared to that of the Gambiae complex or a difference in food metabolism as a consequence of differences in breeding habitat between species. Conclusions: Insecticide resistant and susceptible strains by species proved equally susceptible to black pepper and piperine. It is concluded that black pepper shows potential as a larvicide for the control of certain malaria vector species. [ABSTRACT FROM AUTHOR]

Background: Malaria transmission, primarily mediated by Anopheles gambiae, persists in Dar es Salaam (DSM) despite high coverage with bed nets, mosquito-proofed housing and larviciding. New or improved vector control strategies are required to eliminate malaria from DSM, but these will only succeed if they are delivered to the minority of locations where residual transmission actually persists. Hotspots of spatially clustered locations with elevated malaria infection prevalence or vector densities were, therefore, mapped across the city in an attempt to provide a basis for targeting supplementary interventions. Methods: Two phases of a city-wide population-weighted random sample of cross-sectional household surveys of malaria infections were complemented by two matching phases of geographically overlapping, high-resolution, longitudinal vector density surveys; spanning 2010-2013. Spatial autocorrelations were explored using Moran's I and hotspots were detected using flexible spatial scan statistics. Results: Seven hotspots of spatially clustered elevated vector density and eight of malaria infection prevalence were detected over both phases. Only a third of vectors were collected in hotspots in phase 1 (30 %) and phase 2 (33 %). Malaria prevalence hotspots accounted for only half of malaria infections detected in phase 1 (55 %) and phase 2 (47 %). Three quarters (76 % in phase 1 and 74 % in phase 2) of survey locations with detectable vector populations were outside of hotspots. Similarly, more than half of locations with higher infection prevalence (>10 %) occurred outside of hotspots (51 % in phase 1 and 54 % in phase 2). Vector proliferation hazard (exposure to An. gambiae) and malaria infection risk were only very loosely associated with each other (Odds ratio (OR) [95 % Confidence Interval (CI)] = 1.56 [0.89, 1.78], P = 0.52)). Conclusion: Many small, scattered loci of local malaria transmission were haphazardly scattered across the city, so interventions targeting only currently identifiable spatially aggregated hotspots will have limited impact. Routine, spatially comprehensive, longitudinal entomological and parasitological surveillance systems, with sufficient sensitivity and spatial resolution to detect these scattered loci, are required to eliminate transmission from this typical African city. Intervention packages targeted to both loci and hotspots of transmission will need to suppress local vector proliferation, treat infected residents and provide vulnerable residents with supplementary protective measures against exposure. [ABSTRACT FROM AUTHOR]

Background: On-going malaria transmission is increasingly mediated by outdoor-biting vectors, especially where indoor insecticidal interventions such as long-lasting insecticide treated nets (LLINs) are widespread. Often, the vectors are also physiologically resistant to insecticides, presenting major obstacles for elimination. We tested a combination of electrocuting grids with synthetic odours as an alternative killing mechanism against outdoor-biting mosquitoes. Methods: An odour-baited device, the Mosquito Landing Box (MLB), was improved by fitting it with low-cost electrocuting grids to instantly kill mosquitoes attracted to the odour lure, and automated photo switch to activate attractant-dispensing and mosquito-killing systems between dusk and dawn. MLBs fitted with one, two or three electrocuting grids were compared outdoors in a malaria endemic village in Tanzania, where vectors had lost susceptibility to pyrethroids. MLBs with three grids were also tested in a large semi-field cage (9.6×9.6×4.5m), to assess effects on biting-densities of laboratory-reared Anopheles arabiensis on volunteers sitting near MLBs. Results: Significantly more mosquitoes were killed when MLBs had two or three grids, than one grid in wet and dry seasons (P<0.05). The MLBs were highly efficient against Mansonia species and malaria vector, An. arabiensis. Of all mosquitoes, 99% were non-blood fed, suggesting host-seeking status. In the semi-field, the MLBs reduced mean number of malaria mosquitoes attempting to bite humans fourfold. Conclusion: The improved odour-baited MLBs effectively kill outdoor-biting malaria vector mosquitoes that are behaviourally and physiologically resistant to insecticidal interventions e.g. LLINs. The MLBs reduce human-biting vector densities even when used close to humans, and are insecticide-free, hence potentially antiresistance. The devices could either be used as surveillance tools or complementary mosquito control interventions to accelerate malaria elimination where outdoor transmission is significant. [ABSTRACT FROM AUTHOR]

Background: The SEMOS gold mine's malaria vector control programme forms part of the company's community responsibilities with the programme being managed by the mine's health department since 2005. Methods: Data from approximately 10 years of malaria vector control for the Sadiola District are given: namely malaria vector control methods used by the control programme, positive malaria case data and entomological surveys from 2006, 2011 and 2014. Results: Distribution of pyrethroid-treated bed nets and indoor residual spraying (IRS) with deltamethrin were implemented by the programme from 2005-2011. No IRS was done in 2012. Spraying with the organophosphate, pirimiphos-methyl resumed in 2013 and 2014 and was followed by a 70% drop in malaria cases in 2014. Anopheles arabiensis was the major vector present in 2006 and was susceptible to deltamethrin. In 2011 and 2014, An. gambiae s.s. was the most abundant vector with deltamethrin 24 h mortality of 68% and 19%, respectively. Conclusions: Resistance to the pyrethroid deltamethrin has increased in An. gambiae s.s. since 2011, possibly due to the scale-up in distribution of long-lasting insecticide-treated bed nets. Resistance management strategies are recommended using different classes of insecticides for IRS, and including the distribution of new-generation bed nets. [ABSTRACT FROM AUTHOR]

Background: Malaria vector control relies principally on the use of insecticides, especially pyrethroids. Because of the increasing occurrence of insecticide resistance in target vector populations, the development of new insecticides, particularly those with novel modes of action, is particularly important, especially in terms of managing insecticide resistance. The C8910 formulation is a patented mixture of compounds comprising straight-chain octanoic, nonanoic and decanoic saturated fatty acids. This compound has demonstrated toxic and repellent effects against several arthropod species. The aims of this study were to measure the insecticidal effects of C8910 against an insecticide susceptible (FANG) and a pyrethroid resistant (FUMOZ-R) laboratory strain of An. funestus as well as against wild-caught An. funestus material from Zambia (ZamF), and to investigate the repellent effects of two formulations of C8910 against these strains. Methods: Toxicity against adult females was assessed using a range of concentrations based on the CDC bottle bioassay method and repellence of three different C8910 formulations was assessed using standard choice-chamber bioassays. Results: C8910 proved equally toxic to adult females of the FUMOZ-R and FANG laboratory strains, as well as to adult females of the wild-caught (ZamF) sample. None of the C8910 formulations tested gave any conclusive indication of repellence against any of the strains. Conclusion: C8910 is equally effective as an adulticide against pyrethroid resistant and insecticide susceptible An. funestus. However, the formulations tested did not show any consistent repellence against laboratory reared and wild-caught female samples of this species. Nevertheless, C8910 shows potential as an adulticide that can be used for malaria vector control, particularly in those instances where insecticide resistance management is required. [ABSTRACT FROM AUTHOR]

Background: Malaria control in Africa relies heavily on indoor vector management, primarily indoor residual spraying and insecticide treated bed nets. Little is known about outdoor biting behaviour or even the dynamics of indoor biting and infection risk of sleeping household occupants. In this paper we explore the preferred biting sites on the human body and some of the ramifications regarding infection risk and exposure management. Methods: We undertook whole-night human landing catches of Anopheles arabiensis in South Africa and Anopheles gambiae s.s. and Anopheles funestus in Uganda, for seated persons wearing short sleeve shirts, short pants, and bare legs, ankles and feet. Catches were kept separate for different body regions and capture sessions. All An. gambiae s.l. and An. funestus group individuals were identified to species level by PCR. Results: Three of the main vectors of malaria in Africa (An. arabiensis, An. gambiae s.s. and An. funestus) all have a preference for feeding close to ground level, which is manifested as a strong propensity (77.3% - 100%) for biting on lower leg, ankles and feet of people seated either indoors or outdoors, but somewhat randomly along the lower edge of the body in contact with the surface when lying down. If the lower extremities of the legs (below mid-calf level) of seated people are protected and therefore exclude access to this body region, vector mosquitoes do not move higher up the body to feed at alternate body sites, instead resulting in a high (58.5% - 68.8%) reduction in biting intensity by these three species. Conclusions: Protecting the lower limbs of people outdoors at night can achieve a major reduction in biting intensity by malaria vector mosquitoes. Persons sleeping at floor level bear a disproportionate risk of being bitten at night because this is the preferred height for feeding by the primary vector species. Therefore it is critical to protect children sleeping at floor level (bednets; repellent-impregnated blankets or sheets, etc.). Additionally, the opportunity exists for the development of inexpensive repellent-impregnated anklets and/or sandals to discourage vectors feeding on the lower legs under outdoor conditions at night. [ABSTRACT FROM AUTHOR]

Background Two mitochondrial DNA clades have been described in Anopheles funestus populations from southern Africa. Clade I is common across the continent while clade II is known only from Mozambique and Madagascar. The specific biological status of these clades is at present unknown. We investigated the possible role that each clade might play in the transmission of Plasmodium falciparum and the insecticide resistance status of An. funestus from Zimbabwe and Zambia. Methods Mosquitoes were collected inside houses from Nchelenge District, Zambia and Honde Valley, Zimbabwe in 2013 and 2014. WHO susceptibility tests, synergist assays and resistance intensity tests were conducted on wild females and progeny of wild females. ELISA was used to detect Plasmodium falciparum circumsporozoite protein. Specimens were identified to species and mtDNA clades using standard molecular methods. Results The Zimbabwean samples were all clade I while the Zambian population comprised 80% clade I and 20% clade II in both years of collection. ELISA tests gave an overall infection rate of 2.3% and 2.1% in 2013, and 3.5% and 9.2% in 2014 for Zimbabwe and Zambia respectively. No significant difference was observed between the clades. All populations were resistant to pyrethroids and carbamates but susceptible to organochlorines and organophosphates. Synergist assays indicated that pyrethroid resistance is mediated by cytochrome P450 mono-oxygenases. Resistance intensity tests showed high survival rates after 8-hrs continuous exposure to pyrethroids but exposure to bendiocarb gave the same results as the susceptible control. Conclusions This is the first record of An. funestus mtDNA clade II occurring in Zambia. No evidence was found to suggest that the clades are markers of biologically separate populations. The ability of An. funestus to withstand prolonged exposure to pyrethroids has serious implications for the use of these insecticides, either through LLINs or IRS, in southern Africa in general and resistance management strategies should be urgently implemented. [ABSTRACT FROM AUTHOR]

Adult mosquito survival is strongly temperature and moisture dependent. Few studies have investigated the interacting effects of these variables on adult survival and how this differs among the sexes and with age, despite the importance of such information for population dynamic models. For these reasons, the desiccation tolerance of Anopheles arabiensis Patton and Anopheles funestus Giles males and females of three different ages was assessed under three combinations of temperature and humidity. Females were more desiccation tolerant than males, surviving for longer periods than males under all experimental conditions. In addition, younger adults were more tolerant of desiccation than older groups. Both species showed reduced water loss rate (WLR) as the primary mechanism by which they tolerate desiccation. Although A. arabiensis is often considered to be the more arid-adapted of the two species, it showed lower survival times and higher WLR than A. funestus. The current information could improve population dynamic models of these vectors, given that adult survival information for such models is relatively sparse. [ABSTRACT FROM AUTHOR]

Background: Two mitochondrial DNA clades have been described in Anopheles funestus populations from southern Africa. Clade I is common across the continent while clade II is known only from Mozambique and Madagascar. The specific biological status of these clades is at present unknown. We investigated the possible role that each clade might play in the transmission of Plasmodium falciparum and the insecticide resistance status of An. funestus from Zimbabwe and Zambia. Methods: Mosquitoes were collected inside houses from Nchelenge District, Zambia and Honde Valley, Zimbabwe in 2013 and 2014. WHO susceptibility tests, synergist assays and resistance intensity tests were conducted on wild females and progeny of wild females. ELISA was used to detect Plasmodium falciparum circumsporozoite protein. Specimens were identified to species and mtDNA clades using standard molecular methods. Results: The Zimbabwean samples were all clade I while the Zambian population comprised 80% clade I and 20% clade II in both years of collection. ELISA tests gave an overall infection rate of 2.3% and 2.1% in 2013, and 3.5% and 9.2% in 2014 for Zimbabwe and Zambia respectively. No significant difference was observed between the clades. All populations were resistant to pyrethroids and carbamates but susceptible to organochlorines and organophosphates. Synergist assays indicated that pyrethroid resistance is mediated by cytochrome P450 mono-oxygenases. Resistance intensity tests showed high survival rates after 8-hrs continuous exposure to pyrethroids but exposure to bendiocarb gave the same results as the susceptible control. Conclusions: This is the first record of An. funestus mtDNA clade II occurring in Zambia. No evidence was found to suggest that the clades are markers of biologically separate populations. The ability of An. funestus to withstand prolonged exposure to pyrethroids has serious implications for the use of these insecticides, either through LLINs or IRS, in southern Africa in general and resistance management strategies should be urgently implemented. [ABSTRACT FROM AUTHOR]

Background Malaria vector control in Sudan relies mainly on indoor residual spraying (IRS) and the use of long lasting insecticide treated bed nets (LLINs). Monitoring insecticide resistance in the main Sudanese malaria vector, Anopheles arabiensis, is essential for planning and implementing an effective vector control program in this country. Methods WHO susceptibility tests were used to monitor resistance to insecticides from all four WHOapproved classes of insecticide at four sentinel sites in Gezira state over a three year period. Insecticide resistance mechanisms were studied using PCR and microarray analyses. Results WHO susceptibility tests showed that Anopheles arabiensis from all sites were fully susceptible to bendiocarb and fenitrothion for the duration of the study (2008-2011). However, resistance to DDT and pyrethroids was detected at three sites, with strong seasonal variations evident at all sites. The 1014 F kdr allele was significantly associated with resistance to pyrethroids and DDT (P < 0.001) with extremely high effects sizes (OR > 7 in allelic tests). The 1014S allele was not detected in any of the populations tested. Microarray analysis of the permethrin-resistant population of An. arabiensis from Wad Medani identified a number of metabolic genes that were significantly over-transcribed in the field-collected resistant samples when compared to the susceptible Sudanese An. arabiensis Dongola strain. These included CYP6M2 and CYP6P3, two genes previously implicated in pyrethroid resistance in Anopheles gambiae s.s, and the epsilon-class glutathione-S-transferase, GSTe4. Conclusions These data suggest that both target-site mechanisms and metabolic mechanisms play an important role in conferring pyrethroid resistance in An. arabiensis from Sudan. Identification in An. arabiensis of candidate loci that have been implicated in the resistance phenotype in An. gambiae requires further investigation to confirm the role of these genes. [ABSTRACT FROM AUTHOR]

Background Fungal biopesticides are of great interest to vector control scientists as they provide a novel and environmentally friendly alternative to insecticide use. The aim of this study was to determine whether genes associated with pyrethroid resistance in Anopheles arabiensis from Sudan and South Africa are further induced following exposure to the entomopathogenic fungus, Beauveria bassiana (strain GHA). Methods Following B. bassiana bioassays, RNA was extracted from infected mosquitoes and the transcription of four important insecticide resistance genes, CYP9L1, CYP6M2 and CYP4G16 (cytochrome P450s) and TPX4 (thioredoxin peroxidase) was investigated using quantitative real-time PCR. Results Beauveria bassiana strain GHA was highly infective and virulent against An. arabiensis. In terms of changes in gene transcription, overall, the fold change (FC) values for each gene in the infected strains, were lower than 1.5. The FC values of CYP9L1, CYP6M2 and TPX4, were significantly lower than the FC values of the same genes in uninfected resistant An. arabiensis. Conclusion These data suggest that B. bassiana does not enhance the pyrethroid resistant phenotype on a molecular level as the two An. arabiensis strains used here, with different pyrethroid resistance mechanisms, revealed no increase in pre-existing metabolic transcripts. This supports the fact that fungal pathogens are suitable candidates for vector control, particularly with regard to the development of novel vector control strategies. [ABSTRACT FROM AUTHOR]

Background Malaria control programmes across Africa and beyond are facing increasing insecticide resistance in the major anopheline vectors. In order to preserve or prolong the effectiveness of the main malaria vector interventions, up-to-date and easily accessible insecticide resistance data that are interpretable at operationally-relevant scales are critical. Herein we introduce and demonstrate the usefulness of an online mapping tool, IR Mapper. Methods A systematic search of published, peer-reviewed literature was performed and Anopheles insecticide susceptibility and resistance mechanisms data were extracted and added to a database after a two-level verification process. IR Mapper (www.irmapper.com) was developed using the ArcGIS for JavaScript Application Programming Interface and ArcGIS Online platform for exploration and projection of these data. Results Literature searches yielded a total of 4,084 susceptibility data points for 1,505 populations, and 2,097 resistance mechanisms data points for 1,000 populations of Anopheles spp. collected via recommended WHO methods from 54 countries between 1954 and 2012. For the Afrotropical region, data were most abundant for populations of An. gambiae, and pyrethroids and DDT were more commonly used in susceptibility assays (51.1 and 26.8% of all reports, respectively) than carbamates and organophosphates. Between 2001 and 2012, there was a clear increase in prevalence and distribution of confirmed resistance of An. gambiae s.l. to pyrethroids (from 41 to 87% of the mosquito populations tested) and DDT (from 64 to 91%) throughout the Afrotropical region. Metabolic resistance mechanisms were detected in western and eastern African populations and the two kdr mutations (L1014S and L1014F) were widespread. For An. funestus s.l., relatively few populations were tested, although in 2010-2012 resistance was reported in 50% of 10 populations tested. Maps are provided to illustrate the use of IR Mapper and the distribution of insecticide resistance in malaria vectors in Africa. Conclusions The increasing pyrethroid and DDT resistance in Anopheles in the Afrotropical region is alarming. Urgent attention should be afforded to testing An. funestus populations for metabolic resistance mechanisms. IR Mapper is a useful tool for investigating temporal and spatial trends in Anopheles resistance to support the pragmatic use of insecticidal interventions. [ABSTRACT FROM AUTHOR]

Background Limited information is available on malaria vector composition, feeding habits and malaria transmission in northern Malawi. Evidence of mosquito species diversity in this area was established in 2009, when Anopheles funestus-like, a new member of the An. funestus group was described. Additional biological information is needed to identify this species and to understand its role in malaria transmission. Methods Anopheline mosquitoes were collected in northern Malawi and analyzed for Plasmodium species infection, blood meal source and susceptibility to insecticides. A new hydrolysis probe assay was designed to identify An. funestus-like. Results Anopheles funestus and An. rivulorum predominated in the indoor collections. Most An. funestus-like were collected indoors, mainly fed on animals and were uninfected with P. falciparum. Anopheles funestus showed insecticide resistance to deltamethrin and bendiocarb. A high-precision hydrolysis probe assay was successfully developed to identify An. funestus-like. Discussion Four species in the An. funestus group were collected in Karonga. Resistance to deltamethrin and bendiocarb was observed in An. funestus and further investigation is needed on the insecticide resistance mechanisms. Anopheles funestus-like, while collected indoors, is mainly zoophilic and most likely not a malaria vector. Accession numbers An. funestus (GenBank accession no. KC771136), An. funestus-like (GenBank accession no. KC771137), An. parensis GenBank accession no. KC771138) and An. vaneedeni GenBank accession no. KC771139). [ABSTRACT FROM AUTHOR]

Background: Recent studies presented two clades (clades I and II) within the major malaria vector, Anopheles funestus s.s. on the mitochondrial DNA. We describe a hydrolysis probe analysis (Taqman assay) method for the rapid identification of these two clades. Findings: A total of 53 An. funestus s.s. from Malawi and Mozambique were tested for detection of clade types using the hydrolysis probe analysis. Results were compared to DNA sequence analysis to verify the accuracy of the probes Taqman assay for this vector species. Analysis using the hydrolysis probe revealed that there were 21 individuals from Malawi and 13 individuals from Mozambique for clade I, and 19 individuals from Mozambique for clade II. The results were consistent with the results of DNA sequences. A field sample from northern Zambia revealed the presence of both clade types. Conclusion: A diagnostic method using the hydrolysis probe analysis was developed to identify clade types within An. funestus s.s. This assay will be useful for screening clade types of field-collected An. funestus specimens accurately and efficiently in malaria vector research and control studies. [ABSTRACT FROM AUTHOR]

Background: Anopheles funestus is one of the major malaria vectors in Africa but research on this species has been restricted due to the lack of viable laboratory colonies. The vectorial capacity of natural populations of An. funestus is well known but its ability to host Plasmodium in the laboratory and the development cycle of the parasite within this mosquito species was, until very recently, unknown. In this study we compared laboratory strains of An. funestus that were resistant and susceptible to pyrethroid insecticides, for their receptiveness to infection with Plasmodium berghei and compared development times with other vector species available in our laboratory. Methods: The murine parasite P. berghei was used to infect two base An. funestus colonies (FANG and FUMOZ) and two selected sub-colonies with different degrees of pyrethroid resistance (FUMOZ-BS susceptible and FUMOZ-R resistant). Results were compared with the G3 strain of An. gambiae. Results: While all colonies were able to support the parasite, the development time in An. funestus was generally longer than that recorded in the laboratory strain of An. gambiae. Infected females were able to initiate new rounds of infection when feeding on healthy mice. The pyrethroid resistant strain FUMOZ-R supported the lowest numbers of oocysts and sporozoites while the insecticide susceptible strain FUMOZ-BS produced one of the highest sporozoite indices ever documented in P. berghei research. The FUMOZ base colony, exhibiting partial insecticide resistance was the median in terms of infection intensity. The oocyst number in all colonies did not fully correlate with the sporozoite index, indicating possible factors influencing the sporozoites' transit from the midgut to the salivary glands. Conclusions: The presence of both insecticide resistance and limited parasite infection phenotypes in the same individuals suggests there may be association between the two mechanisms, but further elucidation is required. [ABSTRACT FROM AUTHOR]

Background: Understanding the biology of malaria vector mosquitoes is crucial to understanding many aspects of the disease, including control and future outcomes. The development rates and survival of two Afrotropical malaria vectors, Anopheles arabiensis and Anopheles funestus, are investigated here under conditions of constant and fluctuating temperatures. These data can provide a good starting point for modelling population level consequences of temperature change associated with climate change. For comparative purposes, these data were considered explicitly in the context of those available for the third African malaria vector, Anopheles gambiae. Methods: Twenty five replicates of 20-30 eggs were placed at nine constant and two fluctuating temperatures for development rate experiments and survival estimates. Various developmental parameters were estimated from the data, using standard approaches. Results: Lower development threshold (LDT) for both species was estimated at 13-14°C. Anopheles arabiensis developed consistently faster than An. funestus. Optimum temperature (Topt) and development rate at this temperature (µmax) differed significantly between species for overall development and larval development. However, Topt and µmax for pupal development did not differ significantly between species. Development rate and survival of An. funestus was negatively influenced by fluctuating temperatures. By contrast, development rate of An. arabiensis at fluctuating temperatures either did not differ from constant temperatures or was significantly faster. Survival of this species declined by c. 10% at the 15°C to 35°C fluctuating temperature regime, but was not significantly different between the constant 25°C and the fluctuating 20°C to 30°C treatment. By comparison, previous data for An. gambiae indicated fastest development at a constant temperature of 28°C and highest survival at 24°C. Conclusions: The three most important African malaria vectors all differ significantly in development rates and survival under different temperature treatments, in keeping with known distribution data, though differences among M and S molecular forms of An. gambiae likely complicate the picture. Increasing temperatures associated with climate change favour all three species, but fluctuations in temperatures are detrimental to An. funestus and may also be for An. gambiae. This may have significant implications for disease burden in areas where each species is the main malaria vector. [ABSTRACT FROM AUTHOR]

Background: Long-lasting insecticide treated nets (LLINs) and indoor residual house spraying (IRS) are the main interventions for the control of malaria vectors in Zanzibar. The aim of the present study was to assess the susceptibility status of malaria vectors against the insecticides used for LLINs and IRS and to determine the durability and efficacy of LLINs on the island. Methods: Mosquitoes were sampled from Pemba and Unguja islands in 2010-2011 for use in WHO susceptibility tests. One hundred and fifty LLINs were collected from households on Unguja, their physical state was recorded and then tested for efficacy as well as total insecticide content. Results: Species identification revealed that over 90% of the Anopheles gambiae complex was An. arabiensis with a small number of An. gambiae s.s. and An. merus being present. Susceptibility tests showed thatAn. arabiensis on Pemba was resistant to the pyrethroids used for LLINs and IRS. Mosquitoes from Unguja Island, however, were fully susceptible to all pyrethroids tested. A physical examination of 150 LLINs showed that two thirds were damaged after only three years in use. All used nets had a significantly lower (p < 0.001) mean permethrin concentration of 791.6 mg/m2 compared with 944.2 mg/m² for new ones. Their efficacy decreased significantly against both susceptible An. gambiae s.s. colony mosquitoes and wild-type mosquitoes from Pemba after just six washes (p < 0.001). Conclusion: The sustainability of the gains achieved in malaria control in Zanzibar is seriously threatened by the resistance of malaria vectors to pyrethroids and the short-lived efficacy of LLINs. This study has revealed that even in relatively well-resourced and logistically manageable places like Zanzibar, malaria elimination is going to be difficult to achieve with the current control measures. [ABSTRACT FROM AUTHOR]

Anopheles funestus is one of three major African vectors of malaria. Its distribution extends over much of the tropics and subtropics wherever suitable swampy breeding habitats are present. As with members of the Anopheles gambiae complex, An. funestus shows marked genetic heterogeneity across its range. Currently, two unnamed species are recognized in the group, with molecular and cytogenetic data indicating that more may be present. The control of malaria vectors in Africa has received increased attention in the past decade with the scaling up of insecticide-treated bed nets and indoor residual house spraying. Also in the past decade, the frequency of insecticide-resistant mosquitoes has increased exponentially. Whether this increase is in response to vector control initiatives or because of insecticide use in agriculture is debatable. In this article we examine the progress made on the systematics of the An. funestus group and review research on insecticide resistance and its mechanisms. [ABSTRACT FROM AUTHOR]

Background: Anopheles funestus s.s., one of the major malaria vectors in sub-Saharan Africa, belongs to a group of eleven African species that are morphologically similar at the adult stage, most of which do not transmit malaria. The population structure of An. funestus based on mitochondrial DNA data led to the description of two cryptic subdivisions, clade I widespread throughout Africa and clade II known only from Mozambique and Madagascar. In this study, we investigated five common members of the Anopheles funestus group in southern Africa in order to determine relationships within and between species. Methods: A total of 155 specimens of An. funestus, An. parensis, An. vaneedeni, An. funestus-like and An. rivulorum from South Africa, Mozambique and Malawi were used for the study. The population genetic structure was assessed within and between populations using mitochondrial DNA. Results: The phylogenetic trees revealed three main lineages: 1) An. rivulorum; 2) An. funestus-like clade I and An. parensis clade II; and 3) An. funestus clades I and II, An. funestus-like clade II, An. parensis clade I and An. vaneedeni clades I and II. Within An. funestus, 32 specimens from Mozambique consisted of 40.6% clade I and 59.4% clade II while all 21 individuals from Malawi were clade I. In the analysis of mitochondrial DNA sequences, there were 37 polymorphic sites and 9 fixed different nucleotides for ND5 and 21 polymorphic sites and 6 fixed different nucleotides for COI between the two An. funestus clades. The results for COI supported the ND5 analysis. Conclusion: This is the first report comparing An. funestus group species including An. funestus clades I and II and the new species An. funestus-like. Anopheles funestus clade I is separated from the rest of the members of the An. funestus subgroup and An. funestus-like is distinctly distributed from the other species in this study. However, there were two clades for An. funestus-like, An. parensis and An. vaneedeni. Further investigations are needed to determine what these results mean in terms of the specific status of the clades within each taxon and whether this has any epidemiological implications for malaria transmission. [ABSTRACT FROM AUTHOR]

Abstract: Background: The use of insecticides to control malaria vectors is essential to reduce the prevalence of malaria and as a result, the development of insecticide resistance in vector populations is of major concern. Anopheles arabiensis is one of the main African malaria vectors and insecticide resistance in this species has been reported in a number of countries. The aim of this study was to investigate the detoxification enzymes that are involved in An. arabiensis resistance to DDT and pyrethroids. Methods: The detoxification enzyme profiles were compared between two DDT selected, insecticide resistant strains of An. arabiensis, one from South Africa and one from Sudan, using the An. gambiae detoxification chip, a boutique microarray based on the major classes of enzymes associated with metabolism and detoxification of insecticides. Synergist assays were performed in order to clarify the roles of over-transcribed detoxification genes in the observed resistance phenotypes. In addition, the presence of kdr mutations in the colonies under investigation was determined. Results: The microarray data identifies several genes over-transcribed in the insecticide selected South African strain, while in the Sudanese population, only one gene, CYP9L1, was found to be over-transcribed. The outcome of the synergist experiments indicate that the over-transcription of detoxification enzymes is linked to deltamethrin resistance, while DDT and permethrin resistance are mainly associated with the presence of the L1014F kdr mutation. Conclusions: These data emphasise the complexity associated with resistance phenotypes and suggest that specific insecticide resistance mechanisms cannot be extrapolated to different vector populations of the same species. [ABSTRACT FROM AUTHOR]

Background: New products aimed at augmenting or replacing chemical insecticides must have operational profiles that include both high efficacy in reducing vector numbers and/or blocking parasite transmission and be long lasting following application. Research aimed at developing fungal spores as a biopesticide for vector control have shown considerable potential yet have not been directly assessed for their viability after long-term storage or following application in the field. Methods: Spores from a single production run of the entomopathogenic fungi Beauveria bassiana were dried and then stored under refrigeration at 7°C. After 585 days these spores were sub-sampled and placed at either 22°C, 26°C or 32°C still sealed in packaging (closed storage) or in open beakers and exposed to the 80% relative humidity of the incubator they were kept in. Samples were subsequently taken from these treatments over a further 165 days to assess viability. Spores from the same production run were also used to test their persistence following application to three different substrates, clay, cement and wood, using a hand held sprayer. The experiments were conducted at two different institutes with one using adult female Anopheles stephensi and the other adult female Anopheles gambiae. Mosquitoes were exposed to the treated substrates for one hour before being removed and their survival monitored for the next 14 days. Assays were performed at monthly intervals over a maximum seven months. Results: Spore storage under refrigeration resulted in no loss of spore viability over more than two years. Spore viability of those samples kept under open and closed storage was highly dependent on the incubation temperature with higher temperatures decreasing viability more rapidly than cooler temperatures. Mosquito survival following exposure was dependent on substrate type. Spore persistence on the clay substrate was greatest achieving 80% population reduction for four months against An. stephensi and for at least five months against Anopheles gambiae. Cement and wood substrates had more variable mortality with the highest spore persistence being two to three months for the two substrates respectively. Conclusions: Spore shelf-life under refrigeration surpassed the standard two year shelf-life expected of a mosquito control product. Removal to a variety of temperatures under either closed or open storage indicated that samples sent out from refrigeration should be deployed rapidly in control operations to avoid loss of viability. Spore persistence following application onto clay surfaces was comparable to a number of chemical insecticides in common use. Persistence on cement and wood was shorter but in one assay still comparable to some organophosphate and pyrethroid insecticides. Optimized formulations could be expected to improve spore persistence still further. [ABSTRACT FROM AUTHOR]

Background: Malaria affects large parts of the developing world and is responsible for almost 800,000 deaths annually. As climates change, concerns have arisen as to how this vector-borne disease will be impacted by changing rainfall patterns and warming temperatures. Despite the importance and controversy surrounding the impact of climate change on the potential spread of this disease, little information exists on the tolerances of several of the vector species themselves. Methods: Using a ramping protocol (to assess critical thermal limits -- CT) and plunge protocol (to assess lethal temperature limits -- LT) information on the thermal tolerance of two of Africa's important malaria vectors, Anopheles arabiensis and Anopheles funestus was collected. The effects of age, thermal acclimation treatment, sex and strain (laboratory versus wild adults) were investigated for CT determinations for each species. The effects of age and sex for adults and life stage (larvae, pupae, adults) were investigated for LT determinations. Results: In both species, females are more tolerant to low and high temperatures than males; larvae and pupae have higher upper lethal limits than do adults. Thermal acclimation of adults has large effects in some instances but small effects in others. Younger adults tend to be more tolerant of low or high temperatures than older age groups. Long-standing laboratory colonies are sufficiently similar in thermal tolerance to field-collected animals to provide reasonable surrogates when making inferences about wild population responses. Differences between these two vectors in their thermal tolerances, especially in larvae and pupae, are plausibly a consequence of different habitat utilization. Conclusions: Limited plasticity is characteristic of the adults of these vector species relative to others examined to date, suggesting limited scope for within-generation change in thermal tolerance. These findings and the greater tolerance of females to thermal extremes may have significant implications for future malaria transmission, especially in areas of current seasonal transmission and in areas on the boundaries of current vector distribution. [ABSTRACT FROM AUTHOR]

Rapidly emerging insecticide resistance is creating an urgent need for new active ingredients to control the adult mosquitoes that vector malaria. Biopesticides based on the spores of entomopathogenic fungi have shown considerable promise by causing very substantial mortality within 7-14 days of exposure. This mortality will generate excellent malaria control if there is a high likelihood that mosquitoes contact fungi early in their adult lives. However, where contact rates are lower, as might result from poor pesticide coverage, some mosquitoes will contact fungi one or more feeding cycles after they acquire malaria, and so risk transmitting malaria before the fungus kills them. Critics have argued that 'slow acting' fungal biopesticides are, therefore, incapable of delivering malaria control in real-world contexts. Here, utilizing standard WHO laboratory protocols, we demonstrate effective action of a biopesticide much faster than previously reported. Specifically, we show that transient exposure to clay tiles sprayed with a candidate biopesticide comprising spores of a natural isolate of Beauveria bassiana, could reduce malaria transmission potential to zero within a feeding cycle. The effect resulted from a combination of high mortality and rapid fungal-induced reduction in feeding and flight capacity. Additionally, multiple insecticide-resistant lines from three key African malaria vector species were completely susceptible to fungus. Thus, fungal biopesticides can block transmission on a par with chemical insecticides, and can achieve this where chemical insecticides have little impact. These results support broadening the current vector control paradigm beyond fastacting chemical toxins. [ABSTRACT FROM AUTHOR]

Background: Anopheles funestus is one of the major malaria vectors in Africa and yet there are few genomic tools available for this species compared to An. gambiae. To start to close this knowledge gap, we sequenced the An. funestus transcriptome using cDNA libraries developed from a pyrethroid resistant laboratory strain and a pyrethroid susceptible field strain from Mali. Results: Using a pool of life stages (pupae, larvae, adults: females and males) for each strain, 454 sequencing generated 375,619 reads (average length of 182 bp). De novo assembly generated 18,103 contigs with average length of 253 bp. The average depth of coverage of these contigs was 8.3. In total 20.8% of all reads were novel when compared to reference databases. The sequencing of the field strain generated 204,758 reads compared to 170,861 from the insecticide resistant laboratory strain. The contigs most differentially represented in the resistant strain belong to the P450 gene family and cuticular genes which correlates with previous studies implicating both of these gene families in pyrethroid resistance. qPCR carried out on six contigs indicates that these ESTs could be suitable for gene expression studies such as microarray. 31,000 sites were estimated to contain Single Nucleotide Polymorphisms (SNPs) and analysis of SNPs from 20 contigs suggested that most of these SNPs are likely to be true SNPs. Gene conservation analysis confirmed the close phylogenetic relationship between An. funestus and An. gambiae. Conclusion: This study represents a significant advance for the genetics and genomics of An. funestus since it provides an extensive set of both Expressed Sequence Tags (ESTs) and SNPs which can be readily adopted for the design of new genomic tools such as microarray or SNP platforms. [ABSTRACT FROM AUTHOR]

Background: Malaria vector control programmes that rely on insecticide-based interventions such as indoor house spraying with residual insecticides or insecticide treated bed nets, need to base their decision-making process on sound baseline data. More and more commercial entities in Africa, such as mining companies, are realising the value to staff productivity of controlling malaria transmission in their areas of operation. This paper presents baseline entomological data obtained during surveys conducted for four mining operations in Ghana, West Africa. Results: The vast majority of the samples were identified as Anopheles gambiae S form with only a few M form specimens being identified from Tarkwa. Plasmodium falciparum infection rates ranged from 4.5 to 8.6% in An. gambiae and 1.81 to 8.06% in An. funestus. High survival rates on standard WHO bioassay tests were recorded for all insecticide classes except the organophosphates that showed reasonable mortality at all locations (i.e. > 90%). The West African kdr mutation was detected and showed high frequencies in all populations. Conclusions: The data highlight the complexity of the situation prevailing in southern Ghana and the challenges facing the malaria vector control programmes in this region. Vector control programmes in Ghana need to carefully consider the resistance profiles of the local mosquito populations in order to base their resistance management strategies on sound scientific data. [ABSTRACT FROM AUTHOR]

Background: The dual problems of rising insecticide resistance in the malaria vectors and increasing human malaria cases since 2001 in southern Mozambique are cause for serious concern. The selection of insecticides for use in indoor residual spraying (IRS) programmes is highly dependent on the extent to which local mosquitoes are susceptible to the approved classes of insecticides. The insecticide resistance status and role in malaria transmission of Anopheles funestus was evaluated at the Maragra Sugar Estate in southern Mozambique where an IRS vector control programme has been in operation for seven years using the carbamate insecticide bendiocarb. Results: No Anopheles species were captured inside the sugar estate control area. Anopheles funestus group captured outside of the estate represented 90% (n = 475) of the total collections. Of the specimens identified to species by PCR (n = 167), 95% were An. funestus s.s. One An. rivulorum was identified and seven specimens did not amplify. The Anopheles gambiae complex was less abundant (n = 53) and of those identified (n = 33) 76% were An. arabiensis and 24% An. merus. Insecticide susceptibility tests showed that wild-caught and F-1 family An. funestus were resistant to deltamethrin (32.5% mortality) and lambda-cyhalothrin (14.6% mortality), less so to bendiocarb (71.5% mortality) and fully susceptible to both malathion and DDT (100%). Bendiocarb and pyrethroid resistance was nullified using 4% piperonyl butoxide (Pbo), strongly suggesting that both are mediated by P450 monooxygenase detoxification. ELISA tests of An. funestus for Plasmodium falciparum, gave a sporozoite rate of 6.02% (n = 166). One unidentified member of the An. gambiae complex tested positive for P. falciparum sporozoites. Conclusion: Anopheles funestus was found to be the most abundant and principle vector of malaria in this area, with members of the An. gambiae complex being secondary vectors. Despite the continual use of bendiocarb within the estate for seven years and the level of An. funestus resistance to this insecticide, the IVC programme is still effective against this and other Anopheles in that no vectors were found inside the control area. However, the Mozambique National Malaria Control Programme ceased the use of DDT and bendiocarb in this area of its operations in 2009, and replaced these insecticides with a pyrethroid which will increase insecticide resistance selection pressure and impact on control programmes such as the Maragra IVC. [ABSTRACT FROM AUTHOR]

Summary: A household matched case–control study design was used to explore associations between household characteristics and malaria risk in seven study towns in the hypoendemic area of Mpumalanga Province, South Africa. Controls were identified from neighboring households of each case. Principle component analysis was used to calculate a wealth index for households to allow comparison across socioeconomic groups. Conditional univariate and multiple logistic regression analyses were used to assess associations between household malaria risk and potential risk factors. Univariate analysis demonstrated an increased household malaria risk for people living in mud-walled houses compared with those in brick dwellings (OR=5.10, 95% CI 2.03–12.80, P =0.001). Multivariate analysis confirmed the association between malaria risk and mud-wall construction (OR=6.12, 95% CI 2.26–16.59, P =0.001) and demonstrated an association with opening windows after retiring to sleep (OR=4.01, 95% CI 1.32–12.18, P =0.014). An inverse association between household wealth, third (OR=0.24, 95% CI 0.09–0.65, P =0.005) and fourth quartiles (OR=0.27, 95% CI 0.10–0.79, P =0.016), and malaria risk was observed. Associations found here include increased household malaria risk and mud-wall construction, the practice of opening of windows at night and relative household poverty. Education campaigns targeting risk behavior may reduce malaria risk, but economic development is a more important intervention. [Copyright &y& Elsevier]

Background: This is the second in a series of three articles documenting the geographical distribution of 41 dominant vector species (DVS) of human malaria. The first paper addressed the DVS of the Americas and the third will consider those of the Asian Pacific Region. Here, the DVS of Africa, Europe and the Middle East are discussed. The continent of Africa experiences the bulk of the global malaria burden due in part to the presence of the An. gambiae complex. Anopheles gambiae is one of four DVS within the An. gambiae complex, the others being An. arabiensis and the coastal An. merus and An. melas. There are a further three, highly anthropophilic DVS in Africa, An. funestus, An. moucheti and An. nili. Conversely, across Europe and the Middle East, malaria transmission is low and frequently absent, despite the presence of six DVS. To help control malaria in Africa and the Middle East, or to identify the risk of its re-emergence in Europe, the contemporary distribution and bionomics of the relevant DVS are needed. Results: A contemporary database of occurrence data, compiled from the formal literature and other relevant resources, resulted in the collation of information for seven DVS from 44 countries in Africa containing 4234 georeferenced, independent sites. In Europe and the Middle East, six DVS were identified from 2784 geo-referenced sites across 49 countries. These occurrence data were combined with expert opinion ranges and a suite of environmental and climatic variables of relevance to anopheline ecology to produce predictive distribution maps using the Boosted Regression Tree (BRT) method. Conclusions: The predicted geographic extent for the following DVS (or species/suspected species complex*) is provided for Africa: Anopheles (Cellia) arabiensis, An. (Cel.) funestus*, An. (Cel.) gambiae, An. (Cel.) melas, An. (Cel.) merus, An. (Cel.) moucheti and An. (Cel.) nili*, and in the European and Middle Eastern Region: An. (Anopheles) atroparvus, An. (Ano.) labranchiae, An. (Ano.) messeae, An. (Ano.) sacharovi, An. (Cel.) sergentii and An. (Cel.) superpictus*. These maps are presented alongside a bionomics summary for each species relevant to its control. [ABSTRACT FROM AUTHOR]

Background: Anopheles gambiae is a major vector of malaria in the West African region. Resistance to multiple insecticides has been recorded in An. gambiae S form in the Ahafo region of Ghana. A laboratory population (GAH) established using wild material from this locality has enabled a mechanistic characterization of each resistance phenotype as well as an analysis of another adaptive characteristic - staggered larval time-to-hatch. Methods: Individual egg batches obtained from wild caught females collected from Ghana and the Republic of the Congo were monitored for staggered larval time-to-hatch. In addition, early and late larval time-to-hatch subcolonies were selected from GAH. These selected sub-colonies were cross-mated and their hybrid progeny were subsequently intercrossed and back-crossed to the parental strains. The insecticide susceptibilities of the GAH base colony and the time-to-hatch selected sub-colonies were quantified for four insecticide classes using insecticide bioassays. Resistance phenotypes were mechanistically characterized using insecticide-synergist bioassays and diagnostic molecular assays for known reduced target-site sensitivity mutations. Results: Anopheles gambiae GAH showed varying levels of resistance to all insecticide classes. Metabolic detoxification and reduced target-site sensitivity mechanisms were implicated. Most wild-caught families showed staggered larval time-to-hatch. However, some families were either exclusively early hatching or late hatching. Most GAH larvae hatched early but many egg batches contained a proportion of late hatching larvae. Crosses between the time-to-hatch selected sub-colonies yielded ambiguous results that did not fit any hypothetical models based on single-locus Mendelian inheritance. There was significant variation in the expression of insecticide resistance between the time-to-hatch phenotypes. Conclusions: An adaptive response to the presence of multiple insecticide classes necessarily involves the development of multiple resistance mechanisms whose effectiveness may be enhanced by intra-population variation in the expression of resistance phenotypes. The variation in the expression of insecticide resistance in association with selection for larval time-to-hatch may induce this kind of enhanced adaptive plasticity as a consequence of pleiotropy, whereby mosquitoes are able to complete their aquatic life stages in a variable breeding environment using staggered larval time-to-hatch, giving rise to an adult population with enhanced variation in the expression of insecticide resistance. [ABSTRACT FROM AUTHOR]