To optimize malaria control, WHO has prioritised the need for new indicators to evaluate the efficacy of malaria vector control strategies. The gSG6-P1 peptide from gSG6 protein of Anopheles gambiae salivary glands was previously designed as a specific salivary sequence of malaria vector species. It was shown that the quantification of human antibody (Ab) responses to Anopheles salivary proteins in general and especially to the gSG6-P1 peptide was a pertinent biomarker of human exposure to Anopheles. The present objective was to validate this indicator in the evaluation of the efficacy of Insecticide Treated Nets (ITNs). A longitudinal evaluation, including parasitological, entomological and immunological assessments, was conducted on children and adults from a malaria-endemic area before and after the introduction of ITNs. Significant decrease of anti-gSG6-P1 IgG response was observed just after the efficient ITNs use. Interestingly, specific IgG Ab level was especially pertinent to evaluate a short-time period of ITNs efficacy and at individual level. However, specific IgG rose back up within four months as correct ITN use waned. IgG responses to one salivary peptide could constitute a reliable biomarker for the evaluation of ITN efficacy, at short- and long-term use, and provide a valuable tool in malaria vector control based on a real measurement of human-vector contact.

Anopheles funestus is a major vector of malaria in Africa. It belongs to a group of sibling species that can be identified morphologically only at certain stages of their development. A diagnostic polymerase chain reaction (PCR)-based tool made it possible to differentiate five species of the group. The assay seems to be applicable over all their distribution area for four of these species: An. funestus, An. leesoni, An. parensis, and An. vaneedenip. The fifth species, An. rivulorum, is the second most abundant species of the group and can be mistaken at its adult stage for the major vector of malaria An. funestus. Molecular and morphologic observations of specimens identified as An. rivulorum from Cameroon and South Africa showed that they belong to two different taxa. The species identified in Cameroon, and named here An. rivulorum-like, might extend to western Africa and central Africa. The species-specific PCR assay is supplemented by a primer specific to An. rivulorum-like and thus makes it possible to differentiate the five species of the An. funestus group and the newly defined taxon.

BackgroundIncreasing incidence of DDT and pyrethroid resistance in Anopheles mosquitoes is seen as a limiting factor for malaria vector control. The current study aimed at an in-depth characterization of An. gambiae s.l. resistance to insecticides in Cameroon, in order to guide malaria vector control interventions.MethodsAnopheles gambiae s.l. mosquitoes were collected as larvae and pupae from six localities spread throughout the four main biogeographical domains of Cameroon and reared to adults in insectaries. Standard WHO insecticide susceptibility tests were carried out with 4% DDT, 0.75% permethrin and 0.05% deltamethrin. Mortality rates and knockdown times (kdt50 and kdt95) were determined and the effect of pre-exposure to the synergists DEF, DEM and PBO was assessed. Tested mosquitoes were identified to species and molecular forms (M or S) using PCR-RFLP. The hot ligation method was used to depict kdr mutations and biochemical assays were conducted to assess detoxifying enzyme activities.ResultsThe An. arabiensis population from Pitoa was fully susceptible to DDT and permethrin (mortality rates > 98%) and showed reduced susceptibility to deltamethrin. Resistance to DDT was widespread in An. gambiae s.s. populations and heterogeneous levels of susceptibility to permethrin and deltamethrin were observed. In many cases, prior exposure to synergists partially restored insecticide knockdown effect and increased mortality rates, suggesting a role of detoxifying enzymes in increasing mosquito survival upon challenge by pyrethroids and, to a lower extent DDT. The distribution of kdr alleles suggested a major role of kdr-based resistance in the S form of An. gambiae. In biochemical tests, all but one mosquito population overexpressed P450 activity, whereas baseline GST activity was low and similar in all field mosquito populations and in the control.ConclusionIn Cameroon, multiple resistance mechanisms segregate in the S form of An. gambiae resulting in heterogeneous resistance profiles, whereas in the M form and An. arabiensis insecticide tolerance seems to be essentially mediated by enzyme-based detoxification. Synergists partially restored susceptibility to pyrethroid insecticides, and might help mitigate the impact of vector resistance in the field. However, additional vector control tools are needed to further impact on malaria transmission in such settings.

A large-scale survey of Anopheles gambiae Giles, 1902 susceptibility to DDT, dieldrin, permethrin, and deltamethrin was conducted in the Republic of Cameroon. 15 field populations from various geographical areas were tested using World Health Organization test kits for adult mosquitoes. The laboratory Kisumu susceptible reference strain was tested as a control. Results showed that dieldrin and DDT resistance was still present in some populations, and indicated permethrin or deltamethrin resistance. Within the Anopheles gambiae complex, resistant individuals belonged to An. gambiae s.s. and An. arabiensis species. Both M and S molecular forms of An. gambiae s.s. were found resistant. In most of resistant populations, the knockdown times were 2-5-folds increased. However, none of the surviving mosquitoes was positive to the kdr "Leu-Phe" mutation using polymerase chain reaction (PCR) diagnostic test. These results likely suggested involvement of other resistance mechanism(s), such as enzyme detoxification or kdr "Leu-Ser" mutation. Researches on An. gambiae s.l. resistance should be promoted in Cameroon, to improve malaria vector control programs and to implement resistance management strategies.

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BackgroundPyrethroid insecticides are widely used for insect pest control in Cameroon. In certain insect species, particularly the malaria vector Anopheles gambiae, resistance to this class of insecticides is a source of great concern and needs to be monitored in order to sustain the efficacy of vector control operations in the fields. This study highlights trends in DDT and pyrethroid resistance in wild An. gambiae populations from South Cameroon.MethodsMosquitoes were collected between 2001 and 2007 in four sites in South Cameroon, where insecticides are used for agricultural or personal protection purposes. Insecticide use was documented in each site by interviewing residents. Batches of 2-4 days old adult female mosquitoes reared from larval collections were tested for susceptibility to DDT, permethrin and deltamethrin using standard WHO procedures. Control, dead and survivors mosquitoes from bioassays were identified by PCR-RFLP and characterized for the kdr mutations using either the AS-PCR or the HOLA method.ResultsFour chemical insecticide groups were cited in the study sites: organochlorines, organophosphates, carbamates and pyrethroids. These chemicals were used for personal, crop or wood protection. In the four An. gambiae populations tested, significant variation in resistance levels, molecular forms composition and kdr frequencies were recorded in the time span of the study. Increases in DDT and pyrethroid resistance, as observed in most areas, were generally associated with an increase in the relative frequency of the S molecular form carrying the kdr mutations at higher frequencies. In Mangoum, however, where only the S form was present, a significant increase in the frequency of kdr alleles between 2003 to 2007 diverged with a decrease of the level of resistance to DDT and pyrethroids. Analyses of the kdr frequencies in dead and surviving mosquitoes showed partial correlation between the kdr genotypes and resistance phenotypes, suggesting that the kdr mechanism may act with certain co-factors to be identified.ConclusionThese results demonstrate the ongoing spread of kdr alleles in An. gambiae in Central Africa. The rapid evolution of insecticide resistance in this highly dynamic and genetically polymorphic species remains a challenge for its control.

Presence in Cameroon of the recently introduced Aedes (Stegomyia) albopictus (Skuse) in association with the indigenous Aedes aegypti (L.) raises public heath concerns because it might alter the risk of arbovirus transmission. The breeding site and distribution of the two Stegomyia species are updated and reported following entomological surveys carried out in 22 localities throughout Cameroon, with a total of 1,353 containers with water visited. Ae. aegypti was found in every location sampled, showing higher infestation rates in northern Cameroon. Breeding populations of Ae. albopictus were observed in all 19 southern localities, up to the Adamaoua mountains, but the species was not recorded further north. In the area where both species are present, they were often sampled in the same larval developmental sites, suggesting convergent habitat segregation. The most frequently encountered artificial and natural breeding sites were used tires, discarded tins and plastic containers, abandoned car parts, brick holes, dead leaves on the ground, tree holes, and rock pools. Further monitoring of the demographic as well as geographic expansion of Ae. albopictus in this Afrotropical environment and its relationships with indigenous Ae. aegypti should provide insight into the biology of this highly invasive species and help to implement arboviruses surveillance programs in the area.

A 2-yr longitudinal malaria study was undertaken in a suburb of Yaounde, the capital city of Cameroon, in the village of Simbock, approximately 2 km from the city limits. This study allowed assessment of malaria transmission intensity and dynamics in this region before implementation of pyrethroid impregnated bed nets through the national vector control program. Anophelines were captured on human volunteers by pyrethrum spray collections and in resting sites outdoors. Malaria vectors were Anopheles funestus Giles, Anopheles gambiae s.s. Giles (M and S forms), Anopheles moucheti Evans, and Anopheles nili Theobald. An. moucheti was the most abundant mosquito captured during the study, accounting for >54% of total anophelines caught. The annual Plasmodium falciparum Welch entomological inoculation rates measured by enzyme-linked immunosorbent assay were 277 infected bites per human for the first year and 368 for the second year. An. gambiae s.s., An. funestus, An. moucheti, and An. nili were responsible for 23.8%, 26.8%, 39.2%, and 10.2% of malaria transmission, respectively. Malaria transmission is perennial throughout the year. All these vectors were highly anthropophagous because only two out of 566 mosquitoes blood-meal tested were not taken on humans.