The current distribution and characterization of the L1014F resistance allele of the kdr gene in three malaria vectors (Anopheles gambiae, Anopheles coluzzii, Anopheles arabiensis) in Benin (West Africa).

BACKGROUND: The fight against malaria faces various biological obstacles, including the resistance of parasites to anti-malarial drugs and the resistance of mosquito vectors to insecticides. The resistance of Anopheles gambiae sensu lato (s.l.) to pyrethroids, the only class of insecticides used to impregnate mosquito nets, is known in Benin; the expansion of this resistance is influenced by the existence of gene flow between species, otherwise by the presence or absence of the kdr mutation in them. The objective of this study is to determine the spatial distribution of An. gambiae and the level of expression of the pyrethroid resistance kdr gene in seven agro-ecological zones of Benin. METHODS: The study was conducted in 18 localities belonging to seven agro-ecological zones where environmental parameters varied. The sites represent the main areas of eco-epidemiological malaria in Benin. Anopheles gambiae larvae were collected in natural breeding sites using ladles and dipping method and reared under standard conditions. These larvae were reared under standard conditions of temperature and humidity (26 to 30 °C and 60 to 90%) at the insectarium of the Centre de Recherche Entomologique de Cotonou (CREC). Adult female mosquitoes having emerged are morphologically and molecularly identified. Homozygous resistant (1014F/1014F), homozygous sensitive (1014L/1014L) and heterozygous (1014F/1014L) genotypes of the L1014F kdr gene mutation are determined by PCR. RESULTS: A total of 677 An. gambiae was subjected at the PCR. The results revealed the presence of three vector species of the An. gambiae complex, of which 409 Anopheles coluzzii, 259 An. gambiae, 5 hybrids (An. coluzzii/An. gambiae) and 4 Anopheles arabiensis in the different agro-ecological zones. The four An. arabiensis were only found in Dassa, a locality in the cotton zone of central Benin. The frequency of distribution of the L1014F allele of the kdr gene varies from 84.48 to 100% in An. gambiae, from 80 to 100% in An. coluzzii and from 0 to 75% in An. arabiensis in the different agro-ecological zones. Moreover, a significant difference is generally observed in the distribution of the L1014F allele (P < 0.05). By comparing in pairs the distribution frequencies of this allele in the two species by agro-ecological zone, only a significant difference is noted in the central cotton and fishery zones (P = 0.0496). CONCLUSION: In summary, even if the data are in small portions, the An. Arabiensis species was found only in central Benin and the L1014F allele of the kdr gene is widespread and seems to fix in all the species recorded in the different agro-ecological zones. This situation amplifies the problem of resistance, which could eventually be a significant obstacle for the malaria vectors control. Similarly, a study of their genetic structure via the L1014F allele is necessary in order to put in place strategies to manage this resistance. These strategies will take into account both the ecology and the genetic diversity of the organisms involved to preserve the effectiveness of pyrethroids, the only insecticides used for the impregnation of mosquito nets.

Echocardiographic left atrial remodelling and determinants of left atrial size in the early phase of high blood pressure: a comparative cross-sectional study in Douala, Cameroon.

BACKGROUND: Left atrial remodelling (LAR) has been described in Western populations with chronic hypertension and is associated with a higher risk of adverse cardiovascular events. Although hypertension tends to occur earlier and is more severe in sub-Saharan Africa than in more developed nations, LAR and its associated factors in these African hypertensive subjects have been poorly elucidated. OBJECTIVES: To assess left atrial structural remodelling in black hypertensive patients and determine factors associated with left atrial size. METHODS: This was a cross-sectional, comparative study carried out in two tertiary hospitals in Douala, Cameroon over a period of three months. Fifty-two patients, either newly diagnosed with hypertension or known hypertensives treated for less than a year, were consecutively recruited. These patients were matched (unpaired matching) for age and gender to 40 randomly selected healthy subjects. The posterior-anterior diameter indexed to body surface area (BSA), volume indexed to BSA, and longitudinal and transverse diameters of the left atrium (LA) were measured using transthoracic echocardiography, in accordance with the American Society of Echocardiography guidelines. LAR was defined as increase in LA size, characterised by LA volume ≥ 34 ml/m2. Early morning urine was analysed for microalbuminuria using urine strips to obtain spot albumin/creatinine ratio. Data were analysed using SPSS version 23 and statistical significance was set at p < 0.05. RESULTS: The gender distribution and mean age were similar between the two groups. Hypertensive patients had significantly higher mean body mass index, left ventricular mass and an altered diastolic function. They also had significantly higher LA longitudinal diameter (50.0 vs 47.4 mm; p = 0.045), surface area (17.9 vs 15.5 cm2; p = 0.003) and volume (52.4 vs 43.8 ml; p = 0.002) compared to the non-hypertensive counterparts. Fourteen patients (26.9%) had LA enlargement compared to one (2.5%) in the non-hypertensive group (odds ratio = 9.78, CI: 2.67-35.8, p < 0.0001). Diastolic dysfunction (p = 0.008) was the only independent predictor of LA size in the hypertensive subjects. Microalbuminuria did not significantly correlate with LA size. CONCLUSIONS: Our study shows evidence of LAR in newly diagnosed black African patients with hypertension, characterised by an increase in the LA length, surface area and volume. Future studies are warranted to better elucidate the biological mechanisms underlying the link between the early phase of hypertension and LAR, as well as its prognostic implications in our population.

Dynamics of pyrethroid resistance in malaria vectors in southern Benin following a large scale implementation of vector control interventions.

BACKGROUND: Large-scale implementation of Indoor Residual Spraying and Insecticide Treated Nets has been implemented in Plateau Department, Benin between 2011 and 2014. The purpose of this study was to monitor the frequency and mechanisms of pyrethroid resistance in malaria vectors following the implementation of vector control tools for malaria prevention. METHODS: Anopheles larvae were collected in 13 villages twice a year from 2012 to 2014. WHO tube tests were used to assess the phenotypic resistance of each population to 0.05 % deltamethrin. Sibling species within Anopheles gambiae complex were identified by PCR techniques. Taqman and biochemical assays were performed to identify the presence of kdr mutations in individual mosquitoes and to detect any increase in the activity of enzymes putatively involved in insecticide metabolism (oxidases, esterase and glutathione-S-transferases). Quantitative real time PCR was used to measure the expression of three metabolic genes involved in pyrethroid resistance (CYP6P3, CYP6M2 and GSTD3). RESULTS: Anopheles populations showed < 90 % mortality to deltamethrin in all villages and at all time points. The 1014 F kdr allele frequency was close to fixation (> 0.9) over the sampling periods in both An. gambiae and An. coluzzii. Biochemical assays showed higher activities of alpha esterase and GST in field malaria vector populations compared to susceptible mosquitoes. qPCR assays showed a significant increase of CYP6P3, CYP6M2 GSTD3 expression in An. gambiae after a three-year implementation of LLINs. CONCLUSION: The study confirmed that deltamethrin resistance is widespread in malaria vectors in Southern Benin. We suspect that the increase in deltamethrin resistance between 2012 and 2014 resulted from an increased expression of metabolic detoxification genes (CYP6M2 and CYP6P3) rather than from kdr mutations. It is urgent to evaluate further the impact of metabolic resistance on the efficacy of vector control interventions using pyrethroid insecticides.