Breakdown in the Process of Incipient Speciation in Anopheles gambiae

Understanding genetic causes and effects of speciation in sympatric populations of sexually reproducing eukaryotes is challenging, controversial, and of practical importance for controlling rapidly evolving pests and pathogens. The major African malaria vector mosquito Anopheles gambiae sensu stricto (s.s.) is considered to contain two incipient species with strong reproductive isolation, hybrids between the M and S molecular forms being very rare. Following recent observations of higher proportions of hybrid forms at a few sites in West Africa, we conducted new surveys of 12 sites in four contiguous countries (The Gambia, Senegal, Guinea-Bissau, and Republic of Guinea). Identification and genotyping of 3499 A. gambiae s.s. revealed high frequencies of M/S hybrid forms at each site, ranging from 5 to 42%, and a large spectrum of inbreeding coefficient values from 0.11 to 0.76, spanning most of the range expected between the alternative extremes of panmixia and assortative mating. Year-round sampling over 2 years at one of the sites in The Gambia showed that M/S hybrid forms had similar relative frequencies throughout periods of marked seasonal variation in mosquito breeding and abundance. Genome-wide scans with an Affymetrix high-density single-nucleotide polymorphism (SNP) microarray enabled replicate comparisons of pools of different molecular forms, in three separate populations. These showed strong differentiation between M and S forms only in the pericentromeric region of the X chromosome that contains the molecular form-specific marker locus, with only a few other loci showing minor differences. In the X chromosome, the M/S hybrid forms were more differentiated from M than from S forms, supporting a hypothesis of asymmetric introgression and backcrossing.     

Filarial infection suppresses malaria-specific multifunctional Th1 and Th17 responses in malaria and filarial coinfections

The mechanisms underlying the modulation of both the malaria-specific immune response and the course of clinical malaria in the context of concomitant helminth infection are poorly understood. We used multiparameter flow cytometry to characterize the quality and the magnitude of malaria-specific T cell responses in filaria-infected and -uninfected individuals with concomitant asymptomatic Plasmodium falciparum malaria in Mali. In comparison with filarial-uninfected subjects, filarial infection was associated with higher ex vivo frequencies of CD4(+) cells producing IL-4, IL-10, and IL-17A (p = 0.01, p = 0.001, and p = 0.03, respectively). In response to malaria Ag stimulation, however, filarial infection was associated with lower frequencies of CD4(+) T cells producing IFN-γ, TNF-α, and IL-17A (p < 0.001, p = 0.04, and p = 0.04, respectively) and with higher frequencies of CD4(+)IL10(+)T cells (p = 0.0005). Importantly, filarial infection was associated with markedly lower frequencies of malaria Ag-specific Th1 (p < 0.0001), Th17 (p = 0.012), and "TNF-α" (p = 0.0008) cells, and a complete absence of malaria-specific multifunctional Th1 cells. Filarial infection was also associated with a marked increase in the frequency of malaria-specific adaptive regulatory T/Tr1 cells (p = 0.024), and the addition of neutralizing anti-IL-10 Ab augmented the amount of Th1-associated cytokine produced per cell. Thus, among malaria-infected individuals, concomitant filarial infection diminishes dramatically the frequencies of malaria-specific Th1 and Th17 T cells, and alters the quality and magnitude of malaria-specific T cell responses.     

Expanded numbers of circulating myeloid dendritic cells in patent human filarial infection reflect lower CCR1 expression

APC dysfunction has been postulated to mediate some of the parasite-specific T cell unresponsiveness seen in patent filarial infection. We have shown that live microfilariae of Brugia malayi induce caspase-dependent apoptosis in human monocyte-derived dendritic cells (DCs) in vitro. This study addresses whether apoptosis observed in vitro extends to patent filarial infections in humans and is reflected in the number of circulating myeloid DCs (mDCs; CD11c(-)CD123(lo)) in peripheral blood of infected microfilaremic individuals. Utilizing flow cytometry to identify DC subpopulations (mDCs and plasmacytoid DCs [pDCs]) based on expression of CD11c and CD123, we found a significant increase in numbers of circulating mDCs (CD11c(+)CD123(lo)) in filaria-infected individuals compared with uninfected controls from the same filaria-endemic region of Mali. Total numbers of pDCs, monocytes, and lymphocytes did not differ between the two groups. To investigate potential causes of differences in mDC numbers between the two groups, we assessed chemokine receptor expression on mDCs. Our data indicate that filaria-infected individuals had a lower percentage of circulating CCR1(+) mDCs and a higher percentage of circulating CCR5(+) mDCs and pDCs. Finally, live microfilariae of B. malayi were able to downregulate cell-surface expression of CCR1 on monocyte-derived DCs and diminish their calcium flux in response to stimulation by a CCR1 ligand. These findings suggest that microfilaria are capable of altering mDC migration through downregulation of expression of some chemokine receptors and their signaling functions. These observations have major implications for regulation of immune responses to these long-lived parasites.     

Resistance to DDT and pyrethroids and increased kdr mutation frequency in An. gambiae after the implementation of permethrin-treated nets in Senegal

IntroductionThe aim of this study was to evaluate the susceptibility to insecticides of An. gambiae mosquitoes sampled in Dielmo (Senegal), in 2010, 2 years after the implementation of Long Lasting Insecticide-treated Nets (LLINs) and to report the evolution of kdr mutation frequency from 2006 to 2010.MethodsWHO bioassay susceptibility tests to 6 insecticides were performed on adults F0, issuing from immature stages of An. gambiae s.l., sampled in August 2010. Species and molecular forms as well as the presence of L1014F and L1014S kdr mutations were assessed by PCR. Longitudinal study of kdr mutations was performed on adult mosquitoes sampled monthly by night landing catches from 2006 to 2010.FindingsNo specimen studied presented the L1014S mutation. During the longitudinal study, L1014F allelic frequency rose from 2.4% in year before the implementation of LLINs to 4.6% 0–12 months after and 18.7% 13–30 months after. In 2010, An. gambiae were resistant to DDT, Lambda-cyhalothrin, Deltamethrin and Permethrin (mortality rates ranging from 46 to 63%) but highly susceptible to Fenitrothion and Bendiocarb (100% mortality). There was significantly more RR genotype among An. gambiae surviving exposure to DDT or Pyrethroids. An. arabiensis represented 3.7% of the sampled mosquitoes (11/300) with no kdr resistance allele detected. An. gambiae molecular form M represented 29.7% of the mosquitoes with, among them, kdr genotypes SR (18%) and SS (82%). An. gambiae molecular form S represented 66% of the population with, among them, kdr genotype SS (33.3%), SR (55.6%) and RR (11.1%). Only 2 MS hybrid mosquitoes were sampled and presented SS kdr genotype.ConclusionBiological evidence of resistance to DDT and pyrethroids was detected among An. gambiae mosquitoes in Dielmo (Senegal) within 24 months of community use of LLINs. Molecular identification of L1014F mutation indicated that target site resistance increased after the implementation of LLINs.     

Malaria Morbidity in High and Seasonal Malaria Transmission Area of Burkina Faso

Background

Malariometric parameters are often primary endpoints of efficacy trials of malaria vaccine candidates. This study aims to describe the epidemiology of malaria prior to the conduct of a series of drug and vaccine trials in a rural area of Burkina Faso.

Methods

Malaria incidence was prospectively evaluated over one year follow-up among two cohorts of children aged 0–5 years living in the Saponé health district. The parents of 1089 children comprising a passive case detection cohort were encouraged to seek care from the local health clinic at any time their child felt sick. Among this cohort, 555 children were randomly selected for inclusion in an active surveillance sub-cohort evaluated for clinical malaria during twice weekly home visits. Malaria prevalence was evaluated by cross-sectional survey during the low and high transmission seasons.

Results

Number of episodes per child ranged from 0 to 6 per year. Cumulative incidence was 67.4% in the passive and 86.2% in the active cohort and was highest among children 0–1 years. Clinical malaria prevalence was 9.8% in the low and 13.0% in the high season (p>0.05). Median days to first malaria episode ranged from 187 (95% CI 180–193) among children 0–1 years to 228 (95% CI 212, 242) among children 4–5 years. The alternative parasite thresholds for the malaria case definition that achieved optimal sensitivity and specificity (70–80%) were 3150 parasites/µl in the high and 1350 parasites/µl in the low season.

Conclusion

Clinical malaria burden was highest among the youngest age group children, who may represent the most appropriate target population for malaria vaccine candidate development. The pyrogenic threshold of parasitaemia varied markedly by season, suggesting a value for alternative parasitaemia levels in the malaria case defintion. Regional epidemiology of malaria described, Sapone area field centers are positioned for future conduct of malaria vaccine trials.     

Relationship between Malaria Incidence and IgG Levels to Plasmodium falciparum Merozoite Antigens in Malian Children: Impact of Hemoglobins S and C

Heterozygous hemoglobin (Hb) AS (sickle-cell trait) and HbAC are hypothesized to protect against Plasmodium falciparum malaria in part by enhancing naturally-acquired immunity to this disease. To investigate this hypothesis, we compared antibody levels to four merozoite antigens from the P. falciparum 3D7 clone (apical membrane antigen 1, AMA1-3D7; merozoite surface protein 1, MSP1-3D7; 175 kDa erythrocyte-binding antigen, EBA175-3D7; and merozoite surface protein 2, MSP2-3D7) in a cohort of 103 HbAA, 73 HbAS and 30 HbAC children aged 3 to 11 years in a malaria-endemic area of Mali. In the 2009 transmission season we found that HbAS, but not HbAC, significantly reduced the risk of malaria compared to HbAA. IgG levels to MSP1 and MSP2 at the start of this transmission season inversely correlated with malaria incidence after adjusting for age and Hb type. However, HbAS children had significantly lower IgG levels to EBA175 and MSP2 compared to HbAA children. On the other hand, HbAC children had similar IgG levels to all four antigens. The parasite growth-inhibitory activity of purified IgG samples did not differ significantly by Hb type. Changes in antigen-specific IgG levels during the 2009 transmission and 2010 dry seasons also did not differ by Hb type, and none of these IgG levels dropped significantly during the dry season. These data suggest that sickle-cell trait does not reduce the risk of malaria by enhancing the acquisition of IgG responses to merozoite antigens.     

Effects of management intensity on ant diversity in cocoa plantation (Oume,centre west Côte d’Ivoire)

Anthropogenic habitat modifications, including conversion of forest to agricultural production cause losses of native species. In this study we examined the losses suffered by ant communities in relation to the intensity of management in cocoa plantations established in former tropical forest. An extensive sampling protocol consisting of pitfall trapping, leaf litter sampling, soil sampling and hand sampling was used to characterize ant species richness and composition in the native forest and compare it with three cocoa farms differing in their management intensity. Species richness was negatively correlated with management intensity and differed greatly between management practices and the forest. Two subfamilies (Myrmicinae and Amblyoponinae) showed a significant negative correlation with agricultural intensification probably because their species are constrained to living in forest like habitat. The species composition differed greatly between management practices and the forest. Intensively and moderately managed cocoa plantations were most dissimilar to the forest. In contrast, forest ants were well represented in the least intensively managed plantation. Overall, the findings of this study show that only slightly managed cocoa plantations sustain an ant diversity that comes close to that of the forest. This also holds true for cocoa plantations on former agricultural, and thus previously heavily used, land. The findings may help in the conservation of biodiversity as management practices most likely to sustain forest like ant communities have been identified.     

An Efficient Microinoculation Procedure to Study Plant Virus Multiplication at Predetermined Individual Infection Sites on the Leaves

The microinoculation procedure (inoculated are 0.02—0.05 mm² was carried out using the very thin tip of a stretched Pasteur pipette. It was dipped in a 500 μg/ml suspension of purified TMV, brought into contact with the upper epidermis of the leaf which had been dusted With an abrasive and rotated twice without actually pricking the leaf tissue. Five minutes later, the excess abrasive and virus were removed by rinsing the leaves under water. The procedure was applied to several tobacco-TMV combinations. It induced the formation of single local lesions with over 90% efficiency on hypersensitively reacting hosts. Comparable efficiencies were obtained with systemically reacting hosts, as evidenced by a radiochemical procedure coupled with indirect ELISA (KONATE and FRITIG 1983). It was found that residual virus originating from the inoculum was negligible and could casily be distinguished form newly synthesized virus, even shortly after the microinoculation. This makes it possible to measure the rates of virus cell-to-cell spread and of virus multiplication at various times after inoculation and at various distances form the points of virus entry. These approadies can be extended to the comparison between differently reacting hosts and to the study of interference between different viruses or different virus strains.