Didier Fontenille

Two Chikungunya isolates from the outbreak of La Reunion (Indian Ocean) exhibit different patterns of infection in the mosquito, Aedes albopictus.

Background A Chikungunya (CHIK) outbreak hit La Réunion Island in 2005–2006. The implicated vector was Aedes albopictus. Here, we present the first study on the susceptibility of Ae. albopictus populations to sympatric CHIKV isolates from La Réunion Island and compare it to other virus/vector combinations. Methodology and Findings We orally infected 8 Ae. albopictus collections from La Réunion and 3 from Mayotte collected in March 2006 with two Chikungunya virus (CHIKV) from La Réunion: (i) strain 05.115 collected in June 2005 with an Alanine at the position 226 of the glycoprotein E1 and (ii) strain 06.21 collected in November 2005 with a substitution A226V. Two other CHIKV isolates and four additional mosquito strains/species were also tested. The viral titer of the infectious blood-meal was 107 plaque forming units (pfu)/mL. Dissemination rates were assessed by immunofluorescent staining on head squashes of surviving females 14 days after infection. Rates were at least two times higher with CHIKV 06.21 compared to CHIKV 05.115. In addition, 10 individuals were analyzed every day by quantitative RT-PCR. Viral RNA was quantified on (i) whole females and (ii) midguts and salivary glands of infected females. When comparing profiles, CHIKV 06.21 produced nearly 2 log more viral RNA copies than CHIKV 05.115. Furthermore, females infected with CHIKV 05.115 could be divided in two categories: weakly susceptible or strongly susceptible, comparable to those infected by CHIKV 06.21. Histological analysis detected the presence of CHIKV in salivary glands two days after infection. In addition, Ae. albopictus from La Réunion was as efficient vector as Ae. aegypti and Ae. albopictus from Vietnam when infected with the CHIKV 06.21. Conclusions Our findings support the hypothesis that the CHIK outbreak in La Réunion Island was due to a highly competent vector Ae. albopictus which allowed an efficient replication and dissemination of CHIKV 06.21.

Aedes albopictus, an arbovirus vector: from the darkness to the light.

The Asian tiger mosquito, Aedes albopictus (Skuse, 1894), is an invasive species that can be found on all continents. The species, originally considered a secondary vector of viruses such as Dengue viruses, has recently been suggested to play a role in the transmission of Chikungunya virus in several countries bordering the Indian Ocean, Central Africa and Europe. Here we review the current geographic range and the relevant biological traits of A. albopictus in order to explain its rapid spread. We examine and discuss recent changes in its role as a vector, particularly in the transmission of arboviruses, and its importance in the current and future emergence of pathogens. Finally, we report conventional and innovative ways to control A. albopictus.

Zika Virus in Gabon (Central Africa) – 2007: A New Threat from Aedes albopictus?

Background Chikungunya and dengue viruses emerged in Gabon in 2007, with large outbreaks primarily affecting the capital Libreville and several northern towns. Both viruses subsequently spread to the south-east of the country, with new outbreaks occurring in 2010. The mosquito species Aedes albopictus, that was known as a secondary vector for both viruses, recently invaded the country and was the primary vector involved in the Gabonese outbreaks. We conducted a retrospective study of human sera and mosquitoes collected in Gabon from 2007 to 2010, in order to identify other circulating arboviruses. Methodology/Principal Findings Sample collections, including 4312 sera from patients presenting with painful febrile disease, and 4665 mosquitoes belonging to 9 species, split into 247 pools (including 137 pools of Aedes albopictus), were screened with molecular biology methods. Five human sera and two Aedes albopictus pools, all sampled in an urban setting during the 2007 outbreak, were positive for the flavivirus Zika (ZIKV). The ratio of Aedes albopictus pools positive for ZIKV was similar to that positive for dengue virus during the concomitant dengue outbreak suggesting similar mosquito infection rates and, presumably, underlying a human ZIKV outbreak. ZIKV sequences from the envelope and NS3 genes were amplified from a human serum sample. Phylogenetic analysis placed the Gabonese ZIKV at a basal position in the African lineage, pointing to ancestral genetic diversification and spread. Conclusions/Significance We provide the first direct evidence of human ZIKV infections in Gabon, and its first occurrence in the Asian tiger mosquito, Aedes albopictus. These data reveal an unusual natural life cycle for this virus, occurring in an urban environment, and potentially representing a new emerging threat due to this novel association with a highly invasive vector whose geographic range is still expanding across the globe.

Influence of Temperature on Immature Development, Survival, Longevity, Fecundity, and Gonotrophic Cycles of Aedes albopictus, Vector of Chikungunya and Dengue in the Indian Ocean

ABSTRACT Aedes albopictus is a mosquito originating from Asia, which has extended its range worldwide the last decades. It is a competent vector for several arboviruses. It was first described in La Réunion (an island of the South West part of the Indian Ocean) in 1913. Since then, it has become the dominant Aedes species and a serious threat to public health, especially during the two last arboviruses outbreaks of dengue (1977) and chikungunya (2005–2006). Despite its pest status, data on the biology of this vector are scarce, especially the population present in the Indian Ocean (IO), which has never been studied in detail. Therefore, the immature development, survival, longevity, fecundity, and gonotrophic cycles of Ae. albopictus were studied for an F2 population of the IO. These biological parameters were studied in controlled conditions at eight constant temperatures (5, 10,15, 20, 25, 30, 35, and 40°C). The minimal threshold of immature stages development was found at 10.4°C and its optimum at 29.7°C. The shortest periods for immature development were found at 30°C, with in average of 8.8 d. The optimum intrinsic rate of growth (r) was observed between 25 and 30°C. The gonotrophic cycles were also evaluated, and the shortest cycles were found at 30°C (mean, 3.5 d). Those results are according to the field repartition of this species in La Réunion, allowing Ae. albopictus survival at a large range of temperatures.

Semipermeable species boundaries between Anopheles gambiae and Anopheles arabiensis: Evidence from multilocus DNA sequence variation

Attempts to reconstruct the phylogenetic history of the Anopheles gambiae cryptic species complex have yielded strongly conflicting results. In particular, An. gambiae, the primary African malaria vector, is variously placed as a sister taxon to either Anopheles arabiensis or Anopheles merus. The recent divergence times for members of this complex complicate phylogenetic analysis, making it difficult to unambiguously implicate interspecific gene flow, versus retained ancestral polymorphism, as the source of conflict. Using sequences at four unlinked loci, which were determined from multiple specimens within each of five species in the complex, we found contrasting patterns of sequence divergence between the X chromosome and the autosomes. The isolation model of speciation assumes a lack of gene flow between species since their separation. This model could not be rejected for An. gambiae and An. arabiensis, although the data fit the model poorly. On the other hand, evidence from gene trees supports genetic introgression of chromosome 2 inversions between An. gambiae and An. arabiensis, and also points to more broad scale genetic exchange of autosomal sequences between this species pair. That such exchange has been relatively recent is suggested not only by the lack of fixed differences at three autosomal loci but also by the sharing of full haplotypes at two of the three loci, which is in contrast to several fixed differences and considerably deeper divergence on the X. The proposed acquisition by An. gambiae of sequences from the more arid-adapted An. arabiensis may have contributed to the spread and ecological dominance of this malaria vector.

Genetic differentiation of Anopheles gambiae populations from East and west Africa: comparison of microsatellite and allozyme loci.

Genetic variation of Anopheles gambiae was analysed to assess interpopulation divergence over a 6000 km distance using short tandem repeat (microsatellite) loci and allozyme loci. Differentiation of populations from Kenya and Senegal measured by allele length variation at five microsatellite loci was compared with estimates calculated from published data on six allozyme loci (Miles, 1978). The average Wrights FST of microsatellite loci (0.016) was lower than that of allozymes (0.036). Slatkins RST values for microsatellite loci were generally higher than their FST values, but the average RST value was virtually identical (0.036) to the average allozyme FST. These low estimates of differentiation correspond to an effective migration index (Nm) larger than 3, suggesting that gene flow across the continent is only weakly restricted. Polymorphism of microsatellite loci was significantly higher than that of allozymes, probably because the former experience considerably higher mutation rates. That microsatellite loci did not measure greater interpopulation divergence than allozyme loci suggested constraints on microsatellite evolution. Alternatively, extensive mosquito dispersal, aided by human transportation during the last century, better explains the low differentiation and the similarity of estimates derived from both types of genetic markers.

Ecological niche partitioning between Anopheles gambiae molecular forms in Cameroon: the ecological side of speciation

BackgroundSpeciation among members of the Anopheles gambiae complex is thought to be promoted by disruptive selection and ecological divergence acting on sets of adaptation genes protected from recombination by polymorphic paracentric chromosomal inversions. However, shared chromosomal polymorphisms between the M and S molecular forms of An. gambiae and insufficient information about their relationship with ecological divergence challenge this view. We used Geographic Information Systems, Ecological Niche Factor Analysis, and Bayesian multilocus genetic clustering to explore the nature and extent of ecological and chromosomal differentiation of M and S across all the biogeographic domains of Cameroon in Central Africa, in order to understand the role of chromosomal arrangements in ecological specialisation within and among molecular forms.ResultsSpecies distribution modelling with presence-only data revealed differences in the ecological niche of both molecular forms and the sibling species, An. arabiensis. The fundamental environmental envelope of the two molecular forms, however, overlapped to a large extent in the rainforest, where they occurred in sympatry. The S form had the greatest niche breadth of all three taxa, whereas An. arabiensis and the M form had the smallest niche overlap. Correspondence analysis of M and S karyotypes confirmed that molecular forms shared similar combinations of chromosomal inversion arrangements in response to the eco-climatic gradient defining the main biogeographic domains occurring across Cameroon. Savanna karyotypes of M and S, however, segregated along the smaller-scale environmental gradient defined by the second ordination axis. Population structure analysis identified three chromosomal clusters, each containing a mixture of M and S specimens. In both M and S, alternative karyotypes were segregating in contrasted environments, in agreement with a strong ecological adaptive value of chromosomal inversions.ConclusionOur data suggest that inversions on the second chromosome of An. gambiae are not causal to the evolution of reproductive isolation between the M and S forms. Rather, they are involved in ecological specialization to a similar extent in both genetic backgrounds, and most probably predated lineage splitting between molecular forms. However, because chromosome-2 inversions promote ecological divergence, resulting in spatial and/or temporal isolation between ecotypes, they might favour mutations in other ecologically significant genes to accumulate in unlinked chromosomal regions. When such mutations occur in portions of the genome where recombination is suppressed, such as the pericentromeric regions known as speciation islands in An. gambiae, they would contribute further to the development of reproductive isolation.

Evidence for genetic differentiation between the molecular forms M and S within the Forest chromosomal form of Anopheles gambiae in an area of sympatry.

We studied genetic variation at ten microsatellite DNA loci in Anopheles gambiae populations from the Forest chromosomal form collected in four villages in Cameroon (Central Africa). Both recently described M and S molecular forms occur in sympatry in this area. Geographic differentiation within form was low (Fst < 0.017) despite geographical distance between collection sites ranging from 35 to 350 km. However, higher (Fst > 0.035) and statistically significant levels of genetic differentiation were observed between forms, being the highest between sympatric M and S populations collected within the same village. Results were consistent across all loci spread throughout the genome, therefore reflecting a genome‐wide pattern. Considering previous findings of strong assortative mating within forms and general lack of hybrids in areas of sympatry, we propose that there is now sufficient direct and indirect evidence to consider both M and S molecular forms of An. gambiae as distinct species that have probably speciated recently.

Blood-feeding behavior of Aedes albopictus, a vector of Chikungunya on La Réunion

Chikungunya virus (CHIKV) has long been considered to be transmitted to humans by the human-biting mosquito Aedes aegypti, especially in Africa. However, the recent outbreak of CHIKV involved another vector, Aedes albopictus, and serological data in the literature suggest that several species of domestic or human-related vertebrates can be contaminated by this virus. However, the role of Ae. albopictus mosquitoes as potential enzootic vectors for CHIKV has not yet been evaluated. Here we investigate Ae. albopictus feeding and resting behaviors in an area where a CHIKV epidemic recently occurred, which means deciphering host-seeking and feeding behaviors on several vertebrate species, measuring endophagous/exophagous (activity), endophilic/exophilic (resting) behaviors and its diel (24 h, day/night) biting activity. Ae. albopictus was found to have bimodal daily feeding activities and was found to have exophagic (89%) and exophilic (87%) behaviors. Ae. albopictus showed an opportunistic feeding behavior on a wide range of hosts (from cold-blooded to warm-blooded animals), supporting that it can be implicated in various vertebrate-virus pathosystems. However, with equal availability of one of the four vertebrate hosts (calf, chicken, dog, and goat) proposed against human, Ae. albopictus significantly preferred human, supporting earlier data about its high degree of anthropophily. Multiple blood feeding was also reported in every combination (animal/human) offered to Ae. albopictus, enlightening the higher risks to spread an arbovirus to human population because of interrupted feeding. Such catholic behavior suggests that Ae. albopictus may act as a bridge vector for zoonotic viruses. Further epidemiological implications of this issue are discussed.

Increased susceptibility to malaria during the early postpartum period.

BACKGROUND Pregnancy is associated with increased susceptibility to malaria. It is generally agreed that this increased risk ends with delivery, but the possible persistence of increased susceptibility during the puerperium had not been investigated. METHODS From June 1, 1990, to December 31, 1998, we monitored exposure to malaria, parasitemia, and morbidity among the residents of a village in Senegal in which the rate of transmission of malaria was high. In this population we analyzed 71 pregnancies in 38 women from the year before conception and through one year after delivery. RESULTS Among the 38 women, there were 58 episodes of clinical Plasmodium falciparum malaria during 61,081 person-days of observation. The incidence of malaria was 20.2 episodes per 1000 person-months during the year preceding conception and 12.0 episodes per 1000 person-months during the period from 91 to 365 days after delivery. The incidence of episodes of malaria increased significantly during the second and third trimesters of pregnancy and reached a maximum of 75.1 episodes per 1000 person-months during the first 60 days after delivery. The adjusted relative risk of an episode of malaria was 4.1 (95 percent confidence interval, 1.8 to 9.5) during the first 60 days post partum, as compared with the year preceding pregnancy. The duration of fever during the episodes of malaria was longer and the prevalence and density of asymptomatic malarial parasitemia were significantly higher during pregnancy and the early postpartum period than during the other periods. CONCLUSIONS Among women who live in areas with high rates of transmission of malaria, the susceptibility to malaria is highest during the second and third trimesters of pregnancy and the early postpartum period.