High-resolution species assignment of Anopheles mosquitoes using k-mer distances on targeted sequences.

The ANOSPP amplicon panel is a genus-wide targeted sequencing panel to facilitate large-scale monitoring of Anopheles species diversity. Combining information from the 62 nuclear amplicons present in the ANOSPP panel allows for a more senstive and specific species assignment than single gene (e.g. COI) barcoding, which is desirable in the light of permeable species boundaries. Here, we present NNoVAE, a method using Nearest Neighbours (NN) and Variational Autoencoders (VAE), which we apply to k-mers resulting from the ANOSPP amplicon sequences in order to hierarchically assign species identity. The NN step assigns a sample to a species-group by comparing the k-mers arising from each haplotype's amplicon sequence to a reference database. The VAE step is required to distinguish between closely related species, and also has sufficient resolution to reveal population structure within species. In tests on independent samples with over 80% amplicon coverage, NNoVAE correctly classifies to species level 98% of samples within the An. gambiae complex and 89% of samples outside the complex. We apply NNoVAE to over two thousand new samples from Burkina Faso and Gabon, identifying unexpected species in Gabon. NNoVAE presents an approach that may be of value to other targeted sequencing panels, and is a method that will be used to survey Anopheles species diversity and Plasmodium transmission patterns through space and time on a large scale, with plans to analyse half a million mosquitoes in the next five years.

Genetic structure of the mosquito Aedes aegypti in local forest and domestic habitats in Gabon and Kenya.

BACKGROUND: The mosquito Aedes aegypti is a devastating disease vector transmitting several important human arboviral diseases. In its native range in Africa, the mosquito can be found in both the ancestral forest habitat and anthropogenic habitats such as villages. How do the different habitats impact the population genetic structure of the local mosquito populations? METHODS: To address this question, we simultaneously sampled Ae. aegypti from the forest and local villages in La Lopé, Gabon and Rabai, Kenya. The mosquitoes were genotyped at 12 microsatellite loci and a panel of ~25,000 single nucleotide polymorphisms (SNPs), which allowed us to estimate their genetic ancestries and the population genetic structure related to habitats and sampling sites. RESULTS: In the context of the global population genetic structure of Ae. aegypti, clustering analysis showed that mosquitoes from the same locality (La Lopé or Rabai) have similar genetic ancestry, regardless of their habitats. Further analysis at the local scale also found no strong genetic differentiation between the forest and village mosquitoes in both La Lopé and Rabai. Interestingly, these results from our 2017 samples from Rabai, Kenya contrast to the documentation of genetic differentiation between village and forest mosquito collections from 1975-1976 and 2009. Between-habitat measures of genetic difference (Fst) vary across the genome, with a peak of high divergence observed at the third chromosome only in the La Lopé populations. CONCLUSION: Collectively, these results demonstrated that there is little genetic isolation between forest and village habitats, which suggests possible extensive gene flow between them. From an epidemiological perspective, the forest habitat could act as a refuge for mosquitoes against vector control programmes in the domestic settings. Moreover, sylvatic populations could play a role in zoonotic pathogen transferred to humans. Therefore, future studies on disease transmission and vector control planning in the study area should take natural populations into consideration.

A new species in the major malaria vector complex sheds light on reticulated species evolution.

Complexes of closely related species provide key insights into the rapid and independent evolution of adaptive traits. Here, we described and studied Anopheles fontenillei sp.n., a new species in the Anopheles gambiae complex that we recently discovered in the forested areas of Gabon, Central Africa. Our analysis placed the new taxon in the phylogenetic tree of the An. gambiae complex, revealing important introgression events with other members of the complex. Particularly, we detected recent introgression, with Anopheles gambiae and Anopheles coluzzii, of genes directly involved in vectorial capacity. Moreover, genome analysis of the new species allowed us to clarify the evolutionary history of the 3La inversion. Overall, An. fontenillei sp.n. analysis improved our understanding of the relationship between species within the An. gambiae complex, and provided insight into the evolution of vectorial capacity traits that are relevant for the successful control of malaria in Africa.

Natural Wolbachia infections are common in the major malaria vectors in Central Africa.

During the last decade, the endosymbiont bacterium Wolbachia has emerged as a biological tool for vector disease control. However, for long time, it was believed that Wolbachia was absent in natural populations of Anopheles. The recent discovery that species within the Anopheles gambiae complex host Wolbachia in natural conditions has opened new opportunities for malaria control research in Africa. Here, we investigated the prevalence and diversity of Wolbachia infection in 25 African Anopheles species in Gabon (Central Africa). Our results revealed the presence of Wolbachia in 16 of these species, including the major malaria vectors in this area. The infection prevalence varied greatly among species, confirming that sample size is a key factor to detect the infection. Moreover, our sequencing and phylogenetic analyses showed the important diversity of Wolbachia strains that infect Anopheles. Co-evolutionary analysis unveiled patterns of Wolbachia transmission within some Anopheles species, suggesting that past independent acquisition events were followed by co-cladogenesis. The large diversity of Wolbachia strains that infect natural populations of Anopheles offers a promising opportunity to select suitable phenotypes for suppressing Plasmodium transmission and/or manipulating Anopheles reproduction, which in turn could be used to reduce the malaria burden in Africa.