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      Novel Wolbachia strains in Anopheles malaria vectors from Sub-Saharan Africa

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          Abstract

          Background:  Wolbachia, a common insect endosymbiotic bacterium that can influence pathogen transmission and manipulate host reproduction, has historically been considered absent from the  Anopheles (An.) genera, but has recently been found in  An. gambiae s.l. populations in West Africa.  As there are numerous  Anopheles species that have the capacity to transmit malaria, we analysed a range of species across five malaria endemic countries to determine  Wolbachia prevalence rates, characterise novel  Wolbachia strains and determine any correlation between the presence of  PlasmodiumWolbachia and the competing bacterium  Asaia.

          Methods:  Anopheles adult mosquitoes were collected from five malaria-endemic countries: Guinea, Democratic Republic of the Congo (DRC), Ghana, Uganda and Madagascar, between 2013 and 2017.  Molecular analysis was undertaken using quantitative PCR, Sanger sequencing,  Wolbachia multilocus sequence typing (MLST) and high-throughput amplicon sequencing of the bacterial  16S rRNA gene. 

          Results: Novel  Wolbachia strains were discovered in five species:  An. coluzziiAn. gambiae s.s.,  An. arabiensisAn. moucheti and  An. species A, increasing the number of  Anopheles species known to be naturally infected. Variable prevalence rates in different locations were observed and novel strains were phylogenetically diverse, clustering with  Wolbachia supergroup B strains.  We also provide evidence for resident strain variants within  An. species A.  Wolbachia is the dominant member of the microbiome in  An. moucheti and  An. species A but present at lower densities in  An. coluzzii.  Interestingly, no evidence of  Wolbachia/Asaia co-infections was seen and  Asaia infection densities were shown to be variable and location dependent. 

          Conclusions: The important discovery of novel  Wolbachia strains in  Anopheles provides greater insight into the prevalence of resident  Wolbachia strains in diverse malaria vectors.  Novel  Wolbachia strains (particularly high-density strains) are ideal candidate strains for transinfection to create stable infections in other  Anopheles mosquito species, which could be used for population replacement or suppression control strategies.

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          The genome sequence of the malaria mosquito Anopheles gambiae.

          Anopheles gambiae is the principal vector of malaria, a disease that afflicts more than 500 million people and causes more than 1 million deaths each year. Tenfold shotgun sequence coverage was obtained from the PEST strain of A. gambiae and assembled into scaffolds that span 278 million base pairs. A total of 91% of the genome was organized in 303 scaffolds; the largest scaffold was 23.1 million base pairs. There was substantial genetic variation within this strain, and the apparent existence of two haplotypes of approximately equal frequency ("dual haplotypes") in a substantial fraction of the genome likely reflects the outbred nature of the PEST strain. The sequence produced a conservative inference of more than 400,000 single-nucleotide polymorphisms that showed a markedly bimodal density distribution. Analysis of the genome sequence revealed strong evidence for about 14,000 protein-encoding transcripts. Prominent expansions in specific families of proteins likely involved in cell adhesion and immunity were noted. An expressed sequence tag analysis of genes regulated by blood feeding provided insights into the physiological adaptations of a hematophagous insect.
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            Identification of single specimens of the Anopheles gambiae complex by the polymerase chain reaction.

            A ribosomal DNA-polymerase chain reaction (PCR) method has been developed for species identification of individuals of the five most widespread members of the Anopheles gambiae complex, a group of morphologically indistinguishable sibling mosquito species that includes the major vectors of malaria in Africa. The method, which is based on species-specific nucleotide sequences in the ribosomal DNA intergenic spacers, may be used to identify both species and interspecies hybrids, regardless of life stage, using either extracted DNA or fragments of a specimen. Intact portions of a mosquito as small as an egg or the segment of one leg may be placed directly into the PCR mixture for amplification and analysis. The method uses a cocktail of five 20-base oligonucleotides to identify An. gambiae, An. arabiensis, An. quadriannnulatus, and either An. melas in western Africa or An. melas in eastern and southern Africa.
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              The dominant Anopheles vectors of human malaria in Africa, Europe and the Middle East: occurrence data, distribution maps and bionomic précis

              Background This is the second in a series of three articles documenting the geographical distribution of 41 dominant vector species (DVS) of human malaria. The first paper addressed the DVS of the Americas and the third will consider those of the Asian Pacific Region. Here, the DVS of Africa, Europe and the Middle East are discussed. The continent of Africa experiences the bulk of the global malaria burden due in part to the presence of the An. gambiae complex. Anopheles gambiae is one of four DVS within the An. gambiae complex, the others being An. arabiensis and the coastal An. merus and An. melas. There are a further three, highly anthropophilic DVS in Africa, An. funestus, An. moucheti and An. nili. Conversely, across Europe and the Middle East, malaria transmission is low and frequently absent, despite the presence of six DVS. To help control malaria in Africa and the Middle East, or to identify the risk of its re-emergence in Europe, the contemporary distribution and bionomics of the relevant DVS are needed. Results A contemporary database of occurrence data, compiled from the formal literature and other relevant resources, resulted in the collation of information for seven DVS from 44 countries in Africa containing 4234 geo-referenced, independent sites. In Europe and the Middle East, six DVS were identified from 2784 geo-referenced sites across 49 countries. These occurrence data were combined with expert opinion ranges and a suite of environmental and climatic variables of relevance to anopheline ecology to produce predictive distribution maps using the Boosted Regression Tree (BRT) method. Conclusions The predicted geographic extent for the following DVS (or species/suspected species complex*) is provided for Africa: Anopheles (Cellia) arabiensis, An. (Cel.) funestus*, An. (Cel.) gambiae, An. (Cel.) melas, An. (Cel.) merus, An. (Cel.) moucheti and An. (Cel.) nili*, and in the European and Middle Eastern Region: An. (Anopheles) atroparvus, An. (Ano.) labranchiae, An. (Ano.) messeae, An. (Ano.) sacharovi, An. (Cel.) sergentii and An. (Cel.) superpictus*. These maps are presented alongside a bionomics summary for each species relevant to its control.
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                Author and article information

                Journal
                Wellcome Open Research
                Wellcome Open Res
                F1000 Research, Ltd.
                2398-502X
                2018
                November 27 2018
                : 3
                : 113
                Article
                10.12688/wellcomeopenres.14765.2
                © 2018

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