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Harris County, Texas, remains at continuous risk to mosquito-borne diseases due to its geographic landscape and abundance of medically important mosquito vectors. Targeted mitigation of these mosquitoes requires accurate identification of these mosquitoes taxa. Currently, there is a paucity of genetic information to inform molecular identification and phylogenetic relationships beyond well-studied mosquito species. Here we utilized a genome skimming approach using shallow shot gun sequencing to generate data and assemble the mitochondrial genomes of 37 mosquito species collected in Harris County, Texas. This report includes the complete mitochondrial genome for 25 newly sequenced species spanning 10 genera; the genomes were consistent with reference genomes in the GenBank database having 37 genes (13 protein-coding, 2 rRNA and 22 tRNA), and average AT content of 78.74%. Bayesian and maximum likelihood tree topologies using just the easily aligned 13 concatenated protein coding genes confirmed phylogenetic placement of species for Aedes, Anopheles and Culex genera clustering in single clades as expected. Furthermore, this approach provided more robust phylogenetic placement/identity of study taxa when compared to the use of the traditional cytochrome oxidase I partial gene barcode sequence for molecular identification. This study demonstrates the utility of genome skimming as a cost-effective alternative approach to generate reference sequences for the validation of mosquito identification and taxonomic rectification, knowledge necessary for guiding targeted vector interventions.
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Phylogenetic taxonomy of the Zambian Anopheles coustani group using a mitogenomics approach
Background: Mosquito species belonging to the Anopheles coustani group have been implicated in driving residual malaria transmission in sub-Saharan Africa and are regarded as an established primary vector in Madagascar. The morphological identification of mosquitoes in this group is challenging due to similarity of features and their molecular confirmation is difficult due to a paucity of reference sequence data representing all members of the group. Conventional molecular barcoding with the cytochrome oxidase I (COI) gene and the internal transcribed spacer 2 (ITS2) region targets is limited in their discrimination and conclusive identification of members of species complexes. In contrast, complete mitochondrial genomes (mitogenomes) have demonstrated much improved power over barcodes to be useful in rectifying taxonomic discrepancies in Culicidae. The goal of this study was to characterize the phylogenetic taxonomy of Zambian members of the An. coustani group by generating and then using complete mitochondrial genomes for phylogenetic rectification. Methods: A genome skimming approach was utilized via shallow shotgun sequencing on individual mosquito specimens to generate sequence reads for mitogenome assembly. Bayesian inferred phylogenies and molecular dating estimations were perfomed on the concatenated protein coding genes using the Bayesian Evolutionary Analysis by Sampling Trees 2 (BEAST 2) platform. Divergence estimates were calibrated for members of the An. coustani group based on published calucations for Anopheles-Aedes. Results: This study generated 17 new complete mitogenomes which were comprable to reference An. coustani mitogenomes in the GenBank repository by having 13 protein coding, 22 transfer RNA and 2 ribosomal RNA genes, with an average length of 15,400 bp and AT content of 78.3%. Bayesian inference using the concatenated protein coding genes from the generated and publicly available mitogenomes yielded six clades: one for each of the four taxa targeted in this study, the GenBank references, and a currently unknown species. Divergence times estimated that the An. coustani group separated from the Anopheles gambiae complex approximately 110 million years ago (MYA), and members within the complex diverged at times points ranging from ~ 34 MYA to as recent as ~ 7 MYA. Conclusions: These findings demonstrate the value of mitochondrial genomes in differentiating cryptic taxa and help to confirm morphological identities of An. coustani sensu stricto, Anopheles paludis, Anopheles zeimanni and Anopheles tenebrosus. Divergence estimates within the An. coustani group are similar when compared to species with morphologically distinct features. These analyses also highlight the likely presence of other cryptic An. coustani group members circulating in Zambia.
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- Award ID(s):
- 2134862
- PAR ID:
- 10662420
- Publisher / Repository:
- BioMed Central (BMC)
- Date Published:
- Journal Name:
- Malaria Journal
- Volume:
- 24
- Issue:
- 1
- ISSN:
- 1475-2875
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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- Free Publicly Accessible Full Text
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Accepted Manuscript
- Journal Article:
- https://doi.org/10.1186/s12936-025-05461-z
