Microbiome Simplification During Metamorphosis in Larva and Adults of Armigeres subalbatus (Coquillett, 1898) (Culicidae) Revealed by Shotgun Metagenomics

Abstract

Armigeres subalbatus is medically significant vector for filarial worms and the Japanese encephalitis virus. Shotgun metagenomic sequencing was employed to investigate the bacterial communities in A. subalbatus mosquitoes. The diversity metrics (Shannon H′, Simpson 1-D, Berger-Parker) were calculated for larval and adult stages. De novo assembly and binning were used to recover metagenome-assembled genomes (MAGs) with > 82% completeness and < 4% contamination. Functional profiling assessed gene expression via transcripts per million (TPM) and clusters of orthologous groups (COG) categories. Larval microbiomes showed high alpha diversity (Shannon H′ ≈ 1.336 ± 0.163, Simpson 1-D = 0.684 ± 0.046), dominated by Gammaproteobacteria (Aeromonas, Morganella, and Yersinia) and Bacteroidota, with persistent Shewanella and Acinetobacter. Adult microbiomes exhibited low diversity (Shannon H′ = 0.637 ± 0.100, Berger-Parker = 0.682 ± 0.026), near-monoculture dominated by Aeromonas hydrophila, alongside low-abundance Stenotrophomonas, Pseudomonas, and Microbacterium. Six high-quality MAGs were recovered: larval (Bacteroidota, Shewanella, and Acinetobacter); adult (Acinetobacter, Stenotrophomonas, and Shewanella), confirming persistence of Shewanella and Acinetobacter, absence of Bacteroidota, and emergence of Stenotrophomonas in adults. Adult microbiomes displayed metabolic hyperactivity, with 1.5–4 times higher transcriptional output across COG categories compared to larvae. Chemotaxis [Methyl-accepting chemotaxis protein (MCP), K03406: ~6000 TPM in adults vs. < 1000 TPM in larvae] and ABC transporters (PF00005: > 10,000 TPM in adults) dominated adults, while larval expression was balanced among housekeeping functions. The microbiome undergoes significant restructuring during mosquito development, shifting from diverse larval communities to metabolically active, low-diversity adult assemblages. Recovered MAGs provide a genomic basis for future studies on mosquito microbiota dynamics and functions.