Habitat Type and Locality Structure the Midgut Microbiota of Aedes albopictus

The mosquito midgut microbiome plays a crucial role in nutrition, reproduction, and immunity, yet how locality and urban development shape these communities and their potential relevance to mosquito-pathogen interactions remains poorly understood. Here, we investigated bacterial community composition and diversity in the midguts of adult female Aedes albopictus collected from residential and woodland habitats of Champaign-Urbana and Charleston in Illinois, USA. We sequenced the V4 region of the 16 S rRNA gene from 160 samples and analyzed the data using QIIME 2. After quality and feature filtering, 112 samples were retained, yielding 2,531 unique amplicon sequence variants assigned to 34 bacterial phyla, 246 families, and 404 genera. Because formal contaminant assessment was not possible, findings should be interpreted with appropriate caution. Woodland habitats showed significantly higher Shannon diversity, observed ASV richness, and phylogenetic diversity than residential habitats, with Charleston woodland samples exhibiting the highest richness and phylogenetic diversity. Factorial analyses showed significant associations of both city and habitat type with Shannon diversity, observed richness, and Faith's phylogenetic diversity, with significant interaction terms also detected. Beta diversity analyses revealed distinct clustering of Charleston woodland samples, and factorial PERMANOVA indicated significant associations of both city and habitat type across all four beta diversity metrics, with the strongest interaction effect observed for unweighted UniFrac. In conclusion, these results show that habitat type and locality are strongly associated with the composition and diversity of the Ae. albopictus midgut microbiota, underscoring the importance of habitat-specific microbial patterns in mosquito biology.