Metagenomic studies reveal diverse microbial community in the developmental stages of highly adaptable malarial...

Anopheles stephensi, a highly adaptable malaria vector species, continues to expand its range from South Asia to Sub-Saharan Africa, posing a serious global public health concern. In India, it serves as the principal urban vector of both Plasmodium falciparum and P. vivax. Conventional control measures reliant on chemical insecticides have raised issues of resistance, highlighting the need for alternative strategies such as microbiota-mediated vector control. This study aimed to test the hypothesis that a subset of bacterial taxa persist across developmental stages of An. stephensi, representing potential candidates for transstadial transmission and future paratransgenic manipulation. Using both culture-based data and next-generation sequencing (NGS) approaches targeting the 16 S rRNA gene (V3-V4 region), we characterized bacterial communities from breeding water, larvae, pupae, and adult mosquitoes (male and female) collected in Goa, India. Across all developmental stages, Proteobacteria and Firmicutes were the dominant phyla, while 15 bacterial genera formed the putative core microbiome shared by ≥ 80% of stages at ≥ 0.1% abundance. Among these, Pseudomonas (adult males: 11.5%, pupae: 3.2%), Exiguobacterium, Acinetobacter, Psychrobacter, and Asticcacaulis were consistently detected, together contributing approximately 30% of total microbial composition. Alpha diversity indices indicated higher richness and evenness in pupae and adults than in larvae, suggesting microbial enrichment during metamorphosis. Beta diversity and PCoA analyses clustered pupal and adult stages distinctly from larvae and breeding water, confirming selective microbial retention through development. These findings reveal that An. stephensi harbors a stable, stage-spanning core microbiome dominated by metabolically versatile genera with potential for transstadial persistence. The dominance of Pseudomonas across life stages supports its candidacy for paratransgenic applications aimed at disrupting malaria transmission. This work provides the first integrated culture-NGS baseline of An. stephensi microbiota from India, offering essential insight for microbiome-based vector control strategies.

The online version contains supplementary material available at 10.1007/s13205-026-04739-6.