Exploration of viral diversity in Aedes mosquitoes employing different shotgun metagenomic data analysis pipelines.

Abstract

Metagenomics or metaviromics is emerging as a powerful technology for pathogen surveillance and pandemic preparedness. Mosquitoes are important vectors for transmission of many emerging viruses responsible for numerous outbreaks. Monitoring mosquitoes becomes essential to investigate its virome which leads to understanding of disease dynamics and allow preventive actions. In this study, shotgun metagenomic methodology using Ion GeneStudio S5 System was optimized for exploration of viral diversity. A total of 1913 Aedes larvae were collected from Central India during post monsoon season of 2024. Aedes larvae reared to adulthood and processed for sequencing using Ion Torrent S5 platform. Computational analyses were performed using three bioinformatic pipelines: Chan Zuckerberg ID (CZ ID), Genome Detective Platform and the Galaxy Platform. A mock database of 11 known viruses was created as well as publicly available NCBI Sequence Read Archive (SRA) datasets were used to validate all three pipelines. In terms of detection accuracy, Genome Detective and CZ ID performed exceptionally well and therefore may be suitable for future mosquito virome surveillance studies. We found presence of viruses viz. Alphamesonivirus cavallynense and Phasivirus phasiense dominating in all samples. Dengue virus was detected in one sample by CZ ID, whereas Wenzhou sobemo-like virus, Hubei mosquito virus 2, Cell fusing agent virus found most commonly among samples. Other viruses found like Aedes anphevirus (AeAV; genus Glybovirus), Aedes totivirus, Verdadero virus and Chaq-like virus. Variation in the result among different pipelines are likely attributable to incorporation of different viral reference databases, classification algorithms, metrics and analysis parameters. To the best of our knowledge, this study represents the first metagenomic study of mosquitoes using Ion GeneStudio S5 platform in India. The findings provide a comparative evaluation of the metagenomic pipelines and elucidates detailed information of each pipeline and its working for future studies.