Exploratory metatranscriptomic survey of bat-associated RNA viruses in Quzhou city, China

Bats are recognized as important reservoirs of diverse RNA viruses, yet the bat virome in Quzhou City, Zhejiang Province, remains poorly characterized. Here, we conducted an exploratory metatranscriptomic survey of bat-associated RNA viruses in this region, with particular attention to viral families that include known zoonotic members. Bats were sampled between August and October 2023 from one roost in each of six administrative areas of Quzhou City. Species identification was performed by Cytb sequencing, and pooled intestinal, lung, and other visceral tissues were subjected to high-throughput sequencing, sequence annotation, community comparison, and phylogenetic analysis. A total of 167 bats were collected, representing Hipposideros armiger and Rhinolophus sinicus, and 35 pooled libraries were generated. Viral contigs were assigned to seven RNA viral families: Coronaviridae, Flaviviridae, Astroviridae, Hepeviridae, Hantaviridae, Sedoreoviridae, and Arenaviridae. Coronaviridae was the most frequently detected family (97.14% of pooled libraries), whereas Sedoreoviridae was the least frequent (2.86%). The intestinal pool from Kecheng District (1A) showed the greatest family-level diversity, with five viral families detected. PCoA and PERMANOVA indicated significant regional structuring of viral composition (R ² = 0.315, P < 0.001), whereas tissue-type differences were not statistically significant (R ² = 0.078, P = 0.195). Because host species and sampling region were fully confounded in this design, species-level patterns were interpreted descriptively only and not as independent species effects. Phylogenetic analyses identified several genetically divergent contigs, including Mamastrovirus-related sequences and multiple arenavirid contigs forming distinct branches. However, given the pooled design and the absence of independent laboratory validation, these findings should be interpreted as baseline evidence of viral diversity and geographic structuring rather than confirmation of host association, zoonotic potential, or pathogenicity. This study provides an initial virome profile for batsin Quzhou City and a foundation for future individual-level validation, genome completion, and ecological investigation.