Prevalence, Genetic Diversity, and Landscape Associations of Orthohantavirus puumalaense in Bank Voles (Clethrionomys glareolus) from Northern Sweden

Puumala hantavirus (PUUV, Orthohantavirus puumalaense) is one of the primary causative agents of haemorrhagic fever with renal syndrome in Europe and is maintained in natural populations of the bank vole (Clethrionomys glareolus, also known as Myodes glareolus). Despite public health relevance, we are only starting to understand the molecular properties and interplay between environmental and ecological factors of the pathogen that explain PUUV infection in bank voles. Here, we investigated PUUV occurrence, genetic structure, and environmental associations in bank voles sampled from two boreal forest areas in northern Sweden, during a complete vole population cycle (2020-2023). In total, 519 voles were screened for PUUV RNA using targeted reverse transcription PCR (RT-PCR). PUUV small (S-) segment RNA was detected in both study areas and observed infection patterns varied with sex, body weight, season and year. Specifically, we detected significant interactions between season and area and between season and body weight, with males showing consistently higher infection probabilities. Infection probability was also higher during periods of increased vole abundance and peaked in 2022. Phylogenetic analysis of partial S segment sequences demonstrated that all detected sequences clustered within the North-Scandinavian PUUV lineage, with no apparent spatial differentiation, indicating limited genetic structuring between the sampling areas. Habitat analyses at multiple spatial scales did not identify significant associations between PUUV occurrence and land-use variables, suggesting that infection dynamics were driven primarily by host demographic and temporal factors rather than broad-scale habitat composition. These findings highlight the importance of host demographics and temporal dynamics in shaping PUUV epidemiology in its reservoir, and provide additional insight into the molecular ecology of PUUV in northern Europe.