Metagenomic and gene expression patterns in declining commercial honey bee colonies

Managed honey bee colonies (Apis mellifera) in the US continue to experience high overwinter loss rates driven by parasites, pathogens, poor nutrition, and pesticides. To mitigate these losses, inspection and monitoring are critical for identifying traits of colonies in decline and potential causal factors. In this study, we apply molecular methods to associate potential causative agents with colonies in various stages of decline. Initially, we investigated in-hive bee metagenomic RNA isolated from 15 colonies across seven managed operations in California whose adult bee and brood populations were classified as Strong, Medium, or Weak in strength. We discovered that Weak colonies harbored 2.2- and 3.6- fold more viral species than Medium and Strong colonies, respectively, as well as larger viral read pools despite similar library sizes. They also displayed higher nucleotide variation in Varroa-vectored viruses, indicating associations with high mite populations. When investigating differences in host gene expression, we discovered an upregulation of immune-related pathways in Weak colonies relative to Strong. Specifically, Weak colonies upregulated genes related to wound healing, phagocytosis, oxidative stress resistance, apoptosis, and RNA interference. Most antimicrobial peptides were upregulated in Weak colonies, although defensin1 was significantly higher in Strong colonies, along with several detoxification enzymes and the royal jelly peptide apisimin. Weak colonies also showed an upregulation of transcripts tied to abnormal protein digestion. The low levels of viral replication and fewer species of mite-vectored viruses in Strong colonies may be due to successful Varroa management. Strong colonies also displayed upregulated levels of nine different ubiquinone transcripts, arguably reflecting increasing longevity or a younger in-hive population compared to Weak colonies. Overall, these results provide a detailed account of viral metagenomics and associated host responses, providing new insights into the mechanisms underlying honey bee colony decline under comparable management conditions.