Real-time volatilomics reveals microbiota and pathogen fingerprints in the honey bee

Understanding the complex relationship between gut microbiota and their hosts often relies on invasive sampling techniques. Honey bees provide a tractable model for host-microbe studies. Here, we establish single-bee volatilomics using secondary electrospray ionization-high-resolution mass spectrometry (SESI-HRMS) to examine volatile organic compounds released to the air around an individual live honey bee. Specifically, we focused on the primary gut microbiota metabolites present in gnotobiotic bees. Our findings reveal distinct volatilome profiles in honey bees that depend on their gut bacterial colonization state. We cross-validated our findings using an established metabolomics technique, liquid chromatography-high-resolution mass spectrometry (LC-HRMS), to compare and contrast the metabolites detectable with each method. Finally, we assessed the ability of SESI-HRMS to detect colonization with the bee pathogen Serratia marcescens. By comparing the volatile signature of this bacterium grown in liquid culture with that of infected honey bee headspace, we identified overlapping compounds, including butane-2,3-diol, that were elevated in infected bees relative to uninfected controls. Non-invasive SESI-HRMS volatilomics, paired with LC-HRMS, therefore, have the potential to identify biomarkers of bee microbiome composition and infection at the level of individual insects. These biomarkers represent practical targets for the development of simple, field-ready diagnostic tools for monitoring pollinator health.

Honey bees are vital to various ecosystems and human agriculture as globally present pollinators. Current declines in bee populations threaten crop productivity and biodiversity and are driven by many factors, including habitat loss, pesticides, and pathogenic infections. The findings presented in this work demonstrate how the analysis of volatile species can precisely detect products of metabolic activity coming from the honey bee and its associated gut microbial community. We were able to demonstrate how high-resolution mass spectrometry can be used to study the host-microbiota relationship on the level of an individual insect. This technique, combined with volatilomics, allowed us to outline a volatile signature of infection in honey bees, which could become a target for in-field beehive disease monitoring.