Flow cytometry with cell sorting and sequencing as a tool for the study of the stomach microbiota of the Humboldt Current krill (Euphausiacea)

Abstract

Euphausiids (krill) are important contributors to marine biomass and key players in marine pelagic trophic webs. Euphausiid stomachs represent a specific niche for microbes that participate in the digestion of the host's dietary components. Methods for the study of the diversity and function of these microorganisms remain complex. Bacterial ribosomal sequences obtained from lysates of stomachs are often overrepresented by organisms from the surrounding environment. Flow cytometry with cell sorting (FC-CS) have become a powerful technique to study microbial community structure but also for the study of population genomics of gut-associated bacteria. We compared the performance of the FC-CS-sequencing and total DNA extraction-sequencing to study the stomach microbiota of the Humboldt Current krill. Non-specific amplification was not retrieved in the polymerase chain reaction (PCR) from cells sorted, opposite to the observed using the DNA from the whole lysate. Sequences obtained from the whole stomach DNA were enriched in picocyanobacteria, whereas sequences retrieved from cells sorted belonged almost exclusively to Balneola sp. of the new phylum Balneolaeota. Our results suggest that the stomach-associated microbiota can be successfully characterized by FC-CS and sequencing by manual scraping of the stomach. The implementation of this technique might complement future studies on host-microbes interaction and their implications on the marine pelagic food web.