Metatranscriptomic analysis of autonomously collected and preserved marine bacterioplankton

Community RNA sequencing, or metatranscriptomics, can be used to describe the relative abundances of transcribed genes, as well as the response of microbial populations to environmental stimuli in both observational and experimental settings. Using the Environmental Sample Processor (ESP) platform, we developed and field-tested a protocol for automated collection and fixation of marine microbes that uses an RNALater treatment to preserve RNA in whole cells collected on filters. Replicate microbial samples preserved using this protocol yielded discrete rRNA peaks and stable metatranscriptomic profiles over at least four weeks when stored at room temperature or onboard the ESP at in situ temperatures. In a field test using the validated protocol, we used a morred ESP to remotely and automatically collect seawater samples from Monterey Bay for metatranscriptomic analysis. cDNA libraries were synthesized and successfully sequenced using community RNA extracted from samples collected at four time points over the course of a single day. In all four samples, the oxygenic photoautotrophs were predominantly eukaryotic, while the bacterial community was dominated by proteorhodopsin-containing photoheterotrophs, with the most abundant groups being Polaribacter-like Flavobacteria and a Rhodobacterales bacterium sharing high similarity with Rhodobacterales sp. HTCC2255. However, each time point was associated with distinct species abundance and expressed gene profiles. These laboratory and field tests indicate that autonomous collection and preservation of bacterioplankton for transcriptional analysis is a promising approach for characterizing the expressed genes and environmental responses of marine microbial communities.