Dynamics of marine bacterial and phytoplankton populations using multiplex liquid bead array technology

Abstract

Heterotrophic bacteria and phytoplankton dominate the biomass and play major roles in the biogeochemical cycles of the surface ocean. Here, we designed and tested a fast, high-throughput and multiplexed hybridization-based assay to detect populations of marine heterotrophic bacteria and phytoplankton based on their small subunit ribosomal DNA sequences. The assay is based on established liquid bead array technology, an approach that is gaining acceptance in biomedical research but remains underutilized in ecology. End-labelled PCR products are hybridized to taxon-specific oligonucleotide probes attached to fluorescently coded beads followed by flow cytometric detection. We used ribosomal DNA environmental clone libraries (a total of 450 clones) and cultured isolates to design and test 26 bacterial and 10 eukaryotic probes specific to various ribotypes and genera of heterotrophic bacteria and eukaryotic phytoplankton. Pure environmental clones or cultures were used as controls and demonstrated specificity of the probes to their target taxa. The quantitative nature of the assay was demonstrated by a significant relationship between the number of target molecules and fluorescence signal. Clone library sequencing and bead array fluorescence from the same sample provided consistent results. We then applied the assay to a 37-day time series of coastal surface seawater samples from the Southern California Bight to examine the temporal dynamics of microbial communities on the scale of days to weeks. As expected, several bacterial phylotypes were positively correlated with total bacterial abundances and chlorophyll a concentrations, but others were negatively correlated. Bacterial taxa belonging to the same broad taxonomic groups did not necessarily correlate with one another, confirming recent results suggesting that inferring ecological role from broad taxonomic identity may not always be accurate.