Expanding the Diversity of Bacterioplankton Isolates and Modeling Isolation Efficacy with Large-Scale Dilution-to-Extinction Cultivation

摘要

Cultivated bacterioplankton representatives from diverse lineages and locations are essential for microbiology, but the large majority of taxa either remain uncultivated or lack isolates from diverse geographic locales. We paired large-scale dilution-to-extinction (DTE) cultivation with microbial community analysis and modeling to expand the phylogenetic and geographic diversity of cultivated bacterioplankton and to evaluate DTE cultivation success. Here, we report results from 17 DTE experiments totaling 7,820 individual incubations over 3 years, yielding 328 repeatably transferable isolates. Comparison of isolates to microbial community data for source waters indicated that we successfully isolated 5 of the observed bacterioplankton community throughout the study; 43 and 26 of our isolates matched operational taxonomic units and amplicon single-nucleotide variants, respectively, within the top 50 most abundant taxa. Isolates included those from previously uncultivated clades such as SAR 1 LD12 and Actinobacteria aclV, as well as geographically novel members from other ecologically important groups like SAR11 subclade IIla, SAR116, and others, providing isolates in eight putatively new genera and seven putatively new species. Using a newly developed DTE cultivation model, we evaluated taxon viability by comparing relative abundance with cultivation success. The model (i) revealed the minimum attempts required for successful isolation of taxa amenable to growth on our media and (ii) identified possible subpopulation viability variation in abundant taxa such as SAR11 that likely impacts cultivation success. By incorporating viability in experimental design, we can now statistically constrain the effort necessary for successful cultivation of specific taxa on a defined medium.