Culture-Dependent and Independent Analyses of the Microbial Communities Inhabiting the Giant Duckweed (Spirodela polyrrhiza) Rhizoplane and Isolation of a Variety of Rarely Cultivated Organisms within the Phylum Verrucomicrobia

The microbial communities of the rhizoplane, the surface part of roots, in aquatic plants are not understood at all. In this study, we analyzed microbial communities in the rhizoplane of a floating aquatic plant, giant duckweed (Spirodela polyrrhiza), based on cultivation-dependent and independent analyses. The cultivation-based analysis using agar and gellan gum plates revealed that the rhizoplane isolates were affiliated with four bacterial lineages; the Alphaproteobacteria, Betaproteobacteria, Bacteroidetes and Verrucomicrobia. Interestingly, microbes belonging to the phylum Verrucomicrobia accounted for 24% of all the isolates, suggesting that the rhizoplane of S. polyrrhiza forms a specific habitat for the organisms within this phylum. Culture-independent 16S rRNA gene cloning showed that the clonal sequences were affiliated with eight bacterial classes and phyla: the classes Alphaproteobacteria (14% total clones), Betaproteobacteria (45%), Gammaproteobacteria (2%) and Deltaproteobacteria (2%), and the phyla Bacteroidetes (11%), Verrucomicrobia (2%), Planctomycetes (2%) and Cyanobacteria (22%). Comparative analysis of the microbial communities in the rhizoplane between culture-dependent and independent methods revealed that 33% of the taxonomic groups of bacterial species detected in the molecular analysis were cultivable. Our findings suggest that the microbes in the rhizoplane of giant duckweed are comprised of a diverse array of readily cultured organisms including a variety of strains within the Verrucomicrobia, a well-known phylum that contains a number of yet-to-be cultivated organisms.