ABSTRACT
Exometabolomics is an approach to assess how microorganisms alter, or react to their environments through the depletion and production of metabolites. It allows the examination of how soil microbes transform the small molecule metabolites within their environment, which can be used to study resource competition and cross-feeding. This approach is most powerful when used with defined media that enable tracking of all metabolites. However, microbial growth media have traditionally been developed for the isolation and growth of microorganisms but not metabolite utilization profiling through Liquid Chromatography Tandem Mass Spectrometry (LC-MS/MS). Here, we describe the construction of a defined medium, the Northen Lab Defined Medium (NLDM), that not only supports the growth of diverse soil bacteria but also is defined and therefore suited for exometabolomic experiments. Metabolites included in NLDM were selected based on their presence in R2A medium and soil, elemental stoichiometry requirements, as well as knowledge of metabolite usage by different bacteria. We found that NLDM supported the growth of 108 of the 110 phylogenetically diverse (spanning 36 different families) soil bacterial isolates tested and all of its metabolites were trackable through LC-MS/MS analysis. These results demonstrate the viability and utility of the constructed NLDM medium for growing and characterizing diverse microbial isolates and communities.
ABSTRACT
Endophytic nitrogen-fixing (diazotrophic) bacteria are essential members of the microbiome of switchgrass (Panicum virgatum), considered to be an important commodity crop in bioenergy production. While endophytic diazotrophs are known to provide fixed atmospheric nitrogen to their host plant, there are many other plant growth-promoting (PGP) capabilities of these organisms to be demonstrated. The diversity of PGP traits across different taxa of switchgrass-colonizing endophytes is understudied, yet critical for understanding endophytic function and improving cultivation methods of important commodity crops. Here, we present the isolation and characterization of three diazotrophic endophytes: Azospirillum agricola R1C, Klebsiella variicola F10Cl, and Raoultella terrigena R1Gly. Strains R1C and F10Cl were isolated from switchgrass and strain R1Gly, while isolated from tobacco, is demonstrated herein to colonize switchgrass. Each strain exhibited highly diverse genomic and phenotypic PGP capabilities. Strain F10Cl and R1Gly demonstrated the highest functional similarity, suggesting that, while endophyte community structure may vary widely based on host species, differences in functional diversity are not a clearly delineated. The results of this study advance our understanding of diazotrophic endophyte diversity, which will allow us to design robust strategies to improve cultivation methods of many economically important commodity crops.
