Molecular Analysis of the Microbial Guild Fixing Nitrogen in Ricefield Soils in Missouri

Abstract

Non-symbiotic diazotrophic microbes are important contributors to global N budgets in cereal crops. Knowledge of the biogeography of the organisms in this functional guild increases our understanding of biological N fixation in diverse locations and climates. Here, we describe the diazotrophic community in the previously unstudied, extensive ricefields of southeast Missouri, using restriction fragment length polymorphism (RFLP) analysis and sequencing of nifH gene clones. While nine RFLP patterns were observed in random nifH clones, these groups were not all supported by gene sequencing, suggesting that the RFLP of nifH genes alone is not suitable for describing diazotrophic guilds. Dozens of nifH clones from Missouri ricefield soils were sequenced and analyzed phylogenetically. The nifH genes detected were predominantly from Geobacteraceae, most closely related to Geobacter and Geomonas species. There were substantial clusters of nifH clones most closely related to Desulfovibrionales and other Proteobacteria. Many of the clones did not closely cluster with nifH sequences from known isolates or clades. No cyanobacterial or archaeal sequences were detected in the Missouri ricefield soils. The microbial guild fixing N appeared to be rich in anaerobes and lithotrophs. Organisms in Geobacter and Geomonas seem to be cosmopolitan, but endemism was evident, since nifH clones were recovered that formed clusters not previously reported from ricefields in other locations.