Phylogeny of 16S rRNA and nifH genes and regulation of nitrogenase activity by oxygen and ammonium in the genus Paenibacillus

All Paenibacillus 16S rDNA sequences, except for that of Paenibacillus massiliensis T7, formed a coherent cluster, distinct from Gram-positive nitrogen-fixing Clostridium pasteurianum and Heliobacterium chlorum. All Paenibacillus NifH sequences formed two main clusters. Cluster I encompassing the NifH sequences from most of members of Paenibacillus spp., such as Paenibacillus azotofixans NifH1 and NifH2, Paenibacillus polymyxa and Paenibacillus macerans. Cluster II including only P. azotofixans NifH3. Curiously, three copies of nifH genes of Paenibacillus sabine T27 clustered within P. azotofixans cluster I (NifH1 and NifH2). The effect of O2 and ammonium on nitrogenase activity was studied with 14 different nitrogen-fixing Paenibacillus strains. The optimal oxygen concentration level for all Paenibacillus strains is in the 0 to 0.05% range, similar to that for Klebsiella pneumoniae. In all Paenibacillus strains, the highest nitrogenase activity is obtained in the condition of 0-0.1 mM NH4Cl and the increase of NH4CI from 0.1 to 5 mM caused a rapid inhibition of nitrogenase activity. However, the inhibition was reversible in the presence of 200 mM NH4Cl in some Paenibacillus strains. It is the first time to use almost all of the recognized nitrogen-fixing Paenibacilus spp. to investigate the phylogeny of 16S rRNA and nifH genes. The data that the inhibition of O2 and ammonium on nitrogenase acitivity will provide a base for studying the molecular regulatory mechanism of nitrogen fixation in the genus Paenibacillus.