Assessment of the diversity of Paenibacillus species in environmental samples by a novel rpoB-based PCR-DGGE method

Abstract

A specific PCR system based on the gene encoding the RNA polymerase beta subunit, rpoB, was developed for amplification and denaturing gradient gel electrophoresis (DGGE) fingerprinting of Paenibacillus communities in environmental samples. This gene has been previously proven to be a powerful identification tool for the discrimination of species within the genus Paenibacillus and could avoid the limitations of 16S rRNA-based phylogenetic analysis. Initially, the PCR system based on universal rpoB primers were used to amplify DNAs of different Paenibacillus species. A new reverse primer (rpoBPAEN) was further designed based on an insertion of six nucleotides in the Paenibacillus sequences analyzed. This semi-nested PCR system was evaluated for specificity using DNAs isolated from 27 Paenibacillus species belonging to different 16S rRNA-based phylogenetic groups and seven non-Paenibacillus species. The non-Paenibacillus species were not amplified using this PCR approach and one group of Paenibacillus species consisting of strains without the six-base insert also were not amplified; these latter strains were found to be distinct based on 16S rRNA gene phylogeny. In addition, a clone library was generated from the rpoB fragments amplified from two Brazilian soil types (Cerrado and Forest) and all 62 clones sequenced were closely related to one of the 22 sequences from Paenibacillus previously obtained in this study. To assess the diversity of Paenibacillus species in Cerrado and Forest soils and in the rhizosphere of different cultivars of maize, a PCR-DGGE system was used. The Paenibacillus DGGE fingerprints showed a clear distinction between communities of Paenibacillus in Forest and Cerrado soils and rhizosphere samples clustered along Cerrado soil. Profiles of cultivars CMS22 and CMS36 clustered together, with only 53% of similarity to CMS11 and CMS04. The results presented here demonstrate the potential use of the rpoB-based Paenibacillus-specific PCR-DGGE method for studying the diversity of Paenibacillus populations in natural environments.

Proposed minimal standards for describing new taxa of aerobic, endospore-forming bacteria

“…Zeigler (2005) found recN sequence similarities to yield clustering patterns among Geobacillus very similar to those obtained with 16S rRNA gene sequences, and Mota et al (2005) obtained clustering patterns for Paenibacillus based upon rpoB sequence comparisons that were similar to those obtained with 16S rRNA gene sequences. Wang et al (2007a, b) included gyrB sequence comparisons in studies of the B. subtilis group and demonstrated the synonymy of Bacillus axarquiensis and Bacillus malacitensis with Bacillus mojavensis, and Cerritos et al (2008) included recA sequence comparisons in the work that led to the proposal of Bacillus coalhuilensis.…”

Genome Sequence of the Polymyxin-Producing Plant-Probiotic Rhizobacterium Paenibacillus polymyxa E681

“…Paenibacillus spp. are important members of soil-or plant-associated ecosystems (3,8,20), with Paenibacillus polymyxa as one of the most industrially significant bacteria (13,17,25,31). P. polymyxa E681, an endospore former isolated from the rhizosphere of winter barley in South Korea (14,27), suppresses plant diseases, produces antibiotics and a plant hormone, secretes a variety of hydrolytic enzymes, and has good root-colonizing ability (4,26).…”

Inventory of bats and culturable proteobacteria from Cueva Las Escaleras (Ta´chira, Venezuela): evidence of potential human health risks

“…Another bacterium found in this study, a Firmicutes, was Paenibacillus lactis, an organism originally isolated from raw and heat-treated milk (Schedelman et al, 2004), but that can also be found in bacterial communities in environmental samples (da Mota et al, 2005). We found no reference dealing with pathogenicity to humans of this bacterium or the recently described Raoultella electrica (Kimura et al, 2014).…”