Comparison ofPaenibacillus azotofixansStrains Isolated from Rhizoplane, Rhizosphere, and Non-Root-Associated Soil from Maize Planted in Two Different Brazilian Soils

Abstract

Paenibacillus azotofixans is a nitrogen-fixing bacterium often found in soil and in the rhizospheres of different grasses. In this study, two Brazilian clay soils were planted with cross-hybrid maize (BR-201) and four stages of plant growth were analyzed to characterize the P. azotofixans populations present in the rhizoplanes, rhizospheres, and non-root-associated soils (herein called nonrhizospheres). A total of 106 strains were isolated and identified as P. azotofixans with an API 50CH kit, by classical biochemical tests, and via the use of specific primers based on the 16S rRNA gene in PCRs. To compare the isolated strains, phenotypic characteristics were determined and three different probes were used in hybridization experiments: two nif probes and one probe comprising a 0.58-kb fragment cloned from the P. azotofixans C3L4 genome. These results were used to construct a dendrogram, in which two main clusters could be observed. One cluster contained exclusively strains from Várzea soil, and the other contained the majority of strains from Cerrado soil. The 60 strains from Várzea soil and the 46 strains from Cerrado soil were further analyzed with REP and BOX primers, respectively. Based on the patterns obtained, it was possible to identify 21 different groups among strains from Várzea soil and 4 different groups among strains from Cerrado soil. These different patterns were tested by multivariate analysis of variance, and differences in the populations of P. azotofixans during the four stages of plant growth were demonstrated. Moreover, strains isolated from the rhizoplanes, rhizospheres, and nonrhizospheres of maize planted in Cerrado and Várzea soils were shown to be statistically different; the diversity of P. azotofixans strains was affected by the soil type.

Bacterial community composition in the rhizosphere of a transgenic, herbicide-resistant maize (Zea mays) and comparison to its non-transgenic cultivar Bosphore

“…It cannot be ruled out that the freeze^thaw lysis procedure selected in our study failed to release high amounts of DNA from bacteria with lysis-resistant cell walls and, thus, Gram-positive bacteria may have been underrepresented [47]. In fact, we could not detect the common maize rhizosphere coloniser Paenibacillus polymyxa [48]. In a study where bead-beating was used for lysis, several sequences of Gram-positive bacteria with a low G+C DNA content were recovered from maize rhizospheres after PCR and cloning of partial 16S rRNA genes [27].…”

Ecology and evolution of bacterial microdiversity

“…The analysis of the genotypic (rep-PCR) diversity of the related root-associated species Paenibacillus azoto¢xans populations isolated in Brazilian clay soils evidenced a signi¢cant e¡ect of the soil type [85]. A high phenotypic and genotypic (RAPD, rep-PCR) diversity within P. azo-to¢xans populations isolated from the rhizosphere of wheat, sugar cane, sorghum and maize was found in a study with Brazilian soils [86].…”

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

“…Furthermore, strains of Paenibacillus spp. can degradate polyaromatic hydrocarbons [23], produce phytohormones [24,25], furnish nutrients to plants by nitrogen fixation [13,14,[26][27][28], solubilize phosphate [11] and suppress phytopathogens through antagonistic functions [29,30].…”