Development and application of a selective PCR-denaturing gradient gel electrophoresis approach to detect a recently...

The worldwide presence of a hitherto-nondescribed group of predominant soil microorganisms related to Bacillus benzoevorans was analyzed after development of two sets of selective primers targeting 16S rRNA genes in combination with denaturing gradient gel electrophoresis (DGGE). The high abundance and cultivability of at least some of these microorganisms makes them an appropriate subject for studies on their biogeographical dissemination and diversity. Since cultivability can vary significantly with the physiological state and even between closely related strains, we developed a culture-independent 16S rRNA gene-targeted DGGE fingerprinting protocol for the detection of these bacteria from soil samples. The composition of the B. benzoevorans relatives in the soil samples from The Netherlands, Bulgaria, Russia, Pakistan, and Portugal showed remarkable differences between the different countries. Differences in the DGGE profiles of these communities in archived soil samples from the Dutch Wieringermeer polder were observed over time during which a shift from anaerobic to aerobic and from saline to freshwater conditions occurred. To complement the molecular methods, we additionally cultivated B. benzoevorans-related strains from all of the soil samples. The highest number of B. benzoevorans relatives was found in the soils from the northern part of The Netherlands. The present study contributes to our knowledge of the diversity and abundance of this interesting group of microbes in soils throughout the world.