Abstract
BackgroundMolecular studies of Bacillus diversity in various environments have been reported. However, there have been few investigations concerning Bacillus in steel plant environments. In this study, genotypic and phenotypic diversity and phylogenetic relationships among 40 bacterial isolates recovered from steel plant waste were investigated using classical and molecular methods.Results16S rDNA partial sequencing assigned all the isolates to the Bacillus genus, with close genetic relatedness to the Bacillus subtilis and Bacillus cereus groups, and to the species Bacillus sphaericus. tDNA-intergenic spacer length polymorphisms and the 16S–23S intergenic transcribed spacer region failed to identify the isolates at the species level. Genomic diversity was investigated by molecular typing with rep (repetitive sequence) based PCR using the primer sets ERIC2 (enterobacterial repetitive intergenic consensus), (GTG)5, and BOXAIR. Genotypic fingerprinting of the isolates reflected high intraspecies and interspecies diversity. Clustering of the isolates using ERIC-PCR fingerprinting was similar to that obtained from the 16S rRNA gene phylogenetic tree, indicating the potential of the former technique as a simple and useful tool for examining relationships among unknown Bacillus spp. Physiological, biochemical and heavy metal susceptibility profiles also indicated considerable phenotypic diversity. Among the heavy metal compounds tested Zn, Pb and Cu were least toxic to the bacterial isolates, whereas Ag inhibited all isolates at 0.001 mM.ConclusionIsolates with identical 16S rRNA gene sequences had different genomic fingerprints and differed considerably in their physiological capabilities, so the high levels of phenotypic diversity found in this study are likely to have ecological relevance.
Highlights
Molecular studies of Bacillus diversity in various environments have been reported
Bacillus cereus and Bacillus anthracis, both of which belong to the B. cereus group [2], are amongst those associated with human disease
The type strains of B. subtilis (ATCC 6633T), B. licheniformis (ATCC 14580T), B. pumilus (ATCC 7061T), B. amyloliquefaciens (ATCC 23842), B. cereus (ATCC 11778) and B. sphaericus (ATCC 14577T) were included as reference strains
Summary
Molecular studies of Bacillus diversity in various environments have been reported. there have been few investigations concerning Bacillus in steel plant environments. Genotypic and phenotypic diversity and phylogenetic relationships among 40 bacterial isolates recovered from steel plant waste were investigated using classical and molecular methods. Using 16S rDNA sequence analysis, Ash et al [3] described the presence of five phylogenetically distinct groups in the genus Bacillus, and Nielsen et al [4] subsequently described a sixth group belonging to the alkaliphilic bacilli. Two of these groups are the B. cereus group (B. anthracis, B. cereus, Bacillus thuringiensis, Bacillus mycoides, Bacillus pseudomycoides and Bacillus weihenstephanensis) and the Bacillus subtilis group (B. subtilis, Bacillus pumilus, Bacillus atrophaeus, Bacillus licheniformis and Bacillus amyloliquefaciens) [3]. This method can be used to generate more accurate information because it is capable of screening several parts of the bacterial genome [11,12]
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