Differential identification of Bacillus anthracis from environmental Bacillus species using microarray analysis.
OBJECTIVE
To determine whether microarray analysis could be employed for the differential identification of a range of environmental Bacillus sp. from four strains of Bacillus anthracis.


RESULTS
Oligonucleotide probes were designed that were specific to virulence factor genes of B. anthracis (pag, lef and cap), the variable number tandem repeat region of the B. anthracis vrrA gene and to the 16S-23S rRNA intergenic transcribed spacer region (ITS) and pleiotropic regulator (plcR) regions of the Bacillus cereus subgroup species. Generic probes were also designed to hybridize with conserved regions of the 16S rRNA genes of Bacillus (as a positive control), Neisseria sp., Pseudomonas sp., Streptococcus sp., Mycobacterium sp. and to all members of the Enterobacteriaceae to allow simultaneous detection of these bacteria. Identification of B. anthracis was found to rely entirely on hybridization of DNA specific to regions of the pag, lef and cap genes. Cross-reaction was observed between B. anthracis and other Bacillus species with all the other Bacillus probes tested. Results obtained using microarray hybridizations were confirmed using conventional microbiological techniques and found to have very high comparability.


CONCLUSIONS
Microarray-based assays are an effective method for the identification of B. anthracis from mixed-culture environmental samples without problems of false-positivity that have been observed with conventional PCR assays.


CONCLUSIONS
Identification of environmental Bacillus sp. by conventional PCR is prone to potential for reporting false-positives. This study provides a method for the exclusion of such isolates.
Burton
J E
JE
Novel and Dangerous Pathogens Department, Health Protection Agency, Porton Down, Salisbury, Wiltshire, UK.
Oshota
O J
OJ
Silman
N J
NJ
D001407:Bacillus / Q000235:genetics; D001408:Bacillus anthracis / Q000235:genetics*; D015342:DNA Probes; D004269:DNA, Bacterial / Q000032:analysis*; D005798:Genes, Bacterial; D046228:Microarray Analysis / Q000379:methods*; D009693:Nucleic Acid Hybridization / Q000379:methods; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D016133:Polymerase Chain Reaction / Q000379:methods; D012336:RNA, Ribosomal, 16S / Q000032:analysis
D015342:DNA Probes; D004269:DNA, Bacterial; D012336:RNA, Ribosomal, 16S