Use of Denaturing High-Performance Liquid Chromatography To Identify Bacillus anthracis by Analysis of the 16S-23S rRNA Interspacer Region and gyrA Gene

Abstract

Denaturing high-performance liquid chromatography (DHPLC) was evaluated as a method for identifying Bacillus anthracis by analyzing two chromosomal targets, the 16S-23S intergenic spacer region (ISR) and the gyrA gene. The 16S-23S ISR was analyzed by this method with 42 strains of B. anthracis, 36 strains of Bacillus cereus, and 12 strains of Bacillus thuringiensis; the gyrA gene was analyzed by this method with 33 strains of B. anthracis, 27 strains of B. cereus, and 9 strains of B. thuringiensis. Two blind panels of 45 samples each were analyzed to evaluate the potential diagnostic capability of this method. Our results show that DHPLC is an efficient method for the identification of B. anthracis.

16S rDNA PCR and denaturing gradient gel electrophoresis; a single generic test for detecting and differentiating Mycoplasma species

“…Recently, denaturing HPLC analysis has been used to detect and type bacterial pathogens (Domann et al, 2003;Hurtle et al, 2003) and could theoretically be used as an alternative to DGGE to target single nucleotide polymorphisms in the V3 region of 16S rDNA of Mycoplasma species. However, denaturing-HPLC would require expensive, specialized equipment and more laborious standardization and interpretation compared with DGGE.…”

Detection of Yersinia enterocolitica in milk powders by cross-priming amplification combined with immunoblotting analysis

“…This unique region is transcribed together with ribosomal RNA genes and termed as internal transcribed spacer (ITS). This genomic region shows a significant variation among different species of bacteria, both in sequence and length (Hurtle et al, 2003). Bacteria can harbor multiple alleles of ribosomal operon in their genome, resulting in a significant sequence variation in the spacer region among different species and even strains (Kabadjova et al, 2002;Rachman et al, 2004;Ranka et al, 2004).…”

Rapid and simple detection of bla CTX−M genes by multiplex PCR assay

“…DHPLC can be used to separate PCR products by DNA sequence and size, enabling bacterial identification and molecular characterization of antibiotic-resistance genes (Cooksey et al, 2002;Eaves et al, 2002;Hurtle et al, 2003). Accurate sizing of PCR products has been used for Mycobacterium tuberculosis typing and PCR product analysis to differentiate AmpC â-lactamases (Evans et al, 2004;PerezPerez & Hanson, 2002;Tzouvelekis et al, 2000).…”