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Identification of a novel repetitive DNA element and its use as a molecular marker for strain typing and discrimination of ara^- from ara⁺ Burkholderia pseudomallei isolates

Biomedical Sciences Laboratory, Defence Medical Research Institute and *Department of Microbiology, National University of Singapore, Singapore

Corresponding author: Dr M-A. Lee (e-mail: nmiv13{at}nus.edu.sg).

Received 28 Jan. 2001; revised version received 2 July 2001; accepted 6 July 2001.

Abstract

A novel 10-bp repeat (5'-CGACGCAGGC-3')₃₄ was identified in a strain of Burkholderia pseudomallei, the first repetitive element found in this species. A pair of primers, based on the flanking sequences of the repetitive region, was used in PCR and DNA sequence analysis to determine the genomic structure and distribution of the repetitive element in 76 arabinose^- (ara^-) and 7 ara⁺ B. pseudomallei isolates. DNA fragments of 400–700 bp were amplified in all ara^- isolates. Ara⁺ isolates were characterised by a uniform fragment of 402 bp. Nucleotide sequence analysis of these fragments revealed broad heterogeneity of the variable-number tandem repeats with 26 distinct alleles ranging between (5'-CGACGCAGGC-3')₁₃ and (5'-CGACGCAGGC-3')₄₅ identified in the ara^- isolates. In contrast, a novel non-repetitive sequence was identified in each of the ara⁺ isolates. This was confirmed by Southern blot analysis. Such biotype-specific variable-number tandem repeats may be useful as genetic markers for rapid strain differentiation of ara^- isolates.

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