Identification of melioidosis outbreak by multilocus variable number tandem repeat analysis

Abstract

Endemic melioidosis is caused by genetically diverse Burkholderia pseudomallei strains. However, clonal outbreaks (multiple cases caused by 1 strain) have occurred, such as from contaminated potable water. B. pseudomallei is designated a group B bioterrorism agent, which necessitates rapidly recognizing point-source outbreaks. Pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST) can identify genetically related isolates, but results take several days to obtain. We developed a simplified 4-locus multilocus variable number tandem repeat analysis (MLVA-4) for rapid typing and compared results with PFGE and MLST for a large number of well-characterized B. pseudomallei isolates. MLVA-4 compared favorably with MLST and PFGE for the same isolates; it discriminated between 65 multilocus sequence types and showed relatedness between epidemiologically linked isolates from outbreak clusters and between isolates from individual patients. MLVA-4 can establish or refute that a clonal outbreak of melioidosis has occurred within 8 hours of receipt of bacterial strains.

Figure 1

Comparison of multilocus sequence typing (MLST) and 4-locus multilocus variable number tandem repeat analysis (MLVA-4) dendrograms for 65 Burkholderia pseudomallei isolates. MLST sequence type (ST) is shown for each isolate, with the corresponding isolate number listed for the MLVA-4 profile and shown by the colored lines. The red asterisks indicate 6 isolates that represent diversity of MLVA-4; these isolates were used to calibrate the dendrogram in Figure 3.

Figure 2

Comparison of pulsed-field gel electrophoresis (PFGE) and 4-locus multilocus variable number tandem repeat analysis (MLVA-4) profiles for isolates in 4 clonal groups (see text for details). Isolate number with its MLST sequence type (ST) is listed for each isolate on the PFGE profile, with the corresponding isolate number listed for the MLVA-4 profile. Four unrelated isolates are included for comparison: 0875 (ST115), 1869 (ST337), 1839 (ST336), and 1153 (ST117).

Figure 3

Dendrogram showing 4-locus multilocus variable number tandem repeat analysis profiles for isolates from 3 patients with melioidosis, with isolate number and multilocus sequence typing sequence type (ST) listed (see text for details). Six isolates used to calibrate the dendrogram are indicated by asterisks in Figure 1 and listed in Table 2. CSF, cerebrospinal fluid.