Comparative study of IS6110 restriction fragment length polymorphism and variable-number tandem-repeat typing of Mycobacterium tuberculosis isolates in the Netherlands, based on a 5-year nationwide survey

Abstract

In order to switch from IS6110 and polymorphic GC-rich repetitive sequence (PGRS) restriction fragment length polymorphism (RFLP) to 24-locus variable-number tandem-repeat (VNTR) typing of Mycobacterium tuberculosis complex isolates in the national tuberculosis control program in The Netherlands, a detailed evaluation on discriminatory power and agreement with findings in a cluster investigation was performed on 3,975 tuberculosis cases during the period of 2004 to 2008. The level of discrimination of the two typing methods did not differ substantially: RFLP typing yielded 2,733 distinct patterns compared to 2,607 in VNTR typing. The global concordance, defined as isolates labeled unique or identically distributed in clusters by both methods, amounted to 78.5% (n = 3,123). Of the remaining 855 cases, 12% (n = 479) of the cases were clustered only by VNTR, 7.7% (n = 305) only by RFLP typing, and 1.8% (n = 71) revealed different cluster compositions in the two approaches. A cluster investigation was performed for 87% (n = 1,462) of the cases clustered by RFLP. For the 740 cases with confirmed or presumed epidemiological links, 92% were concordant with VNTR typing. In contrast, only 64% of the 722 cases without an epidemiological link but clustered by RFLP typing were also clustered by VNTR typing. We conclude that VNTR typing has a discriminatory power equal to IS6110 RFLP typing but is in better agreement with findings in a cluster investigation performed on an RFLP-clustering-based cluster investigation. Both aspects make VNTR typing a suitable method for tuberculosis surveillance systems.

Fig 1

Distribution of cluster sizes obtained on the basis of RFLP (gray bars) and VNTR (black bars) typing.

Fig 2

Composition of RFLP clusters by VNTR typing. In total, 69% of the RFLP clusters showed a completely identical (n = 256) or similar (n = 47; with only one or more single cases split off by a distinct VNTR pattern) composition by VNTR typing. For 24% (n = 107) of the RFLP clusters, VNTR typing resulted in sets of only unique patterns, whereas 6% (n = 28) of the RFLP clusters were subdivided into 2 or 3 VNTR clusters.

Fig 3

(Top) RFLP-clustered isolates (n = 1,462) divided by cluster investigation results, classified as confirmed, presumed, and no epidemiological link. For each segment, the distribution of confirmed clustering by VNTR typing is shown. (Bottom) Percentages of confirmed or presumed epidemiological links for RFLP-clustered cases. Isolates with and without VNTR-supported clustering.

Fig 4

(Top) Number of loci that differ in VNTR typing patterns (n = 300) split off from their RFLP cluster. (Bottom) Distribution of detected differences by locus in 24-locus VNTR genotypes of samples clustered by RFLP typing and split off by VNTR typing, separated by single-locus variation (SLV) cases and multilocus variation cases (MLV).

Fig 5

Percentages of single-locus variants (SLV) and multilocus variants (MLV) for the isolates clustered by RFLP and split off by VNTR typing in relation to the epidemiological information.