First Worldwide Proficiency Study on Variable-Number Tandem-Repeat Typing of Mycobacterium tuberculosis Complex Strains

Abstract

f Although variable-number tandem-repeat (VNTR) typing has gained recognition as the new standard for the DNA fingerprinting of Mycobacterium tuberculosis complex (MTBC) isolates, external quality control programs have not yet been developed. Therefore, we organized the first multicenter proficiency study on 24-locus VNTR typing. Sets of 30 DNAs of MTBC strains, including 10 duplicate DNA samples, were distributed among 37 participating laboratories in 30 different countries worldwide. Twenty-four laboratories used an in-house-adapted method with fragment sizing by gel electrophoresis or an automated DNA analyzer, nine laboratories used a commercially available kit, and four laboratories used other methods. The intra-and interlaboratory reproducibilities of VNTR typing varied from 0% to 100%, with averages of 72% and 60%, respectively. Twenty of the 37 laboratories failed to amplify particular VNTR loci; if these missing results were ignored, the number of laboratories with 100% interlaboratory reproducibility increased from 1 to 5. The average interlaboratory reproducibility of VNTR typing using a commercial kit was better (88%) than that of in-house-adapted methods using a DNA analyzer (70%) or gel electrophoresis (50%). Eleven laboratories using in-house-adapted manual typing or automated typing scored inter-and intralaboratory reproducibilities of 80% or higher, which suggests that these approaches can be used in a reliable way. In conclusion, this first multicenter study has documented the worldwide quality of VNTR typing of MTBC strains and highlights the importance of international quality control to improve genotyping in the future.

High-Throughput Mycobacterial Interspersed Repetitive-Unit–Variable-Number Tandem-Repeat Genotyping for Mycobacterium tuberculosis Epidemiological Studies

“…A few studies have investigated the intra-and interlaboratory reliability of MIRU-VNTR typing. In particular, they confirmed that manual methods can present discrepancies and allele-calling failures (9). These factors hinder the wider implementation of this method.…”

Mycobacterium tuberculosis: Evolution, Host–Pathogen Interactions, and Implications for Tuberculosis Control

“…The introduction of this genotyping method replaced IS6110-RFLP as the gold standard and allowed VNTR patterns to be compared in international databases. Because of the ease of use and the improved performance, VNTR typing has been integrated in TB control programs on an international scale in the United States, Europe, and other parts of the world [30,44]. A global epidemiological database is available that has led to insights into the geographical distribution and evolution of M. tuberculosis [26,44].…”

“…The determination of the tandem repeat sizes can be performed in an agarose gel-based electrophoresis method or on DNA sequencing machines. Moreover, the results are expressed as numerical codes and are therefore very easy to compare and exchange [44,52]. Among different sets of MIRU-VNTR loci described for typing M. tuberculosis isolates, a new international standard typing method system based on 24 loci was proposed in 2006, by a group of international institutions [17].…”

Comparative Study of IS 6110 Restriction Fragment Length Polymorphism and Variable-Number Tandem-Repeat Typing of Mycobacterium tuberculosis Isolates in the Netherlands, Based on a 5-Year Nationwide Survey

“…The first round of worldwide proficiency testing initiated by our laboratory within the framework of an ECDC project revealed that both the intralaboratory and interlaboratory reproducibilities are to be improved in a number of laboratories, indicating the need for careful implementation (20). Moreover, new patient cluster identification numbers and compositions resulting from the discordances with RFLP typing introduced a major challenge for the TB units of the Municipal Health Services.…”