Use of the genotype MTBDR assay for rapid detection of rifampin and isoniazid resistance in Mycobacterium tuberculosis complex isolates

Abstract

A commercially available DNA strip assay (Genotype MTBDR; Hain Lifescience, Nehren, Germany) was evaluated for its ability to detect mutations conferring resistance to rifampin (RMP) and isoniazid (INH) in clinical Mycobacterium tuberculosis complex isolates. A total of 103 multidrug-resistant (MDR; i.e., at least resistant to RMP and INH) and 40 fully susceptible strains isolated in Germany in 2001 in which resistance mutations have been previously defined by DNA sequencing and real-time PCR analysis were investigated. The Genotype MTBDR assay identified 102 of the 103 MDR strains with mutations in the rpoB gene (99%) and 91 strains (88.4%) with mutations in codon 315 of katG. All 40 susceptible strains showed a wild-type MTBDR hybridization pattern. The concordance between the MTBDR assay and the DNA sequencing results was 100%. Compared to conventional drug susceptibility testing, the sensitivity and specificity were 99 and 100% for RMP resistance and 88.4 and 100% for INH resistance, respectively. In conclusion, the MTBDR assay is a rapid and easy-to-perform test for the detection of the most common mutations found in MDR M. tuberculosis strains that can readily be included in a routine laboratory work flow.

FIG. 1.

Locations of probes within the 81-bp hot spot cluster of the rpoB gene.

FIG. 2.

Representative results for patterns obtained with the Genotype MTBDR assay. The positions of the oligonucleotides and the marker line are given on the left side. The specificity and targeted genes of the lines are shown from top to bottom as follows: conjugate control; amplification control (23S rRNA); M. tuberculosis complex-specific control (23S rRNA); control of rpoB amplification 5 to 9; rpoB wild-type probes located in the 81-bp hot spot region (the localization is shown in Fig. 1); rpoB mutant (Mut) probes with mutations in codons 516, 526, and 531; control of katG amplification; katG codon 315 wild-type probe; and katG codon 315 mutation probes (sequences in parenthesis). Along the bottom is a band for orientation of the strip. Patterns of the strips for MDR and susceptible M. tuberculosis strains are shown on the right. Lane 1, negative control; lane 2, H37Rv (wild type); lane 3, rpoB D516V, katG S315T1; lane 4, rpoB H526Y, katG S315T1; lane 5, rpoB H526D, katG S315T1; lane 6, rpoB S531L, katG S315T1; lane 7, rpoB H526N, katG S315T1; lane 8, rpoB 514-to-516 deletion, katG S315T1.