This Week in TB R&D – 13 July 2010

12 Jul 2010
by Working Group

Genotype MTBDRplus assay kit

Last week we highlighted the Xpert® MTB/RIF assay for diagnostic testing. This week, we will briefly describe the GenoType® MTBDRplus assay. The GenoType® MTBDRplus assay uses PCR to amplify biotin-labeled amplicons (single-stranded DNA sequences) of M. tb from sputum. The amplicons are then incubated with strips of membrane-bound probes to allow for hybridization. After hybridization, the strips are washed and reagent containing streptavidin-alkaline phosphatase conjugate is added to the strip for an alkaline phosphatase staining reaction to occur for visualization of colorimetric bands. The membrane-bound probes are comprised of rifampin (RIF) rpoB wild-type and the most commonly observed rpoB mutations, as well as isoniazid (INH) katG and inhA wild-type and their most commonly observed respective mutations. There are also several amplification and hybridization probes included on the strips for assay control. Indication of a possible resistant sample is made when hybridization of a wild-type probe is absent or when a mutant probe is present.

In 2007, Hillemann, Rusch-Gerdes, and Richter published a paper which evaluated the GenoType® MTBDRplus assay for its sensitivity and specificity to detect RIF resistance or INH resistance in comparison to the old GenoType® MTBDR assay. They observed that the GenoType® MTBDRplus assay had similar sensitivity for RIF resistance, but increased sensitivity for INH resistance over the old MTBDR assay; whereas the specificity for both assays were ~ 98% and 100% for RIF resistance or INH resistance detection, respectively.

In 2008, the WHO endorsed the use of line-probe assays, such as the GenoType® MTBDRplus assay, for the rapid detection of MDR-TB. Since then several papers have attempted to evaluate its use in various geographical regions and settings.

Schematic diagram of the genes and possible banding patterns of the line-probe assay.

A paper by Huang and colleagues compared the assay to standard DNA testing on Taiwanese patient samples. Their results for the GenoType® MTBDRplus assay suggested a decreased overall performance in accuracy of RIF, INH, and MDR-TB isolates (95.4%, 81.8%, and 78.5%) compared to previously published data, however it was comparable to that of drug susceptibility testing (DST) using conventional culture-based systems. Although the authors highlight the value of having a rapid DST, they raised a concern about the low accuracy of INH resistance detection and note they ‘were unable to replace conventional DST.’

Results from two papers this year, one by Bwanga and colleagues and the other by Huyen and colleagues, which evaluated the GenoType® MTBDRplus assay in Uganda and South Vietnam, respectively, also noted the advantage of the quick determination of RIF or INH, resistance and MDR detection. Bwanga and colleagues evaluated the assay in the context of a resource-limited setting in comparison with six other recognized MDR detection assays. Although the GenoType® MTBDRplus assay demonstrated almost 100% specificity in RIF, INH, and MDR detection, the authors noted its lower INH sensitivity. Huyen and colleagues report support the 100% specificity for RIF, INH, and MDR detection observed with this assay, however sensitivity was lower for MDR detection (88.9%).

Collectively, all of the reports referenced here recommend using the GenoType® MTBDRplus assay for quick analysis. Furthermore, they suggest that this is assay is useful for the determination of RIF, INH, or MDR resistance to initiate early second-line drug treatment in association with conventional DST for confirmation of resistance and the type of resistance.

Should the MTBDRplus assay replace other conventional culture-based DST? What are the pros? Cons? Is the MTBDRplus assay a more useful tool in resource-limited countries compared to the Xpert MTB/RIF assay if both assay can provide MDR status in a day? Why or why not? Please share your thoughts below.

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