IngentaConnect A low cost, home-made, reverse-line blot hybridisation assay for ...

Authors: Morcillo N.¹; Zumarraga M.²; Alito A.²; Dolmann A.³; Schouls L.⁴; Cataldi A.²; Kremer K.⁴; van Soolingen D.⁴

Source: The International Journal of Tuberculosis and Lung Disease, Volume 6, Number 11, November 2002 , pp. 959-965(7)

Publisher: International Union Against Tuberculosis and Lung Disease

Abstract:

SETTING: Cetrangolo Hospital, Vicente Lopez, Argentina, 1995–1999.

OBJECTIVE: To describe a home-made reverse-line blot hybridisation assay for the detection of rifampicin resistance-associated mutations in the rpoB gene of Mycobacterium tuberculosis, and to evaluate the usefulness of this rifampicin oligonucleotide, or ‘RIFO’ assay, to predict rifampicin resistance.

DESIGN: A total of 135 M. tuberculosis isolates from the Cetrangolo Hospital were tested using the RIFO assay, the proportion method and the Mycobacterial Growth Indicator Tube (MGIT 960). In addition, 120 drug-susceptible isolates from the Netherlands were included.

RESULTS: The results obtained with the proportion method and the MGIT 960 system were in full agreement. In the RIFO assay, 90 of the 97 rifampicin-resistant isolates were correctly identified (sensitivity 92.8%, positive predictive value 100%). All of the drug-susceptible isolates were correctly predicted by the RIFO assay.

CONCLUSIONS: With this home-made molecular test, rifampicin resistance in M. tuberculosis can be predicted in colonies isolated in culture in only 1 day, and can therefore shorten the laboratory turn around time for rifampicin susceptibility testing by weeks. In principle the test can also be applied directly to Ziehl-Neelsen slides and clinical material, as has been demonstrated for another reverse-line blot-based assay for M. tuberculosis, spoligotyping.

Keywords: tuberculosis; PCR; reverse-line blot hybridisation; rifampicin; resistance; MDR-TB

Language: English

Document Type: Regular paper

Affiliations: 1: Mycobacterial Regional Reference Laboratory, Tuberculosis Control Programme of Buenos Aires Province, Argentina 2: National Institute of Agricultural Technology (INTA/CICV), Moron, Argentina 3: Cetrangolo Hospital, Vicente Lopez, Argentina 4: National Institute of Public Health and the Environment (RIVM), Bilthoven, The Netherlands

A low cost, home-made, reverse-line blot hybridisation assay for rapid detection of rifampicin resistance in Mycobacterium tuberculosis