Masala, Speranza and Molicotti, Paola and Bua, Alessandra and Zumbo, Antonella and Delogu, Giovanni and Sechi, Leonardo Antonio and Zanetti, Stefania Anna Lucia (2010) Molecular characterization of Sardinian Mycobacterium tuberculosis isolates by IS6110 restriction fragment length polymorphism, MIRU-VNTR and rep-PCR. New Microbiologica, Vol. 33 (2), p. 155-162. ISSN 1121-7138. Article.
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An evaluation of the utility of rep PCR typing compared to the 15 loci discriminatory set of MIRU-VNTR was undertaken. Twenty-nine isolates of Mycobacterium tuberculosis from patients were examined. Genomic DNA was extracted from the isolates by standard method. The number of copies of tandem repeats of the 15 MIRU-VNTR loci was determined by PCR amplification and agarose gel electrophoresis of the amplicons. M. tuberculosis outbreak-related strains were distinguished from other isolates. MIRU-VNTR typing identified 4 major clusters of strains. The same isolates clustered together after RFLP typing, but rep-PCR identified only 3 of them. The concordance between RFLP and MIRU-VNTR typing was complete, with the exception of two isolates with identical RFLP patterns that differed in the number of tandem repeat copies at two MIRU-VNTR alleles. A further isolate, even sharing the same RFLP pattern, differed by one repeat from the rest of its cluster. We also tested the use of an automated rep-PCR for clinical laboratory applications but it failed to identify the link between two pairs of epidemiologically related strains clustered by the other 2 techniques. For superior discrimination, ease of comparison of results and lower cost, MIRU-VNTR typing should be the favored PCR-based typing tool.
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