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Journal of Clinical Microbiology, July 2009, p. 2026-2032, Vol. 47, No. 7
0095-1137/09/$08.00+0     doi:10.1128/JCM.02308-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Prospective Universal Application of Mycobacterial Interspersed Repetitive-Unit-Variable-Number Tandem-Repeat Genotyping To Characterize Mycobacterium tuberculosis Isolates for Fast Identification of Clustered and Orphan Cases{triangledown}

Noelia Alonso-Rodriguez,1,{dagger} Miguel Martínez-Lirola,3,{dagger} M. Luisa Sánchez,6 Marta Herranz,1,2 Teresa Peñafiel,6 Magdalena del Carmen Bonillo,5 Milagros Gonzalez-Rivera,7 Juan Martínez,8 Teresa Cabezas,4 Luis Felipe Diez-García,3 Emilio Bouza,1,2 and Darío García de Viedma1,2*

Servicio de Microbiología, Hospital General Universitario Gregorio Marañón, Madrid, Spain,1 CIBER Enfermedades Respiratorias (CIBERES), Spain,2 Complejo Hospitalario Torrecárdenas, Almería, Spain,3 Empresa Publica Hospital de Poniente, El Ejido, Almería, Spain,4 Distrito Atención Primaría Levante-Alto Almanzora, Huercal-Overa, Almería, Spain,5 Unidad de Tuberculosis de Poniente, El Ejido, Almería, Spain,6 LI Secuenciación, Hospital General Universitario Gregorio Marañón, Madrid, Spain,7 Centro Penitenciario El Acebuche, Almería, Spain8

Received 2 December 2008/ Returned for modification 9 February 2009/ Accepted 8 May 2009

The use of molecular tools for genotyping Mycobacterium tuberculosis isolates in epidemiological surveys in order to identify clustered and orphan strains requires faster response times than those offered by the reference method, IS6110 restriction fragment length polymorphism (RFLP) genotyping. A method based on PCR, the mycobacterial interspersed repetitive-unit-variable-number tandem-repeat (MIRU-VNTR) genotyping technique, is an option for fast fingerprinting of M. tuberculosis, although precise evaluations of correlation between MIRU-VNTR and RFLP findings in population-based studies in different contexts are required before the methods are switched. In this study, we evaluated MIRU-VNTR genotyping (with a set of 15 loci [MIRU-15]) in parallel to RFLP genotyping in a 39-month universal population-based study in a challenging setting with a high proportion of immigrants. For 81.9% (281/343) of the M. tuberculosis isolates, both RFLP and MIRU-VNTR types were obtained. The percentages of clustered cases were 39.9% (112/281) and 43.1% (121/281) for RFLP and MIRU-15 analyses, and the numbers of clusters identified were 42 and 45, respectively. For 85.4% of the cases, the RFLP and MIRU-15 results were concordant, identifying the same cases as clustered and orphan (kappa, 0.7). However, for the remaining 14.6% of the cases, discrepancies were observed: 16 of the cases clustered by RFLP analysis were identified as orphan by MIRU-15 analysis, and 25 cases identified as orphan by RFLP analysis were clustered by MIRU-15 analysis. When discrepant cases showing subtle genotypic differences were tolerated, the discrepancies fell from 14.6% to 8.6%. Epidemiological links were found for 83.8% of the cases clustered by both RFLP and MIRU-15 analyses, whereas for the cases clustered by RFLP or MIRU-VNTR analysis alone, links were identified for only 30.8% or 38.9% of the cases, respectively. The latter group of cases mainly comprised isolates that could also have been clustered, if subtle genotypic differences had been tolerated. MIRU-15 genotyping seems to be a good alternative to RFLP genotyping for real-time interventional schemes. The correlation between MIRU-15 and IS6110 RFLP findings was reasonable, although some uncertainties as to the assignation of clusters by MIRU-15 analysis were identified.

* Corresponding author. Mailing address: Servicio de Microbiología y Enfermedades Infecciosas, Hospital Gregorio Marañón, C/ Dr. Esquerdo 46, 28007 Madrid, Spain. Phone: 91 4265104. Fax: 91 5044906. E-mail: dgviedma2{at}gmail.com

{triangledown} Published ahead of print on 20 May 2009.

{dagger} Both authors contributed equally to the study.

Journal of Clinical Microbiology, July 2009, p. 2026-2032, Vol. 47, No. 7
0095-1137/09/$08.00+0     doi:10.1128/JCM.02308-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.





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