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Journal of Clinical Microbiology, August 2002, p. 3021-3024, Vol. 40, No. 8
0095-1137/02/$04.00+0     DOI: 10.1128/JCM.40.8.3021-3024.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Simple and Rapid Differentiation of Mycobacterium tuberculosis H37Ra from M. tuberculosis Clinical Isolates through Two Cytochemical Tests Using Neutral Red and Nile Blue Stains

Carlos Y. Soto,¹ Núria Andreu,² Isidre Gibert,^1,2 and Marina Luquin^1,,2*

Departamento de Genética y de Microbiología, Facultad de Ciencias,¹ Instituto de Biotecnología y Biomedicina, Universidad Autónoma de Barcelona, 08193 Bellaterra (Barcelona), Spain²

Received 20 November 2001/ Returned for modification 21 January 2002/ Accepted 5 May 2002

The attenuated Mycobacterium tuberculosis strain H37Ra is one of the most commonly used controls for M. tuberculosis identification in the clinical laboratory and is a source of false-positive results for M. tuberculosis as a consequence of cross-contamination. Therefore, the ability to discriminate between H37Ra and real clinical isolates has important public health implications. To date, differentiation of H37Ra from M. tuberculosis clinical isolates is possible only by IS6110 genotyping and spoligotyping. In the 1950s, some authors reported that the virulent strain H37Rv and M. tuberculosis clinical isolates were able to fix basic dyes in their anionic forms, while H37Ra was not. We have studied the different techniques described for M. tuberculosis cytochemical staining and have chosen the best of these, introducing certain modifications in order to increase their discriminative power and reproducibility. We describe cytochemical staining of M. tuberculosis cells with neutral red and Nile blue, which differentiates H37Ra from virulent strains. This method could be used as an easy laboratory tool for distinguishing between H37Ra and real M. tuberculosis clinical isolates.

Present address Departamento de Microbiología, Medicina Preventiva y Salud Pública, Universidad de Zaragoza, 50009 Zaragoza, Spain.

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