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Journal of Clinical Microbiology, April 2001, p. 1396-1401, Vol. 39, No. 4
0095-1137/01/$04.00+0   DOI: 10.1128/JCM.39.4.1396-1401.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

Detection of Phospholipase C in Nontuberculous Mycobacteria and Its Possible Role in Hemolytic Activity

Arley Gomez,1,2 Armand Mve-Obiang,1 Bernard Vray,3 Wieslawa Rudnicka,4 Isdore C. Shamputa,1 Françoise Portaels,1,* Wayne M. Meyers,5 Pierre-Alain Fonteyne,1 and Laurence Realini1

Mycobacteriology Unit, Institute of Tropical Medicine, B 2000 Antwerp,1 and Laboratoire d'Immunologie Expérimentale, Faculté de Medecine, Université Libre de Bruxelles, Brussels,3 Belgium; Faculty of Medicine, Universidad del Quindio, Quindio, Colombia2; Department of Infectious Biology, Institute of Microbiology and Immunology, University of Łódz 90-237 Łódz, Lodowa 106, Poland4; and Armed Forces Institute of Pathology, Washington, D.C. 20306-60005

Received 25 September 2000/Returned for modification 12 October 2000/Accepted 26 December 2000

Phospholipase C plays a key role in the pathogenesis of several bacterial infections, for example, those caused by Clostridium perfringens and Listeria monocytogenes. Previous studies have reported multiple copies of plc genes homologous to Pseudomonas aeruginosa plcH and plcN genes encoding the hemolytic and nonhemolytic phospholipase C enzymes in the genomes of Mycobacterium tuberculosis, M. marinum, M. bovis, and M. ulcerans. In this study we analyzed the possible relationship between phospholipase C and hemolytic activity in 21 strains of nontuberculous mycobacteria representing nine different species. Detection of phospholipase C enzymatic activity was carried out using thin-layer chromatography to detect diglycerides in the hydrolysates of radiolabeled phosphatidylcholine. DNA sequences of M. kansasii and M. marinum homologous to the genes encoding phospholipase C from M. tuberculosis and M. ulcerans were identified by DNA-DNA hybridization and sequencing. Finally, we developed a direct and simple assay to detect mycobacterial hemolytic activity. This assay is based on a modified blood agar medium that allows the growth and expression of hemolysis of slow-growing mycobacteria. Hemolytic activity was detected in M. avium, M. intracellulare, M. ulcerans, M. marinum, M. tuberculosis, and M. kansasii mycobacteria with phospholipase C activity, but not in M. fortuitum. No hemolytic activity was detected in M. smegmatis, M. gordonae, and M. vaccae. Whether or not phospholipase C enzyme plays a role in the pathogenesis of nontuberculous mycobacterial diseases needs further investigation.

* Corresponding author. Mailing address: Mycobacteriology Unit, Institute of Tropical Medicine, Nationalestraat 155, 2000 Antwerp, Belgium. Phone: (323) 2476317. Fax: (323) 2476333. E-mail: portaels{at}itg.be.

Journal of Clinical Microbiology, April 2001, p. 1396-1401, Vol. 39, No. 4
0095-1137/01/$04.00+0   DOI: 10.1128/JCM.39.4.1396-1401.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.


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