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Journal of Clinical Microbiology, September 2001, p. 3272-3278, Vol. 39, No. 9
0095-1137/01/$04.00+0 DOI: 10.1128/JCM.39.9.3272-3278.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.
Evaluation of PCR-Restriction Profile Analysis and
IS2404 Restriction Fragment Length Polymorphism and
Amplified Fragment Length Polymorphism Fingerprinting for
Identification and Typing of Mycobacterium ulcerans and
M. marinum
K.
Chemlal,*,1
G.
Huys,2
P.-A.
Fonteyne,1
V.
Vincent,4
A. G.
Lopez,1
L.
Rigouts,1
J.
Swings,2,5
W. M.
Meyers,3 and
F.
Portaels1
Department of Microbiology, Mycobacteriology Unit,
Institute of Tropical Medicine, B-2000 Antwerp,1
and Laboratorium Voor Microbiologie2
and BCCM/LMG Culture Collection,5
Universiteit Gent, B-9000 Gent, Belgium; Armed Forces
Institute of Pathology, Washington, D.C. 203063;
and Laboratoire de Référence des
Mycobactéries, Institut Pasteur, 75724 Paris Cedex,
France4
Received 11 December 2000/Returned for modification 28 March
2001/Accepted 29 May 2001
Mycobacterium ulcerans and M. marinum are
emerging necrotizing mycobacterial pathogens that reside in common
reservoirs of infection and exhibit striking pathophysiological
similarities. Furthermore, the interspecific taxonomic relationship
between the two species is not clear as a result of the very high
phylogenetic relatedness (i.e., >99.8% 16S rRNA sequence similarity),
in contrast to only 25 to 47% DNA relatedness. To help understand the
genotypic affiliation between these two closely related species, we
performed a comparative analysis including PCR restriction profile
analysis (PRPA), IS2404 restriction fragment length
polymorphism (RFLP), and amplified fragment length polymorphism (AFLP)
on a set of M. ulcerans (n = 29) and
M. marinum (n = 28) strains recovered from
different geographic origins. PRPA was based on a triple restriction of
the 3' end region of 16S rRNA, which differentiated M. ulcerans into three types; however, the technique could not distinguish M. marinum from M. ulcerans
isolates originating from South America and Southeast Asia. RFLP based
on IS2404 produced six M. ulcerans types
related to six geographic regions and did not produce any band with
M. marinum, confirming the previous findings of Chemlal et
al. (K. Chemlal, K. DeRidder, P. A. Fonteyne, W. M. Meyers,
J. Swings, and F. Portaels, Am. J. Trop. Med. Hyg. 64:270-273,
2001). AFLP analysis resulted in profiles which grouped M. ulcerans and M. marinum into two separate clusters.
The numerical analysis also revealed subgroups among the M. marinum and M. ulcerans isolates. In conclusion, PRPA
appears to provide a rapid method for differentiating the African
M. ulcerans type from other geographical types but is
unsuitable for interspecific differentiation of M. marinum
and M. ulcerans. In comparison, whole- genome techniques such as IS 2404-RFLP and AFLP appear to be far more useful
in discriminating between M. marinum and M. ulcerans, and may thus be promising molecular tools for the
differential diagnosis of infections caused by these two species.
*
Corresponding author. Mailing address: Department of
Microbiology, Mycobacteriology Unit, Institute of Tropical Medicine, Nationalestraat 155, B-2000 Antwerp, Belgium. Phone: 32(3)247-63-36. Fax: 32(3)247-63-33. E-mail: kchemlal{at}itg.be.
Journal of Clinical Microbiology, September 2001, p. 3272-3278, Vol. 39, No. 9
0095-1137/01/$04.00+0 DOI: 10.1128/JCM.39.9.3272-3278.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.
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