Comparison of a variable-number tandem-repeat (VNTR) method for typing Mycobacterium avium with mycobacterial interspersed repetitive-unit-VNTR and IS1245 restriction fragment length polymorphism typing

Abstract

Mycobacterium avium complex (MAC) infections are increasing annually in various countries, including Japan, but the route of transmission and pathophysiology of the infection remain unclear. Currently, a variable-number tandem-repeat (VNTR) typing method using the Mycobacterium avium tandem repeat (MATR) loci (MATR-VNTR) is employed in Japan for epidemiological studies using clinical isolates of M. avium. In this study, the usefulness of this MATR-VNTR typing method was compared with that of the IS1245-restriction fragment length polymorphism (IS1245-RFLP) typing method and a mycobacterial interspersed repetitive-unit (MIRU)-VNTR typing method reported previously (V. C. Thibault, M. Grayon, M. L. Boschiroli, C. Hubbans, P. Overduin, K. Stevenson, M. C. Gutierrez, P. Supply, and F. Biet, J. Clin. Microbiol. 45:2404-2410, 2007). Seventy clinical isolates identified as M. avium from human immunodeficiency virus-negative patients with MAC infections were used. MATR-VNTR typing using 15 loci distinguished 56 patterns of different allele profiles, yielding a Hunter-Gaston discriminatory index (HGDI) of 0.990. However, IS1245-RFLP and MIRU-VNTR typing yielded HGDIs of 0.960 and 0.949, respectively, indicating that MATR-VNTR has an excellent discriminatory power compared with MIRU-VNTR and IS1245-RFLP typing. Moreover, concomitant use of the MATR-VNTR method and IS1245-RFLP typing increased the HGDI to 0.999. MATR-VNTR typing is inexpensive and easy to perform and could thus be useful in establishing a digital multifacility database that will greatly contribute to the clarification of the transmission route and pathophysiology of M. avium infections.

FIG. 1.

Dendrogram of IS1245-RFLP patterns of M. avium isolates, including 70 clinical strains and reference strains, as determined by the unweighted-pair group method using average linkages, using the different band calculation and a 1.0% position tolerance. The scale at the top shows the genetic similarity among the profiles, expressed as percentages. Faint bands were included in the comparison. MAA (M. avium subsp. avium), strain GTC00603; MAH (M. avium subsp. hominissuis), strain ATCC 19978; MAS (M. avium subsp. silvaticum), strain ATCC49884.

FIG. 2.

Dendrogram and allele profiles constructed from MATR-VNTR typing results for M. avium isolates, including 70 clinical strains and reference strains. The dendrogram was created from distance matrix files by Fitch-Margoliash analysis according to MATR-VNTR markers. Allele profiles analyzed using MATR-VNTR loci from isolates showing 100% similarity are boxed. The Manhattan distance is indicated at the bottom. MAA (M. avium subsp. avium), strain GTC00603; MAH (M. avium subsp. hominissuis), strain ATCC 19978; MAS, (M. avium subsp. silvaticum) strain ATCC 49884; MAP (M. avium subsp. paratuberculosis), strain ATCC 19698.

FIG. 3.

Dendrogram and allele profiles constructed from MIRU-VNTR typing results for M. avium isolates, including 70 clinical strains and reference strains. The dendrogram was created from distance matrix files by Fitch-Margoliash analysis according to MIRU-VNTR markers. Allele profiles analyzed using MIRU-VNTR loci from isolates showing 100% similarity are boxed. The Manhattan distance is indicated at the bottom. MAA (M. avium subsp. avium), strain GTC00603; MAH (M. avium subsp. hominissuis), strain ATCC 19978; MAS (M. avium subsp. silvaticum), strain ATCC 49884; MAP (M. avium subsp. paratuberculosis), strain ATCC 19698.