Genetic diversity of human isolates of Mycobacterium bovis assessed by spoligotyping and Variable Number Tandem Repeat genotyping

Abstract A collection of clinical isolates including 9 Mycobacterium bovis bacille Calmette–Guerin (BCG), 37 M. bovis and 1 isolate identified as M. bovis / caprae intermediate, recovered from humans in Tuscany, Italy, from 1990 to 2009, was genotyped by spoligotyping and Variable Number Tandem Repeat (VNTR) typing. Spoligotyping detected 15 unique profiles; the “BCG-like” SIT482/SB0120 spoligotype was largely prevalent accounting for 63.8% of isolates. VNTR typing, based on the 15 VNTR loci commonly tested for Mycobacterium tuberculosis , detected 29 unique profiles; only 8 VNTR loci (VNTR 43, MIRU 04, QUB-11b, ETR-A, VNTR 47, MIRU 31, QUB-26 and VNTR 53) provided a satisfactory allelic diversity in the VNTR analysis. Combined together, spoligotyping and VNTR typing yielded 33 unique patterns and 5 clusters including a total of 19 isolates. Clustered isolates, further typed for additional 9 VNTR loci, finally yielded 3 distinct clusters including 3 M. bovis BCG isolates each, and 1 cluster of 6 M. bovis isolates. Minimum spanning tree analysis showed that, in spite of the many distinct VNTR profiles, most M. bovis isolates displayed a high phylogenetic proximity, due to the variation of a single VNTR allele, thus indicating that the population of human M. bovis isolates in our setting is relatively homogeneous and conserved.