Genetic diversity of Mycobacterium tuberculosis in Sicily based on spoligotyping and variable number of tandem DNA repeats and comparison with a spoligotyping database for population-based analysis

In a previous study, we proposed to associate spoligotyping and typing with the variable number of tandem DNA repeats (VNTR) as an alternative strateg y to IS6110-restriction fragment length polymorphism (RFLP) for molecular e pidemiological studies on tuberculosis, The aim of the present study was to further evaluate this PCR-based typing strategy and to describe the popula tion structure of Mycobacterium tuberculosis in another insular setting, Si cily, A collection of 106 DNA samples from M. tuberculosis patient isolates was characterized by spoligotyping and VNTR typing. All isolates were inde pendently genotyped by the standard IS6110-RFLP method, and clustering resu lts between the three methods were compared. The totals for the clustered i solates were, respectively, 15, 60, and 82% by 1S6110-RFLP, spoligotyping, and VNTR typing. The most frequent spoligotype included type 42 that missed spacers 21 to 24 and spacers 33 to 36 and derived types 33, 213, and 273 t hat, together represented as much as 26% of all isolates, whereas the Haarl em clade of strains (types 47 and 50, VNTR allele 32333) accounted for 9% o f the total strains. The combination of spoligotyping and VNTR typing resul ts reduced the number of clusters to 43% but remained superior to the level of IS6110-RFLP clustering (ca. 15%). All but one IS6110-defined cluster we re identified by the combination of spoligotyping and VNTR clustering resul ts, whereas 9 of 15 spoligotyping-defined clusters could be further subdivi ded by TS6110-RFLP. Reinterpretation of previous IS6110-RFLP results in the light of spoligotyping-VNTR typing results allowed us to detect an additio nal cluster that was previously missed, Although less discriminative than I S6110-RFLP, our results suggest that the use of the combination of spoligot yping and VNTR typing is a good screening strategy fur detecting epidemiolo gical links for the study tuberculosis epidemiology at tile molecular level .