to investigate the miru polymorphism , whole genomic dna was prepared from bacterial cultures or clinical specimens . the specimens were tissue fragments from patients with nonulcerated ( plaques and edematous forms ) or ulcerated forms . dna extraction from pure cultures was performed by heating the colonies in tris - edta at 95c for 10 minutes . clinical specimens from laboratory - confirmed cases of bu were decontaminated by using the reversed petroff method , and mycobacterial dna was extracted from the decontaminated solution as previously described ( 6 ) . the agilent 2100 bioanalyzer system ( agilent technologies , waldbronn , germany ) was used to separate 1 l of pcr product electrophoretically . m. ulcerans isolates from drc and uganda and tissue extracts from patients from sudan ( nzara ) and uganda ( nakasongola ) showed distinct profiles ( central africa : 1222 and east africa : 4111 ) , different from the originally homogeneous african genotype ( atlantic africa : 3113 ; table 1 ) . in drc , 3 different genotypes exist , corresponding to 3 different provinces : bas - congo , maniema ( kasongo ) , and orientale ( bunia ) . isolates from gabon , congo - brazzaville , and cameroon had the typical african genotype , now designated the atlantic african genotype . identical miru - vntr profiles were observed by using dna extracted from tissues or cultures from patients residing in the same area . the specificity of the miru - vntr method was tested on 14 different mycobacterium spp . only m. marinum , m. shottsii , and m. liflandii tested positive , but they were distinguished from m. ulcerans by exhibiting different profiles ( data not shown ) . sequencing of the concerned loci showed the conserved miru sequence at locus 1 and 9 in m. ulcerans . locus 6 ( 3 ) and locus 33 contain respectively a 56-bp and a 58-bp tandem repeat ( table 2 ) . although m. ulcerans isolates from africa are relatively homogeneous , this study demonstrates more heterogeneity between strains than previously reported . all isolates from west africa ( cte d'ivoire , ghana , togo , benin ) and central africa ( cameroon ; gabon ; congo - brazzaville ; drc bas - congo ; angola ) have the identical miru - vntr profile , and all originate from regions ( i.e. , bas - congo ) or countries that border the atlantic ocean . the isolates that come from regions or countries in the nile river basin ( i.e. , orientale in drc , sudan , and uganda ) or the congo river basin ( i.e. , maniema ) have distinct profiles . these results demonstrate for the first time heterogeneity among m. ulcerans from different african countries . the 3 african profiles are the atlantic african profile , the central african congo river basin profile , and the east african nile river basin profile . this is also the first detection of mirus and vntrs in clinical specimens , even in smear - negative specimens . further detailed differentiation of this etiologic agent will lead to an understanding of the epidemiology of bu . as in tuberculosis , better discriminatory typing methods help assess the efficacy of antimycobacterial treatment of bu patients by differentiating reactivation from reinfection . although m. ulcerans appears to be quite monomorphic , full sequencing of this organism will permit detection of genes specific for m. ulcerans , and more discriminatory vntr should become available .
mycobacterium ulcerans causes buruli ulcer , an ulcerative skin disease in tropical and subtropical areas . despite restricted genetic diversity , mycobacterial interspersed repetitive unit 
 variable - number tandem repeat analysis on m. ulcerans revealed 3 genotypes from different african countries . 
 it is the first time this typing method succeeded directly on patient samples .