Spoligotyping in molecular epidemiology of tuberculosis in Ghana

Objectives: molecular epidemiological studies of Mycobacterium tuberculosis in high prevalence areas in sub-Saharan Africa are hampered by the difficu lty of culturing organisms from clinical samples. This study aimed to evalu ate for application in a developing country, a modification of a never poly merase chain reaction (PCR) based molecular epidemiological typing method, termed spoligotyping. Methods: DIVA extraction from sputum was followed by PCR amplification of s pacers between direct repeats in the M. tuberculosis genome, and hybridizat ion to a range of the 53 known spacer sequences. Results: sputum from 175 patients in the Ashanti region of Ghana were colle cted, and satisfactory spoligotyping results were obtained in 159. A total of 100 different spoligotype patterns were observed with 84 patients having unique patterns and the remainder falling into 16 clusters. A number of ep idemiologically linked cases were shown to be unrelated on the basis of dif ferent spoligotype patterns, but epidemiological links were not found to ex plain clusters, Comparison of spoligotyping of DNA extracted from sputum wi th restriction fragment length polymorphism (RFLP) from mycobacterial cultu re in a subset of 25 patients, indicated that spoligotyping was less discri minatory than RFLP, Sixteen spoligotype patterns were shown to comprise 23 different RFLP patterns, Conclusions: this study suggests that the PCR based technique of spoligotyp ing can be applied successfully to DNA extracted from sputum collected in t he setting of a developing country but that this is less discriminatory tha n RFLP, Spoligotyping is particularly useful when used to support conventio nal epidemiology since a proportion of false epidemiological associations c an be identified.