Objectives: Molecular epidemiological studies of Mycobacterium tuberculosis in high prevalence areas in sub-Saharan Africa are hampered by the difficulty of culturing organisms from clinical samples. This study aimed to evaluate for application in a developing country, a modification of a novel polymerase chain reaction (PCR) based molecular epidemiological typing method, termed spoligotyping.
Methods: DNA extraction from sputum was followed by PCR amplification of spacers between direct repeats in the M. tuberculosis genome, and hybridization to a range of the 53 known spacer sequences.
Results: Sputum from 175 patients in the Ashanti region of Ghana were collected, 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 epidemiologically linked cases were shown to be unrelated on the basis of different spoligotype patterns, but epidemiological links were not found to explain clusters. Comparison of spoligotyping of DNA extracted from sputum with restriction fragment length polymorphism (RFLP) from mycobacterial culture in a subset of 25 patients, indicated that spoligotyping was less discriminatory than RFLP, Sixteen spoligotype patterns were shown to comprise 2 3 different RFLP patterns.
Conclusions: This study suggests that the PCR based technique of spoligotyping can be applied successfully to DNA extracted from sputum collected in the setting of a developing country, but that this is less discriminatory than RFLP. Spoligotyping is particularly useful when used to support conventional epidemiology since a proportion of false epidemiological associations can be identified.