Evolution of Drug Resistance in Mycobacterium tuberculosis

This thesis examines the contrasting roles of genetic drift and selection on the emergence of drug resistance in Mycobacterium tuberculosis. In clinical practice some alleles of rifampicin resistance are isolated more frequently than others. To identify if this variation is due to genetic drift or selection, the mutation rate to rifampicin resistance in M. tuberculosis (H37Rv) was determined. PCR-SSCP analysis revealed only three patterns from the rifampicin resistant isolates, each pattern arising at the same mutation rate (Mann-Whitney U test P>0.5). Fitness, defined as the ratio of generations of resistant and susceptible cells formed in mixed culture, of the differing rifampicin resistant alleles was determined relative to the parent fully susceptible strain. There was a significant correlation between fitness and the clinical isolation rate of each allele (regression analysis P=0.026). The fitness of two isolates, with identical IS6110 RFLP pattern isolated from 2 siblings, was determined. One isolate had developed multi drug resistance, the second isolate had remained fully drug susceptible. The fitness of the drug resistant isolate was significantly lower than the drug susceptible isolate (matched pair t test p=0.002). The decreased relative fitness of the resistant isolate implied a physiological cost for the development of drug resistance. Isolates of M. tuberculosis from three patients involved in a hospital outbreak of multi drug resistant tuberculosis were obtained. The fitness of these isolates was determined