Unraveling Epistatic Interactions Between Sites Under Drug-Dependent Selection in the <i>Mycobacterium tuberculosis</i> Genome

Many Mycobacterium tuberculosis genome sites experience different selective forces depending on whether a patient is treated with antibiotics. Here, we searched for pairs of such sites that evolve interdependently. We reconstructed the phylogeny of more than 11,000 Mycobacterium tuberculosis clinical isolates with known phenotypes for at least one of the 13 antitubercular drugs. By analyzing the distributions of substitutions and phenotypic state changes on the phylogeny, we identified sites where substitutions were associated with the acquisition of drug resistance or occurred preferentially in resistant or susceptible lineages. Among these sites, we searched for concordantly and discordantly evolving site pairs, carefully accounting for the presence of drug-associated selection and other coordinated selective forces. We identified one concordantly evolving site pair and 14 discordantly evolving site pairs between sites that are known to be strongly associated with resistance to antitubercular drugs. The concordantly evolving site pair and five out of 14 discordantly evolving site pairs were between sites whose substitutions were associated with resistance to different drugs, while the other nine discordantly evolving site pairs were between sites located either in the same genes or in different genes involved in alternative adaptive pathways to the same drugs. Overall, our findings emphasize the dual role of epistasis, which can both promote and limit the acquisition of resistance to multiple drugs.