The molecular clock of Mycobacterium tuberculosis

The molecular clock and its phylogenetic applications to genomic data have changed how we study and understand one of the major human pathogens, Mycobacterium tuberculosis (MTB), the etiologic agent of tuberculosis. Genome sequences of MTB strains sampled at different times are increasingly used to infer when a particular outbreak begun, when a drug-resistant clone appeared and expanded, or when a strain was introduced into a specific region. Despite the growing importance of the molecular clock in tuberculosis research, there is a lack of consensus as to whether MTB displays a clocklike behavior and about its rate of evolution. Here we performed a systematic study of the molecular clock of MTB on a large genomic data set (6,285 strains), covering different epidemiological settings and most of the known global diversity. We found that sampling times below 15-20 years were often insufficient to calibrate the clock of MTB. For data sets where such calibration was possible, we obtained a clock rate between 1x10(-8) and 5x10(-7) nucleotide changes per-site-per-year (0.04-2.2 SNPs per-genome-per-year), with substantial differences between clades. These estimates were not strongly dependent on the time of the calibration points as they changed only marginally when we used epidemiological isolates (sampled in the last 40 years) or three ancient DNA samples (about 1,000 years old) to calibrate the tree. Additionally, the uncertainty and the discrepancies in the results of different methods were sometimes large, highlighting the importance of using different methods, and of considering carefully their assumptions and limitations. Author summary One of the major recent advancements in evolutionary biology is the development of statistical methods to infer the past evolutionary history of species and populations with genomic data. In the last five years, many researchers have used the molecular clock (i.e. the observation that genomes accumulate mutations at an approximately constant pace) to study the evolution and epidemiology of Mycobacterium tuberculosis, a bacterial pathogen that causes tuberculosis and is responsible for 1.6 million human deaths ever year. Applications of the molecular clock are used to understand when tuberculosis emerged as a pathogen, the evolution of drug resistance, how different strains transmit and spread across the world and how MTB populations are affected by control programs. Here, we performed a systematic analysis of the molecular clock of MTB, analyzing several whole genome sequence data sets with the same set of methodologies. We characterized the rate of molecular evolution (the pace of the clock), and its variation between different MTB populations and lineages. Our results provide an important guideline for future analyses of tuberculosis and other organisms.