Predicting evolutionary paths to antibiotic resistance is key for understanding and controlling drug resistance. When considering a single final resistant genotype, epistatic contingencies among mutations restrict evolution to a small number of adaptive paths. Less attention has been given to multi-peak landscapes, and while specific peaks can be favoured, it is unknown whether and how early a commitment to final fate is made. Here we characterize a multi-peaked adaptive landscape for trimethoprim resistance by constructing all combinatorial alleles of seven resistance-conferring mutations in dihydrofolate reductase. We observe that epistatic interactions increase rather than decrease the accessibility of each peak; while they restrict the number of direct paths, they generate more indirect paths, where mutations are adaptively gained and later adaptively lost or changed. This enhanced accessibility allows evolution to proceed through many adaptive steps while delaying commitment to genotypic fate, hindering our ability to predict or control evolutionary outcomes.
We are grateful to N.D. Lord for gift of strain NDL47, and to D.M. Weinreich, D.L. Hartl and D. Landgraf for helpful discussions. This work was supported by the Novartis Institutes for Biomedical Research, US National Institutes of Health grant R01-GM081617 and the European Research Council FP7 ERC Grant 281891. A.C.P. is a James S. McDonnell Foundation Postdoctoral Fellow.
Table of growth measurements for every strain at every trimethoprim concentration, where relative growth is quantified by the integral of Optical Density from 0 to 30 hours. Each row specifies a single replicate set of growth measurements over a range of trimethoprim concentrations. In each row the strain genotype is listed in full (for example, 'A26' indicates wildtype allele at position 26, 'A26T' indicates a mutation at this position). Rows are grouped by threes, for the three replicate measurements, excepting wildtype which was measured in twelve replicates and is described in twelve rows. In cases where growth is 0 (strain unviable) or never exceeds the IC75 growth threshold, Log10(IC75) is described by the value of '-2'.