A bacterial tragedy of the commons that masks the actual frequency of mutants

The frequency of mutants in a population is central to the understanding of evolution. Mutant frequency is usually assessed by plating a bacterial culture on selective medium in which only specific rare mutants can grow, assuming that all mutant cells present on the plate are able to form colonies. Here we show an exception to this rule. Wild-type Escherichia coli cells are unable to grow with glycerol-2-phosphate (G2P) as a carbon source. In contrast, PHO-constitutive mutants can hydrolyse G2P to glycerol and form colonies on plates having G2P as their sole carbon source. However, the frequency of PHO-constitutive colonies on the selective plate is exceptionally low. Here we show that such mutations occur at a relatively high rate, but the growth of the existing mutants is inhibited due to a competition with the surrounding wild-type cells for the limited amounts of glycerol produced by the mutants. This scenario in which neither the wild-type nor the majority of the mutants are able to grow constitutes an unavoidable case of the tragedy of the commons. Evidence shows that the few mutants that do form colonies derive from micro-clusters of mutants on the selective plate. In addition, a mathematical model describes the fate of the wild-type and mutant populations on the selective plate.