Molecular pet or parasite? Exploring selection for vertical and horizontal plasmid transfer

Understanding the environmental conditions that drive selection for increased horizontal plasmid transfer is crucial for predicting the spread of plasmid-encoded antibiotic resistance. In natural systems, plasmids exhibit diverse lifestyles, ranging from "host-centric" strategies, which favor vertical gene transfer (VGT) from mother to daughter cell at the expense of horizontal mobility, to "parasitic" strategies, which favor horizontal gene transfer (HGT) by conjugation at the expense of host fitness. However, laboratory evolution experiments are biased towards host-centric evolution, highlighting a gap in our ability to consistently select for horizontal mobility. To understand this experimental bias, we developed a mathematical model to explore the invasion of pleiotropic transfer mutations. Using local linear stability analysis, we derived an invasion criterion establishing that, for a given pleiotropic cost, the availability of plasmid-free cells determines whether increased transfer is selected. We expanded this model to better represent our previous evolution experiment, in which selection for a host-centric mutant occurred despite the addition of plasmid-free cells and periodic selection for transconjugants. We found that standard batch culture protocols inherently impose strong selective pressure on VGT, heavily limiting the laboratory observation of increases in HGT. We experimentally and theoretically demonstrated that a simple protocol modification--minimizing excess growth by eliminating batch culture passages--effectively tips selection towards HGT. Finally, we performed a parameter sweep to predict the invasion success of hypothetical mutants across HGT-VGT phenotypic space. Our predictive framework can be used to further explore the evolution of plasmid transfer under conditions more representative of natural environments where medically and environmentally relevant plasmids evolve.