Mutators can drive the evolution of multi-resistance to antibiotics.

Antibiotic combination therapies are an approach used to counter the evolution of resistance; their purported benefit is they can stop the successive emergence of independent resistance mutations in the same genome. Here, we show that bacterial populations with 'mutators', organisms with defects in DNA repair, readily evolve resistance to combination antibiotic treatment when there is a delay in reaching inhibitory concentrations of antibiotic-under conditions where purely wild-type populations cannot. In populations of Escherichia coli subjected to combination treatment, we detected a diverse array of acquired mutations, including multiple alleles in the canonical targets of resistance for the two drugs, as well as mutations in multi-drug efflux pumps and genes involved in DNA replication and repair. Unexpectedly, mutators not only allowed multi-resistance to evolve under combination treatment where it was favoured, but also under single-drug treatments. Using simulations, we show that the increase in mutation rate of the two canonical resistance targets is sufficient to permit multi-resistance evolution in both single-drug and combination treatments. Under both conditions, the mutator allele swept to fixation through hitch-hiking with single-drug resistance, enabling subsequent resistance mutations to emerge. Ultimately, our results suggest that mutators may hinder the utility of combination therapy when mutators are present. Additionally, by raising the rates of genetic mutation, selection for multi-resistance may have the unwanted side-effect of increasing the potential to evolve resistance to future antibiotic treatments.

Decolonizing drug-resistant E. coli with phage and probiotics: breaking the frequency-dependent dominance of residents.

Widespread antibiotic resistance in commensal bacteria creates a persistent challenge for human health. Resident drug-resistant microbes can prevent clinical interventions, colonize wounds post-surgery, pass resistance traits to pathogens or move to more harmful niches following routine interventions such as catheterization. Accelerating the removal of resistant bacteria or actively decolonizing particular lineages from hosts could therefore have a number of long-term benefits. However, removing resident bacteria via competition with probiotics, for example, poses a number of ecological challenges. Resident microbes are likely to have physiological and numerical advantages and competition based on bacteriocins or other secreted antagonists is expected to give advantages to the dominant partner, via positive frequency dependence. Since a narrow range of Escherichia coli genotypes (primarily those belonging to the clonal group ST131) cause a significant proportion of multidrug-resistant infections, this group presents a promising target for decolonization with bacteriophage, as narrow-host-range viral predation could lead to selective removal of particular genotypes. In this study we tested how a combination of an ST131-specific phage and competition from the well-known probiotic E. coli Nissle strain could displace E. coli ST131 under aerobic and anaerobic growth conditions in vitro. We showed that the addition of phage was able to break the frequency-dependent advantage of a numerically dominant ST131 isolate. Moreover, the addition of competing E. coli Nissle could improve the ability of phage to suppress ST131 by two orders of magnitude. Low-cost phage resistance evolved readily in these experiments and was not inhibited by the presence of a probiotic competitor. Nevertheless, combinations of phage and probiotic produced stable long-term suppression of ST131 over multiple transfers and under both aerobic and anaerobic growth conditions. Combinations of phage and probiotic therefore have real potential for accelerating the removal of drug-resistant commensal targets.