Contrasted coevolutionary dynamics between a bacterial pathogen and its bacteriophages

Many antagonistic interactions between hosts and their parasites result in coevolution. Although coevolution can drive diversity and specificity within species, it is not known whether coevolutionary dynamics differ among functionally similar species. We present evidence of coevolution within simple communities of Pseudomonas aeruginosa PAO1 and a panel of bacteriophages. Pathogen identity affected coevolutionary dynamics. For five of six phages tested, time-shift assays revealed temporal peaks in bacterial resistance and phage infectivity, consistent with frequency-dependent selection (Red Queen dynamics). Two of the six phages also imposed additional directional selection, resulting in strongly increased resistance ranges over the entire length of the experiment (ca. 60 generations). Cross-resistance to these two phages was very high, independent of the coevolutionary history of the bacteria. We suggest that coevolutionary dynamics are associated with the nature of the receptor used by the phage for infection. Our results shed light on the coevolutionary process in simple communities and have practical application in the control of bacterial pathogens through the evolutionary training of phages, increasing their virulence and efficacy as therapeutics or disinfectants.Many host–parasite associations coevolve, and patterns in this antagonistic interaction are influenced by biology and environment (1–3). In single-species host–parasite interactions, parasite genotypes show differences in their host ranges and specificities on host genotypes, providing the basis for such coevolution (4–8). Arms race dynamics (ARD) driven by directional selection favors a broader resistance range in the host against a greater number of parasite genotypes and an increased host range in the parasite allowing more host genotypes to be infected (2, 9). In contrast, fluctuating selection dynamics (FSD), in which there is no directional change in the evolution of the host resistance range, is governed by negative frequency-dependent selection, favoring hosts that resist the most frequently encountered parasite genotypes and parasites that infect the most common host genotypes (9–11). It has been suggested that ARD predominates during the initial stages of coevolution, when adaptations to the coevolving opponent are largely cost-free, whereas FDS is more significant at later stages, when attack/defense alleles accumulate in the genome and impose costs (12). Thus, when systems shift from ARD to FSD, dynamic coevolutionary equilibria may arise, with constant numbers of attack/defense alleles at the individual level (13) and the continuous frequency-dependent (re)cycling of alleles at the population level (14).Coevolutionary dynamics between hosts and parasites is increasingly investigated using experimental evolution (15) and more specifically time-shift assays (16–18), in which hosts or parasites from a given time point are compared with their counterparts from the past or future, enabling the examination of putative reciprocal adaptations (19, 20). Some of the prime experimental models are bacteria and their lytic phages, which exhibit rapid evolution and are amenable to time-shift tests. Recent study indicates that coevolving populations may exhibit either ARD or FSD (12, 21–24), and there is some evidence for the genetic mechanism involved [e.g., mutations at tail fiber genes (12, 25)] and for the expansion of the phage host range as coevolution proceeds (6). Most studies of bacteria–phage coevolution involve the model system Pseudomonas fluorescens SBW25 and its lytic phage ϕ2 (15). Although these studies are important for an in-depth understanding of this process, their restriction to a single host–parasite pair is unfortunate, given the immense diversity of bacteria and phages in both terrestrial and aquatic ecosystems (26–28) and the importance of many of these organisms in human health (29). Thus, the generality of previous results to other bacterial species and to different phages parasitizing a given bacterium remains an open question.Coevolution may be particularly important in an applied context, namely when predators or parasitic organisms are used for biocontrol (30). For instance, Conrad et al. (31) recently argued that a community perspective with its ecological and evolutionary underpinnings is needed to explore the usefulness of phages as antimicrobial agents to treat cystic fibrosis patients infected with several bacteria and notably strains of Pseudomonas aeruginosa, a congeneric to the model organism P. fluorescens. Different phages naturally have different host ranges (8, 32, 33), but whether phage taxonomic origin influences impacts on P. aeruginosa is not known. Despite the study of phage mixtures to control P. aeruginosa infections (34), the nature of bacterial cross-resistance to phages other than that with which the bacterium evolved has not been addressed.Previous studies are inconclusive regarding whether P. aeruginosa coevolves with bacteriophages (35, 36). Given the ubiquity of this model organism in natural habitats (37) and as a widespread pathogen in hospitals (38, 39), it is important to know whether phage parasitism influences P. aeruginosa population biology and adaptation, whether coevolution between these antagonists actually occurs, and, if so, whether there are general patterns shared by different phages. Here, we test coevolutionary dynamics and their consistency in a panel of lytic bacteriophages and their host P. aeruginosa PAO1. We allowed this bacterium to interact and potentially coevolve with each of six different phage isolates separately, four from the Podoviridae and two from the Myoviridae (Table S1), for a total of 10 serial transfers (∼60 generations). Using bacteria and phages isolated from different time points, we conducted time-shift assays of resistance to infer patterns of the coevolutionary process. Finally, to assess specificity, we performed a cross-resistance assay with evolved bacteria and the six ancestral phage isolates to compare resistance of the bacteria to their “own” phage and to “foreign” phages with which they had not coevolved.