Capturing dynamic phage–pathogen coevolution by clinical surveillance | Nature

Download PDF Subjects Bacterial infection Bacteriophages Coevolution Abstract Bacteria harness diverse defence systems that protect against phage predation 1 , many of which are encoded on horizontally transmitted mobile genetic elements 2 . In turn, phages evolve counter-defences 3 , driving a dynamic arms race that remains underexplored in human disease contexts. For the diarrhoeal pathogen Vibrio cholerae , a higher burden of its lytic phage ICP1 in patient stool correlates with reduced disease severity 4 . However, direct molecular evidence of lytic phages driving selection of epidemic V. cholerae has not been demonstrated. Here, through clinical surveillance in cholera-endemic Bangladesh, we capture the acquisition of a parasitic anti-phage mobile genetic element, PLE11, that initiated a selective sweep coinciding with the largest cholera outbreak in recent records. PLE11 showed potent anti-phage activity against cocirculating ICP1, explaining its rapid and dominating emergence. We identify PLE11-encoded Rta as the defence responsible and provide evidence that Rta restricts phage tail assembly. Using experimental evolution, we predict phage counteradaptations against PLE11 and document the eventual emergence and selection of clinical ICP1 that achieve a convergent evolutionary outcome. Finally, we discover how PLEs balance their dependence on ICP1 tail proteins for horizontal transmission with the restriction of phage tail assembly by Rta: PLEs construct chimeric tails composed of both mobile genetic element-encoded and phage-encoded proteins to ensure their transmission. Collectively, our findings reveal the molecular basis of the natural selection of a globally important pathogen and its virus in a clinically relevant context. Main The infectious diarrhoeal disease cholera, caused by the pathogen Vibrio cholerae , remains a major threat to global public health. The Bay of Bengal is considered the source of the continuing seventh cholera pandemic, where seventh pandemic El Tor (7PET) sublineages evolve and spread to vulnerable nations across the globe 5 . Unravelling the factors influencing the evolution and selection of 7PET strains is challenging, as they result from the complex interplay of genetic changes, particularly the flux of new mobile genetic elements (MGEs), and selective advantages that can be difficult to ascertain. Recent metagenomic analyses in cholera-endemic Bangladesh indicate that higher ratios of ICP1, the predominant lytic phage preying on V. cholerae in the context of disease 6 , correlate with reduced risk of severe disease in patients 4 . This suggests that the acquisition of phage resistance could contribute to outbreak severity and influence the evolution of pandemic lineages. However, direct molecular evidence of this link is lacking, as are mechanistic insights into how the dynamic oscillations of phage susceptibility and resistance unfold in nature. Among the fluctuating MGEs found in 7PET V. cholerae are a family of phage satellites called phage-inducible chromosomal island-like elements (PLEs) that play a crucial role in defending against ICP1 predation 7 . PLEs have evolved intricate defence mechanisms that disrupt the phage life cycle while exploiting phage machinery and structural components to package themselves into modified viral particles, thereby blocking phage transmission 7 , 8 , 9 , 10 . PLEs are highly specialized in safeguarding V. cholerae populations from predation by ICP1, but phage counteradaptations can neutralize their potent anti-phage activity. Analyses of sparsely collected ICP1 isolates have revealed three anti-PLE mechanisms the phage uses to overcome PLE-mediated hijacking and restore phage propagation. These mechanisms vary among phage isolates 6 and mediate nucleolytic degradation of the PLE genome through distinct mechanisms (Fig. 1a ). The phage-encoded origin-directed nuclease (Odn) 11 and attachment-directed inhibitor (Adi) 12 antagonize specific subsets of the ten PLE variants discovered so far 13 , targeting sequence variants of the PLE origin of replication and integrase, respectively. A remarkable adaptation in ICP1 is its co-option of CRISPR–Cas, which replaces odn in the same genomic locus and provides broad-spectrum counter-defence against all PLEs tested so far 11 , 14 . Genomic analyses have documented the temporal flux of distinct PLE variants in epidemic V. cholerae 13 . However, without knowledge of cocirculating ICP1 genotypes, the molecular factors driving such epidemiological patterns remain unclear. Building on the foundational understanding of PLE–ICP1 conflict, we harnessed high-resolution clinical surveillance in Bangladesh to capture and understand the emergence and selection of epidemic V. cholerae and ICP1 in a region critical to the dynamics of pandemic cholera. Fig. 1: Clinical surveillance reveals the acquisition and selection of PLE11 in V. cholerae that restricts ICP1 despite anti-PLE mechanisms. a , Known infect