Coevolutionary phage training leads to greater bacterial suppression and delays the evolution of phage resistance

Abstract

The evolution of antibiotic resistant bacteria threatens to become the leading cause of worldwide mortality. This crisis has renewed interest in the practice of phage therapy. Yet, bacterial capacity to evolve resistance is likely to debilitate this therapy as well. To combat the evolution of phage resistance and improve treatment outcomes, many have suggested leveraging phages ability to counter resistance by evolving phages on target hosts before using them in therapy (phage training). We found that during in vitro experiments, a phage trained for 28 days suppressed bacteria ~1000-fold for 3-8 times longer than its untrained ancestor. This extension was due to a delay in the evolution of resistance. Several factors contributed to this prolonged suppression. Mutations that confer resistance to trained phages are ~100x less common and, while the target bacterium can evolve complete resistance to the untrained phage in a single step, multiple mutations are required to evolve complete resistance to trained phages. Mutations that confer resistance to trained phages are more costly than mutations for untrained phage resistance. And when resistance does evolve, trained phages are better able to suppress these forms of resistance. One way the trained phage improved was through recombination with a gene in a defunct prophage in the host genome, which doubled phage fitness. This direct transfer of information encoded by the host but originating from a relict phage provides a previously unconsidered mode of training phage. Overall, we provide a case study for successful phage training and uncover mechanisms underlying its efficacy.

Leapfrog dynamics in phage-bacteria coevolution revealed by joint analysis of cross-infection phenotypes and whole genome sequencing

“…Acquiring such large effect mutations could also help lineages leap ahead. Lastly, these models typically do not incorporate recombination, which was observed within this phage population to cause sudden increases in fitness (Borin et al 2020) and could contribute to the phage's ability to leap ahead. In contrast, evidence of recombination was not observed in the bacterial genome sequences, consistent with earlier findings in which recombination is not known to occur in this strain of E. coli (Souza et al 1997).…”

“…Acquiring such large effect mutations could also help lineages leap ahead. Lastly, these models typically do not incorporate recombination, which was observed within this phage population to cause sudden increases in fitness (Borin et al . 2020) and could contribute to the phage’s ability to leap ahead.…”

Comparison of bacterial suppression by phage cocktails, dual-receptor generalists, and coevolutionarily trained phages

“…We also compared our cocktail against a trained generalist (λtgen) that previously demonstrated strong suppressive capabilities (Borin et al, 2021). Although both of our treatments use the same two receptors, λtgen was significantly more suppressive.…”

“…In previous work, we demonstrated that the ability to infect through 2 distinct receptors substantially improves bacterial suppression (Borin et al 2021). We did this by comparing phages that were "trained" via coevolution with their host for 20 d. Half of the phages we compared had evolved the ability to use an additional receptor during training and were far more suppressive (~1000-fold) than the phage ancestor.…”

Phage therapy for One Health approach: current status, challenges and opportunities

“…菌，这种识别特异性甚至达到细菌"株"的水平。噬菌体的局限性首先体现在侵染裂解谱范 围上。Zhao 等人发现，当土壤中有多种类型病原菌时，宿主特异性噬菌体抑制病原细菌的 能力显著低于宽宿主范围的噬菌体(多价噬菌体) [90] 。其次，噬菌体侵染病原菌是频率依 赖型的，当环境中的宿主细菌数量足够多且生理上和遗传上适合噬菌体感染时，才能有利于 噬菌体侵染增殖 [91] 。如果目标细菌密度太低，噬菌体与病原细菌相遇的几率会大大下降， 噬菌体的裂解速率也会降低，甚至会转为溶原状态 [92] 。此外，单一噬菌体疗法与抗生素类 似，其作用效果会因病原菌的抗性进化而削弱。为解决这些局限性，除了筛选裂解谱广的噬 菌体，还需要定向驯化出具有更强侵染能力的噬菌体 [93] ，或构建噬菌体鸡尾酒等策略提高 噬菌体疗法的稳定性。 值得注意的是， 有研究发现噬菌体是环境中抗生素抗性基因 (ARGs) 的储存库和潜在的传播载体 [94][95] ，建议避免使用编码溶原性、毒力因子或携带 ARGs 的噬菌 体。评估农业环境中噬菌体-宿主病原菌-ARGs 之间的关系对噬菌体疗法的应用推广、规避 其环境风险具有重要意义。…”