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<p>The clinical translation of phage therapy for multidrug-resistant infections is constrained by the lack of rapid, standardized therapeutic phage selection. Here, we introduce digital phage susceptibility testing (dPhaST), an automated droplet digital PCR workflow that quantifies phage-induced DNA release as a molecular signature of lysis. By targeting conserved 16<i>S</i> rRNA regions, dPhaST measures lytic activity across diverse bacterial pathogens within 3 h. Across 122 phage–host combinations involving 19 bacterial strains from six species, dPhaST shows 95.9% concordance with spot tests while resolving weak and heterogeneous lytic activities that are not readily distinguished phenotypically. It remains robust during the early infection window despite phage-encoded nuclease activity and tolerates phage cross-contamination better than spot tests. The method captures defense-mediated interactions involving CRISPR–Cas and Sir2-HerA systems. In this work, we show that automated digital quantification enables rapid and mechanistically informative profiling of early phage lytic efficacy across Gram-positive and Gram-negative pathogens.</p>

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Rapid and quantitative measurement of bacteriophage infectivity via fully automated droplet digital PCR

  • 期刊论文

  • 出版模式：

    开放获取

  • 发表日期：

    2026年7月30日

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  • Yanfei Liu,
  • Huiwei Zhao,
  • Xinyue Cao,
  • Shize Jiang,
  • Jun Fu,
  • Junjing Xue,
  • Caiming Li,
  • Zhangrun Xu,
  • Ming Li,
  • Wenbin Du

摘要

The clinical translation of phage therapy for multidrug-resistant infections is constrained by the lack of rapid, standardized therapeutic phage selection. Here, we introduce digital phage susceptibility testing (dPhaST), an automated droplet digital PCR workflow that quantifies phage-induced DNA release as a molecular signature of lysis. By targeting conserved 16S rRNA regions, dPhaST measures lytic activity across diverse bacterial pathogens within 3 h. Across 122 phage–host combinations involving 19 bacterial strains from six species, dPhaST shows 95.9% concordance with spot tests while resolving weak and heterogeneous lytic activities that are not readily distinguished phenotypically. It remains robust during the early infection window despite phage-encoded nuclease activity and tolerates phage cross-contamination better than spot tests. The method captures defense-mediated interactions involving CRISPR–Cas and Sir2-HerA systems. In this work, we show that automated digital quantification enables rapid and mechanistically informative profiling of early phage lytic efficacy across Gram-positive and Gram-negative pathogens.