School of Life Science and Technology, Tokyo Institute of Technology, 4259 J2-15 Nagatsuta-cho, Midori-ku, Yokohama 226-8501, Japan.
Division of Bacteriology, Department of Infection and Immunity, Faculty of Medicine, Jichi Medical University, Yakushiji, Shimotsuke 329-0498, Japan.
J Biosci Bioeng. 2020 Jun;129(6):693-699. doi: 10.1016/j.jbiosc.2020.02.001. Epub 2020 Feb 24.
Pseudomonas aeruginosa is an opportunistic pathogen that causes nosocomial disease among immunocompromised and chronic cystic fibrosis (CF) patients. We characterized two newly isolated Pseudomonas phages, ϕPA01 and ϕPA02, with different host spectra, and examined their effect as a cocktail with antibiotics against P. aeruginosa, to indicate the possibility of combining a phage cocktail and antibiotics in treating pseudomonal infection. Phages ϕPA01 (66,220 bp) and ϕPA02 (279,095 bp) belong to the genus Pbunalikevirus and Phikzlikevirus, respectively. No virulence or lysogenic associated gene was found in their genomes, thus they are potentially safe for phage therapy. We generated respective phage-resistant strains to investigate cross-resistance between two phages. Slight cross-resistance to ϕPA02 in ϕPA01-resistant strain was observed, while ϕPA02-resistant strain remained susceptible to ϕPA01. A ϕPA01 resistant strain that was cross-resistant to ϕPA02 appeared in round 5 (R5-PA01R), revealed frameshift mutation in phosphoglucomutase (algC), which is important for the synthesis of core lipopolysaccharide (LPS). Knockout of algC was resistant to both phages. Complementation of ΔalgC restored phages' infectivity, suggesting that LPS as host receptor. Phage cocktail suppressed the growth of P. aeruginosa for longer (20 h) hour compared with single phage (8-9 h), further suggesting their potential to be used as a phage cocktail. Furthermore, application of the phage cocktail with ciprofloxacin (0.25 μg/ml) and meropenem (2 μg/ml), managed to suppress the growth of P. aeruginosa up to 96 h. Our results show the potential application of ϕPA01 and ϕPA02 as phage cocktail together with antibiotics for treatment of P. aeruginosa.
铜绿假单胞菌是一种机会性病原体，会导致免疫功能低下和慢性囊性纤维化 (CF) 患者发生医院获得性疾病。我们对两种新分离的具有不同宿主谱的铜绿假单胞菌噬菌体ϕPA01 和 ϕPA02 进行了表征，并研究了它们与抗生素联合使用对铜绿假单胞菌的作用，以表明将噬菌体鸡尾酒与抗生素联合用于治疗假单胞菌感染的可能性。噬菌体 ϕPA01（66220 bp）和 ϕPA02（279095 bp）分别属于 Pbunalikevirus 和 Phikzlikevirus 属。在它们的基因组中未发现毒力或溶原相关基因，因此它们对噬菌体治疗具有潜在的安全性。我们生成了各自的噬菌体抗性菌株，以研究两种噬菌体之间的交叉抗性。在噬菌体抗性菌株中观察到对 ϕPA02 的轻微交叉抗性，而噬菌体抗性菌株仍对 ϕPA01 敏感。在第 5 轮（R5-PA01R）中出现了对 ϕPA02 交叉抗性的噬菌体抗性菌株，其磷酸葡萄糖变位酶 (algC) 发生移码突变，这对于核心脂多糖 (LPS) 的合成很重要。algC 缺失突变株对两种噬菌体均有抗性。algC 的互补恢复了噬菌体的感染性，表明 LPS 是宿主受体。噬菌体鸡尾酒抑制铜绿假单胞菌生长的时间比单一噬菌体（8-9 h）更长（20 h），这进一步表明它们有可能被用作噬菌体鸡尾酒。此外，噬菌体鸡尾酒联合环丙沙星（0.25 μg/ml）和美罗培南（2 μg/ml）的应用能够抑制铜绿假单胞菌的生长长达 96 h。我们的研究结果表明，噬菌体 ϕPA01 和 ϕPA02 具有作为噬菌体鸡尾酒与抗生素联合应用于治疗铜绿假单胞菌的潜力。
