Shanghai Institute of Phage, Shanghai Public Health Clinical Centre, Fudan University, Shanghai, China.
Centre for Tuberculosis Research, Shanghai Public Health Clinical Centre, Fudan University, Shanghai, China.
Clin Microbiol Infect. 2023 Dec;29(12):1601.e1-1601.e7. doi: 10.1016/j.cmi.2023.08.022. Epub 2023 Aug 29.
Phage-resistant bacteria often emerge rapidly when performing phage therapy. However, the relationship between the emergence of phage-resistant bacteria and improvements in clinical symptoms is still poorly understood.
An inpatient developed a pulmonary infection caused by multidrug-resistant Klebsiella pneumoniae. He received a first course of treatment with a single nebulized phage (ΦKp_GWPB35) targeted at his bacterial isolate of Kp7450. After 14 days, he received a second course of treatment with a phage cocktail (ΦKp_GWPB35+ΦKp_GWPA139). Antibiotic treatment was continued throughout the course of phage therapy. Whole-genome analysis was used to identify mutations in phage-resistant strains. Mutated genes associated with resistance were further analysed by generating knockouts of Kp7450 and by measuring phage adsorption rates of bacteria treated with proteinase K and periodate. Bacterial virulence was evaluated in mouse and zebrafish infection models.
Phage-resistant Klebsiella pneumoniae strains emerged after the second phage treatment. Comparative genomic analyses revealed that fabF was deleted in phage-resistant strains. The fabF knockout strain (Kp7450ΔfabF) resulted in an altered structure of lipopolysaccharide (LPS), which was identified as the host receptor for the therapeutic phages. Virulence evaluations in mice and zebrafish models showed that LPS was the main determinant of virulence in Kp7450 and alteration of LPS structure in Kp7450ΔfabF, and the bacteriophage-resistant strains reduced their virulence at cost.
This study may shed light on the mechanism by which some patients experience clinical improvement in their symptoms post phage therapy, despite the incomplete elimination of pathogenic bacteria.
在进行噬菌体治疗时，噬菌体耐药细菌通常会迅速出现。然而，噬菌体耐药细菌的出现与临床症状的改善之间的关系仍不清楚。
一名住院患者因多重耐药肺炎克雷伯菌引起肺部感染。他接受了一次单独雾化噬菌体（ΦKp_GWPB35）治疗，针对他的 Kp7450 细菌分离株。14 天后，他接受了第二次噬菌体鸡尾酒治疗（ΦKp_GWPB35+ΦKp_GWPA139）。在噬菌体治疗过程中，继续使用抗生素治疗。全基因组分析用于鉴定噬菌体耐药株的突变。通过生成 Kp7450 的敲除突变体，并测量经蛋白酶 K 和过碘酸钠处理的细菌的噬菌体吸附率，进一步分析与耐药相关的突变基因。在小鼠和斑马鱼感染模型中评估细菌的毒力。
第二次噬菌体治疗后出现了噬菌体耐药肺炎克雷伯菌菌株。比较基因组分析表明，fabF 在噬菌体耐药菌株中缺失。fabF 敲除菌株（Kp7450ΔfabF）导致脂多糖（LPS）结构发生改变，LPS 被鉴定为治疗噬菌体的宿主受体。在小鼠和斑马鱼感染模型中的毒力评估表明，LPS 是 Kp7450 毒力的主要决定因素，LPS 结构的改变在 Kp7450ΔfabF 中，噬菌体耐药菌株以降低其毒力为代价。
本研究可能阐明了一些患者在噬菌体治疗后症状改善的机制，尽管未能完全消除病原菌。
