Analysis of Selection Methods to Develop Novel Phage Therapy Cocktails Against Antimicrobial Resistant Clinical Isolates of Bacteria.
Analysis of Selection Methods to Develop Novel Phage Therapy Cocktails Against Antimicrobial Resistant Clinical Isolates of Bacteria.
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DOI：
10.3389/fmicb.2021.613529
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发表时间：
2021
影响因子：
5.2
通讯作者：
Clokie MRJ
中科院分区：
文献类型：
--
作者：
Haines MEK;Hodges FE;Nale JY;Mahony J;van Sinderen D;Kaczorowska J;Alrashid B;Akter M;Brown N;Sauvageau D;Sicheritz-Pontén T;Thanki AM;Millard AD;Galyov EE;Clokie MRJ
抗生素耐药性（AMR）是全球面临的一个主要问题。与尿路感染（UTI）相关的脓毒症的主要细菌是大肠杆菌。大肠杆菌和克雷伯菌属沿着以及肠杆菌属。这些都有AMR菌株被称为ESBL（超广谱β-内酰胺酶），这是世界卫生组织优先病原体名单上的特点，作为“关键”的研究。噬菌体（Bacteriophages，简称BPV）作为一种可以感染和杀死细菌的病毒，可以提供一种有效的工具来对付这些AMR菌株。目前还没有开发噬菌体鸡尾酒的“金标准”。在这里，我们描述了一种新的方法来开发一种有效的噬菌体鸡尾酒对一组产ESBL大肠杆菌。大肠杆菌和克雷伯氏菌主要从英国医院的患者中分离出来。通过比较噬菌体功效的不同测量，我们显示了哪些是最稳健的，并提出了一种有效的筛选级联，可用于开发噬菌体鸡尾酒以靶向其他AMR细菌物种。对从尿液样本中分离的38株产ESBL临床菌株的目标面板进行整理并用于测试噬菌体效力。在初步筛选68个菌株后，确定了6个菌株，并对这38个菌株进行了测试，以确定其临床覆盖率和杀灭效率。为了实现这一点，我们评估了四种不同的方法来评估这些细菌分离株的噬菌体毒力。这些是直接斑点试验（DST）、平板效率（EOP）测定、嗜酸性杀伤测定（PKA）和生物膜测定。最终的ESBL混合液对38株菌有23株（61%）有效杀灭，对克雷伯菌13株（68%）有效，对大肠埃希菌13株（68%）有效。coli 10/19（53%）。ESBL E.大肠杆菌收集物具有来自流行UTI相关ST 131序列类型的六个分离物，其中五个被最终混合物有效靶向。在用于评估噬菌体毒力的四种方法中，数据表明PKA与更耗时的EOP一样有效，并且两种测定的数据相关性良好。这表明，在确定鸡尾酒中应使用哪种药物时，促渗性杀灭是一个很好的替代方法。当与毒力指数组合时，该测定还允许“噬菌体协同作用”告知混合物设计。
Antimicrobial resistance (AMR) is a major problem globally. The main bacterial organisms associated with urinary tract infection (UTI) associated sepsis are E. coli and Klebsiella along with Enterobacter species. These all have AMR strains known as ESBL (Extended Spectrum Beta-Lactamase), which are featured on the WHO priority pathogens list as “critical” for research. Bacteriophages (phages), as viruses that can infect and kill bacteria, could provide an effective tool to tackle these AMR strains. There is currently no “gold standard” for developing a phage cocktail. Here we describe a novel approach to develop an effective phage cocktail against a set of ESBL-producing E. coli and Klebsiella largely isolated from patients in United Kingdom hospitals. By comparing different measures of phage efficacy, we show which are the most robust, and suggest an efficient screening cascade that could be used to develop phage cocktails to target other AMR bacterial species. A target panel of 38 ESBL-producing clinical strains isolated from urine samples was collated and used to test phage efficacy. After an initial screening of 68 phages, six were identified and tested against these 38 strains to determine their clinical coverage and killing efficiency. To achieve this, we assessed four different methods to assess phage virulence across these bacterial isolates. These were the Direct Spot Test (DST), the Efficiency of Plating (EOP) assay, the planktonic killing assay (PKA) and the biofilm assay. The final ESBL cocktail of six phages could effectively kill 23/38 strains (61%), for Klebsiella 13/19 (68%) and for E. coli 10/19 (53%) based on the PKA data. The ESBL E. coli collection had six isolates from the prevalent UTI-associated ST131 sequence type, five of which were targeted effectively by the final cocktail. Of the four methods used to assess phage virulence, the data suggests that PKAs are as effective as the much more time-consuming EOPs and data for the two assays correlates well. This suggests that planktonic killing is a good proxy to determine which phages should be used in a cocktail. This assay when combined with the virulence index also allows “phage synergy” to inform cocktail design.
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