Data_Sheet_1_Impact of a phage cocktail targeting Escherichia coli and Enterococcus faecalis as members of a gut bacterial consortium in vitro and in vivo.docx

Escherichia coli and Enterococcus faecalis have been implicated as important players in human gut health that have been associated with the onset of inflammatory bowel disease (IBD). Bacteriophage (phage) therapy has been used for decades to target pathogens as an alternative to antibiotics, but the ability of phage to shape complex bacterial consortia in the lower gastrointestinal tract is not clearly understood. We administered a cocktail of six phages (either viable or heat-inactivated) targeting pro-inflammatory Escherichia coli LF82 and Enterococcus faecalis OG1RF as members of a defined community in both a continuous fermenter and a murine colitis model. The two target strains were members of a six species simplified human microbiome consortium (SIHUMI-6). In a 72-h continuous fermentation, the phage cocktail caused a 1.1 and 1.5 log (log10 genome copies/mL) reduction in E. faecalis and E. coli numbers, respectively. This interaction was accompanied by changes in the numbers of other SIHUMI-6 members, with an increase of Lactiplantibacillus plantarum (1.7 log) and Faecalibacterium prausnitzii (1.8 log). However, in germ-free mice colonized by the same bacterial consortium, the same phage cocktail administered twice a week over nine weeks did not cause a significant reduction of the target strains. Mice treated with active or inactive phage had similar levels of pro-inflammatory cytokines (IFN-y/IL12p40) in unstimulated colorectal colonic strip cultures. However, histology scores of the murine lower GIT (cecum and distal colon) were lower in the viable phage-treated mice, suggesting that the phage cocktail did influence the functionality of the SIHUMI-6 consortium. For this study, we conclude that the observed potential of phages to reduce host populations in in vitro models did not translate to a similar outcome in an in vivo setting, with this effect likely brought about by the reduction of phage numbers during transit of the mouse GIT.

大肠杆菌和粪肠球菌已被确认为影响人类肠道健康的重要微生物，它们与炎症性肠病（IBD）的发生密切相关。噬菌体（phage）治疗作为一种替代抗生素的途径，已应用数十年以靶向病原体，但其对下消化道中复杂细菌群落（bacterial consortia）的塑造能力尚未得到充分理解。本研究中，我们向连续发酵器和小鼠结肠炎模型中，以一特定群落中的成员为目标，分别施用了包含六种噬菌体（活性或热失活的）的混合物，这些噬菌体针对促炎的大肠杆菌LF82和粪肠球菌OG1RF。这两种目标菌株属于一个六种物种简化的人类微生物组联合体（SIHUMI-6）。在72小时的连续发酵过程中，噬菌体混合物导致粪肠球菌和的大肠杆菌数量分别减少了1.1和1.5个对数单位（log10基因组拷贝/mL）。这一相互作用伴随着其他SIHUMI-6成员数量的变化，其中植物乳杆菌（Lactiplantibacillus plantarum）和普拉梭菌（Faecalibacterium prausnitzii）的数量分别增加了1.7和1.8个对数单位。然而，在同一细菌联合体殖民的无菌小鼠中，每周两次、连续九周施用相同的噬菌体混合物并未导致目标菌株数量的显著减少。接受活性或非活性噬菌体治疗的小鼠，在未刺激的结肠直肠结肠片培养中，促炎细胞因子（IFN-y/IL12p40）的水平相似。然而，接受活性噬菌体治疗的小鼠的小鼠下消化道（盲肠和远端结肠）的组织学评分较低，这表明噬菌体混合物确实影响了SIHUMI-6联合体的功能。对于本研究，我们得出结论，噬菌体在体外模型中减少宿主种群的能力并未在体内环境中产生类似的效应，这种效果可能是由噬菌体在通过小鼠肠道过程中的数量减少所引起的。