Liang Bingchun, Zhao Wenpeng, Han Bo, Barkema Herman W, Niu Yan D, Liu Yongxia, Kastelic John P, Gao Jian
Department of Clinical Veterinary Medicine, College of Veterinary Medicine, China Agricultural University, Beijing, China.
Department of Production Animal Health, Faculty of Veterinary Medicine, Hospital Drive NW, University of Calgary, Calgary, AB, Canada.
Front Microbiol. 2022 Oct 13;13:943279. doi: 10.3389/fmicb.2022.943279. eCollection 2022.
Bovine mastitis caused by multi-drug resistant (MDR) is difficult to treat with antibiotics, whereas bacteriophages may be a viable alternative. Our objective was to use 2 . strains, 1 MDR and the other non-MDR, to isolate phages from sewage samples and compare their biological and genomic characteristics. Additionally, phage infected mouse mammary gland was also analyzed by H&E staining and ELISA kits to compare morphology and inflammatory factors, respectively. Based on assessments with double agar plates and transmission electron microscopy, phage CM_Kpn_HB132952 had clear plaques surrounded by translucent halos on the bacterial lawn of . KPHB132952 and belonged to , whereas phage CM_Kpn_HB143742 formed a clear plaque on the bacterial lawn of . KPHB143742 and belonged to . In 1-step growth curves, CM_Kpn_HB132952 and CM_Kpn_HB143742 had burst sizes of 0.34 and 0.73 log PFU/mL, respectively. The former had a latent period of 50 min and an optimal multiplicity of infection (MOI) of 0.01, whereas for the latter, the latent period was 30 min (MOI = 1). Phage CM_Kpn_HB132952 had better thermal and acid-base stability than phage CM_Kpn_HB143742. Additionally, both phages had the same host range rate but different host ranges. Based on Illumina NovaSeq, phages CM_Kpn_HB132952 and CM_Kpn_HB143742 had 140 and 145 predicted genes, respectively. Genomic sequencing and phylogenetic tree analysis indicated that both phages were novel phages belonging to the family. Additionally, the histopathological structure and inflammatory factors TNF-α and IL-1β were not significantly different among phage groups and the control group. In conclusion, using 1 MDR and 1 non-MDR strain of . , we successfully isolated two phages from the same sewage sample, and demonstrated that they had distinct biological and genomic characteristics.
由多重耐药菌（MDR）引起的牛乳腺炎难以用抗生素治疗，而噬菌体可能是一种可行的替代方法。我们的目标是使用2株菌，1株多重耐药菌和另1株非多重耐药菌，从污水样本中分离噬菌体，并比较它们的生物学和基因组特征。此外，还通过苏木精-伊红（H&E）染色和酶联免疫吸附测定（ELISA）试剂盒对噬菌体感染的小鼠乳腺进行分析，分别比较其形态和炎症因子。基于双层琼脂平板评估和透射电子显微镜观察，噬菌体CM_Kpn_HB132952在肺炎克雷伯菌KPHB132952的菌苔上形成了周围有半透明晕圈的清晰噬菌斑，属于[噬菌体所属类别未给出]，而噬菌体CM_Kpn_HB143742在肺炎克雷伯菌KPHB143742的菌苔上形成了清晰噬菌斑，属于[噬菌体所属类别未给出]。在一步生长曲线中，CM_Kpn_HB132952和CM_Kpn_HB143742的裂解量分别为0.34和0.73 log PFU/mL。前者的潜伏期为50分钟，最佳感染复数（MOI）为0.01，而后者的潜伏期为30分钟（MOI = 1）。噬菌体CM_Kpn_HB132952比噬菌体CM_Kpn_HB143742具有更好的热稳定性和酸碱稳定性。此外，两种噬菌体的宿主范围率相同，但宿主范围不同。基于Illumina NovaSeq测序，噬菌体CM_Kpn_HB132952和CM_Kpn_HB143742分别有140个和145个预测基因。基因组测序和系统发育树分析表明，两种噬菌体都是属于[噬菌体所属类别未给出]科的新型噬菌体。此外，噬菌体组和对照组之间的组织病理学结构以及炎症因子肿瘤坏死因子-α（TNF-α）和白细胞介素-1β（IL-1β）没有显著差异。总之，使用1株多重耐药菌和1株非多重耐药菌的肺炎克雷伯菌，我们成功地从同一污水样本中分离出两种噬菌体，并证明它们具有不同的生物学和基因组特征。
