Peng Xiaocui, Chang Jianliang, Zhang Hongxia, Li Xiaoyu, Zhang Changhong, Jiao Shiyan, Lv Chengxiu, Wang Na, Zhao Jun, Wang Bu, Zhang Wei, Zhang Zhihua
Department of Postgraduate, Hebei North University, Zhangjiakou, China.
Respiratory and Critical Care Medicine Department, The First Affiliated Hospital of Hebei North University, Zhangjiakou, China.
Front Microbiol. 2025 Mar 7;16:1491961. doi: 10.3389/fmicb.2025.1491961. eCollection 2025.
Hypervirulent and multidrug-resistant (hvKP and MDR-KP) are significant public health threats. This study aimed to isolate a lytic bacteriophage targeting these high-risk strains, systematically characterize its biological properties, genomic features, and therapeutic efficacy, and establish a foundation for clinical phage therapy and novel antimicrobial development.
The phage vB_Kp_XP4 was isolated from river water using the double-layer agar plate method with the clinically isolated strain P4 as the host. Morphology was analyzed via transmission electron microscopy (TEM). Host range, pH, and thermal stability were assessed using spot assays and OD measurements. One-step growth curves determined the latent period and burst size. Whole-genome sequencing and phylogenetic analysis were performed. Therapeutic efficacy and safety were evaluated in a infection model.
TEM revealed Phage vB_Kp_XP4 as a tailed phage with an icosahedral head and a long, flexible tail. It lysed an hvKP strain (carrying , , , genes) and an MDR-KP strain (resistant to carbapenems, fluoroquinolones, etc.), with an optimal MOI of 0.1 and latent period <10 minutes. Stability was maintained at pH 4-11 and ≤70°C. Whole-genome sequencing revealed a linear double-stranded DNA genome of 44,344 bp with a G+C content of 53.80%. The genome comprised 54 coding sequences and lacked lysogenic, virulence, or antibiotic resistance genes. Phylogenetic analysis positioned phage vB_Kp_XP4 as a novel species within the genus , family . In the model, vB_Kp_XP4 prolonged survival of P4-infected larvae ( < 0.001).
Phage vB_Kp_XP4 exhibits high stability, specificity, potent lytic activity, and no undesirable genes, demonstrating effective in vivo therapeutic efficacy, suggest its potential for clinical applications against infections. The presence of multiple halos during plaque formation further enhances its research value. The complete genome sequence has been submitted to GenBank under accession number PP663283.
高毒力和多重耐药性（hvKP和MDR-KP）是重大的公共卫生威胁。本研究旨在分离一种靶向这些高风险菌株的裂解性噬菌体，系统地表征其生物学特性、基因组特征和治疗效果，并为临床噬菌体治疗和新型抗菌药物开发奠定基础。
以临床分离菌株P4为宿主，采用双层琼脂平板法从河水中分离噬菌体vB_Kp_XP4。通过透射电子显微镜（TEM）分析其形态。使用点滴试验和OD测量评估宿主范围、pH值和热稳定性。一步生长曲线确定潜伏期和裂解量。进行全基因组测序和系统发育分析。在感染模型中评估治疗效果和安全性。
TEM显示噬菌体vB_Kp_XP4是一种具有二十面体头部和长而灵活尾部的有尾噬菌体。它可裂解一株hvKP菌株（携带、、、基因）和一株MDR-KP菌株（对碳青霉烯类、氟喹诺酮类等耐药），最佳感染复数为0.1，潜伏期<10分钟。在pH 4-11和≤70°C条件下保持稳定。全基因组测序显示其线性双链DNA基因组为44344 bp，G+C含量为53.80%。该基因组包含54个编码序列，无溶原性、毒力或抗生素抗性基因。系统发育分析将噬菌体vB_Kp_XP4定位为科属内的一个新物种。在模型中，vB_Kp_XP4延长了P4感染幼虫的存活时间（<0.001）。
噬菌体vB_Kp_XP4具有高稳定性、特异性、强大的裂解活性且无不良基因，在体内显示出有效的治疗效果，表明其在临床应用中对抗感染的潜力。噬菌斑形成过程中多个晕圈的出现进一步提高了其研究价值。完整的基因组序列已提交至GenBank，登录号为PP663283。
