Morphological, biological, and genomic characterization of Klebsiella pneumoniae phage vB_Kpn_ZC2
Mohamed S. Fayez 1*, Toka A. Hakim 1, Bishoy Maher Zaki 1,2, Salsabil Makky 1, Mohamed Abdelmoteleb 3, Kareem Essam 1, Anan Safwat 1, Abdallah S. Abdelsattar 1, Ayman El-Shibiny 1,4
- Center for Microbiology and Phage Therapy, Zewail City of Science and Technology, Giza 12578, Egypt
- Microbiology and Immunology Department, Faculty of Pharmacy, October University for Modern Sciences and Arts (MSA), Giza 11787, Egypt
- Department of Botany, Faculty of Science, Mansoura University, Mansoura 35516, Egypt
- Faculty of Environmental Agricultural Sciences, Arish University, Arish 45511, Egypt
Mohamed S. Fayez and Mshokry@zewailcity.edu.eg
Background: Bacteriophages (phages) are an extremely promising alternative to conventional antibiotic therapies, particularly when combating bacteria that have developed resistance to multiple antibiotics. Klebsiella pneumoniae is recognized as an opportunistic pathogen capable of causing potentially fatal infections. Thus, this study aims at the characterization of a novel isolated phage vB_Kpn_ZC2 (ZCKP2, for short).
Methods: Using the clinical isolate KP/08 as a host strain, the phage ZCKP2 was isolated from sewage water. The isolated bacteriophage was purified, amplified, and subjected to various tests, including Pulse-Field Gel Electrophoresis (PFGE) to determine its molecular weight, transmission electron microscopy to examine its structure, analysis of its antibacterial activity against a range of Klebsiella pneumoniae hosts, stability studies, and whole genome sequencing.
Results: Phage ZCKP2 has been categorized as a siphovirus based on the results of Transmission Electron Microscopy. The phage genome size was estimated to be 48.2 kbp using Pulsed Field Gel Electrophoresis and phage sequencing. Additionally, the absence of genes related to lysogeny, antibiotic resistance, and virulence in the annotated genome suggests that Phage ZCKP2 is safe for therapeutic use. Based on the genome-based taxonomic analysis, Phage ZCKP2 belongs to a novel family that has not yet been formally classified.
Furthermore, Phage ZCKP2 was found to remain stable at different temperatures and pH ranges (-20 − 70 °C and pH 4 – 9). In terms of antibacterial activity, Phage ZCKP2 consistently produced clear zones on KP/08 bacteria and other hosts and effectively killed bacteria over time at different MOIs (0.1, 1, and 10). Additionally, the genome annotation predicted the presence of antibacterial lytic enzymes. The predicted topology of class II holins in some putative proteins with dual transmembrane domains indicates their significant contribution to antibacterial activity. The characterization of Phage ZCKP2 demonstrates its safety and effectiveness against multidrug-resistant K. pneumoniae, thus making it a strong candidate for further clinical applications in vivo and phage therapy.