Characterization of Pseudomonas lytic phages and their application as a cocktail with antibiotics in controlling Pseudomonas aeruginosa
Section snippets
Bacteria, culture media and growth condition
Standard strain P. aeruginosa PAO1 was used in all experiments, unless otherwise stated. All experiments were conducted in Luria–Bertani (LB) broth (10 g polypeptone, 10 g sodium chloride and 5 g yeast extract per liter) at 37 °C, with shaking at 120 rpm, unless otherwise stated.
Isolation and preparation of phage stock
Two lytic phages, ϕPA01 and ϕPA02, were isolated from sewage influent obtained from the municipal wastewater treatment plant in Tokyo using P. aeruginosa PAO1 as the host by double layer agar plating method. Phages were
Morphology of phage ϕPA01 and ϕPA02
Based on the morphology of ϕPA01 and ϕPA02 observed (Fig. 1A–D), they were classified to the Myoviridae family. Both phages displayed a capsid head connected to a long contractile tail (Fig. 1B and D, indicated by black arrows). ϕPA02 showed a distinct morphology with a large capsid with the diameter and height of 121 ± 3 nm and 134 ± 4 nm, respectively. This feature is similar to reported giant phages (37). The relaxed tail fiber of ϕPA02 was 206 ± 7 nm long and 25 ± 1 nm wide. Meanwhile, the
Genomic characterization of novel phages
A well characterized phage is one of the key factors determining the success of a phage therapy (18). Phages from the genera Pbunalikevirus and Phikzlikevirus were able to control the growth and biofilm formation of P. aeruginosa in vitro (42) and in vivo (43,44) in previous studies. Genomic analysis has revealed the absence of integrase, genes associated with toxin and virulence in ϕPA01 and ϕPA02, suggesting that they are potentially safe candidates for phage therapy (18). Previous study
Acknowledgments
We are thankful to Professor Longzhu Cui, Dr. Shinya Watanabe and Dr. Kitaro Kiga (Jichi Medical Hospital University, Department of Bacteriology) for kindly supporting us in conducting the experiment using the clinical strains collection from Jichi Medical University Hospital. S.P. Ong received scholarship from Ministry of Education, Culture, Sports, Science and Technology of Japan during her post graduate study. All authors declared that there are no conflicts of interest in this article. This
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