耐碳青霉烯类肠炎沙门氏菌噬菌体解聚酶Dpo52的鉴定与表征

Abstract

Salmonella enteritidis , a gram-negative bacterium, is a common cause of human foodborne diseases. Multidrug-resistant strains are increasing year by year, especially the carbapenem resistant isolates. One of the reasons for antibiotic resistance is the formation of bacterial biofilm. But reports on phage depolymerases against carbapenem resistant S. enteritidis are scanty, which underlines the importance of the study. In this study, we characterized a phage that can infect carbapenem resistant S. enteritidis , vB_Sen_S 3 P. A phage was isolated from sewage water in Changchun, China, its genome was analyzed, and its depolymerases were expressed and purified. The host specificity and activity of the depolymerases were determined. The isolated carbapenem resistant S. enteritidis phage, vB_Sen_S 3 P, infects 22 out of 30 tested clinical isolates of S. enteritidis , including carbapenem-resistant S. enteritidis S 25 and S 27 . The highest phage titer was observed at a multiplicity of infection of 10 − 5 and a burst size of approximately 16,931 PFU/infection. Transmission electron microscope micrographs indicated that vB_Sen_S 3 P had an icosahedral head and a short non-shrinkable tail. Based on a comparative analysis, the complete genome of vB_Sen_S 3 P was 43,620 bp with a GC content of 49.76%. PSI-BLAST analysis indicated that ORF52 of vB_Sen_S 3 P (Dpo52) shares low identity (≤ 41.4%) with other reported Klebsiella phage depolymerases. Dpo52 was expressed and purified as a soluble protein using E. coli BL21. Dpo52 was stable at pH 4–11 and in a temperature range of 4 to 60℃.Through the cytotoxicity test, Dpo52 was observed non-cytotoxic to macrophages. Dpo52 inhibited the biofilm formation of S. enteritidis S 25 and S 32 via its enzymatic activity of extracellular polysaccharide degradation. To our knowledge, this is the first study of a S. enteritid