SELECTION OF THE ACTIVE PHAGES AGAINST B.FRAGILIS FOR FURTHER STUDY OF THRAPEUTIC PERPECTIVES

B.fragilis is an obligate anaerobic commensal colonizing human intestines and carries number of physiological functions. At the same time B.fragilis is commonly isolated from the septic clinical samples and due to its capsule represents one of the provoking agents for abscess development. Enterotoxigenic B.fragilis (ETBF) strains also increase the likelihood of developing colon cancer. Increasing incidence of antibiotic-resistant pathogens led to the high demand to alternative antimicrobials. Bacteriophage (phage) therapy already practiced for a century in some of the Post-Soviet countries including Georgia has been suggested as a substitute of antibiotics. It should be noted that this study is the first attempt to isolate virulent B.fragilis phages for further therapeutic application as all phages known up until now were used for detection of fecal water contamination only. The aim of the study was to isolate B.fragilis specific phages for their further use against infections caused by this bacteria Eighteen B.fragilis strains were isolated from human feces using conventional microbiological methods and precise identification was done via MULDI-TOF mass spectrometry. Three ETBF strains were provided by the University of Ghent (Belgium). Three lytic phages (ФVA7, ФMTK and ФUZ-1) of Siphoviridae family were isolated from the waste water samples collected in Tbilisi and in Ghent using conventional phage isolation and enumeration techniques. Electron microscopy was used for the visualization of the phage particles. To determine lytic activity of the isolated phages and estimate their antimicrobial efficacy the spot test assay and efficiency of plating (EOP) were studied using 18 clinical strains of B.fragilis and 12 intestinal commensal strains related to Bacterioides spp. and Parabacterioides spp.. Although according to the spot test results two of the isolated phages expressed high specificity to B.fragilis demonstrating broad host range within this species, however EOP results showed that only ФVA7 can be selected as the best candidate for the model in vitro tissue culture experiments aiming demonstration of the therapeutic and prophylactic potential of phages against ETBF and/or NETBF.