Isolation of a broad spectrum of bacteriophage action on enteropathogenic E. coli and study of the effect of food decontamination in model experiments

Abstract The incidence of diseases caused by enteropathogenic Escherichia coli has significantly increased. In addition, the emergence and worldwide spread of antibiotic-resistant strains has become the most important problem for mankind. Therefore, the search for new alternative drugs is inevitable, timely and unavoidable. Bacteriophages - bacterial viruses are the most optimal and harmless means against infectious processes caused by microorganisms. Besides, phage therapy has many advantages compared to antibiotics. For example, the effectiveness against antibiotric resistant pathogens, strict specificity to target bacteria, so that they do not to damage the normal microflora, etc. Along with the therapeutic use of bacteriophages, they are widely used as for bio-sanitary purposes. For example, they are used for the decontamination of meat and seafood, raw fruit-vegetable and other foods, as well as for disinfection of the surfaces, which are in contact with contaminated food. After the FDA approval the first phage-based product (ListShield) on August 18, 2006, the interest towards development of new bacteriophages preparations which insure food safety has gradually increased. Consequently, the research has been actively ongoing which is aiming isolation of the new phages and their uses for different purposes, such as therapeutic, biocontrolling and decontaminating means. The aim of our study was isolation of the broad spectrum phages against enteropathogenic E.coli and evaluation of their decontaminating effect in the model experiments. For this purpose the following tasks have been accomplished: 1) Contamination level of the products (cheese, matsoni, sausages, minced meat, etc.) existing on the Georgian market has been assessed; 2) The new E.coli- strains have been isolated, identified and their antibiotic resistance profiles was determined; 3) The new bacteriophages specific to these strains have been isolated and characterized; 4) The newly isolated bacteriophages have been used in the model surface biocontrol studies. As a result these experiments out of 25 samples we have isolated 20 bacterial isolates among which due to traditional bacteriology (growth on selective media) and molecular identification methods (PCR, gel electrophoresis) 10 strains have been identified as E.coli. According to antibiotic susceptibility tests, it was determined that out of 10 strains 1 (10%) appeared to be resistant to spectinomycin, 5 (50%) – to tetracycline, 4 (40%) – amoxicillin, 2 (20%) – kanamycin, 6 (60%) - ampicillin, 1 (10%) - amikacin, 1 (10%) – gentamicin, 4 (40%) - chloramphenicol, 4 (40%) – ofloxacin, 4 (40%) – erythromycin, 3 (30%) – ciprofloxacin. Thus, it was revealed that all 10 E.coli strains showed multiple drug resistance and were only partly susceptible in one or more antibiotics. In course of research we have isolated three E,.coli specific phages, two oyt of them appeared to belong to Myoviridae morphological family, and one to - Podoviridae-ს. Phage host range activity was evaluated using streak method. Based on the obtained results the selected phages have been used in the model experiments to evaluate their bio-controlling effectiveness against contaminated surface and packaging materials. Thus, we can conclude that the E.coli strains isolated in the scope of this research are characterized with multiple drug-resistance. At the same time the newly isolated phages demonstrated broad activity against these antibiotic resistant strains. Consequently, it is possible to develop a commercial product prototype, which would be directed to elimination of E.coli associated contamination and biocontrol of the food products, packaging materials and/or surfaces. The presented study was performed at the R & D Department of the George Eliava Institute of Bacteriophages, Microbiology and Virology.