Isolation And Dentification Of Enterococcus Faecalis In Cattle Pyometra Pus

Recently, by investigating the causes of uterine infections of cows, many studies showedmultiple pathogens were isolated from endometrial secretion and pyometra, including mostfrequently isolated strains such as Enterococcus faecalis（Ef）, Streptococcus, E.coli andStaphylococcus aureus. And this indicated Mixed infection of all these four pathogens was themain significant reason of pyometra. But Enterococcus faecalis is opportunistic andparticularly resistant to environment as resident flora of intestinal tract of humans and animals,so it was difficult to prevent and cure in clinic. In this research, as a case study ofenterococcus isolate of cattle pyometra, which have been confirmed as Enterococcus faecalisby biological characteristics and16SrDNA testing, infected mice to find out virulence andpathogenesis, and analyzed difference of cell wall proteins by SDS-PAGE after extracting cellwall, it provide study foundation of pathogenesis and immune response.Firstly in this experiment the isolates was gram-positive enterococcus and negative inperoxidase from pyometra of infected cattle farm in Tianjin. The isolated were identified as Dgroup by serology and physiological biochemical characters, and the characters conform toEnterococcus faecalis, so it iwas preliminary decided to Enterococcus faecalis. Meanwhilethe result of16SrDNA testing showed it was99.6%homologous identity to Enterococcusfaecalis OG1RF strains in Genbank. The sequence analysis determined that the isolated isEnterococcus faecalis. The drug sensitivity test showed that the strains were resisted topenicillin, vancomycin, tetracycline, but were sensitive to ciprofloxacin, ampicillin andcephalosporin. Mice were challenged to compare to clinical symptoms, necropsy, mortalityand the number of isolated bacteria from infected tissues, then the optimal route of infectionwas determined as intraperitoneal injection. And infected mouse models with intraperitonealinjection were established for exploring the virulence of Enterococcus faecalis to mice. Theresult showed mice died within20h to7d after infected, and there were symptoms ofhypopnea, diarrhea, twitch. The characteristic changes at necropsy were bleeding under theskin, liver, spleen, and kidney causing varying degrees of swelling, gallbladder filling withbile, but no significant changes were observed in lung and brain. One of the mice wasobviously congested in liver, two of them has pericardial effusion and spotted and focalnecrosis in spleen. Meanwhile, LD50of cattle is5.40×10¹⁰CFU through mortality of infectedmice, and there was little difference to the mutton and duck stains. The cell wall proteins were analyzed by SDS-PAGE after purifying by ultrasonic wave and ammonium sulphateprecipitation. The result showed there were dozens of different bands on the cell wall proteingel map of Enterococcus faecalis, obviously a single band with molecular weight of67kDwas in common with different animal-derived strains.