Shiga toxin producing Escherichia coli is a contaminant of food and water that causes a diarrheal syndrome followed by more severe disease of the kidneys and symptoms of the central nervous system in humans. The isolation of Shiga toxin producing Escherichia coli (STEC) from diarrheic and apparently health calves is difficult due to lack of differential phenotypic characteristics from nonpathogenic Escherichia coli. The improvement of molecular technology allows identification of both toxin and serogroup specific genetic determinants. In this study, 300 fecal samples from diarrheic and apparently healthy calves were screened for STEC using PCR targeting Shiga toxin determinants. In addition routine culture methods for isolating O157 and non O157 STEC were also performed. The screening assays of serotyping isolates revealed 7 (4.1%) of O157H7, 156 (92.8%) of non O157 and 5 (3.1%) for untypable strains. These included STEC serotypes of O157H7 and O26 from diarrheic samples, and O78, O55 and O126 from apparently healthy calves. The high rate of STEC isolation and the diversity of STEC serogroup from calves focus the light on the importance of calves as reservoir of E. coli as well as motivate us to improve biosecurity measures in dairy farms.
Brucellosis, Leptospirosis, Mycoplasmosis and Listeriosis are important zoonosis and represent an important cause of reproductive losses in animals worldwide, especially in Mediterranean countries and Egypt. Aiming at improvement in the diagnostic scheme, a quick method for the simultaneous detection of these microorganisms in different clinical samples using a SYBR green multiplex real-time polymerase chain reaction (m RT-PCR) has been developed. The m PCR has been standardized by using 4-pairs of primers to amplify 31kDa gene encoding protein in Brucella spp., lig gene in pathogenic Leptospira, hlyA gene in Listeria monocytogenes and 16S rDNA in Mycoplasma spp. This study was applied to 161 different clinical samples (milk, blood, fetal fluids, semen, tissue and vaginal discharges). Real time PCR assay revealed a specific dissociation peak at Tm=79.0ºC, 80.0ºC, 85.2ºC and 88.0ºC (± 0.5) for Listeria, mycoplasma, brucella and Leptospira respectively. The real time assay neither revealed interferences between primers nor nonspecific florescence. In conclusion the high output, time saving and cost effective SYBR green m RT-PCR method established, could offer an effective tool for simultaneous, quick and reliable identification of these microbial agents in different clinical samples.
A total of 150 milk samples were collected from both acute and chronically infected cattle from both Giza and Minoufia governorates. Incidence of mycotic mastitis was at rate of 72% and 88%for Giza and Minoufia governorates respectively. Different mold and yeast species were isolated. The most isolated mold species from Giza samples were Aspergillus species (64.4%) followed by Geotrichum (15.2%)then Penicillium (10.2%), while in Minoufia governorate, the most isolated mold species were Penicillium at a percentage of (50.6%)followed by Aspergillus at a percentage of (40.2%). About yeast species isolation, C. parapsillosisis was the most isolated yeast species from Giza samples with percentage of 33.4%,while in Minoufia samples, C. tropicalis was the most isolated yeast species with a percentage of 22.7%. All of the isolates are identified depending on macroscopical and microscopical identification. Twelve isolated yeast strains were biochemically identified depending on rapaid yeast plus identification system, all of the tested strains were correctly identified except for C. parapsillosis strains, only 83%of the tested strains were correctly identified. Six negative examined milk samples with culture on Sabaroud dextrose agar media were subjected to Rt-PCR and Propidium mono azide stain, four of them were positive although they were negative on culture as they are considered as samples containing VBNC(Viable but non culturable) strains.
Salmonella is one of the most important causative agents of food poisoning and gastroenteritis in humans. This study spot highlights on isolation, identification and molecular characterization of salmonella serovars from imported frozen meat using the convential and modern molecular tools. Methods: A cross-sectional study was carried out on 100 samples of frozen meat collected from different supermarkets from Minufiya governorate, Egypt. Results: The prevalence of Salmonella were 6%. Serotyping of the obtained salmonella isolated revealed that Salmonella enteritidis, Salmonella typhimurium and Salmonella .paratyphi were the prevalent serotypes in the examined samples. S. typhimurium, only 3 samples (3%) ,Salmonella enteritidis was isolated from only 2 sample (2%) and S.paratyphi only 1 samples (1%). The application of conventional PCR for the six obtained isolates of Salmonella serotypes using universal gene (invA) was effective tool for identification and genotypic of pathogenic Salmonella serotypes. Conclusions: This study concluded that Salmonella is among the most important food borne pathogens worldwide contaminating a wide range of animal products including meat products. Also indicated that the cPCR was specific and rapid method for identification and genotyping of pathogenic salmonella serotypes.
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