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Molecular detection of Staphylococcus aureus and its toxins in refrigerated milk from dairy cows and at room temperature

Grant number: 06/05125-2
Support Opportunities:Scholarships in Brazil - Master
Effective date (Start): September 01, 2007
Effective date (End): May 31, 2009
Field of knowledge:Agronomical Sciences - Veterinary Medicine - Preventive Veterinary Medicine
Principal Investigator:Helio Langoni
Grantee:Patrícia Yoshida Faccioli Martins
Host Institution: Faculdade de Medicina Veterinária e Zootecnia (FMVZ). Universidade Estadual Paulista (UNESP). Campus de Botucatu. Botucatu , SP, Brazil


Mastitis is the most important disease in dairy herds because has high prevalence, causes damages on production, modifies the gland mammary function and the animal health. It is a threat to public health because can transmit pathogens to human. The principal bovine mastitis agent is Staphylococcus aureus. These bacteria are very important to public health because produces enterotoxins related to toxin shock, food poisoning, allergies and autoimmune diseases. These enterotoxins are not inactived by proteolytic enzymes, and are active after ingestion. They are not affected by microorganism or food enzyme. The enterotoxins are heat-resistant, resisting to pasteurization and ultrapasteurization, remaining in milk and milk products. The significant rise of genetic methods to detection and characterization of pathogenic bacteria in food allowed the establishment of a viable alternative regarding traditional culture methods to microbiology diagnosis. This because it has more typifying and description capacity, speed, good limit of detection, good selectivity, specificity, potential to automation and possibility to work with no cultivable bacteria. One hundred samples will be collected from bulk tanks of infected herds with Staphylococcus aureus. These samples will be isolated on Baird-Parker agar to counting CFU and later biochemical identification. The positive milk to Staphylococcus aureus will be divided in two parts. One will be refrigerated and another will be stored at 25ºC, at bacteriological chamber, for 5 hours. In both samples, it will be detected by research, using Polymerase Chain Reaction PCR, encoding genes to enterotoxins sea, seb, sec and sed, and by Reverse Passive Latex Agglutination (RPLA) the enterotoxins A, B, C and D. The present study has as goal to compare the PCR sensitivity and specificity with microbiology exams in identification of Staphylococcus aureus in bulk tank milk. Results obtained will provide important data to standardization the use of PCR and RPLA directly from milk, without be necessary microbiologic isolation to Staphylococcus aureus. Moreover, it will be determinated risks to public health with the growth of enterotoxigenic Staphylococcus aureus in milk stored on field on bad conditions of refrigeration.

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