Bovine mastitis agents identification and counting using polimerase chain reaction

• Grant number: 07/08557-3
• Support Opportunities: Regular Research Grants
• Duration: April 01, 2008 - November 30, 2010
• Field of knowledge: Agronomical Sciences - Veterinary Medicine - Preventive Veterinary Medicine
• Principal Investigator: Marcos Veiga dos Santos
• Grantee: Marcos Veiga dos Santos
• Host Institution: Faculdade de Medicina Veterinária e Zootecnia (FMVZ). Universidade de São Paulo (USP). São Paulo , SP, Brazil

Abstract

Bovine mastitis is considered the most important disease in dairy cattle worldwide, and determines the major economic losses either to dairy producers and industry, as it negatively impacts milk yield and quality. Most of intramamary infections are caused by bacteria pathogens and, once affected, it might undergo to a chronic end. 80% of mastitis' bacteria causing pathogens are Escherichia coli, Streptococcus uberis, Streptococcus dysgalatiae, Streptococcus agalactiae and Staphylococcus aureus, as so S. agalactiae and S. aureus are contagious, which require precise identification in order to establish adequate means to control spread of pathogens in livestock and treatment of infected cows. Identification methodology nowadays are based on bacterial plate culturing, followed by phenotypic characterizing, biochemical tests and enzymatic profile. However, main disadvantages rely on false negative tests, which might be a result of the pathogen growth inhibition due to an antibiotic residue in sample; low detection level of pathogens in milk sample or leukocyte presence of clinical cases' samples. Additionally, the suitability of a pathogen detection methodology is dependent on factors such as specificity, sensitivity, cost and analysis time consuming, and its applicability to routine large quantity of milk samples. Though, reasons for carrying the project rely on the need of a low-cost, rapid, specific and sensitive methodology to diagnose subclinical mastitis due to contagious pathogens, S. aureus and S. agalactiae, by quantitative polymerase chain reaction. For this reason, the aims of this study are to establish a methodology routine assay for subclinical mastitis causing pathogens counting and identification (S. aureus and S. agalactiae) from milk sampled on three different ways (aseptically sampled from the affected mammary gland, aseptically sampled from glands, and non-aseptically sampled), comparing to traditional plate culture for pathogen identification. Expected results are on determining specificity and sensitivity of proposed quantitative PCR methodology, which might allow precise decision on means to control spread of pathogens in livestock and treatment of infected cows. (AU)