engleski

Use of modern molecular biology method to identify contagious mastitis-causing organisms: comparison of Real-Time PCR Assay and traditional method.

Mastitis is the most frequently occurring and economically the most important infectious disease in dairy cattle. Usually it is caused by one primary pathogen, and approximately 10 bacterial species or species groups account for more than 95% of all clinical and subclinical infections. Reliable identification of the causal bacteria is important for targeting antimicrobial therapy. This is only possible with accurate identification of bacteria in the mastitic milk samples. According to the literature, no bacterial growth is detected in at least 20 to 30% of milk samples taken from udder quarters with clinical mastitis and even more from those with subclinical mastitis. This study was a field trial on 68 milk samples from clinically healthy cows that were analyzed with both bacterial culture method and real-time PCR-based assay (PathoProof Mastitis Complete-16 assay) that identifies 15 pathogen species or species groups responsible for intramammary infections, and a gene coding for staphylococcal β-lactamase production (penicillin resistance). The aim was to provide a validation of the analytical specificity and sensitivity of the real-time PCR-based assay in comparison to the traditional method. With the bacterial culture method, from 68 milk samples, 8 were positive on Staphylococcus aureus. According to the real-time PCR-based assay, from this 8 samples 4 were detected positive for S. aureus, while the other 4 were positive for Staphylococcus spp. Out of the 60 samples that showed negative on bacterial culture, with the real-time PCR-based assay S. aureus was detected in 9 more samples. Other than S. aureus, the most commonly detected were Staphylococcus spp., Enterococcus spp., yeast and the gene coding for staphylococcal β-lactamase production. The real-time PCR assay shows excellent analytical specificity and sensitivity and holds much promise as a routine tool in diagnosing mastitis pathogens. All targeted species are detected simultaneously, overcoming the common problem of mixed growth results of bacterial culturing. Studies indicate superior analytical specificity over traditional methods, even for samples that show “no growth” in culture. It is also a useful tool to resolve bacterial etiology of milk samples with no growth in conventional culturing.

mastitis pathogen; real-time PCR-based mastitis assay; bacterial culture method; Staphylococcus aureus

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