Use of PCR for detection of bovine tuberculosis bacillus in milk of positive skin test cows
| Autor: | Vinicius Silva Castro, Vânia Margaret Flosi Paschoalin, Dandara Virginia Guia Semedo Fernandes, Greika Ferreira Moura, Elis Santos Soares, Eduardo Eustáquio de Souza Figueiredo, Ricardo César Tavares Carvalho |
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| Rok vydání: | 2014 |
| Předmět: |
Detection limit
medicine.medical_specialty Mycobacterium bovis Tuberculosis General Veterinary food and beverages Gold standard (test) Biology biology.organism_classification Isolation (microbiology) medicine.disease Proteinase K Surgery Microbiology chemistry.chemical_compound chemistry In vivo medicine biology.protein Lysozyme |
| Zdroj: | Brazilian Journal of Veterinary Research and Animal Science. 51:42 |
| ISSN: | 1678-4456 1413-9596 |
| DOI: | 10.11606/issn.2318-3659.v51i1p42-48 |
| Popis: | The causative agent of bovine tuberculosis (BTB) is Mycobacterium bovis, a bacterium belonging to the M. tuberculosis complex (MTC). The definitive diagnosis is achieved through isolation and identification of M. bovis from clinical samples, using a combination of traditional culture and biochemical methods, which is considered the “gold standard”. This procedure is cumbersome and time-consuming. We evaluated a PCR assay for the direct detection of MTC DNA in milk of positive skin test cows, using primers that were previously tested and proven reliable to target the IS6110 element. Milk previously seeded with M. bovis was used as the starting material, for standardization of the technique. The procedure involved extracting the DNA by enzymatic lysis (proteinase K and lysozyme), phenol, chloroform, isoamyl alcohol, followed by ethanol precipitation and PCR. The PCR assay allowed us to detect BTB in artificially contaminated milk, with a detection limit of 100 CFU/mL, and was also able to detect the bacillus in 50% (75/150) of the milk samples tested. This procedure could be used to assist the in vivo diagnosis of BTB, complementing sorological or microbiological tests, becoming an alternative option for epidemiological studies of BTB transmission and preventing contaminated milk from entering the food supply. |
| Databáze: | OpenAIRE |
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