Molecular detection of Sarcocystis spp. in tissue samples can be useful for experimental and diagnostic purposes. However, the parasite spreads unevenly through tissues, forming tissue
cysts, and the cystic wall is an obstacle in
DNA extraction protocols. Therefore, adequate sampling and effective disruption of the
cysts are essential to improve the accuracy of
DNA detection by PCR. The aims of this study were to evaluate the suitability of four protocols for
DNA extraction from
cysts of Sarcocystis spp. present in bovine myocardium samples or after their harvest in
phosphate-buffered saline (PBS)
solution as well as determine the effects of single or multiple sampling on the accuracy of molecular diagnosis of
sarcocystosis in cattle hearts.
Cysts and myocardium samples from nine bovine hearts were randomly distributed to four
DNA extraction protocols: kit, kit with modification, DNAzol, and cetyl-trimethyl
ammonium bromide (
CTAB). Samples were submitted to
DNA extraction and PCR as replicates of each heart (simplicate, duplicate, and triplicate), and the probability of a true positive diagnostic was calculated. Among the protocols tested, the kit with modification was determined to be the most suitable for
DNA extraction from
cysts in PBS
solution (92.6 % of
DNA detection by PCR); DNAzol resulted in higher
DNA detection frequency from bovine myocardium samples (48.1 %). Multiple sampling improved the molecular diagnosis of Sarcocystis spp.
infection in cattle hearts, increasing at 22.2 % the rate of true positive diagnostic.