Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2013 Oct 18;197(1-2):85-94.
doi: 10.1016/j.vetpar.2013.04.024. Epub 2013 Apr 25.

Development of a multiplex real time PCR to differentiate Sarcocystis spp. affecting cattle

Gastón Moré  1 , Susann Schares, Aline Maksimov, Franz J Conraths, María C Venturini, Gereon Schares
Affiliations

Development of a multiplex real time PCR to differentiate Sarcocystis spp. affecting cattle

Gastón Moré et al. Vet Parasitol. .

Abstract

Cattle are intermediate hosts of Sarcocystis cruzi, Sarcocystis hirsuta and Sarcocystis hominis which use canids, felids or primates as definitive hosts (DH), respectively, and in addition of Sarcocystis sinensis from which the DH is unknown. The aims of the present study were to develop and optimize a multiplex real time PCR for a sensitive and specific differentiation of Sarcocystis spp. affecting cattle and to estimate the prevalence of Sarcocystis spp. in Argentinean cattle. The 18S rRNA genes from individual sarcocysts were amplified and cloned to serve as controls. For the amplification of bovine Sarcocystis spp. a total of 3 primers were used in combination with specific individual probes. Each assay was evaluated and optimized individually and subsequently combined in a multiplex assay (BovSarcoMultiplex real time PCR). The analytical specificity of the multiplex assay was assessed using 5 ng of DNA of heterologous Sarcocystis spp. and other apicomplexan parasites, and no positive reactions were observed other than for the species the PCR targeted. The analytical sensitivity ranged between 0.0125 and 0.125 fg of plasmid DNA (equivalent to the DNA of 2-20 plasmid DNA copies) or resembling DNA of 0.1-0.3 bradyzoites. A total of 380 DNA loin samples from Argentina were tested and 313, 29, 14 and 2 were positive for S. cruzi, S. sinensis, S. hirsuta and S. hominis, respectively. S. sinensis was the most prevalent species among thick walled Sarcocystis spp. in Argentinean cattle. Mixed infections were detected in 8.9% of all samples. Diagnostic sensitivity and specificity for the BovSarcoMultiplex real time PCR relative to previous microscopic examination for thin and thick-walled cyst were 91.5% and 41.7%, 36.3% and 95.9% respectively. Improved DNA extraction methods may allow to further increase the specific and sensitive detection of Sarcocystis spp. in meat samples.

Keywords: Argentina; Beef; Multiplex real time PCR; Sarcocystis spp..

PubMed Disclaimer

Publication types

LinkOut - more resources