A real-time PCR to detect and analyze virulent EMCV loads in sows and piglets

• Published in: Molecular biology reports Vol. 39; no. 12; pp. 10013 - 10017
• Main Authors: Wang, Zhao, Liu, Yebing, Lin, Wencheng, Cui, Shangjin
• Format: Journal Article
• Language: English
• Published: Dordrecht Springer Netherlands 01-12-2012; Springer Nature B.V
• Subjects: Animal Anatomy; Animal Biochemistry; Animals; Biomedical and Life Sciences; Cardiovirus Infections - diagnosis; Cardiovirus Infections - veterinary; Cardiovirus Infections - virology; DNA, Viral - genetics; Encephalomyocarditis virus - genetics; Female; Histology; Hogs; Life Sciences; Molecular biology; Morphology; Organ Specificity; Polymerase chain reaction; Pregnancy; Pregnancy Complications - diagnosis; Pregnancy Complications - veterinary; Pregnancy Complications - virology; Real-Time Polymerase Chain Reaction - standards; Reference Standards; Reproducibility of Results; Sensitivity and Specificity; Sus scrofa; Swine; Swine Diseases - diagnosis; Swine Diseases - virology; Viral infections; Viral Load; Virus distribution; Sow; EMCV; Piglets; Real-time PCR
• ISSN: 0301-4851; 1573-4978
• DOI: 10.1007/s11033-012-1870-y

A real-time polymerase chain reaction with SYBR Green was developed for the detection and quantification of encephalomyocarditis virus (EMCV) in porcine tissues; the method uses two primers specific for the 3D gene. The detection limit of this assay was 22 gene copies/reaction, equivalent to 0.001 TCID
50
/ml. The assay was linear over a 10
7
dilution range of template concentrations and was specific for EMCV; it did not amplify other porcine pathogens (porcine circovirus 2, porcine reproductive and respiratory virus, classical swine fever virus, pseudorabies virus, or porcine teschovirus). This assay detected EMCV titers at least 10
4
smaller than the routine PCR assay. To increase our understand of EMCV pathogenesis, the new method was used to quantify levels of EMCV genome in various tissues of artificially challenged sows and piglets. The virus was found mainly in the heart, lung, spleen, kidney, and endometrium of sows, and mainly in the heart, spleen, lung, and testis of fetuses. The real-time PCR method described here should be useful for the study of EMCV infection and distribution in pigs.