Real-time PCR to detect and analyze virulent PPV loads in artificially challenged sows and their fetuses.
To establish a real-time polymerase chain reaction with SYBR Green for detection and quantification of porcine parvovirus (PPV) in porcine tissues, two primers specific for the non-structural protein 1 gene were designed. The detection limit of this assay was 3-23 gene copies/reaction, equivalent to 0.001 TCID(50)/ml. The assay was linear over a 10(6) dilution range of template concentrations. Other porcine pathogens involved in reproductive disorders (porcine circovirus 2, porcine reproductive and respiratory virus, pseudorabies virus, classical swine fever virus) were negative by this assay. This assay could detect PPV titres at least 10(5) smaller than the hemagglutination assay. To better understand the pathogenesis of PPV, the levels of viral DNA in various tissues of artificially challenged sows and their fetuses were quantified with this method. 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. This study provides a new tool for the study of PPV infection and distribution in sows and their fetuses.
Division of Swine Infectious Disease, State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute of Chinese Academy of Agricultural Sciences, Harbin 150001, China.
Lan-fei
LF
Miao
Chao-fan
CF
Zhang
Chang-mu
CM
Chen
Shang-jin
SJ
Cui
D000818:Animals; D000956:Antigens, Viral / Q000032:analysis; D017931:DNA Primers; D005260:Female; D005313:Fetal Death / Q000662:veterinary / Q000821:virology; D005784:Gene Amplification; D005814:Genes, Viral; D006801:Humans; D018445:Infectious Disease Transmission, Vertical / Q000662:veterinary*; D008168:Lung / Q000196:embryology / Q000821:virology; D008431:Maternal-Fetal Exchange; D010322:Parvoviridae Infections / Q000635:transmission / Q000662:veterinary*; D028323:Parvovirus, Porcine / Q000235:genetics / Q000472:pathogenicity*; D016133:Polymerase Chain Reaction / Q000379:methods / Q000662:veterinary; D011247:Pregnancy; D011251:Pregnancy Complications, Infectious / Q000662:veterinary* / Q000821:virology; D013552:Swine; D013553:Swine Diseases / Q000175:diagnosis / Q000196:embryology / Q000821:virology*
D000956:Antigens, Viral; D017931:DNA Primers