Laboratory diagnosis of contagious ecthyma: Comparison of different PCR protocols with virus isolation in cell culture

• Published in: Journal of virological methods Vol. 134; no. 1-2; pp. 119 - 124
• Main Authors: Kottaridi, Christine, Nomikou, Kiki, Lelli, Rossella, Markoulatos, Panayotis, Mangana, Olga
• Format: Journal Article
• Language: English
• Published: London Elsevier B.V 01-06-2006; Amsterdam Elsevier; New York, NY
• Subjects: Animals; Biological and medical sciences; Cell culture; Contagious ecthyma; Diagnosis; DNA Primers; Drug Resistance, Viral; Ecthyma, Contagious - diagnosis; Ecthyma, Contagious - virology; Fundamental and applied biological sciences. Psychology; Greece; Interferons - pharmacology; Microbiology; Open Reading Frames - genetics; Orf virus; Parapox virus; Polymerase chain reaction; Polymerase Chain Reaction - methods; Poxviridae - drug effects; Poxviridae - genetics; Poxviridae - isolation & purification; Sensitivity and Specificity; Sheep; Techniques used in virology; Viral Envelope Proteins - genetics; Virology; Greece; Polymerase chain reaction; Parapox virus; Cell culture; Diagnosis; Contagious ecthyma; Virus; Microbiology; Isolation; Method; Virology
• ISSN: 0166-0934; 1879-0984
• DOI: 10.1016/j.jviromet.2005.12.005

A new polymerase chain reaction (PCR) assay for rapid diagnosis of contagious ecthyma was designed and applied to 21 clinical samples from Greece. This assay, which detects a highly conserved gene from the parapox genome, was evaluated for its sensitivity and specificity in order to be considered as a useful diagnostic tool. A comparative study with two published PCR protocols one using primers PPP1–PPP3, PPP1–PPP4 which targets putative virion envelope gene B2L and the other using VIR1–VIR2 primers which amplifies ORF virus interferon resistant (VIR) gene, as well as cell culture virus neutralization assay was carried out. All samples tested were amplified successfully with the PCR protocol established in the laboratory. The combination of primers PPP1–PPP3 and PPP1–PPP4 in a semi-nested PCR gave a positive result in 20 of 21 samples while primers VIR1–VIR2 failed to amplify successfully 7 of 21 samples. The diagnostic value of parapox viral DNA amplification was also compared with the results of virus isolation by cell culture and was positive in three samples that the virus isolation was obtained.