Detection of sheep poxvirus in skin biopsy samples by a multiplex polymerase chain reaction.
The development of a multiplex polymerase chain reaction (PCR) method with amplification of capripoxvirus in a single-step procedure from skin biopsies using three primer pairs, two specific for capripoxvirus and one specific for alpha-tubulin is described. A sensitive multiplex PCR was achieved by optimization of parameters such as the primer concentrations, magnesium and dNTPs concentrations. False negative results that sometimes arise due to inhibitors of DNA amplification may be avoided by the inclusion in the assay of alpha-tubulin primers. The results reported on 42 skin biopsies from sheep suspected to have poxvirus infection, indicated that the assay could monitor simultaneously DNA extraction from skin biopsy samples and allow improved detection of capripoxvirus within 24 h of specimen receipt in the laboratory.
Markoulatos
P
P
Department of Virology, Hellenic Pasteur Institute, Athens, Greece. vresearch@hol.gr
Mangana-Vougiouka
O
O
Koptopoulos
G
G
Nomikou
K
K
Papadopoulos
O
O
D000818:Animals; D001706:Biopsy; D018151:Capripoxvirus / Q000235:genetics / Q000302:isolation & purification*; D017931:DNA Primers; D004279:DNA, Viral / Q000032:analysis; D016133:Polymerase Chain Reaction / Q000379:methods*; D011213:Poxviridae Infections / Q000175:diagnosis / Q000662:veterinary* / Q000821:virology; D012680:Sensitivity and Specificity; D012756:Sheep; D012757:Sheep Diseases / Q000175:diagnosis* / Q000821:virology; D012867:Skin / Q000821:virology*
D017931:DNA Primers; D004279:DNA, Viral