Avian Pathol.: Quantification of pigeon circovirus in serum, blood, semen, and different tissues of naturally infected pigeons using a real-time polymerase chain reaction.
Article Type: Brief article
Subject: Pigeons (Health aspects)
Polymerase chain reaction (Usage)
DNA viruses (Health aspects)
DNA viruses (Research)
Authors: Duchatel, J.P.
Todd, D.
Willeman, C.
Losson, B.
Pub Date: 09/01/2009
Publication: Name: Journal of Avian Medicine and Surgery Publisher: Association of Avian Veterinarians Audience: Academic Format: Magazine/Journal Subject: Health Copyright: COPYRIGHT 2009 Association of Avian Veterinarians ISSN: 1082-6742
Issue: Date: Sept, 2009 Source Volume: 23 Source Issue: 3
Topic: Event Code: 310 Science & research
Geographic: Geographic Scope: United States Geographic Code: 1USA United States
Accession Number: 252006993
Full Text: The development of a real-time polymerase chain reaction (PCR) based on SYBR Green chemistry is described for the quantification of pigeon circovirus (PiCV) DNA in various samples. Plasmid containing a fragment of the PiCV genome was used to create a standard curve and to estimate the viral DNA copies in analyzed samples. Both primers were designed in highly conserved regions to avoid false negatives, and amplified a 139-base-pair amplicon. When the amplifications were performed in the presence of cellular DNA extracted from PCR-negative liver, bursa, and spleen samples, the detection limits were respectively 20, 20, and 60 copies of genome per milligram of tissue. These limits were 10, 160, and 25 copies/[micro]l for control blood, sera, and semen, respectively. For cloacal swab, the detection limit was 200 copies. The assay showed a linear detection over a 6-log range ([R.sup.2] > .99) and displayed reliable interassay and intra-assay reproducibility. Application of the test to sera samples indicated the presence of the virus in Belgium in 1991, 6 years before PiCV infections were histologically diagnosed. Testing of samples from pigeons with "young pigeon sickness" showed that the viral loads were high in the bursa of Fabricius (up to 2.07 x [10.sup.9] copies/mg), the liver (up to 2.88 x [10.sup.8]copies/mg) and spleen (up to 5.57 x [10.sup.8]copies/mg). For liver, the viral load was significantly higher in sick pigeons than in apparently healthy pigeons. Detection of high quantities of PiCV DNA (up to 1.6 x [10.sup.9]copies/[micro]l) in the sera or blood of some young healthy pigeons indicated that the viral load in this sample type would not be useful as predictive indicator of disease. This work also showed that PiCV DNA can be detected in relatively large amounts in semen (up to 1.0 x [10.sup.7]copies/ejaculate) and cloacal swabs (up to 3.6 x [10.sup.10]copies/swab), confirming that PiCV may be transmitted by vertical and horizontal routes.

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