Diagnosis of duck circovirus infections by conventional and real-time polymerase chain reaction tests.
Development of the first conventional and real-time polymerase chain reaction (PCR) tests for the diagnoses of duck circovirus (DuCV) infections is described. Both tests amplified a 230 bp fragment specific to the DuCV Rep gene. Although both tests had the same detection limit (13 x 10(3) target DNA/ml) when the target DNA was diluted in water, the detection limit of the real-time test (13 x 10(4) target DNA/ml) was 10-fold less than the conventional test (13 x 10(5) target DNA/ml) when the amplifications were performed in the presence of cellular DNA. Using the conventional PCR test, DuCV DNA was detected in 85 (84%) of 101 bursa of Fabricius samples from dead or sick ducks, aged between 1 and 12 weeks, and in samples from 35 (94%) of 37 flocks. Application of the SYBR Green-based real-time PCR test to 54 selected bursa of Fabricius samples indicated that more samples were positive by real-time PCR than by conventional PCR, allowed the numbers of genome copies to be estimated and showed that some bursa of Fabricius samples contained over 10(13) genome copies/g tissue. Although DuCV infections were detected in birds aged from 1 to 12 weeks, higher virus DNA levels were detected in ducks aged older than 5 weeks than in ducks younger than 5 weeks. An in situ hybridization method for the detection of DuCV in histological samples was also developed. Additional work is required to determine the clinicopathological significance of DuCV infections.
Dipartimento di Scienze Biopatologiche ed Igiene delle Produzioni Animali e Alimentari, Università degli Studi di Perugia, 06126, Perugia, Italy.
E
E
Fringuelli
A N J
AN
Scott
A
A
Beckett
J
J
McKillen
J A
JA
Smyth
V
V
Palya
R
R
Glavits
E
E
Ivanics
A
A
Mankertz
M P
MP
Franciosini
D
D
Todd
D000818:Animals; D002060:Bursa of Fabricius / Q000821:virology; D018173:Circoviridae Infections / Q000662:veterinary* / Q000821:virology; D004372:Ducks / Q000821:virology*; D016133:Polymerase Chain Reaction / Q000379:methods / Q000662:veterinary*; D011201:Poultry Diseases / Q000175:diagnosis* / Q000821:virology*; D012015:Reference Standards; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity