Diagnosis of Marek's disease by real time PCR from samples ...
Diagnosis of Marek's disease by real time PCR from samples ...
Diagnosis of Marek's disease by real time PCR from samples ...
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Materials and Methods• Samples were collected <strong>from</strong> three differentchallenge studies at different <strong>time</strong>s after infection• Collection <strong>of</strong> <strong>samples</strong>: Original and in FTA® cardsBlood Feather Pulp Tumors56 36 14
Material and Methods• Real <strong>time</strong> <strong>PCR</strong>:SYBR® green (GAPDH)Taq Man Probe (gB, pol2, HVT)• Primers:MDV serotype 1 (gB)MDV serotype 2 (pol2)MDV serotype 3 HVT gene (spanning internal repeat regions)Chicken genome (GAPDH)• Statistical analysis: Correlation test <strong>of</strong> Pearson
Material and Methods
Real <strong>time</strong> <strong>PCR</strong>Ratio=Spt1Ratio=Stp2Ratio=Stp3Ct(GAPDH)Ct(gB)Ct(GAPDH)Ct(pol2)Ct(GAPDH)Ct(HVT)ThresholdCt
Use <strong>of</strong> FTA cards to measure viral DNA load(serotype 1) in blood and feather pulpFTA®GAPDH/gB FTA = .01209 + .99970 * GAPDH/gB orgnl.Correlation: r = .99987 p
Use <strong>of</strong> FTA cards to measure viral DNA load(serotype 3) in blood and feather pulpFTA®1.21.00.80.60.40.20.0GPDH/HVT FTA = .29615 + .09697 * GAPDH/HVT org.Correlation: r = .0852856 56 <strong>samples</strong> <strong>samples</strong>1.41.21.00.80.60.40.20.0GAPDH/HVT FTA = -.1043 + .94784 * GAPDH/HVT orgCorrelation: r = .49598 p
Use <strong>of</strong> FTA cards to measure viral DNA load(serotype 1) in tumors2.2GPDH/gB FTA = .10263 + .80534 * GPDH/gB orgCorrelation: r = .80333 p
Conclusion• Correlation between the load <strong>of</strong> MDV DNA inoriginal <strong>samples</strong> and <strong>samples</strong> taken in FTA®cards is statistically significant for all threeserotypes. Exception HVT in blood
Practical applications• Imprints <strong>of</strong> tumors taken in FTA® cards can beused for the diagnosis <strong>of</strong> MD• Samples <strong>of</strong> blood taken in FTA® cards can be usedfor early diagnosis <strong>of</strong> MD (wild type virus) and formonitoring serotype 2 MD vaccines• Samples <strong>of</strong> feather pulp taken in FTA® cards canbe used for monitoring serotypes 1, 2, and 3
AcknowledgmentsRicardo CortesTom CiminoThis study was funded <strong>by</strong> Merial Select Inc.
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