用于通过实时聚合酶链反应诊断马立克氏病及监测马立克氏病疫苗接种的血液和羽毛髓样本比较

Comparison of blood and feather pulp samples for the diagnosis of Marek's disease and for monitoring Marek's disease vaccination by real time-PCR.

作者信息

Cortes Aneg L, Montiel Enrique R, Lemiere Stephane, Gimeno Isabel M

机构信息

Population Health and Pathobiology Department, College of Veterinary Medicine. North Carolina State University, 4700 Hillsborough Street, Raleigh, NC 27606, USA.

出版信息

Avian Dis. 2011 Jun;55(2):302-10. doi: 10.1637/9578-101510-ResNote.1.

DOI:10.1637/9578-101510-ResNote.1

PMID:21793449

Abstract

Comparison of blood and feather pulp (FP) samples for the diagnosis of Marek's disease (MD) and for monitoring Marek's diseases vaccination in chickens (serotypes 2 and 3 vaccines) by real time-PCR was evaluated. For diagnosis of MD, quantification of serotype 1 Marek's disease virus (MDV) DNA load was evaluated in 21 chickens suffering from MD. For each chicken, samples of blood and FP were collected and MDV DNA load was quantified. Solid tumors are the sample of choice for MD diagnosis by real time-PCR and, hence, 14 solid tumors were included in the study as positive controls. Load of MDV DNA in FP was equivalent to that detected in solid tumors (threshold cycle [Ct] ratio above 1.7). MDV DNA load in blood samples was lower than in solid tumors and FP samples. Nonetheless, there was a statistically significant correlation of the results obtained from FP and blood (r = 0.92). Results of the Pearson correlation test showed that Ct ratio values of 1.7 in FP correspond to Ct ratio values of 1.2 in peripheral blood. For monitoring vaccines, serotypes 2 and 3 MDV DNA load was evaluated in blood and FP samples of vaccinated chickens. Serotype 2 MDV DNA load was evaluated in samples of blood and FP from 34 chickens vaccinated with SB-1 strain. Serotype 3 MDV DNA load was evaluated in blood and FP samples from 53 chickens vaccinated with HVT strain. For both serotypes, frequency of positive samples and load of vaccine DNA was higher in FP than in blood samples. There was not a statistically significant correlation between the load of SB-1 DNA (r = 0.17) or HVT DNA (r = -0.04) in FP and blood. Our results show that the load of serotypes 1, 2, and 3 DNA is higher in FP than in blood. Diagnosis of MD could be done using both FP and blood samples. Monitoring of MD vaccination by real time-PCR required the use of FP samples. There were a high percentage of false negative samples when using blood to detect serotypes 2 and 3 MDV by real time-PCR.

摘要

评估了通过实时荧光定量聚合酶链反应（real time-PCR），比较血液和羽毛髓（FP）样本用于诊断鸡马立克氏病（MD）以及监测鸡马立克氏病疫苗接种情况（2型和3型疫苗）。为诊断MD，对21只患MD的鸡评估了1型马立克氏病病毒（MDV）DNA载量的定量。对每只鸡采集血液和FP样本并对MDV DNA载量进行定量。实体瘤是通过实时荧光定量聚合酶链反应诊断MD的首选样本，因此，本研究纳入14个实体瘤作为阳性对照。FP中MDV DNA载量与实体瘤中检测到的相当（熔解曲线分析比值[Ct]高于1.7）。血液样本中MDV DNA载量低于实体瘤和FP样本。尽管如此，FP和血液样本所得结果存在统计学显著相关性（r = 0.92）。Pearson相关检验结果显示，FP中Ct比值为1.7对应外周血中Ct比值为1.2。为监测疫苗，对接种疫苗鸡的血液和FP样本评估2型和3型MDV DNA载量。对34只接种SB-1株疫苗鸡的血液和FP样本评估2型MDV DNA载量。对53只接种火鸡疱疹病毒（HVT）株疫苗鸡的血液和FP样本评估3型MDV DNA载量。对于两种血清型，FP中阳性样本频率和疫苗DNA载量均高于血液样本。FP和血液中SB-1 DNA载量（r = 0.17）或HVT DNA载量（r = -0.04）之间无统计学显著相关性。我们的结果表明，1型、2型和3型DNA载量在FP中高于血液。可使用FP和血液样本进行MD诊断。通过实时荧光定量聚合酶链反应监测MD疫苗接种需要使用FP样本。通过实时荧光定量聚合酶链反应使用血液检测2型和3型MDV时存在高比例假阴性样本。