Gimeno Isabel M, Cortes Aneg L, Silva R F
Population Health and Pathobiology Department, College of Veterinary Medicine, North Carolina State University, 4700 Hillsborough Street, Raleigh, NC 27606, USA.
Avian Dis. 2008 Jun;52(2):203-8. doi: 10.1637/8089-081407-Reg.1.
Outbreaks of Marek's disease (MD) in vaccinated flocks still occur sporadically and lead to economic losses. Unfortunately, adequate methods to predict MD outbreaks are lacking. In the present study, we have evaluated whether high load of challenge MD virus (MDV) DNA in peripheral blood could aid in the early diagnosis of MD and in monitoring efficacy of vaccines against MD. One experiment was conducted to simulate field conditions by combining various vaccines (turkey herpesvirus [HVT] and HVT + MDV serotype 2 [SB1]) and challenge viruses (GA, Md5, and 648A). Vaccine efficacy among our experimental groups ranged from 13.3% to 94.2%. Each chicken was sampled three times during the length of the experiment (3, 5, and 15 wk postchallenge [wpc]), and gross lesions were evaluated in chickens that died and at termination of the experiment. DNA was extracted from whole blood and buffy coats from each sample, and the load of challenge MDV DNA and HVT DNA were quantified by real-time polymerase chain reaction. Chickens that developed MD by the end of the experiment had higher load of challenge MDV DNA (threshold cycle [Ct] glyceraldehyde-3-phosphate dehydrogenase [GAPDH]/Ct glycoprotein B [gB] ratios of 1.0, 1.04, and 1.05 at 3, 5, and 15 wpc, respectively) than those that did not develop MD (Ct GAPDH/Ct gB ratios of 0.7, 0.69, and 0.46 at 3, 5, and 15 wpc, respectively). However, load of HVT DNA in blood was not correlated with the development of tumors (Ct GAPDH/Ct HVT ratios from 0.04 to 0.10 in both groups). Vaccinated groups with >75% protection had statistically significant less challenge DNA virus (Ct GAPDH/Ct gB ratios of 0.76, 0.70, and 0.45 at 3, 5, and 15 wpc, respectively) than less protected groups (Ct GAPDH/Ct gB ratios of 0.92, 0.97, and 0.85 at 3, 5, and 15 wpc, respectively). No differences in the load of HVT DNA could be found between protected and nonprotected groups at any time point of the study (Ct GAPDH/Ct HVT from 0.05 to 0.09 in both groups). Our results showed that load of challenge MDV DNA but not load of HVT DNA in blood can be used as criterion for early diagnosis of MD.
接种疫苗的鸡群中仍偶尔会爆发马立克氏病（MD），并导致经济损失。不幸的是，目前缺乏预测MD爆发的有效方法。在本研究中，我们评估了外周血中高负荷的MD强毒（MDV）DNA是否有助于MD的早期诊断以及监测MD疫苗的效力。进行了一项实验，通过组合多种疫苗（火鸡疱疹病毒 [HVT] 和HVT + MDV 2型 [SB1]）和强毒（GA、Md5和648A）来模拟田间条件。我们实验组的疫苗效力范围为13.3%至94.2%。在实验期间（攻毒后3、5和15周 [wpc]）对每只鸡进行三次采样，并对死亡鸡和实验结束时的鸡进行大体病变评估。从每个样本的全血和血沉棕黄层中提取DNA，并通过实时聚合酶链反应定量攻毒MDV DNA和HVT DNA的负荷。实验结束时发生MD的鸡比未发生MD的鸡具有更高的攻毒MDV DNA负荷（攻毒后3、5和15周时，甘油醛-3-磷酸脱氢酶 [GAPDH] 的阈值循环 [Ct]/糖蛋白B [gB] 的Ct比值分别为1.0、1.04和1.05）（攻毒后3、5和15周时，Ct GAPDH/Ct gB比值分别为0.7、0.69和0.46）。然而，血液中HVT DNA的负荷与肿瘤的发生无关（两组的Ct GAPDH/Ct HVT比值均为0.04至0.10）。保护率>75%的接种组的攻毒DNA病毒负荷在统计学上显著低于保护率较低的组（攻毒后3、5和15周时，Ct GAPDH/Ct gB比值分别为0.76、0.70和0.45）（攻毒后3、5和15周时，Ct GAPDH/Ct gB比值分别为0.92、0.97和0.85）。在研究的任何时间点，保护组和未保护组之间的HVT DNA负荷均未发现差异（两组的Ct GAPDH/Ct HVT均为0.05至0.09）。我们的结果表明，血液中攻毒MDV DNA的负荷而非HVT DNA的负荷可作为MD早期诊断的标准。
