Anchun Cheng, Mingshu Wang, Hongyi Xin, Dekang Zhu, Xinran Li, Haijuen Chen, Renyong Jia, Miao Yang
Key Laboratory of Animal Disease and Human Health of Sichuan Province, Yaan 625014, Sichuan Province, China.
J Microbiol Methods. 2009 Jun;77(3):332-6. doi: 10.1016/j.mimet.2009.02.002.
We developed a reverse transcriptase polymerase chain reaction (RT-PCR) method for the detection of duck hepatitis virus type 1 (DHV-1) in the tissues of infected and clinically affected ducks and in chick and duck embryos. We found the assay to be effective in detecting the virus in China, where it is being used in studies on the epidemiology of the disease. We applied this simple and rapid diagnostic method to the detection of DHV isolates grown in chick and duck embryos and in tissues obtained from infected birds. The assay also proved useful for the differentiation of DVH from the duck plague virus (DPV), muscovy parvovirus (MPV), gosling parvovirus (GPV), avian influenza virus (AIV/H5N1), Pasteurella multocida (PA/5:A), Riemerella anatipestifer (RA/serotype 1), and Salmonella enteritidis (SE). The limit of the sensitivity of this method for the detection of DHV-1 RNA was 3 pg/10 microl. As compared to Dot-ELISA and virus isolation, the rate of agreement for the detection of experimentally infected livers was 100%; moreover, the RT-PCR method was also capable of detecting DHV-1 RNA from the livers that had been infected and stored at -20 degrees C for 22 years; in contrast, Dot-ELISA and virus isolation method could only detect DHV-1 from the livers that had been infected and stored at -20 degrees C for 13 and 11 years, respectively. The rate of positivity in 185 clinically suspected diseased livers subjected to detection by RT-PCR, Dot-ELISA, and virus isolation was 89.2%, 69.2%, and 55.7%, respectively. These results indicated that the RT-PCR approach is rapid, sensitive, and reliable for the detection and differentiation of DHV-1 from the other clinical samples and suspected isolates.
我们开发了一种逆转录酶聚合酶链反应（RT-PCR）方法，用于检测感染鸭和出现临床症状鸭的组织以及鸡胚和鸭胚中的1型鸭肝炎病毒（DHV-1）。我们发现该检测方法在中国检测该病毒有效，目前正用于该疾病的流行病学研究。我们将这种简单快速的诊断方法应用于检测在鸡胚和鸭胚中培养的DHV分离株以及从感染禽类获得的组织中的病毒。该检测方法还被证明有助于区分DHV与鸭瘟病毒（DPV）、番鸭细小病毒（MPV）、雏鹅细小病毒（GPV）、禽流感病毒（AIV/H5N1）、多杀性巴氏杆菌（PA/5:A）、鸭疫里氏杆菌（RA/血清型1）和肠炎沙门氏菌（SE）。该方法检测DHV-1 RNA的灵敏度极限为3 pg/10微升。与斑点酶联免疫吸附测定（Dot-ELISA）和病毒分离相比，检测实验感染肝脏时的符合率为100%；此外，RT-PCR方法还能够检测出在-20℃下保存22年的感染肝脏中的DHV-1 RNA；相比之下，Dot-ELISA和病毒分离方法分别只能检测出在-20℃下保存13年和11年的感染肝脏中的DHV-1。对185份临床疑似患病肝脏进行RT-PCR、Dot-ELISA和病毒分离检测的阳性率分别为89.2%、69.2%和55.7%。这些结果表明，RT-PCR方法在检测DHV-1并将其与其他临床样本和疑似分离株区分开来方面快速、灵敏且可靠。
