在濒死和健康金鱼中检测疱疹病毒造血坏死病原体（鲤疱疹病毒2型）：定量PCR诊断方法的验证

Detection of the herpesviral hematopoietic necrosis disease agent (Cyprinid herpesvirus 2) in moribund and healthy goldfish: validation of a quantitative PCR diagnostic method.

作者信息

Goodwin A E, Merry G E, Sadler J

机构信息

University of Arkansas at Pine Bluff, Aquaculture/Fisheries Center, Arkansas 71601, USA.

出版信息

Dis Aquat Organ. 2006 Apr 6;69(2-3):137-43. doi: 10.3354/dao069137.

DOI:10.3354/dao069137

PMID:16724557

Abstract

Cyprinid herpesvirus 2 (CyHV-2) is a pathogen of goldfish Carassius auratus auratus L. that causes herpesviral hematopoietic necrosis (HVHN) disease. The disease is associated with necrosis of hematopoietic tissues and anemia with high mortality. We have developed a real time 5'-nuclease PCR method (Taqman) that quantitatively detects CyHV-2 with a linear response over 8 logs of target concentration. The coefficient of variability on replicate samples tested on different days was 13% and the calculated sensitivity approached 1 target molecule per reaction. The assay does not cross-react with other similar fish herpesviruses, including CyHV-1 (carp pox) and CyHV-3 (koi herpesvirus), but reliably detects known CyHV-2 positive fish. The assay detects CyHV-2 not just in clinical cases of HVHN but also in apparently healthy 1 yr old goldfish fingerlings and even in 3 to 5 yr old broodfish.

摘要

鲤疱疹病毒2型（CyHV-2）是金鱼（Carassius auratus auratus L.）的一种病原体，可引发疱疹病毒性造血坏死（HVHN）疾病。该疾病与造血组织坏死及贫血相关，死亡率很高。我们开发了一种实时5'-核酸酶PCR方法（Taqman法），可定量检测CyHV-2，在8个对数级的靶标浓度范围内呈线性响应。在不同日期测试的重复样本的变异系数为13%，计算得出的灵敏度接近每个反应1个靶标分子。该检测方法不与其他类似的鱼类疱疹病毒发生交叉反应，包括鲤疱疹病毒1型（鲤痘）和鲤疱疹病毒3型（锦鲤疱疹病毒），但能可靠地检测出已知的CyHV-2阳性鱼。该检测方法不仅能在HVHN临床病例中检测到CyHV-2，还能在看似健康的1岁金鱼幼鱼甚至3至5岁的亲鱼中检测到。