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首页> 外文期刊>The Internet journal of veterinary medicine >Comparison Of Ear Notch Immunohistochemistry Against E2 (Gp53) Protein And Antigen-Capture Elisa In Sera, For The Detection Of Calves Persistently Infected With Bovine Diarrhea Virus
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Comparison Of Ear Notch Immunohistochemistry Against E2 (Gp53) Protein And Antigen-Capture Elisa In Sera, For The Detection Of Calves Persistently Infected With Bovine Diarrhea Virus

机译：血清中针对E2（Gp53）蛋白和抗原捕获酶联免疫吸附ELISA的耳朵缺口免疫组织化学的比较，用于检测持续感染牛腹泻病毒的犊牛

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The objective of this study was to compare two methods of detection of calves persistently infected (PI) with bovine viral diarrhea virus (BVDV): immunohistochemistry (IHC), using ear skin biopsies, and antigen capture enzyme-linked immunosorbent assay (ELISAAg) using sera. We also aimed to determine the site of immunopositivity. Samples were taken from 80 Holstein calves of up to 3 months of age. For IHC, the streptavidin-biotin-peroxidase complex method was used, along with a monoclonal antibody binding to E2 (gp53) protein of BVDV type 1 and 2. For ELISAAg, a commercial kit was used based on the Erns (gp44–48) protein of BVDV type 1 and 2. Twelve of the 80 skin biopsies (15%) were IHC positive. Immunopositivity was observed in the epidermis, hair follicles and dermis mononuclear cells being similar to the reported in previous studies. This study confirms that the anti-BVDV monoclonal antibody types 1 and 2, E2 (gp53), were able to detect viral antigens in the skin biopsies, previously fixed in 10% formalin. All sera were negative in ELISAAg. Statistical analysis showed a significant difference between the two methods in their capacity to detect PI animals, P (<0.01), indicating that IHC was more sensitive than ELISAAg, identifying 15% of infected animals. Introduction Bovine viral diarrhea virus (BVDV) is classified in the Pestivirus genus within the Flaviviridae family. It is an important pathogen of cattle that can cause reproductive failure, weak born or persistently infected (PI) calves and mucosal disease. BVDV also contributes to the bovine respiratory disease complex (Fray et al., 2000; Liebler-Tenorio et al., 2000; Cornish et al., 2005). BVDV has two genotypes, BVDV 1a 1b and BVDV 2, with each genotype presenting two biotypes, cytopathic (cp) and non-cytopathic (ncp), based on whether or not they cause cellular alterations. Persistent infection results from cows being exposed to the ncp variant of the virus before day 125 of gestation, allowing the fetus to develop immunotolerance to the virus and letting the virus persist after birth (Fulton et al., 2000; Brock, 2004; Bolin and Grooms, 2004; Zimmer et al., 2004). PI animals continuously shed great quantities of the virus even when they are clinically healthy and are therefore considered an important source of viral dissemination among the herd. One of the main strategies that has been used to eliminate BVDV in herds is to identify and remove PI calves (Fray et al., 2000; Cornish et al., 2005). For this, several diagnostic tests have been used which involve viral isolation (VI), inverse transcription, polymerase chain reaction (PCR) and real time (RT)-PCR, antigen capture enzyme-linked immunosorbent assay (ELISAAg), viral genome identification in peripheral blood leucocytes (PBL) and immunohistochemistry (IHC) (Liebler-Tenorio et al., 2000; Njaa et al., 2000; Malhum et al., 2002; Brodersen, 2004; Luzzago et al., 2006; Hilbe et al., 2007). In order to opportunely detect PI calves, a sensitive, economical, easy to perform diagnostic test is required, to facilitate prevention and control strategies of this infection in herds. The IHC test is based on viral antigen detection in infected animal skins. This test has been effective for PI animal detection since only a small portion of the skin, generally obtained from the ear, is required, numerous samples can be analyzed simultaneously and the results have been satisfactory. The aim of this study was to detect PI calves of up to 3 months old, by IHC in skin biopsies and ELISAAg in serum samples, compare the results of both tests and determine immunopositive localization in skin biopsies. Material and methods Sample collection Ear skin biopsies and sera were taken simultaneously from 80 Holstein calves of up to 3 months old, from herds with a BVDV infection background in the Complejo Agropecuario Industrial de Tizayuca, Sociedad Anonima (CAITSA), located on the Mexico-Pachuca highway in the state of Hidalgo, Mexico.Immunoh
机译：这项研究的目的是比较两种检测被牛病毒性腹泻病毒（BVDV）持续感染（PI）的小牛的方法：使用耳皮肤活检的免疫组织化学（IHC）和使用以下方法的抗原捕获酶联免疫吸附测定（ELISAAg）：血清我们还旨在确定免疫阳性的位置。样本取自80个3个月大的荷斯坦牛犊。对于IHC，使用了抗生蛋白链菌素-生物素-过氧化物酶复合物法，以及与BVDV 1型和2型E2（gp53）蛋白结合的单克隆抗体。对于ELISAAg，使用了基于Erns（gp44-48）的商业试剂盒1型和2型BVDV的蛋白。80例皮肤活检中有12例（15％）是IHC阳性。在表皮，毛囊和真皮单核细胞中观察到免疫阳性，与先前研究中报道的相似。这项研究证实，抗BVDV单克隆抗体1和2，E2（gp53）能够检测皮肤活检中的病毒抗原，以前已用10％福尔马林固定。 ELISAAg所有血清均为阴性。统计分析表明，两种方法在检测PI动物方面的能力之间存在显着差异，P（<0.01），表明IHC比ELISAAg更为灵敏，可识别15％的感染动物。简介牛病毒性腹泻病毒（BVDV）属于黄病毒科的瘟病毒属。它是牛的重要病原体，可导致生殖衰竭，弱势出生或持续感染（PI）犊牛和粘膜病。 BVDV也有助于牛呼吸系统疾病（Fray等，2000; Liebler-Tenorio等，2000; Cornish等，2005）。 BVDV具有两种基因型，即BVDV 1a 1b和BVDV 2，每种基因型根据其是否引起细胞改变而表现出两种细胞型，即细胞性（cp）和非细胞性（ncp）。持续感染是由于母牛在妊娠第125天之前暴露于病毒的ncp变体，导致胎儿对病毒产生免疫耐受，并在出生后使病毒持续存在（Fulton等，2000; Brock，2004; Bolin和Grooms，2004; Zimmer等，2004）。即使在临床上健康的PI动物也不断散发大量病毒，因此被认为是畜群中病毒传播的重要来源。消除牛群中BVDV的主要策略之一是鉴定和去除PI犊牛（Fray等，2000; Cornish等，2005）。为此，已经使用了几种诊断测试，包括病毒分离（VI），逆转录，聚合酶链反应（PCR）和实时（RT）-PCR，抗原捕获酶联免疫吸附测定（ELISAAg），病毒基因组鉴定外周血白细胞（PBL）和免疫组织化学（IHC）（Liebler-Tenorio等，2000; Njaa等，2000; Malhum等，2002; Brodersen，2004; Luzzago等，2006; Hilbe等。 ，2007）。为了及时检测PI犊牛，需要灵敏，经济，易于执行的诊断测试，以促进牛群中这种感染的预防和控制策略。 IHC测试基于感染动物皮肤中的病毒抗原检测。该测试对PI动物的检测非常有效，因为只需要通常从耳朵获得的一小部分皮肤，就可以同时分析大量样品，并且结果令人满意。这项研究的目的是通过皮肤活检中的IHC和血清样品中的ELISAAg检测最多3个月大的PI犊牛，比较两个测试的结果并确定皮肤活检中的免疫阳性定位。材料和方法样本采集同时从位于墨西哥州Sociedad Anonima的Complejo Agropecuario Industrial de Tizayuca工业区（CAITSA）的80头3个月大的荷斯坦牛犊进行了耳朵皮肤活检和血清，这些牛犊感染了BVDV。墨西哥伊达尔戈州的帕丘卡高速公路。

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