重组冷休克结构域蛋白是一种潜在的抗原，可用于检测山羊感染捻转血矛线虫早期和晚期的特异性抗体。

Recombinant cold shock domain containing protein is a potential antigen to detect specific antibody during early and late infections of Haemonchus contortus in goat.

机构信息

MOE Joint International Research Laboratory of Animal Health and Food Safety, College of Veterinary Medicine, Nanjing Agricultural University, No.1, Weigang, Nanjing, Jiangsu Province, People's Republic of China, 210095.

出版信息

BMC Vet Res. 2020 Feb 3;16(1):36. doi: 10.1186/s12917-020-2261-6.

DOI:10.1186/s12917-020-2261-6

PMID:32013987

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC6998305/

Abstract

BACKGROUND

Haemonchus contortus (H. contortus) is one of the most important parasites that cause huge economic losses to small ruminant industry worldwide. Effective prognosis and treatment depend upon the early diagnosis of H. contortus infection. To date, no widely-approved methods for the identification of prepatent H. contortus infection are available to identify prepatent H. contortus infection properly. The aim of this study was to evaluate the diagnostic potential of recombinant cold shock H. contortus protein (rHc-CS) during early and late infections of H. contortus in goat.

RESULTS

Purified rHc-CS exhibited a clear band, with a molecular weight about 38 kDa. H. contortus eggs were not detected by fecal egg count technique from feces collected at 0 to 14 days post infection (D.P.I). However, eggs were detected at 21, 28 and 35 D.P.I. Hence, results of immunoblotting assay showed specific anti rHc-CS antibody detection in all goat sera collected at early stage (14 D.P.I) and late stage (21-103 D.P.I) of H. contortus infection. Furthermore, no cross reactivity was observed against Trichinella spiralis, Fasciola hepatica and Toxoplasma gondii or uninfected goats. Among several evaluated rHc-CS indirect-ELISA format variables, favorable antigen coating concentration was found 0.28 μg/well at 37 °C 1 h and overnight at 4 °C. Moreover, optimum dilution ratio of serum and rabbit anti-goat IgG was recorded as 1:100 and 1:4000, respectively. The best blocking buffer was 5% Bovine Serum Albumin (BSA) while the best time for blocking, serum incubation and TMB reaction were recorded as 60, 120 and 10 min, respectively. The cut-off value for positive and negative interpretation was determined as 0.352 (OD). The diagnostic specificity and sensitivity of the rHc-CS, both were recorded as 100%.

CONCLUSION

These results validated that rHc-CS is a potential immunodiagnostic antigen to detect the specific antibodies during early and late H. contortus infections in goat.

摘要

背景

捻转血矛线虫（H. contortus）是一种重要的寄生虫，给全世界的小反刍动物产业造成了巨大的经济损失。有效的预测和治疗依赖于捻转血矛线虫感染的早期诊断。迄今为止，尚无广泛认可的方法可用于正确识别潜伏期捻转血矛线虫感染。本研究旨在评估重组冷休克捻转血矛线虫蛋白（rHc-CS）在山羊捻转血矛线虫早期和晚期感染中的诊断潜力。

结果

纯化的 rHc-CS 表现出一条清晰的带，分子量约为 38kDa。在感染后 0 至 14 天（D.P.I.）采集的粪便中，未通过粪便虫卵计数技术检测到捻转血矛线虫卵。然而，在 21、28 和 35 D.P.I.时检测到了卵。因此，免疫印迹分析的结果显示，在捻转血矛线虫感染的早期（14 D.P.I.）和晚期（21-103 D.P.I.）阶段，所有山羊血清中均检测到特异性抗 rHc-CS 抗体。此外，与旋毛虫、肝片吸虫和刚地弓形虫或未感染的山羊无交叉反应。在评估的几种 rHc-CS 间接 ELISA 格式变量中，发现 37°C 1 小时和 4°C 过夜的最佳抗原包被浓度为 0.28μg/孔。此外，血清和兔抗山羊 IgG 的最佳稀释比例分别为 1:100 和 1:4000。最佳封闭缓冲液为 5%牛血清白蛋白（BSA），封闭、血清孵育和 TMB 反应的最佳时间分别为 60、120 和 10 分钟。阳性和阴性解释的截断值确定为 0.352（OD）。rHc-CS 的诊断特异性和敏感性均为 100%。