Detection of feline immunodeficiency virus by neutral red-based loop-mediated isothermal amplification assay

IF 1.7 Q2 AGRICULTURE, DAIRY & ANIMAL SCIENCE

Veterinary World Pub Date : 2024-01-01 DOI:10.14202/vetworld.2024.72-81

Wichayet Saejung, Kotchaporn Khumtong, Witsanu Rapichai, Siriluk Ratanabunyong, Amonpun Rattanasrisomporn, K. Choowongkomon, Oumaporn Rungsuriyawiboon, Jatuporn Rattanasrisomporn

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Abstract

Background and Aim: Feline immunodeficiency virus (FIV) is a retroviral pathogen globally responsible for immunodeficiency disease in cats. However, the current diagnosis based on antibody detection has limitations and can also produce false-positive results. This study aimed to develop a one-pot loop-mediated isothermal amplification (LAMP) process integrated with neutral red (NR-LAMP) assay for detection of FIV proviral DNA. Materials and Methods: We developed a one-pot, gag gene-based NR-LAMP for convenient, rapid, specific, and sensitive colorimetric inspection of FIV proviral DNA. Results: The developed NR-LAMP was capable of amplifying at an optimum temperature of 65°C for 40 min. No cross-amplification was detected between FIV and other feline viruses tested, indicating the high specificity (98.44%) of the novel FIV-LAMP primer. Our NR-LAMP assay has a detection limit of 4.2 × 101 copies/μL. A total of 80 clinical samples with a background of FIV infection were collected and tested using the proposed method. The NR-LAMP assay showed a high sensitivity of 100% compared to conventional polymerase chain reaction assay. Conclusion: These results support the suitability of NR-LAMP as a potential future alternative clinical molecular approach for further use in the diagnosis of FIV-infected cats. Keywords: feline immunodeficiency virus, loop-mediated isothermal amplification, molecular diagnosis, neutral red.

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通过基于中性红的环介导等温扩增测定法检测猫免疫缺陷病毒

背景和目的：猫免疫缺陷病毒（FIV）是一种逆转录病毒病原体，是全球猫免疫缺陷病的罪魁祸首。然而，目前基于抗体检测的诊断方法存在局限性，也可能产生假阳性结果。本研究旨在开发一种与中性红（NR-LAMP）检测相结合的一次性环介导等温扩增（LAMP）过程，用于检测 FIV 前病毒 DNA：我们开发了一种基于gag基因的NR-LAMP，用于方便、快速、特异、灵敏地比色检测FIV前病毒DNA：结果：所开发的 NR-LAMP 能够在 65°C 的最佳温度下扩增 40 分钟。在 FIV 和其他猫科动物病毒之间没有检测到交叉扩增，这表明新型 FIV-LAMP 引物具有很高的特异性（98.44%）。我们的 NR-LAMP 检测方法的检测限为 4.2 × 101 拷贝/μL。我们共收集了 80 份具有 FIV 感染背景的临床样本，并使用所提出的方法进行了检测。与传统的聚合酶链反应检测法相比，NR-LAMP 检测法的灵敏度高达 100%：这些结果支持了 NR-LAMP 作为一种潜在的临床分子方法的适用性，可进一步用于诊断受 FIV 感染的猫。

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来源期刊

Veterinary World Multiple-

CiteScore

3.60

自引率

12.50%

发文量

317

审稿时长

16 weeks

期刊介绍： Veterinary World publishes high quality papers focusing on Veterinary and Animal Science. The fields of study are bacteriology, parasitology, pathology, virology, immunology, mycology, public health, biotechnology, meat science, fish diseases, nutrition, gynecology, genetics, wildlife, laboratory animals, animal models of human infections, prion diseases and epidemiology. Studies on zoonotic and emerging infections are highly appreciated. Review articles are highly appreciated. All articles published by Veterinary World are made freely and permanently accessible online. All articles to Veterinary World are posted online immediately as they are ready for publication.