A fast, accurate strategy based on LAMP–NALF for detecting human adenovirus serotype 3 in children

IF 2.7 3区 化学 Q2 CHEMISTRY, ANALYTICAL
Qi Yun, Wen Hui Xia, Si Fei Ma, Dong Ming Lu, Meng Gu and Ao Shuang Zhu
{"title":"A fast, accurate strategy based on LAMP–NALF for detecting human adenovirus serotype 3 in children","authors":"Qi Yun, Wen Hui Xia, Si Fei Ma, Dong Ming Lu, Meng Gu and Ao Shuang Zhu","doi":"10.1039/D4AY01973C","DOIUrl":null,"url":null,"abstract":"<p >\r\n <em>Objective</em>: To establish a rapid, convenient and accurate visual detection protocol for human adenovirus serotype 3 in children's respiratory infections based on LAMP–NALF. <em>Methods</em>: Adenovirus serotypes prevalent in the region (Changzhou City, Jiangsu Province, China) were analyzed by assessing the results of tNGS analysis of respiratory pathogens in children. On this basis, multiple sets of LAMP primers were designed for the adenovirus hexon gene as the detection target. Optimal primer combinations were further screened, and the composition of the reagent system and amplification procedures were improved to achieve efficient amplification. To realize an instrument-independent, fast-testing strategy, by combining NALF, which is based on the principle of lateral chromatography, small molecule antigens were modified for primers. Moreover, amplification products were bound to the corresponding antibodies on nucleic acid chromatography test strips for visualization. Finally, a series of tests were performed to validate the sensitivity, specificity, and methodological consistency of the LAMP–NALF assay. <em>Results</em>: Adenovirus infections in children in this region during this period were mainly caused by serotype 3. The optimal amplification temperature for optimal primers of LAMP was 65 °C. The combination of LAMP and NALF could detect femtogram-grade positive plasmid of the hexon gene; besides, there was no cross-reactivity with five other types of common respiratory pathogens in children. The agreement between the LAMP–NALF assay and tNGS/qPCR for detecting the adenovirus in 100 respiratory clinical specimens was 98%. <em>Conclusion</em>: The combination of LAMP and NALF can achieve rapid, convenient and accurate detection of human adenovirus type 3, which can provide a powerful support for the pathogenic diagnosis of adenoviral infections in children, thus contributing to the promotion of the development of point-of-care testing for pathogens.</p>","PeriodicalId":64,"journal":{"name":"Analytical Methods","volume":" 10","pages":" 2295-2301"},"PeriodicalIF":2.7000,"publicationDate":"2025-02-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Analytical Methods","FirstCategoryId":"92","ListUrlMain":"https://pubs.rsc.org/en/content/articlelanding/2025/ay/d4ay01973c","RegionNum":3,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q2","JCRName":"CHEMISTRY, ANALYTICAL","Score":null,"Total":0}
引用次数: 0

Abstract

Objective: To establish a rapid, convenient and accurate visual detection protocol for human adenovirus serotype 3 in children's respiratory infections based on LAMP–NALF. Methods: Adenovirus serotypes prevalent in the region (Changzhou City, Jiangsu Province, China) were analyzed by assessing the results of tNGS analysis of respiratory pathogens in children. On this basis, multiple sets of LAMP primers were designed for the adenovirus hexon gene as the detection target. Optimal primer combinations were further screened, and the composition of the reagent system and amplification procedures were improved to achieve efficient amplification. To realize an instrument-independent, fast-testing strategy, by combining NALF, which is based on the principle of lateral chromatography, small molecule antigens were modified for primers. Moreover, amplification products were bound to the corresponding antibodies on nucleic acid chromatography test strips for visualization. Finally, a series of tests were performed to validate the sensitivity, specificity, and methodological consistency of the LAMP–NALF assay. Results: Adenovirus infections in children in this region during this period were mainly caused by serotype 3. The optimal amplification temperature for optimal primers of LAMP was 65 °C. The combination of LAMP and NALF could detect femtogram-grade positive plasmid of the hexon gene; besides, there was no cross-reactivity with five other types of common respiratory pathogens in children. The agreement between the LAMP–NALF assay and tNGS/qPCR for detecting the adenovirus in 100 respiratory clinical specimens was 98%. Conclusion: The combination of LAMP and NALF can achieve rapid, convenient and accurate detection of human adenovirus type 3, which can provide a powerful support for the pathogenic diagnosis of adenoviral infections in children, thus contributing to the promotion of the development of point-of-care testing for pathogens.

Abstract Image

基于lamp - alf的快速、准确的儿童血清3型人腺病毒检测策略
目的:建立一种快速、方便、准确的基于lamp - alf的儿童呼吸道感染人腺病毒3型目视检测方案。方法:对江苏省常州市某地区儿童呼吸道病原菌tNGS检测结果进行分析。在此基础上,以腺病毒六邻体基因为检测靶点,设计了多套LAMP引物。进一步筛选最佳引物组合,改进试剂体系组成和扩增流程,实现高效扩增。为了实现独立于仪器的快速检测策略,结合基于侧层析原理的NALF,对小分子抗原进行了引物修饰。扩增产物与相应抗体结合在核酸层析试纸上显示。最后,进行了一系列试验来验证lamp - alf测定的敏感性、特异性和方法学一致性。结果:该地区儿童腺病毒感染以血清3型为主。LAMP引物的最佳扩增温度为65℃。LAMP与alf联合检测飞图级六邻体基因阳性质粒;与其他5种儿童呼吸道常见致病菌无交叉反应。lamp - alf法与tNGS/qPCR法检测100例呼吸道临床标本腺病毒的一致性为98%。结论:LAMP与alf联合应用可实现人3型腺病毒快速、便捷、准确的检测,可为儿童腺病毒感染的病原诊断提供有力支持,有助于促进病原体即时检测的发展。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
求助全文
约1分钟内获得全文 求助全文
来源期刊
Analytical Methods
Analytical Methods CHEMISTRY, ANALYTICAL-FOOD SCIENCE & TECHNOLOGY
CiteScore
5.10
自引率
3.20%
发文量
569
审稿时长
1.8 months
期刊介绍: Early applied demonstrations of new analytical methods with clear societal impact
×
引用
GB/T 7714-2015
复制
MLA
复制
APA
复制
导出至
BibTeX EndNote RefMan NoteFirst NoteExpress
×
提示
您的信息不完整,为了账户安全,请先补充。
现在去补充
×
提示
您因"违规操作"
具体请查看互助需知
我知道了
×
提示
确定
请完成安全验证×
copy
已复制链接
快去分享给好友吧!
我知道了
右上角分享
点击右上角分享
0
联系我们:info@booksci.cn Book学术提供免费学术资源搜索服务,方便国内外学者检索中英文文献。致力于提供最便捷和优质的服务体验。 Copyright © 2023 布克学术 All rights reserved.
京ICP备2023020795号-1
ghs 京公网安备 11010802042870号
Book学术文献互助
Book学术文献互助群
群 号:481959085
Book学术官方微信