Multiple accurate and sensitive arrays for Capripoxvirus (CaPV) differentiation

Gaihua Cao; Yifan Xiong; Meimei Shi; Yue Qiu; Yu Wang; Fuping Nie; Danqun Huo; Changjun Hou

doi:10.1016/j.aca.2023.341391

Multiple accurate and sensitive arrays for Capripoxvirus (CaPV) differentiation

Anal Chim Acta. 2023 Aug 1:1267:341391. doi: 10.1016/j.aca.2023.341391. Epub 2023 May 17.

Authors

Gaihua Cao¹, Yifan Xiong¹, Meimei Shi², Yue Qiu¹, Yu Wang², Fuping Nie³, Danqun Huo⁴, Changjun Hou⁵

Affiliations

¹ Key Laboratory for Biorheological Science and Technology of Ministry of Education, Bioengineering College of Chongqing University, Chongqing, 400044, PR China.
² State Key Laboratory of Cattle Diseases Detection (Chongqing) of Customs, Diagnosis and Testing Laboratory of Lumpy Skin Disease, Chongqing Customs Technology Center, Chongqing, 400020, PR China.
³ State Key Laboratory of Cattle Diseases Detection (Chongqing) of Customs, Diagnosis and Testing Laboratory of Lumpy Skin Disease, Chongqing Customs Technology Center, Chongqing, 400020, PR China. Electronic address: nie1626@163.com.
⁴ Key Laboratory for Biorheological Science and Technology of Ministry of Education, Bioengineering College of Chongqing University, Chongqing, 400044, PR China. Electronic address: huodq@cqu.edu.cn.
⁵ Key Laboratory for Biorheological Science and Technology of Ministry of Education, Bioengineering College of Chongqing University, Chongqing, 400044, PR China; Chongqing Key Laboratory of Bio-perception & Intelligent Information Processing, School of Microelectronics and Communication Engineering, Chongqing University, Chongqing, 400044, PR China. Electronic address: houcj@cqu.edu.cn.

PMID: 37257965
DOI: 10.1016/j.aca.2023.341391

Abstract

Capripoxvirus (CaPV) contains three viruses that have caused massive losses in the livestock and dairy industries. Accurate CaPV differentiation has far-reaching implications for effectively controlling outbreaks. However, it has a great challenge to distinguishing three viruses due to high homology of 97%. Here, we established a sensitive CRISPR/Cas12a array based on Multiple-recombinase polymerase amplification (M-RPA) for CaPV differentiation, which provided a more comprehensive and accurate differentiation mode targeting VARV B22R and RPO30 genes. By sensitive CRISPR/Cas12a and M-RPA, the actual detection limits of three viruses were as low as 50, 40 and 60 copies, respectively. Moreover, Lateral flow dipstick (LFD) array based on CRISPR/Cas12a achieved portable and intuitive detection, making it suitable for point-of-care testing. Therefore, CRISPR/Cas12a array and LFD array paved the way for CaPV differentiation in practice. Additionally, we constructed a real-time quantitative PCR (qPCR) array to fill the qPCR technical gap in differentiation and to facilitate the quarantine departments.

Keywords: CRISPR/Cas12a; Goatpox virus (GTPV); Lumpy skin disease virus (LSDV); Multiple-recombinase polymerase amplification (M-RPA); Real-time quantitative PCR (qPCR); Sheeppox virus (SPPV).

MeSH terms

Animals
Capripoxvirus* / genetics
Goats / genetics
Nucleic Acid Amplification Techniques
Poxviridae Infections* / diagnosis
Real-Time Polymerase Chain Reaction
Sensitivity and Specificity