1. A novel approach for detection of brucella using a real-time recombinase polymerase amplification assay
- Author
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Yong Wang, Yuqi Chen, Kairong Mao, Wenlong Nan, Yueyong Zhang, Pengfei Tan, Qin Lide, and Chen Yiping
- Subjects
DNA, Bacterial ,Recombinase Polymerase Amplification ,Brucella ,Real-Time Polymerase Chain Reaction ,complex mixtures ,Sensitivity and Specificity ,Recombinases ,03 medical and health sciences ,Plasmid ,Bacterial Proteins ,medicine ,Molecular Biology ,Gene ,Pathogen ,030304 developmental biology ,DNA Primers ,0303 health sciences ,Brucella Vaccine ,biology ,030306 microbiology ,Zoonosis ,Brucellosis ,Cell Biology ,bacterial infections and mycoses ,medicine.disease ,biology.organism_classification ,Virology ,enzymes and coenzymes (carbohydrates) - Abstract
Brucella, the etiological agent of brucellosis, is an important zoonosis pathogen worldwide. Brucella infects humans and various domestic and wild animals, and represents a great threat to public health and animal husbandry. In the present study, we developed a real-time recombinase polymerase amplification (RPA) assay for the detection of Brucella. The assay targeted the bcsp31 gene of Brucella, and an RPA exo probe and a pair of primers were selected for assay validation. RPA sensitivity and specificity were evaluated using plasmid standards, Brucella representative strains, and non-Brucella strains. The RPA assay achieved a detection limit of 17 molecules in 95% of cases based on probit analysis, and could successfully distinguish 18 representative Brucella strains (B. abortus biovars 1, 2, 3, 4, 5, 6, 7 and 9, B. melitensis biovars 1, 2 and 3, B. suis biovars 1, 2, 3 and 4, B. canis, B. neotomae and B. ovis), and four Brucella vaccine strains (A19, S19, S2 and M5). A total of 52 Brucella field strains were detected by real-time PCR and RPA in parallel, and compared with real-time PCR, the sensitivity of the RPA assay was 94% (49/52). Thus, this RPA assay may be a rapid, sensitive, and specific tool for the prevention and control of Brucellosis.
- Published
- 2019