Establishment and application of ultra-fast real-time PCR for Brucella detection
10.3969/j.issn.1002-2694.2025.00.40
- VernacularTitle:布鲁氏菌超快速荧光PCR方法建立与临床应用
- Author:
Zhen-na XU
1
;
Zhi-peng WU
;
Wei-bin HONG
;
Zhi-shen GUAN
;
Qi-ming LIN
;
Zuan-lan MO
;
Yi-fei YE
;
Hai-yan XIE
;
Min LI
;
Yan-qiu ZHU
;
Xiao-jun LI
;
Xian-peng ZHANG
Author Information
1. 东莞市动物疫病预防控制中心,东莞 523086
- Publication Type:Journal Article
- Keywords:
Brucella;
ultra-fast quantitative PCR;
exogenous reference;
real-time PCR
- From:
Chinese Journal of Zoonoses
2025;41(3):278-283
- CountryChina
- Language:Chinese
-
Abstract:
This study was aimed at establishing a method of ultra-fast quantitative PCR for Brucella detection.We used an exogenous recombinant plasmid as the internal reference and targeted the T4SS secretion system,an important Brucella viru-lence factor,to design specific primers and probes.The sensitivity,specificity,and repeatability of this method were evaluated,and a standard curve was constructed.The coincidence rate of detection findings with this method versus quantitative PCR was determined.This method markedly decreased the detection time to only 10 minutes.The standard curve demonstrated a good linear relationship(Y=-3.410 7x+38.357,R2=0.998 5)with a low minimum detection limit of 10 copies/μL.The method exhibited good specificity and did not specifically amplify several common clinical bacteria other than Brucella.The de-tection of three concentrations of positive plasmids yielded coefficients of variation(CVs)of 0.20％to 0.91％,thus demonstra-ting the method's excellent repeatability.Furthermore,140 clinical samples were analyzed concurrently with the fluorescence PCR method,which yielded a 100％compliance rate and consistent results.Our findings indicated that the Brucella ultra-fast quantitative PCR was ultrafast;had high sensitivity,high specificity,and good specificity;and can be used for the clinical de-tection of Brucella and emergency investigation of epidemics.Therefore,this method is valuable for the early diagnosis of Bru-cella.
