Application of multiplex fluorescence quantitative PCR for the detection of Brucella from different samples
LIU Jia-yin1,2, TIAN Guo-zhong2, YU Gao-wa3, DU Song-nan3, CHEN Jun-jie3, MENG Ying3,YANG Xiao-wen2, Fan Yu2, ZHAO Hong-yan2, PIAO Dong-ri2, YANG Wen-jie1, JIANG Hai1,2
1.School of Public Health, Baotou Medical College Inner Mongolia University of Science and Technology, Baotou 014060, China; 2.State Key Laboratory for Infections Disease Prevention Control, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Disease,National Institute for Communicable Disease Prevention and Control, Chinese Center for Disease Control and Prevention, Beijing 102206,China; 3.Tongliao Center for Disease Control and prevention, Inner Mongolia Autonomous Region, Tongliao 028000,China; 4.School of Medical Technology and Anesthesia,Baotou Medical College Inner Mongolia University of Science and Technology, Baotou 014060, China
Abstract:To explore the value of multiplex fluorescence quantitative PCR in the detection of Brucella in blood, serum and environmental samples. 149 subjects were classified according to their medical conditions, including 75 in the suspected group and 74 in the treatment group. The collected samples of environment and subjects were detected by multiplex fluorescence quantitative PCR, and the results were statistically analyzed. The target genes of three pairs of primers and probes were cloned into PUC57 vector to make positive standard for sensitivity detection and standard curve drawing.The positive rates of single, double and triple methods were 79.2%, 34.2% and 27.5% respectively. The positive rates of single and double, single and triple fluorescence quantitative methods were statistically different, and the chi-square values were 55.42 and 73.42, respectively (P<0.05). The standard curve generated by single fluorescence quantitative PCR of Brucella is y=-3.773 2x+43.188,R2=0.995 3, the linear relationship is good, and the minimum detection range is 101 copies/μl .All the environmental samples detected by multiple fluorescence quantitative method were positive.Multiple fluorescence quantitative PCR can be used to detect blood, serum and environmental samples. Triple fluorescence quantitative method can not only complete the identification of genus level, but also distinguish Brucella from bovine and sheep species in one experiment, but also realize rapid quantification. It plays an important role in the prevention and control of brucellosis.
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