Establishment and evaluation of polymerase chain reaction for detection of Clonorchis sinensis eggs in human feces
Objective This paper aims to establish a rapid,specific and sensitive method for the detection of Clonorchis sinensis eggs in human feces,evaluate the method,and provide basis for the rapid,accurate and effective diagnosis of Clonorchis sinen-sis.Methods The cytochrome c oxidase subunit 1 gene of Clonorchis sinensis was used as the target site,and Primer Express 3.0 software was used to design primers.After searching similar sequences by Blast software,an optimal set of primers without cross-reaction with common parasites was selected.The DNA of Feline opisthorchis,Strongylus faecalis,Schistosoma japoni-cum,Echinococcus granulosus,Paragonimus welsoni and other parasites that can infect human beings and whose egg morphology is easily confused and the species relationship is relatively close were used for specific experiments.The positive plasmid contai-ning the target fragment was diluted and amplified by PCR to determine the sensitivity of the detection method.The method was used to detect fecal samples around Baicheng City,Jilin Province,and verify the feasibility of the method.Results The prim-ers designed by using the cytochrome c oxidase subunit 1 gene only had amplified bands on the DNA of Clonorchis sinensis or eggs,while other controls and negative controls did not have obvious amplified bands.The sensitivity of the plasmid DNA of Clonorchis sinensis egg reached 2.47 × 102 copies/μl.The established method can effectively amplify the eggs of Clonorchis sinensis in the actual feces,and the results are consistent with the traditional picture microscopy method.Conclusion PCR method has strong specificity and sensitivity,which can detect Clonorchis sinensis eggs in human feces quickly and accurately,and can provide technical support for the rapid diagnosis of Clonorchis sinensis.
NETL
NSTL
万方数据
维普
