鹦鹉热衣原体数字PCR检测方法的建立与应用OA

Chlamydia psittaci(C.psittaci)is a zoonotic pathogen that primarily causes respiratory tract infections.Due to the absence of specific clinical symptoms,it is prone to missed diagnosis and misdiagnosis,which may lead to severe illness,and its harmfulness is often underestimated.In this study,a pair of primers and a probe were designed based on the nucleotide sequence of the C.psittaci ompA gene(GenBank accession number:PQ511242.1)published in GenBank.A digital PCR(dPCR)detection method for C.psittaci was established,and the reaction system was optimized.The results showed that the optimal concentrations of primers and probe for dPCR were 900 nmol/L and 250 nmol/L,respectively,and the optimal annealing temperature was 56℃.The method achieved a lower limit of detection of 0.58 copies/μL,demonstrated a good linear relationship,and showed no cross-reaction with seven other pathogens,indicating high specificity and good reproducibility.In the clinical detection of 72 bovine and ovine blood and vaginal swab samples,the positive detection rate of dPCR was 19.04%,while that of real-time quantitative PCR(qPCR)was 18.06%.The dPCR method established in this study is more sensitive and efficient than the qPCR method and can be used for the quantitative detection of C.psittaci.