Kösters Katrin, Reischl Udo, Schmetz Johanna, Riffelmann Marion, Wirsing von König Carl Heinz
Institut für Hygiene und Laboratoriumsmedizin, Klinikum Krefeld, Lutherplatz 40, 47805 Krefeld, Germany.
J Clin Microbiol. 2002 May;40(5):1719-22. doi: 10.1128/JCM.40.5.1719-1722.2002.
Real-time PCR assays based on the LightCycler technology were developed for individual (simplex PCR) and simultaneous (duplex PCR) detection and discrimination of Bordetella pertussis and Bordetella parapertussis in clinical samples. The assays were evaluated with 113 specimens from patients with and without symptoms of pertussis. Results were compared to those from conventional culture and TaqMan real-time PCR. The analytical sensitivity ranged from 0.1 to 10 CFU for B. pertussis and B. parapertussis, and intra- and interassay variations were less than 7%. Results were available within 2 h. With the simplex format, 21 of 100 samples from patients with clinical symptoms of pertussis were positive for B. pertussis and/or B. parapertussis. With the duplex format, 18 of 100 samples were positive. LightCycler PCR increased the diagnostic sensitivity over that of culture by 2.0-fold (duplex PCR) (P = 0.08) to 2.3-fold (simplex PCR) (P = 0.02). Our data suggest that duplex PCR in this format showed good analytical sensitivity but lost some sensitivity on clinical samples compared with the simplex format.
基于LightCycler技术开发了实时PCR检测方法，用于临床样本中百日咳博德特氏菌和副百日咳博德特氏菌的单独（单重PCR）和同时（双重PCR）检测及鉴别。用来自有和无百日咳症状患者的113份标本对这些检测方法进行了评估。将结果与传统培养法和TaqMan实时PCR的结果进行了比较。百日咳博德特氏菌和副百日咳博德特氏菌的分析灵敏度范围为0.1至10 CFU，批内和批间变异小于7%。2小时内可获得结果。采用单重检测法，100份有百日咳临床症状患者的样本中有21份百日咳博德特氏菌和/或副百日咳博德特氏菌呈阳性。采用双重检测法，100份样本中有18份呈阳性。LightCycler PCR使诊断灵敏度比培养法提高了2.0倍（双重PCR）（P = 0.08）至2.3倍（单重PCR）（P = 0.02）。我们的数据表明，这种形式的双重PCR显示出良好的分析灵敏度，但与单重形式相比，在临床样本上失去了一些灵敏度。
