DPO multiplex PCR as an alternative to culture and susceptibility testing to detect Helicobacter pylori and its resistance to clarithromycin
DPO multiplex PCR as an alternative to culture and susceptibility testing to detect Helicobacter pylori and its resistance to clarithromycin
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DOI：
10.1186/1471-230x-11-112
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发表时间：
2011-10-17
影响因子：
2.4
通讯作者：
Megraud, Francis
中科院分区：
文献类型：
--
作者：
Lehours, Philippe;Siffre, Elodie;Megraud, Francis
背景资料：幽门螺杆菌对大环内酯类耐药是使用含质子泵转运蛋白-克拉霉素治疗时治疗失败的主要危险因素。大环内酯类耐药是由于23 S rRNA基因编码的23 S核糖体亚基上的一些突变。本研究旨在研究双引物寡核苷酸（DPO）-PCR试剂盒Seeplex（R）ClaR-H的性能。pylori ACE检测旨在检测H. pylori和两种引起克拉霉素耐药的点突变。方法：采用Seeplex（R）ClaR-H.在127例胃活检中评价了幽门螺杆菌ACE检测，并与常规细菌培养进行了比较，随后通过E-test以及使用荧光共振能量转移（FRET）技术的内部实时PCR确定了对克拉霉素的敏感性。以培养物为参照，DPO-PCR和荧光定量FRET-PCR的敏感性分别为97.7%和100%，特异性分别为83.1%和80.7%。但两种PCR检测14 H的结果一致。pylori阳性者，培养阴性者。E-test和DPO-PCR在克拉霉素敏感性方面的一致性为95.3%（41/43），而荧光定量FRET-PCR和DPO-PCR的一致性为95%（57/60）。幽门螺杆菌的胃活检，并研究其敏感性克拉霉素在实验室，不能进行实时PCR检测。
Background: Macrolide resistance in Helicobacter pylori is the major risk factor for treatment failure when using a proton pump inhibitor-clarithromycin containing therapy. Macrolide resistance is due to a few mutations on the 23S ribomosal subunit encoded by the 23S rRNA gene. The present study aimed at investigating the performance of the dual priming oligonucleotide (DPO)-PCR kit named Seeplex (R) ClaR-H. pylori ACE detection designed to detect H. pylori and two types of point mutations causing clarithromycin resistance in H. pylori.Methods: The performance of Seeplex (R) ClaR-H. pylori ACE detection was evaluated on 127 gastric biopsies in comparison to conventional bacterial culture followed by the determination of susceptibility to clarithromycin by E-test, as well as by an in-house real-time PCR using a fluorescence resonance energy transfer (FRET) technology.Results: Considering culture as the reference test, the sensitivity of DPO-PCR and real-time FRET-PCR was 97.7% and 100% while specificity was 83.1% and 80.7%, respectively. However, both PCR were concordant in detecting 14 H. pylori positive cases which were negative by culture. Globally, E-test and DPO-PCR were concordant with regard to clarithromycin susceptibility in 95.3% of the cases (41/43), while real-time FRET-PCR and DPO-PCR were concordant in 95% (57/60).Conclusion: The DPO-PCR is an interesting tool to detect H. pylori on gastric biopsies and to study its susceptibility to clarithromycin in laboratories that cannot perform real-time PCR assays.