Lawson Andrew J, Elviss Nicola C, Owen Robert J
Campylobacter and Helicobacter Reference Unit, Laboratory of Enteric Pathogens, Health Protection Agency Centre for Infections, 61 Colindale Avenue, London NW9 5HT, UK.
J Antimicrob Chemother. 2005 Aug;56(2):282-6. doi: 10.1093/jac/dki199. Epub 2005 Jun 15.
To investigate the occurrence of 16S rDNA mutations associated with resistance or reduced susceptibility to tetracycline in Helicobacter pylori isolated in England and Wales, and to develop a real-time PCR assay to detect these DNA polymorphisms from culture and gastric biopsies.
Tetracycline susceptibility was determined by disc diffusion. The MIC of isolates with reduced susceptibility was determined by Etest and agar dilution methods. The 16S rDNA of these isolates was sequenced and resistance-associated mutations identified. A LightCycler assay developed to detect these mutations was applied to DNA extracted from culture and gastric biopsies.
From 1006 isolates of H. pylori examined, 18 showed reduced susceptibility to tetracycline. Of these, three were resistant (>or=4 mg/L). Mutations in 16S rDNA were detected in 10 of the reduced susceptibility isolates: one double base mutation of A926T/A928C, one A926C, one A928C; and seven A926G. The first two polymorphisms were novel and had not been reported from clinical isolates previously. The LightCycler assay identified each of the 10 isolates with 16S rDNA mutations, but did not detect polymorphisms in 100 tetracycline-susceptible H. pylori isolates. The assay correctly determined the tetracycline susceptibility of H. pylori in 20 gastric biopsy samples.
Mutations in 16S rDNA were detected in H. pylori isolated in England and Wales with reduced susceptibility to tetracycline, but resistance to this antibiotic was uncommon. We show molecular-based susceptibility testing for tetracycline is possible direct from biopsy material.
调查在英格兰和威尔士分离出的幽门螺杆菌中与对四环素耐药或敏感性降低相关的16S rDNA突变的发生情况，并开发一种实时PCR检测方法，以从培养物和胃活检组织中检测这些DNA多态性。
采用纸片扩散法测定四环素敏感性。采用Etest法和琼脂稀释法测定敏感性降低的分离株的最低抑菌浓度（MIC）。对这些分离株的16S rDNA进行测序，并鉴定与耐药相关的突变。将开发用于检测这些突变的LightCycler检测方法应用于从培养物和胃活检组织中提取的DNA。
在检测的1006株幽门螺杆菌分离株中，18株对四环素敏感性降低。其中，3株耐药（≥4mg/L）。在10株敏感性降低的分离株中检测到16S rDNA突变：1株A926T/A928C双碱基突变、1株A926C、1株A928C；7株A926G。前两种多态性是新发现的，此前尚未在临床分离株中报道。LightCycler检测方法鉴定出了10株具有16S rDNA突变的分离株，但未在100株四环素敏感的幽门螺杆菌分离株中检测到多态性。该检测方法正确地确定了20份胃活检样本中幽门螺杆菌对四环素的敏感性。
在英格兰和威尔士分离出的幽门螺杆菌中检测到16S rDNA突变，这些菌株对四环素敏感性降低，但对该抗生素耐药并不常见。我们表明，直接从活检材料进行基于分子的四环素敏感性检测是可行的。
