聚合酶链反应快速检测胃粘膜幽门螺杆菌

本研究的目的是了解简单的PCR方法诊断幽门螺杆菌感染的价值。选用互补于幽门螵杆菌尿素酶A基因片段的一对引物，建立PCR方法扩增幽门螺杆菌DNA．扩增产物经琼脂糖电泳显示一条411bp区带。用PCR扩增41株HP分离菌均阳性，而空肠弯曲菌等8种肠道细菌均阴性．显示100%特异，系列稀释试验显示PCR能检测0．1pg的HP DNA；126例胃粘膜标本用PCR、尿素酶试验、培养和涂片检查，HP检出率分别为70．6%、56．3%、32．5%和56．3%，这些结果提示简单快速的PCR方法是检测FIP的有价值的方法。     

胃黏膜组织中恶臭假单胞菌临床意义的探讨

目的探索恶臭假单胞菌在幽门螺杆菌感染相关胃肠疾病中的意义。方法采集14 C尿素呼气试验阳性患者病变胃黏膜354例,进行细菌的分离培养。根据菌落形态、革兰染色、尿素酶试验及幽门螺杆菌特异性16SrRNA基因片段的PCR进行幽门螺杆菌的鉴定,同时提取胃黏膜组织DNA,通过幽门螺杆菌特异性PCR进行快速诊断。将非幽门螺杆菌转种于营养琼脂培养基,进行快速尿素酶试验。尿素酶阳性的非幽门螺杆菌染色体DNA利用细菌16SrRNA基因通用引物进行PCR扩增、测序及序列比对。使用全自动细菌鉴定仪对经测序比对为恶臭假单胞菌的菌株进行鉴定。采用K-B纸片扩散法进行药物敏感性试验。结果从354例样本中分离出革兰阴性、尿素酶阳性的恶臭假单胞菌10株,经16SrRNA基因测序和序列比对,与GenBank中恶臭假单胞菌的相似性≥98%。10株恶臭假单胞菌的胃黏膜标本中,6例标本幽门螺杆菌特异性PCR为阳性,4例为阴性。传代存活的7株恶臭假单胞菌的K-B法药物敏感试验显示对左氧氟沙星均敏感,1株对四环素敏感,5株对阿莫西林耐药,6株对克拉霉素耐药,7株对甲硝唑、氨苄青霉素均耐药。7株菌经全自动细菌生化鉴定仪鉴定结果均为恶臭假单胞菌。结论病变胃黏膜中分离出尿素酶阳性且对治疗幽门螺杆菌感染的多种抗生素耐药;可能造成临床上14 C-尿素呼气试验假阳性,并继发感染影响胃肠疾病的进展。     

猫胃粘膜螺旋菌的调查与鉴定

本研究应用多种分子生物学和病理学技术,对18只猫胃粘膜的螺旋样细菌进行了调查和鉴定。15/18的猫胃粘膜组织尿素酶阳性,5只猫胃培养出弯曲样和HP样细菌,后者及8/11只猫胃粘膜DNA与螺杆菌属细菌特异寡核苷酸探针发生阳性杂交。且该株细菌与及7/11的猫胃粘膜组织DNA应用幽门螺杆菌特异PCR引物扩增阳性.涂片及组织切片W-S银染发现三种类型的螺旋样菌:幽门螺杆菌样细菌;人胃螺旋菌样细菌及弯曲菌样细菌。PCR对照研究发现切片发现的HP样菌即为HP,而GH-LOs也为螺杆菌属细菌。提示猫也是幽门螺杆菌的动物宿主,其与人类HP感染之间的关系尚待进一步阐明。     

猫胃粘膜螺旋菌的调查与鉴定

本研究应用多种分子生物学和病理学技术．对18只猫胃粘膜的螺旋样细菌进行了调查和鉴定。l5/18的猫胃牯膜组织尿素酶阳性．5只猫胃培养出弯曲样和HP样细菌．后及8／11只猫胃粘膜DNA与螺杆菌属细菌特异寡棱昔酸探针发生阳性杂交。且该株细菌与及7/11的猫胃粘膜组织DNA应用幽门螺杆菌特异PCR引物扩增阳性。涂片及组织切片W-S银染发现三种类型的螺旋样菌：幽门螺杆菌样细菌；人胃螺旋菌样细菌及弯曲菌样细菌。PCR对照研究发现切片发现的HP拌菌即为HP，而GH-LOs也为螺杆菌属细菌。提示猫也是幽门螺杆菌的动物宿主，其与人类HP感染之间的关系尚待进一步阐明。     

^14C—尿素呼气试验测幽门螺杆菌感染

幽门螺杆菌(HP)感染的检测方法多种，有创伤性和非创伤性，大多具有相似的准确度，其敏感性和特异性约在90％或以上，创伤性的检查有胃粘膜组织切片染色、细菌培养、快速尿素酶试验(HPUT)和PCR扩增等方法，临床以胃粘膜HPUT更简便、快捷，得到较普遍推广，但需胃镜检查。无创伤性检验     

慢性胃炎、胃癌组织中幽门螺杆菌基因分型的临床研究

目的 探讨幽门螺杆菌(HP)基因分型与慢性胃炎、胃癌的关系。方法 应用PCR—RFLP方法对36例慢性胃炎及45例胃癌组织中的幽门螺杆菌菌株进行检测。并用HindⅡ酶切尿毒酶B(UreB)扩增基因产物进行多态性分析。结果 36例慢性胃炎HP阳性24例(66．7％)，45例胃癌组织中HP阳性23例(51．1％)。二者差异无显著性。47例HP阳性标本均显示不同的RFLP图谱。胃癌组织中未发现特异性条带。结论 慢性胃炎、胃癌与HP感染密切相关。HP基因具有多态性，HP菌株存在基因型差异。PCR—RFLP方法可对HP菌株进行鉴定并将其分型。     

13C尿素呼气试验与快速尿素酶试验检测幽门螺旋杆菌感染的比较

目的通过比较幽门螺旋杆菌（HP）感染的两种检测方法,探讨13c尿素呼气试验在诊断幽门螺旋杆菌感染中的价值。方法对252例胃病患者,先行胃镜检查,行快速尿素酶试验检测HP感染,作为标准对照,同时进行13C尿素呼气试验。并将两种方法所得的阳性率、敏感性、特异性、正确性比较。结果快速尿素酶试验阳性率、敏感性、特异性、正确性分别是62．7％、82．2％、90．5％、91．2％,13C尿素呼气试验阳性率、敏感性、特异性、正确性分别是70．6％、94．4％、96．5％、94．4％,两种检测方法无显著性差异。结论快速尿素酶试验与13C尿素呼气试验检测幽门螺旋杆菌感染差异小,但13C尿素呼气试验操作简单,患者容易接受,是一种较理想的非侵入性诊断幽门螺旋杆菌的方法。     

消化系统疾病

,84001 PCR检测胃粘膜幽门螺旋杆菌/陈曦…//河南医科大学学报一1998,33(2)一58一60 60例上消化道疾病患者,用PCR法胃粘膜中HP总阳性率为86.7%,显著高于尿素酶试验68.3%(尸<0.01)、切片W一S银染和涂片HE染色法法均65%仁尸<0.05)。2()例HP阳性消化性溃疡患者经根除治疗后进行检测,15例用3种传统检测方法均为阴性,似可认为HP已被根除,但用PCR法检测却有4例HP仍为阳性。示PCR法检测HP感染或用于治疗后的评价较其他方法敏感、可靠。表2参5(周璐)984002幽门螺杆菌感染可视化诊断的临床研究/刘 宾…//中华内科杂志一19蛇.37(4)一247一2…     

13C-尿素呼气试验在蒙古沙鼠幽门螺杆菌感染检测中的应用

目的　应用13 C -尿素呼气试验检测蒙古沙鼠幽门螺杆菌感染 ,从而建立一长期监控小型试验动物幽门螺杆菌感染无创检测技术。方法　分别在蒙古沙鼠感染幽门螺杆菌后第 2 0 ,5 0 ,10 0及 2 0 0d用13 C -尿素呼气试验进行检测 ,并对检测后沙鼠以细菌分离培养、ELISA、PCR、快速尿素酶试验、病理切片等五种常规方法检测幽门螺杆菌。结果　在上述不同检测时期 ,用13 C -尿素呼气试验所得蒙古沙鼠幽门螺杆菌感染阳性率分别为 77 78% ,83 33% ,84 2 1%和 80 0 0 % ,而应用常规方法检测结果阳性率为 83 33% ,88 88% ,94 2 1%和 85 0 0 %。比较两组试验结果发现 ,13 C -尿素呼气试验比常规检测方法检出阳性率相对偏低 ,但统计学分析发现其具有一致性。结论　13 C -尿素呼气试验用于幽门螺杆菌感染蒙古沙鼠模型检测是一种可行的无创检测方法 ,可作为评价Hp感染的重要参考指标之一     

~(13)C-尿素呼气试验和血清抗CagA IgG检测对幽门螺杆菌感染的诊断价值

幽门螺杆菌(Hp)是慢性胃炎,消化性溃疡的主要致病菌,并与胃癌、MALT淋巴瘤的发生发展密切相关。在我国,有报道Hp的感染率可高达70％。Hp感染的诊断方法有血清抗Hp抗体检测,~(13)C-或~(14)C-尿素呼气试验、胃粘膜快速尿素酶试验、胃粘膜组织学染色检查、细菌分离培养及PCR扩增检测等。血清学检查和尿素呼气试验(UBT)具有非侵入性、无痛苦和检查相对简单等优点。为了进一步比较这两种诊断方法对Hp感染的临床应用价值,我们用双盲法对195例患者进行双重检测,现将结果报道如下。     

Detection of Helicobacter pylori cagA gene by polymerase chain reaction in faecal samples.

OBJECTIVE: The polymerase chain reaction (PCR) has been extensively and successfully used to detect Helicobacter pylori in gastric juice and gastric biopsies. In contrast, the results obtained using faeces as biological samples for PCR are rather conflicting. This may be due to the presence of faecal inhibitory compounds (polysaccharides) which can inhibit the amplification reaction. The aim of this study was to characterize the H. pylori genotype in faecal samples by using specific primers for the cagA gene. To overcome the problem of contamination by polysaccharides, we used a filter-based extraction technique already applied in a previous study. METHODS: Antral and body biopsies were obtained from 30 symptomatic patients undergoing upper endoscopy. PCR was used to detect the presence of H. pylori organisms in faecal samples by using primers selected for the urease gene A. In addition, H. pylori organisms were characterized both in faecal samples and paraffin-embedded biopsies by PCR with specific primers for the cagA gene. RESULTS: All patients showed a positive CLO test (rapid urease test) and evidence of H. pylori by Warthin-Starry stain. PCR detected the urease A gene in the faecal samples of all patients. The cagA gene was detected in the faecal and biopsy samples of 18 subjects (60%). Duodenal ulcer and/or antral erosions were observed in 15 of the 18 cagA-positive patients (83.3%) and in five of the 12 cagA-negative patients (41.7%). Endoscopic features of normal mucosa or gastritis were observed in three cagA-positive patients (16.7%) and in seven cagA-negative patients (56.3%). cagA-positive status was found to be significantly related to the endoscopic features of duodenal ulceration and/or antral erosions. CONCLUSIONS: Our findings prove that faeces are suitable samples for the detection of cagA status. Moreover, they confirm the existence of a significant relationship between cagA-positive status and duodenal ulcer and/or antral erosions.     

Nested polymerase chain reaction for detection of Helicobacter pylori in gastric biopsy specimens

Sensitivity and specificity are important for tests used to defect Helicobacter pylori infection from gastric biopsy specimens. Molecular methods, such as PCR and nested PCR, are sensitive methods for H. pylori detection. The objective of this study was to evaluate the performance of PCR and nested PCR compared to culture, the rapid urease test (RUT) and histology for the diagnosis of H. pylori in 130 gastric biopsy specimens from symptomatic dyspeptic patients. Sensitivity and specificity with PCR were 91 and 100% and with nested PCR were 95 and 97%, respectively. H. pylori was detected by PCR and nested PCR at levels as low as 125 fg (70 cells) and 25 fg (14 cells), respectively. These results suggest nested PCR is a highly sensitive direct method to detect H. pylori infection from biopsy specimens.     

PCR法检测胃粘膜中的幽门螺杆菌

目的建立检测幽门螺杆菌（Ｈｐ）的更为敏感的聚合酶链反应（ＰＣＲ），并探讨它在Ｈｐ感染的诊断及根除效果判定中的应用．方法以一对合成的与Ｈｐ１６ＳｒＲＮＡ基因互补的寡核苷酸为引物（ＣＰ１／ＣＰ２），建立了从胃粘膜中检测Ｈｐ的ＰＣＲ反应，并与常规检测方法进行比较．结果ＰＣＲ方法检测Ｈｐ标准菌株及５０株临床分离菌株均产生５００ｂｐ片段，检出的最小ＤＮＡ量为０１ｐｇ，相当于１００个细菌细胞；所有１３株其它细菌及无Ｈｐ感染的人胃粘膜则无扩增产物出现．用该方法检测９６名初诊患者Ｈｐ感染情况及２１名药物治疗后患者Ｈｐ根除情况，并与胃粘膜活组织尿素酶试验、细菌培养及银染色方法比较，证实ＰＣＲ方法可以检出常规方法不能检出的少量Ｈｐ．结论ＰＣＲ是检测Ｈｐ的最为敏感的方法，有助于Ｈｐ感染的诊断和Ｈｐ根除的精确判断     

Rapid urease test from non-ulcer part of stomach is superior to histology from ulcer in detection of Helicobacter pylori infection in patients with gastric ulcer

BACKGROUND/AIMS: Helicobacter pylori is the major pathogenesis of peptic ulcer disease. It is important for the endoscopists to detect H. pylori infection during endoscopy. Only one endoscopic diagnostic method can be used due to the limitation of health insurance payments in Taiwan. Most endoscopists use the tissue obtained from the ulcer margins for histopathological examination and detection of H. pylori infection in patients with gastric ulcer. Whether this is suitable deserves study. METHODOLOGY: A total of 103 consecutive subjects with gastric ulcer were recruited. Biopsy specimens from the margins of gastric ulcer were sent for histological examination with modified Giemsa stain. Two biopsy specimens from the antrum and greater curvature site of mid body were embedded in rapid urease test (CLOtest). A patient was classified as H. pylori positive if either CLOtest or histology were positive. RESULTS: 57 patients had H. pylori infection. The detection rate of rapid urease test and histological examination was 96.5% (55/57) and 59.6% (35/57), respectively. Of the 3 patients who had positive histological examinations and negative urease test, only one was confirmed again to have H. pylori infection. The detection rates of rapid urease test and histological examination in different locations of ulcer (antrum/angularis/proximal stomach) were 92.6%/100%/100% and 81.5%/42.1%/36.4%, respectively. CONCLUSIONS: Our study shows that rapid urease test has higher detection rate than histological examination of the biopsy specimens obtained from the margins of gastric ulcer in diagnosis of H. pylori infection. Under the consideration of the health insurance payments limitation and elimination of false-negative detection rate of H. pylori infection, we strongly recommend the rapid urease test from the antrum and body specimens rather than from the ulcer margins for detection of the bacteria in patients with gastric ulcer disease.     

胃粘膜石蜡切片PCR法检测幽门螺杆菌

目的建立从石蜡包埋的胃粘膜标本中扩增ＨｐＤＮＡ的聚合酶链反应（ＰＣＲ）．方法以一对Ｈｐ特异的寡核苷酸为引物，采用双扩增ＰＣＲ扩增Ｈｐ１６ＳｒＲＮＡ基因，并与常规检测方法进行了比较．结果双扩增ＰＣＲ检出的最小ＨｐＤＮＡ量为００１ｐｇ．用该方法检测十二指肠溃疡患者２０例石蜡包埋的胃粘膜标本，并与尿素酶试验、细菌培养、Ｗａｒｔｈｉｎ＿Ｓｔａｒｒｙ银染色及ＰＣＲ对新鲜胃粘膜标本的检测作比较，结果１８例患者上述五项检测结果一致，２例尿毒酶试验、银染色及ＰＣＲ对新鲜胃粘膜标本的检测呈阳性的患者，采用双扩增ＰＣＲ对石蜡包埋标本的检测未发现ＨｐＤＮＡ．ＰＣＲ对石蜡包埋标本及新鲜胃粘膜标本的检测有显著相关性．结论双扩增ＰＣＲ为分子水平上Ｈｐ感染的回顾性研究提供一有利工具．     

Helicobacter pylori in dental plaque and stomach of patients from Northern Brazil

AIM: To establish whether virulence factor genes vacA and cagA are present in Helicobacter pylori (H. pylori) retrieved from gastric mucosa and dental plaque in pa-tients with dyspepsia. METHODS: Cumulative dental plaque specimens and gastric biopsies were submitted to histological exami-nation, rapid urease test and polymerase chain reac-tion (PCR) assays to detect the presence of cagA and vacA polymorphisms.RESULTS: Detection of H. pylori from dental plaque and gastric biopsy samples was greater by PCR co...     

Diagnosis and genotyping of Helicobacter pylori by polymerase chain reaction of bacterial DNA from gastric juice

BACKGROUND: Efficient and accurate detection of Helicobacter pylori infection as well as identification of virulence-associated alleles are important for the treatment of gastroduodenal diseases caused by this gastric pathogen. The present study was performed to test the efficiency of gastric juice polymerase chain reaction (PCR) method for the rapid detection of H. pylori infection and to determine the bacterial genotypes without the need for culture, which is often not feasible especially in developing countries. METHODS: DNA was extracted from gastric juice samples collected from 45 subjects and was used to amplify urease B gene (ureB) for H. pylori. Results obtained from this method were further confirmed by rapid urease test (RUT), histology and culture. Genotypes of the infected strains predicted from gastric juice PCR were compared to the genotype data obtained from the isolated strains. RESULTS: Among 45 cases, 32 were positive by RUT, 37 by histological examination, 25 by gastric juice PCR method, while culture yielded positive results for 19 samples. Except for one case, all the 19 culture-positive strains gave the same genotype with the gastric juice PCR result. It was found that the gastric juice PCR is more efficient for detection of multiple-strain infection as compared to genotype data obtained from strains isolated as pooled culture. CONCLUSIONS: This moderately sensitive technique could be employed with good efficiency, particularly in cases where it is difficult to obtain biopsy. Moreover, with this method bacterial genotype could be obtained.     

患者胃肠引流液中幽门螺杆菌的检出及VitC含量测定

目的探讨胃肠引流液中幽门螺杆菌(Helicobacterpylori,Hp)的检出率及患者VitC水平。方法用PCR技术、细菌培养、快速尿素酶试验检测胃肠引流液中的Hp,用反相液相色谱法检测患者胃肠引流液及血浆中VitC含量。结果282例患者胃肠引流液中Hp检出率为411%(116/282),PCR、细菌培养及快速尿素酶试验阳性率分别为429%(121/282)、113%(32/282)、411%(116/282)。胃溃疡、十二指肠溃疡、复合溃疡、胃癌、急性肠梗阻、急性胆囊炎及门静脉高压合并食管胃底静脉曲张患者Hp检出率分别为546%(30/55)、522%(24/46)、583%(7/12)、482%(27/56)、222%(8/36)、276%(16/58)、211%(4/19),差异具显著性(P<001)。胃溃疡、十二指肠溃疡、复合溃疡、胃癌Hp阳性患者与Hp阴性患者血浆及胃肠引流液VitC浓度相比,差异均具有显著性(P<001)。结论胃肠引流液中可检出Hp,且能分离培养出活菌;PCR技术是一种敏感、有效的检测胃引流液中Hp的方法。Hp感染人体后引起胃肠引流液及血浆中VitC含量的减少,可能与胃十二指肠疾病的发生有关。     

Gastric juice for the diagnosis of H pylori infection in patients on proton pump inhibitors

AIM： To determine the efficacy of gastric juice polymerase chain reaction （PCR） for the detection of H pylori infection in comparison with histology and gastric antral biopsy PCR in patients on a proton pump inhibitor （PPI）. METHODS： Eighty-five consecutive patients with dyspeptic symptoms were enrolled. Gastric biopsies for histology, PCR and gastric juice were collected at endoscopy for PCR of the H pylori urease C gene （ure C）. Sensitivity, specificity, positive predictive value （PPV）, negative predictive value （NPV）, accuracy, positive and negative likelihood ratio for PCR of gastric juice for the H pylori ure C gene was compared to histology and gastric antral biopsy H pylori ure C PCR in patients with and without PPI. RESULTS： Gastric juice PCR was positive in 66 （78%） patients. Histology showed H pylori associated gastritis in 57 （67%）. Gastric biopsy PCR was positive in 72 （85%）. In patients not taking PPI, the sensitivity, specificity, PPV, NPV, accuracy and positive and negative likelihood ratio for gastric juice PCR were 89%, 72%, 91%, 67%, 90%, 85%, 3.1 and 0.1 respectively. In patients on PPI these values were 86%, 100%%, 100%, 29%, 86%, 9.5 and 1.4, respectively. CONCLUSION： Gastric juice PCR for the diagnosis of H pylori infection has increased sensitivity compared to histology with PPI. The use of gastric juice PCR is recommended to confirm H pylori status in patients taking PPIs.     

Conventional Cleaning and Disinfection Techniques Eliminate the Risk of Endoscopic Transmission of Helicobacter pylori

Objective : To determine whether endoscopes serve as a reservoir for Helicobacter pylori and whether two commonly used cleaning and disinfection methods eliminate the risk of H. pylori transmission. Methods : A prospective study was carried out in 107 patients who were undergoing upper gastrointestinal endoscopy for routine clinical indications. H. pylori DNA was assayed by polymerase chain reaction (PCR) of endoscope washes before and after procedure, in gastric aspirates and in endoscope washes after cleaning and disinfection of endoscopes. Gastric biopsies were assayed by rapid urease test (CLOtest, Tri-Med Specialties Inc., Lenexa, KS) of two antral biopsies. Results : Forty-one of 107 (38%) patients were H. pylori-positive by PCR. Endoscopes were contaminated after 25 of 41 (61%) H. pylori-positive procedures. However, 107 of 107 pre-endoscopy and postcleaning aspirates were negative, indicating that decontamination was 100% effective. The urease test was positive in 25 of 41 H. pylori-positive patients, a sensitivity of 61%. PCR was positive in 41 of 41 H. pylori-positive patients, a sensitivity of 100%. In 5 of 16 PCR-positive/urease-negative patients, the identification of H. pylori was clinically relevant. Conclusion : Endoscopes are frequently contaminated with H. pylori after endoscopy in H. pylori-infected patients, but conventional cleaning and disinfection techniques are highly effective in eliminating H. pylori . When appropriate negative control samples are obtained from the endoscope, PCR of endoscopic gastric aspirates appears to be a sensitive test that can detect clinically relevant H. pylori infection that is missed when only a rapid urease test is used.