聚合敏链反应对胃活检标本中幽门螺杆菌DNA的检测

应用聚合酶链反应（ＰＣＲ）对幽门螺杆菌（ＨＰ）特异性的尿素酶Ａ基因进行扩增，４０个循环可以检测出１０个ＨＰ，加用非同位素地高辛探针杂交，可以检出单个ＨＰ。用本法对１００例胃镜活检标本进行ＰＣＲ检测，其阳性率达８２％，而尿素酶水解试验，细菌涂片及培养的阳性率则分别为６８％、４６％、１８％。ＰＣＲ方法能快速、敏感、特异地检测出临床标本中ＨＰ，对于研究ＨＰ与上消化道疾病之间的关系，指导抗菌治疗均具有重要     

聚合酶链反应检测幽门螺杆菌的临床评价

应用聚合酶链反应（ＰＣＲ）对６４例患者的１２８份胃组织和胃液标本中的幽门螺杆菌（ＨＰ）特异性的尿素酶Ａ基因进行检测，其阳性率分别为８１．３％和７８．１％。胃组织标本尿素酶水解试验、涂片染色和培养的阳性率分别为６７．２％、４５．３％和１７．２％，胃液中涂片和培养的阳性率分别为２５．０％和４．７％，表明以ＰＣＲ方法检测临床标本中的ＨＰ快速、敏感、特异，对于研究ＨＰ与上消化道疾病之间的关系及指导治疗有着重要的意义。而且胃液标本和组织标本两者ＰＣＲ阳性率无明显差异，这对于临床检测的重复性和减少活检的创伤性，有着一定的意义。     

聚合酶链反应对脑脊液标本中结核杆菌DNA重复序列的检测

采用聚台酶链反应（ＰＣＲ）对３４例结核性脑膜炎（结脑）患者脑脊液（ＣＳＦ）进行检测，其中５份结核杆菌培养阳性的标本，ＰＣＲ检测均阳性；２９例ＣＳＦ结核杆菌培养阴性的标本，１９例ＰＣＲ检测阳性，总的阳性率为７０．５％。而作为对照的２０例非结脑患者的ＣＳＦ标本则无阳性。利用育法微量结核杆菌与对照菌测试，可以反证本方法的准确性，其符合率为１００％，提示本方法可以用于临床标本中结核杆菌ＤＮＡ的快速检测，不失为一快速，灵敏而又特异的诊断手段。关键词     

用聚合酶链反应检测血清标本诊断阿米巴肝脓肿的价值

用聚合酶链反应（ＰＣＲ）检测阿米巴肝脓肿患者血清标本中的溶组织内阿米巴３００００蛋白基因,结果４２例患者中有３５例呈阳性反应,阳性率８３．３％,低于脓标本ＰＣＲ阳性率（１００％）（Ｐ〈０．０１）。３例细菌性肝脓肿、１例肝癌及１０例其它部位脓肿患者的血清和脓对照标本ＰＣＲ均呈阴性反应。对ＰＣＲ检测血标本诊断阿米巴肝脓肿的价值进行了讨论。     

聚合酶链反应试剂对检测痰结核分支杆菌的影响

目的观察聚合酶链反应（ＰＣＲ）的试剂对检测痰结核分支杆菌的敏感性和特异性的影响。方法对结核病组２０５例和非结核病组１０５例的痰标本同时进行三个厂家生产的ＰＣＲ试剂检测、普通细菌培养和抗酸染色法涂片检测。再对ＰＣＲ假阳性的痰普通细菌培养结果进行分析，试找出ＰＣＲ假阳性的其它因素。结果三种不同引物序列和扩增产物片段长度的ＰＣＲ试剂对结核性痰标本的阳性率分别为８２．４％、７１．７％和６１．０％，组间差异有非常显著意义（Ｐ＜０．００５）。结论ＰＣＲ试剂的引物序列和扩增产物长度对检测痰结核分支杆菌的阳性率和假阳性率有很大的影响     

PCR检测对男性尖锐湿疣HPV_（6．11）的诊断价值

ＰＣＲ检测对男性尖锐湿疣ＨＰＶ＿（６．１１）的诊断价值黄世明，李广云，山东省千佛山医院（２５００１４）赵庆利，魏学斌，白强近年来，我们采用聚合酶链反应（ＰＣＲ）技术检测了３５例男性尖锐湿疣患者的人乳头瘤病毒（ＨＰＶ）６．１１型。阳性率为１００％。资料...     

聚合酶链反应诊断军团菌感染的实验研究

目的早期诊断军团菌感染。方法采用聚合酶链反应（ＰＣＲ）检测嗜肺军团菌（Ｌｐ）ｍｉｐ基因的特异性片段，并应用于豚鼠米克戴德军团菌（Ｌｍ）感染后的组织标本检测。结果可以扩增出ＬｐＩ型和Ｌｍ６３０ｂｐ的特异性片段，其他临床分离株不被测得，检测灵敏度为１５０ｆｇ军团菌基因组ＤＮＡ（相当于１５～３０个军团菌）。对２２份Ｌｍ感染后不同时间获取的豚鼠组织标本，应用该法与培养法检测结果比较显示，在感染后３．５天培养法阳性率为８８．９％，ＰＣＲ法１００％，在感染后７．５天培养法阳性率为０，而ＰＣＲ法仍有２２．２％的阳性率。结论本方法敏感、特异，既能检出嗜肺军团菌，又能测得Ｌｍ，大大提高了军团病的诊断率     

唾液抗幽门螺杆菌IgG检测诊断幽门螺杆菌感染的评价

目的 评价唾液抗幽门螺杆菌（ＨＰ）抗体诊断ＨＰ感染的价值。方法 ９３例病人同时进行唾液抗ＨｐＩｇＡ测定、快速尿素酶试验（ＲＵＴ）、组织涂片ＨＥ染色及病理ＷＳ银染色法诊断ＨＰ感染结果 唾液抗ＨｐＩｇＧ检测阳性率近似于ＲＵＴ法，高于组织涂片ＨＥ染色法和病理ＷＳ银染色法，其诊断ＨＰ感染的敏感性与９６．９７％，特异性８８．２３％，准确性９５．１８％，阳性预测值９７％，阴性预测值８８．２４％，结论唾液抗Ｈｐ     

三种快速检测结核分支杆菌方法的评价

用聚合酶链反应（ＰＣＲ）技术、单克隆抗体（ＭｃＡｂ）ＴＢ１５－Ｃ＿３经酶联免疫吸附试验（ＥＬＩＳＡ）及抗酸染色等３种方法，对１０种抗酸杆菌及２种非分支杆菌进行检测。ＰＣＲ仅对结核分支杆菌复合体扩增出２４５ｂｐ特异性条带，ＭｃＡｂ－ＥＬＩＳＡ检测除人型结核分支杆菌外，还与ＢＣＧ、鸟型及瘰疬分支杆菌反应阳性，抗酸染色则所有分支杆菌均呈阳性。用ＰＣＲ、ＭｃＡｂ－ＥＬＩＳＡ及抗酸染色检测了１２４份临床标本，ＰＣＲ的检出率高于抗酸染色涂片的阳性率（Ｐ＜０．０５），ＭｃＡｂ－ＥＬＩＳＡ检测阳性率明显高于抗酸染色涂片和ＰＣＲ的阳性率（Ｐ＜０．０１）。认为三种方法在使用中各自有其优点，不可偏废。     

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．     

Clinical significance of PCR in Helicobacter pylori DNA detection in human gastric disorders

ＣｌｉｎｉｃａｌｓｉｇｎｉｆｉｃａｎｃｅｏｆＰＣＲｉｎＨｅｌｉｃｏｂａｃｔｅｒｐｙｌｏｒｉＤＮＡｄｅｔｅｃｔｉｏｎｉｎｈｕｍａｎｇａｓｔｒｉｃｄｉｓｏｒｄｅｒｓＸＵＧｕｏＭｉｎｇ，ＪＩＸｕＨｕａｉ，ＬＩＺｈａｏＳｈｅｎ，ＭＡＮＸｉａｏＨｕａ...     

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。选用互补于幽门螺杆菌尿素酶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HP的有价值的方法。     

Helicobacterpylori in Dental Plaque (A Comparison of Different PCR Primer Sets)

This study was designed to compare differentprimer sets for PCR analysis of H. pylori in the sameseries of 40 dental plaque samples. Three pairs ofprimers, HPU1/HPU2, HP1/HP2, and EHC-U/EHC-L, directed to the urease A gene, 16S rRNA gene, or 860-bpDNA of H. pylori, respectively, were used. Our resultsdemonstrate that EHC-L/EHC-U were more specific andsensitive for H. pylori added to saliva or dental plaque than HPU1/HPU2 and HP1/HP2. Thedetection rates for H. pylori DNA in dental plaquesamples from randomly selected adult patients from theDental Clinic of the University of Ulm were 26.5% (9/34) for HPU1/HPU2, 78.9% (30/38) for HP1/HP2, and100% (40/40) for EHC-U/EHC-L (P < 0.001). Nested PCRusing primers directed to the 860-bp DNA of H. pylorifurther confirmed the presence of H. pylori DNA (40/40) in all these samples. Our resultsindicate that primers EHC-U/EHC-L are to be recommendedfor PCR detection of H. pylori in the oralcavity.     

Use of polymerase chain reaction in the diagnosis of abdominal tuberculosis

BACKGROUND: Early diagnosis and prompt treatment of abdominal tuberculosis is vitally important as it greatly reduces disease and treatment related morbidity and even mortality in extreme cases. A polymerase chain reaction (PCR) test was evaluated for its feasibility as a diagnostic tool in abdominal tuberculosis (TB) in the Indian scenario. METHODS: PCR for the identification of M. tuberculosis amplified a 340 bp nucleotide sequence located within the 38 kDa protein gene of M. tuberculosis. Tissues for processing were obtained from patients suspected to have abdominal TB. These were from various sources such as abdominal lymph nodes, segments of intestine and bowel obtained at various times and in different ways such as laparoscopy, colectomy, bowel and lymph node resection. Fifty such patients had their tissues sent for PCR. RESULTS: PCR results were compared with histopathology (HP). Of the 50 samples, 31 were positive for abdominal TB by HP whereas 30 were positive by PCR. Twenty-four of these were positive for both HP and PCR while of the seven samples positive for HP, five were negative and two gave inhibition by PCR. Six samples negative by HP were positive by PCR. CONCLUSION: This study demonstrates that PCR can be used as an effective tool to diagnose abdominal TB.     

Evaluation of polymerase chain reaction for detection of Mycobacterium tuberculosis in pleural fluid

OBJECTIVES: Tuberculosis, a reemergent killer, is threatening to assume serious proportions all over the world, particularly in view of the AIDS pandemic. The detection of mycobacterial DNA by polymerase chain reaction (PCR) in clinical samples is a promising approach for the rapid diagnosis of tuberculous infections. The aims of this study were to evaluate PCR for detection of Mycobacterium tuberculosis in pleural fluids and to correlate the results with adenosine deaminase activity (ADA) estimation and acid-fast bacilli (AFB) screening. METHODS: The sensitivity and specificity of PCR in detection of mycobacterial DNA in 20 samples of tuberculous pleural effusion were evaluated using 40 samples of nontubercular pleural effusion as controls. The results were correlated with the ADA in all 60 pleural fluids. In addition, AFB detection by Ziehl-Neelsen staining on cytospin smears of all pleural fluids was also compared. RESULTS: Of the 20 samples of tuberculous pleural effusion, mycobacterium could be detected by AFB staining in 4 samples. Fourteen samples were PCR positive. None of the samples from the control group were AFB or PCR positive. The sensitivity of PCR, therefore, was 70.0% with specificity of 100% (positive predictive value, 100%; negative predictive value, 86.95%). The sensitivity of AFB screening was at best 20%. The mean of ADA values in tubercular pleural effusions was 63.21 U/L (SD, 33.01), and the mean in the control samples was 51.1 U/L (SD, 29.71). Taking a cut-off value of 50 U/L, both the sensitivity and specificity of ADA estimation in diagnosing tuberculosis were only 55%. CONCLUSION: PCR represents a rapid and sensitive method for the detection of mycobacterial DNA in tuberculous pleural effusions. AFB screening has low sensitivity, and ADA estimation has both low sensitivity and specificity. Therefore, when the clinical suspicion is high and smear result is negative, but the signs and symptoms of M tuberculosis are apparent, PCR is the method of choice for identifying the infection.     

基于TaqMan探针的猪繁殖与呼吸障碍综合症病毒实时荧光定量PCR方法的建立

目的 建立一种能检测猪繁殖与呼吸障碍综合症病毒（PRRSV）的TaqMan探针荧光定量PCR方法。方法 根据PRRSV的ORF7基因保守区的核苷酸序列设计引物和TaqMan探针,通过探针浓度的优化,建立检测PRRSV的TaqMan探针荧光定量PCR方法。用该方法对30份临床疑似病料进行检测, 并与常规RT-PCR方法和病毒分离方法进行比较。结果 TaqMan荧光PCR检测PRRSV的最佳探针浓度为0.4μmol,检测灵敏度可达3.51拷贝/μl。检测的30份样品与病毒分离结果的符合率为100%,与普通PCR的检测结果（25/30）比较,本方法对临床样品的检出率（28/30）更高。结论 建立的方法特异性强、敏感性高、重复性好,可用于临床样品的检测。     

Detection of M. tuberculosis in clinical samples of diversified nature by IS6110 based PCR

Performance of the polymerase chain reaction technique based on IS6110 sequence was evaluated in clinical samples obtained from pulmonary and extrapulmonary cases of tuberculosis. One hundred and seventy two samples were processed for detection of M. tuberculosis by ZN stained smear examination, LJ medium culture, BACTEC radiometric culture and PCR tests amplifying 123bp region of IS6110 sequence. A significant difference was seen in the sensitivities of different tests, the figures being 83% for PCR test, 35.2% for smear examination, 47.16% for LJ culture and 53.45% for BACTEC culture (p < 0.05). However, no significant difference was found as far as specificity was concerned. PCR test sensitivity in. pulmonary and extrapulmonary clinical samples were 90.14% and 77.27% respectively and found to be significantly higher (p < 0.05) when compared with those of other tests. The mean detection time for M. tuberculosis was 24.03 days by LJ medium culture, 12.89 days by BACTEC culture and less than one day by PCR test. PCR based on IS6100 sequence is highly sensitive method for the early diagnosis of pulmonary and extrapulmonary tuberculosis.     

一种直接检测结核病痰标本的PCR技术的建立与评价

目的建立并评价聚合酶链式反应(polymerase chain reaction,PCR)在结核病痰标本检测中的应用价值。方法根据结核分枝杆菌复合体IS6110序列设计引物INS1和INS2,并建立PCR反应体系和反应条件。运用PCR方法分别检测标准菌株、结核分枝杆菌PCR检测标准品和拟诊结核病患者痰标本,采用痰涂片和细菌培养为对照。结果比较的统计学分析采用卡方检验。结果PCR方法对结核分枝杆菌、牛分枝杆菌、卡介苗标准株的最小检出浓度分别达到102,103,103个细菌/毫升,能够特异地检出结核分枝杆菌复合体。在PCR检测的574例拟诊病例中,PCR检测阳性病例241例,42%;痰涂片和细菌培养的阳性率分别为19.69%和26.31%,PCR检测阳性率高于传统细菌学检验方法,经χ2检验,差异具有显著统计学意义(χ2=103.67,P<0.01)。以痰培养结果为标准,计算PCR检测方法的敏感度为67.53%。结论PCR检测方法与传统的细菌学检测方法相比可以提高阳性标本检出率,而且具有快速、特异、简便的特点,有望成为结核病大规模筛查和临床快速检测方法。     

Development and evaluation of a multiplex polymerase chain reaction for the detection of Mycobacterium tuberculosis from pulmonary specimens

Abstract Background: The diagnosis of pulmonary tuberculosis is still a major challenge. Using a polymerase chain reaction (PCR), one can detect Mycobacterium tuberculosis in clinical samples within a few hours. However, single gene targets may result in false negativity due to the absence of target DNA in some M. tuberculosis isolates. The objective of this study was to develop and evaluate a multiplex PCR (M-PCR) using IS6110 and devR primers for the detection of M. tuberculosis in sputum samples. Methods: Sputum samples were collected from: (1) 200 confirmed cases of tuberculosis; (2) 100 suspected cases of tuberculosis diagnosed on the basis of clinical and radiological findings; (3) 200 non-tubercular patients suffering from respiratory diseases other than tuberculosis, in whom tuberculosis had been excluded. All 500 sputum samples were subjected to PCR using IS6110 primers, and M-PCR using IS6110 and devR primers; results were compared with conventional techniques. Results: It was found that M-PCR was 97.5% successful in detecting the presence of tuberculosis in the confirmed tuberculosis group as compared to 84.5% by IS6110-based PCR. In the suspected tuberculosis group, M-PCR could detect 45% of cases as compared to 40% by IS6110-based PCR. Overall, the specificities of both the PCR and M-PCR were found to be 96.5%. Conclusions: This study demonstrated that the M-PCR assay is more sensitive than the IS6110-based PCR for the detection of M. tuberculosis in sputum specimens and could be applied in situations of highly suspected tuberculosis when all others tests including IS6110 PCR are negative.