套式聚合酶链反应快速诊断结核性心包炎

快速诊断结核性心包炎以便及早抗痨治疗防止缩窄性心包炎的发生。应用套式聚合酶链反应（Ｎｅｓｔｅｄ－ ＰＣＲ） 、直接涂片、培养对８３ 例心包积液标本进行结核菌检测。结果：结核性心包积液Ｎｅｓｔｅｄ － ＰＣＲ阳性率为７２ ．５ ％ 、培养阳性率为１９ ．６ ％ 、涂片镜检阳性率为２３ ．５ ％ 。非结核性心包积液无１ 例阳性。Ｎｅｓｔｅｄ－ ＰＣＲ特异性好、敏感性强并可能避免一般聚合酶链反应（ＰＣＲ） 所出现的假阳性     

聚合酶链反应诊断结核性腹膜炎的评价

目的评价聚合酶链反应（ＰＣＲ）方法对结核性腹膜炎的诊断价值。方法用ＰＣＲ技术检测３０例结核腹水中结核分支杆菌ＤＮＡ，并与腹水涂片抗酸染色及酶联免疫吸附试验（ＥＬＩＳＡ）检测腹水中抗ＰＰＤ抗体进行比较。结果ＰＣＲ的阳性率为６０％，特异性９４．４％；ＥＬＩＳＡ法阳性率６３．３％，特异性７２．２％；涂片镜检均为阴性。结论ＰＣＲ在诊断结核性腹膜炎具有较高的敏感性和特异性，优于ＥＬＩＳＡ法及涂片镜检，如与ＥＬＩＳＡ技术结合可进一步提高检测的敏感性和特异性。     

抗酸杆菌荧光染色镜检的应用研究

目的对荧光显微镜检测抗酸杆菌的应用进行评价。方法5个结核病专业诊疗机构共收集2157份标本,每份标本均采用荧光染色（FS）、萋-尼染色（Z-N染色）镜检。其中1245份标本同时进行传统培养检查,并且以培养结果为最终的金标准,计算FS和Z-N染色镜检方法的敏感度、特异度和正确指数[正确指数-（敏感度＋特异度）-11（假阴性率＋假阳性率）]。所有结果采用SPSS17．0统计软件完成配对7。检验,以P〈O．05为差异有统计学意义。结果全部标本荧光染色镜检阳性率为10．8％（234／2157）,萋-尼染色镜检阳性率为8．8％（190／2157）,两种镜检方法阳性率差异有统计学意义（X2=25．473,P〈0．001）。初诊标本经荧光染色镜检的阳性率为15．9％（179／1127）,萋一尼染色阳性率为13．2％（149／1127）,荧光染色镜检检出抗酸杆菌的能力较萋尼染色高20．1％;两种镜检方法检查初诊标本的阳性率,差异有统计学意义（X2=18．000,P〈0．001）。同时完成涂片和培养的标本,荧光染色镜检阳性率13．8％（172／1245）,培养阳性率15．2％（189／1245）,两者比较差异无统计学意义（X2=2．173,P=0．140）。同时完成涂片和培养的初诊标本,荧光染色镜检阳性率16．4％（128／781）,培养阳性率19．7％（154／781）,分离培养阳性率较荧光染色阳性率高20．3％,差异有统计学意义（X2=7．861,P〈0．01）。荧光染色镜检的敏感度为60．3％（114／189）、特异度为94．5％（998／1056）,假阴性率为39．7％（75／189）、结果正确指数为0．548,萋-尼染色镜检相应指标分别是56．6％（107／189）,97．1％（1025／1056）,43．4％（82／189）和0．537。结论与萋一尼染色镜检比较,荧光染色镜检具有更高的敏感度和很好的特异度;能够提高工作效率,降低工作强度,适合在工作量较大、人力资源紧张的基层实验室开展。     

巢式聚合酶链反应检测支气管内膜结核患者活检组织中结核 …

目的 了解巢式聚合酶链反应（ＮＰＣＲ）技术对支气管内膜结核的诊断价值。方法 应用巢式聚合酶链反应（ＮＰＣＲ）对６７份支气管内膜活检组织进行结核分支杆菌ＤＮＡ检测。并与病理检查,刷检涂片,支气管镜检后痰涂片和痰培养结果比较,对照为４３例支气管肺癌患者,结果６７例支气管内膜结核活检组织病理检查,刷检涂片,支气管镜检“激惹”后痰涂片,镜检术后痰培养及ＮＰＣＲ检测阳性率分别为１３％,１９％,２２％,１５％     

聚合酶链反应在诊断结核性胸腔积液中的应用

应用聚合酶链反应（ＰＣＲ）技术对６２例胸水进行结核菌ＤＮＡ检则，并同时与胸水涂片及培养结果进行比较。结果显示：３４例结核性胸水涂片抗酸染色、结核菌培养和ｐＣＲ检测阳性率分别为５．９％、８．８％和５２．９％，后者显著高于前二者（Ｐ均＜０．０１）。２８例非结核性胸水涂片和培养结果均呈阴性，但ＰＣＲ有４例（１４．３％）出现阳性。结果表明ＰＣＲ直接检测胸水中结核菌显示出快速、敏感和高效等优点。同时还对影响ＰＣＲ检测结核菌的某些因素作了分析。     

应用PCR检测结核杆菌的临床分析

本文总结了一个病区１９９３～１９９５年５２例住院结核病人与３７例住院肺癌病人的ＰＣＲ结果，并与同期的痰涂片和／或ＢＡＣＴＥＣ培养结果进行了比较，结果显示ＰＣＲ阳性检出率为６１．５％，高于痰涂片（４６．１％）亦稍高于ＢＡＣＴＥＣ（５３．８％）在涂片阳性者结核病人中ＰＣＲ阴性检出率竟达４１．７％，而诊断未合并肺结核的肺癌病人中ＰＣＲ阳性率达３０．５％，作者分析了造成不一致的原因。     

不同温度下应用荧光染色法检测抗酸杆菌的研究

荧光染色法检测抗酸杆菌是结核病实验室诊断的重要方法之一，我们对此进行了改进。本研究应用双盲法比较３７℃和室温下荧光染色法对抗酸杆菌检出率的影响，选取１１７例住院结核病人痰标本涂片染色镜检并培养，结果发现３７℃染色时涂片阳性率为６２．４％，而室温染阳性率分别为３９．３％，３７℃染色明显提高了是性检出率，实验同时发现培养阳性率仅为１６．２％，明显低于涂片镜检，特别是３７℃染色镜检的阳性率，涂（＋）培（     

内脏利什曼病患者骨髓及血内病原体特异性kDNA片…

目的：建立简易、准确的诊断内脏利什曼病和病原体鉴定技术。方法：采用作者设计的杜氏利什曼原虫（Ｌ．ｄ．）种特异引物Ｉ和Ⅱ，经ＰＣＲ扩增样品内病原体ｋＤＮＡ２９７ｂｐ片段，检测２２例确诊内脏利什曼病（ＶＬ）患者骨髓、血、血清共５５份样品和４例临床疑诊为黑热病患者的骨髓（共２６例，５９份样品）。结果：（１）ＰＣＲ法与骨髓涂片镜检符合率为９６．２％（２５／２６）；（２）ＰＣＲ法检测骨髓、血及血清的总阳性率     

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。     

聚合酶链反应在诊断结核性胸膜炎中的应用

目的：应用ＰＣＲ技术对７０例胸水进行结核菌ＤＮＡ检测。方法：７０例胸水做ＴＢ－ＰＣＲ、胸水抗酸菌涂片，皮肤结素试验及血结核菌抗体检测，并与临床诊断标准进行比较。结果：７０例胸水中，结核性胸腔积液４６例，ＰＣＲ阳性率４５．７％，涂片镜检、结素强阳性、结核菌抗体（＋）则分别为２．２％，８．７％，１０．９％，前者明显高于后三者（Ｐ值均＜０．００５），但其ＰＣＲ敏感性与临床诊断标准比较相关性仍低，非结核性胸水２４例中，ＰＣＲ阳性率８．３％。结论：ＰＣＲ检测结核菌尚有一些条件需进一步完善，目前临床尚不能将其作为诊断结核病包括结核性胸膜炎的标准，仍需结合病人的临床表现及其它检查结果来综合判断。     

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。     

对肺结核可疑者采用症状查痰法提高患者发现率

目的探讨更全面的发现肺结核病例的检查诊断程序。方法在采用通用的胸透筛查法发现肺结核患者的基础上，采用症状查痰法，对有症状且≥3周者，直接做痰结核分枝杆菌检查，并拍摄X线胸片，对按胸透筛查法和症状查痰法发现的患者进行比较；对肺结核诊断程序改进后的结核病患者的发现情况进行分析。结果采用症状查痰法发现活动性肺结核新病例900例，比胸透筛查法多发现活动性肺结核患者73例，患者发现率提高8．8％(73／827)；发现涂阳肺结核病例262例，比胸透筛查法多发现活动性肺结核患者30例，患者发现率提高12．9％(30／232)；培养阳性肺结核病例360例，比胸透筛查法多发现患者63例，患者发现率提高21．2％(63／297)。症状查痰法发现新病例的细菌学检查阳性者的比例高于胸透筛查法，胸透筛查法发现新病例中，涂片阳性者占28．1％(232／827)，痰菌培养阳性者占35．9％(297／827)；而症状查痰法发现新病例中，涂片阳性者占29．1％(262／900)，痰菌培养阳性者占40．0％(360／900)。症状查痰法多检出的活动性肺结核患者73例中，涂阳和菌阳患者的比例均很高，分别为41．1％(30／73)和86．3％(63／73)，较胸透筛查法的检出涂阳比例(28．1％)和菌阳比例(35．9％)高。结论采用症状查痰法比胸透筛查法发现的活动性肺结核患者增加，特别是菌阳肺结核患者的发现率增加21．2％，使这些患者得到及时诊断，有利于结核病传染源的控制。     

Value of smear and PCR in bronchoalveolar lavage fluid in culture positive pulmonary tuberculosis.

At present, further investigations are needed in patients with suspected pulmonary tuberculosis (TB) and either negative sputum smear or without sputum. The aim of the present study was to analyse the yield of bronchoalveolar lavage fluid (BALF) smear and PCR in patients with confirmed pulmonary TB. Patients with a positive culture for Mycobacterium tuberculosis complex in sputum or BALF were analysed over 5 yrs. In total, 90 out of 230 (39%) patients with culture-positive pulmonary TB had a positive sputum smear, and 120 patients underwent bronchoscopy. BALF smear was positive in 56 (47%), BALF PCR in 93 (78%) patients, and BALF smear and/or PCR was positive in 83%. In total, 71 patients who underwent bronchoscopy and had complete clinical records were further analysed. BALF (smear or Mycobacterium tuberculosis complex-PCR) allowed a rapid diagnosis in 10 (59%) out of 17 patients who had a negative sputum smear, and 49 (91%) out of 54 patients without sputum production. Of these 71 patients, 12 (17%) were only culture positive. Rapid diagnosis of pulmonary TB by smear and/or PCR was made in 190 out of 210 patients (90%) in sputum or BALF. In conclusion, combined use of bronchoalveolar lavage fluid smear and Mycobacterium tuberculosis complex-PCR has a good diagnostic yield in patients with sputum smear-negative tuberculosis or without sputum production.     

Short-term bleach digestion of sputum in the diagnosis of pulmonary tuberculosis in patients co-infected with HIV

The bleach digestion of sputum may improve the yield of smear microscopy but has not been validated in patients with HIV. Therefore we assessed the performance of bleach-digested smear microscopy among patients with HIV. One thousand three hundred and twenty one patients with chronic cough submitted three sputum samples for direct smear microscopy and were offered HIV tests. One sample was selected for a bleach-digested smear and another one was cultured. Patients were classified as having 'definite' (>or=2 positive smears), 'very likely' (smear-negative, culture- positive), 'less likely' (one smear-positive, culture-negative) and 'unlikely' (smear and culture negative) tuberculosis (TB). In all, 566/1045 (54%) patients were HIV positive and 731/1186 (62%) were culture positive. The digested smears were positive in 123/125 (98%) 'definite', 4/118 (3%) 'very likely' and 1/174 'unlikely' TB patients with HIV and in 125/127 (98%) 'definite', 2/74 (3%) 'very likely', 4/4 'less likely' and 2/127 'unlikely' TB without HIV. Three direct smears identified 252 (57%) and one digested smear 254 (57%) of the 444 patients with 'definite' or 'very likely' TB. One bleach-digested smear performed similarly to three direct smears. Both methods were less sensitive in HIV-positive patients. Further studies are needed to compare the performance of the two methods under operational conditions.     

Laboratory diagnosis of pulmonary tuberculosis in TB and HIV endemic settings and the contribution of real time PCR for M. tuberculosis in bronchoalveolar lavage fluid

Background Tuberculosis (TB) in Africa is increasing because of the human immunodeficiency virus (HIV) epidemic, and in HIV/AIDS patients it presents atypically. Pulmonary tuberculosis (PTB) in Africa is mainly diagnosed clinically, by chest radiograph or by sputum smear for acid fast bacilli (AFB). Methods We evaluated in 120 HIV‐infected patients with chest infection the diagnostic accuracy of AFB smear of sputum and bronchoalveolar lavage (BAL) fluid, sputum Mycobacterium tuberculosis (MTB) culture, real‐time PCR and MycoDot^® serological test, using MTB culture of BAL fluid as gold standard. We correlated PCR cycle threshold values (C_T) to the culture results. Retrospectively, we evaluated the development of active TB in patients with positive PCR but negative culture. Results Culture of BAL fluid identified 28 patients with PTB. Fifty‐six patients could not produce adequate sputum. Sputum AFB smear and the serological test had sensitivities of 66.7% and 0%, respectively. PCR with C_T 40 was positive in 73 patients, 27 of whom were also TB culture positive (96.4% sensitivity and 52.3% specificity of PCR). PCR with C_T 32 had sensitivity of 85.7% and specificity of 90.9% to diagnose PTB in BAL. No patients with positive PCR but negative culture developed active TB during 18 months follow‐up. Conclusion In these HIV‐infected patients, AFB smear and serology had very low sensitivities. PCR of BAL with C_T value 32 had improved specificity to diagnose active PTB. A prospective follow‐up study is warranted in TB/HIV endemic settings, applying real time PCR to both sputum and BAL.     

交叉引物恒温扩增检测技术在疑似肺结核患者临床诊断中的价值

目的 评估交叉引物恒温扩增(cross priming amplification, CPA)检测技术在肺结核早期临床诊断中的应用价值。 方法 以2016年1—10月广东省佛山市和江门市及所辖7个县(区)级结核病防治机构(简称“结防机构”)就诊的初诊疑似肺结核患者为研究对象,共纳入患者6507例;所有患者均进行了胸部影像学检查并送检痰标本进行涂片镜检、固体培养和CPA检测,培养阳性菌株进行菌种鉴定,项目点临床医生结合实验室和临床检查结果对纳入患者进行初诊诊断,国家级临床专家对初诊临床诊断结果进行现场复核。分析项目地区确诊活动性肺结核患者中病原学阳性患者所占比例,并与专家复核后的临床诊断结果进行比较,分析涂片镜检、固体培养和CPA诊断肺结核的敏感度及特异度。 结果 6507例疑似肺结核患者中,涂片镜检、固体培养和CPA检测阳性率分别为18.2%(1187/6507)、25.0%(1629/6507)和23.9%(1555/6507)。3199例临床确诊为活动性肺结核患者中,涂片联合培养阳性检出率为48.5%(1550/3199),涂片联合CPA阳性检出率为47.9%(1531/3199),涂片、培养和CPA三种方法联合检测阳性率为54.4%(1740/3199)。以临床诊断结果为标准,涂片镜检、固体培养和CPA检测诊断活动性肺结核的敏感度分别为35.0%(1120/3199)、46.4%(1485/3199)和44.6%(1428/3199),特异度分别为98.0%(3241/3308)、95.7%(3164/3308)和96.2%(3181/3308)。 结论 县(区)级结防机构可应用CPA检测技术用于疑似肺结核患者的快速诊断,多种诊断技术联合应用可显著提高活动性肺结核患者的病原学诊断阳性率。     

联检脑脊液中LAM-IgG和TB-DNA在诊断结核性脑膜炎中的应用

目的　了解脑脊液 (CSF)中阿拉伯糖甘露糖脂IgG抗体 (LAM-IgG)和TB-DNA指标对结脑的诊断价值。方法 以CSF为标本,用酶联免疫吸附试验 (ELISA)检测LAM-IgG,用聚合酶链反应(PCR)检测TB-DNA。结果 102份结脑病人CSF标本,LAM-IgG阳性率51.0% (52102),TB-DNA阳性率81.4% (83102),LAM-IgG阳性及 或TB-DNA阳性共93例 (91.2%)。40份非结核性的中枢神经系统疾病病例的CSF标本,均未检出LAM-IgG和TB-DNA.结论 LAM-IgG和TB-DNA均是诊断结脑的较好的指标,两者联检可进一步提高检测敏感性。     

Comparative study of PCR, smear examination and culture for diagnosis of cutaneous tuberculosis

Diagnosis of tuberculosis (TB), especially cutaneous TB by conventional laboratory method is unreliable and time consuming. We assessed the utility of Polymerase Chain Reaction (PCR) test vis a vis other laboratory tests in 37 clinical samples of skin biopsy from equal number of patients with different variants of cutaneous TB. The PCR test amplifying 165bp region of 65kDa antigen coding gene specific for M. tuberculosis was performed on skin biopsy samples obtained from cases with a strong clinical evidence of cutaneous TB. The samples were also subjected to other laboratory tests e.g. smear examination, conventional (LJ based culture) and rapid BACTEC culture and histopathological examination for mycobacteria. Significant difference (p<0.05) was observed in the sensitivity of PCR test vis-a-vis other tests e.g. smear examination, LJ and BACTEC culture. PCR test showed a higher sensitivity than histopathological examination but the difference was not found to be statistically significant (p>0.05). PCR test showed the maximum positivity of 79.4% followed by histopathology (73.5%), BACTEC culture (47.5%), LJ media culture (29.4%) and smear examination (5.8%). The sensitivity and specificity of PCR test employing culture as the "gold standard" were 95.2% and 100%. The mean time taken for a positive result in different tests were less than 24 hours for smear examination, 1 day for PCR test, 23.42 days for BACTEC culture and 38.02 days for LJ culture. These results show that PCR amplification of 165bp region of 65kDa antigen coding gene of M. tuberculosis is a rapid and sensitive test for diagnosis of cutaneous TB using skin biopsy samples.     

昆明地区结核分枝杆菌标本培养检测及耐药情况分析

目的 了解昆明地区结核病患者和疑似结核病患者中Mtb培养检测情况和耐药情况。方法 收集1928例结核病患者和疑似结核病患者2046份标本,包括痰、支气管刷检物或肺泡灌洗液、胸腹腔积液、脑脊液、尿液、脓液、其他标本（淋巴液、心包积液、分泌物、穿刺液等）。使用BACTEC MGIT 960全自动分枝杆菌快速培养仪和中性罗氏培养基进行培养,分别记录初始抗酸涂片结果,BACTEC MGIT 960全自动分枝杆菌快速培养仪、中性罗氏培养基报告阳性后的涂片结果。分别对培养时间、培养阳性率、培养污染率及5种一线抗结核药物（S、INH、RFP、EMB和PZA）的耐药情况作统计分析。结果 BACTEC MGIT 960全自动分枝杆菌快速培养仪和中性罗氏培养基阳性率分别为38.2%（782/2046）和32.6%（666/2046）;结核诊断率37.3%（719/1928）;抗结核一线药耐药率为41.9%（212/506）,耐多药率（至少同时耐INH和RFP）为20.0%（101/506）。结论 昆明地区结核病患者对一线抗结核药物耐药情况严重。     

结核抗体IgG检测辅助诊断结核病的应用价值

目的 探讨结核抗体IgG检测(简称“IgG检测”)辅助诊断结核病的应用价值。方法 收集黑龙江省传染病防治院2015年7月至2016年5月期间,具有IgG检测、抗酸杆菌涂片(简称“涂片”)镜检、结核分枝杆菌液体培养(简称“液体培养”)、CT扫描及临床诊断等资料的住院及门诊患者,共计1494例,其中继发性肺结核1020例(肺结核组)、肺外结核54例(肺外结核组)和排除结核病的其他肺部疾病420例(其他肺病组),对比分析不同患者的临床资料。结果 1020例肺结核患者和54例肺外结核患者的IgG检测阳性率分别为73.33%(748/1020)、62.96%(34/54),高于涂片镜检[分别为43.24%(441/1020)、20.37%(11/54)]和液体培养[分别为61.37%(626/1020)、29.63%(16/54)](χ ²=190.02,P<0.001;χ ²=20.15, P<0.001; χ ²=33.18,P<0.001; χ ²=12.07,P=0.001);IgG检测肺结核与其他肺病患者的阳性率(6.90%,29/420)差异有统计学意义(χ ²=553.47,P<0.001)。IgG检测1440例肺部疾病患者的敏感度、特异度、总符合率分别为73.33%(748/1020)、93.10%(391/420)、79.10%(1139/1440)。1020例肺结核患者中,涂片镜检阳性者的IgG检测阳性率(86.85%,383/441)与涂片镜检阴性者的IgG检测阳性率(63.04%,365/579),以及液体培养阳性者的IgG检测阳性率(80.51%,504/626)与液体培养阴性者的IgG检测阳性率(61.93%,244/394)的差异均有统计学意义(χ ²值分别为72.56、42.70,P值均<0.001)。分别以涂片镜检和液体培养为标准,IgG检测1020例肺结核组患者的敏感度、特异度、总符合率分别为86.85%(383/441)和80.51%(504/626) 、36.96%(214/579)和38.07%(150/394)、58.53%(597/1020)和64.12%(654/1020)。1020例肺结核组患者中涂阴培阴(菌阴)肺结核患者为372例(36.47%),其IgG检测阳性率为61.29%(228/372),阳性患者的CT扫描表现以斑片、条索状阴影(83.77%,191/228)多见。420例其他肺病患者IgG检测假阳性率为6.90%(29/420),与肺部感染(62.07%,18/29)和肿瘤(13.79%,4/29)患者有少量交叉反应。 结论 结核抗体IgG检测具有较高的敏感度和特异度,对结核病,尤其是对菌阴肺结核和肺外结核检出具有较高的辅助诊断价值。