Objective:To evaluate the clinical value of pathogen detection of pulmonary tuberculosis using bronchial lavage fluid as sample.Methods:The data of laboratory information system from Guangzhou Chest Hospital between January and December, 2019 were used; and the results of auramine O fluorescent stain smear and mycobacterial isolation of sputum and bronchial lavage fluid were comparatively analyzed.Results:The positive rates of 115 bronchial lavage fluid smears or 307 sandwich cup collection smears of bronchial lavage fluid were significantly lower than those of the sputum smears ( P<0.001 and P=0.049). The positive rates of Mycobacteria isolated from bronchial lavage fluid and sputum with the MGIT 960 system from the 882 patients were 37.53% and 33.90%, respectively. 89 patients who were negative in sputum culture were positive in the culture of bronchial lavage fluid, accounting for 10.09%, with a statistical difference ( χ2=353.555, P<0.001). Conclusion:Isolating mycobacterial from bronchial lavage fluid is an important method for the diagnosis of pulmonary tuberculosis, but the value of stain smear test is small.

Objective:To evaluate the value of detecting DNA and RNA of Mycobacterium tuberculosis in bronchial lavage fluid for diagnosis of tuberculosis.Methods:The data of 1 243 cases from Guangzhou Chest Hospital in 2019, who were performed bronchoscopy and their samples of bronchial lavage fluid were tested for isolation of Mycobacteria and DNA and RNA detection of Mycobacterium tuberculosis at the same time, were collected and analyzed respectively.Results:The four combinations of detection including "isolation positive plus DNA positive plus RNA positive" , "isolation positive plus DNA positive plus RNA negative" , "isolation negative plus DNA negative plus RNA positive" , and "isolation negative plus DNA negative plus RNA negative" were presented. And their percentage composition were 20.51% (255/1 243), 13.27% (165/1 243), 8.53% (106/1 243), 57.68% (717/1 243) among all of 1 243 cases, while they were 33.16% (255/769), 21.46% (165/769), 10.40% (80/769), 34.98% (269/769) among 769 patients with pulmonary tuberculosis. Compared with 50.00% (290/580) positive ratio of both isolation and DNA detection among patients with non-cavernous pulmonary tuberculosis, it was 68.78% (130/189) among patients with cavernous pulmonary tuberculosis, the latter was significantly higher than the former (χ 2=20.293, P<0.01). Conclusion:DNA detection of Mycobacterium tuberculosis in bronchial lavage fluid is worthy of extensive promotion and application in clinical diagnosis and treatment because its performance is equivalent to isolation of Mycobacteria and it is especially rapid for getting test result compared with isolation of Mycobacteria.

目的：深入了解中国结核杆菌利福平耐药株rpoB基因531、526、516三个高突变率发生位点的碱基突变特点，为全面理解结核杆菌利福平耐药机制提供科学依据。方法计算机检索PubMed数据库，对截止2015年3月31日发表的中国结核杆菌利福平耐药株rpoB基因突变研究的文献数据进行整合分析。结果纳入分析文献49篇，获得3319株利福平耐药结核杆菌 rpoB 基因各位点碱基突变的详细信息，其中257株（7．74％）未检测到突变、3062株（92．26％）检测到rpoB基因突变。在3062株 rpoB 基因突变株中：531、526、516位点突变的发生率分别为54．44％（1667／3062）、27．07％（829／3062）和8．92％（273／3062）；531位点碱基突变以TCG／TTG突变为主，占93．58％（1560／1667）；526位点碱基突变主要有 CAC／GAC、CAC／TAC、CAC／CGC、CAC／CTC、CAC／AAC 等类型，分别占33．53％（278／829）、26．30％（218／829）、13．27％（110／829）、10．13％（84／829）、8．08％（67／829）；516位点碱基突变主要有GAC／GGC、GAC／GTC、GAC／TAC等类型，分别占35．53％（97／273）、35．53％（97／273）、16．85％（46／273）。结论中国结核杆菌利福平耐药株 rpoB 基因531、526、516位点碱基突变以单碱基突变为主，突变率最高的531位点碱基突变主类型较为单一，而526、516位碱基突变主类型则相对分散。

目的 了解结核分枝杆菌、鸟-胞内分枝杆菌复合群、堪萨斯分枝杆菌、戈登分枝杆菌、瘰疬分枝杆菌液体培养早期的蛋白质代谢与分泌活动,以进一步加深对它们致病性的理解,并初步分析由此途径寻找准确、快速鉴定这些菌种的新型标志物的可能性.方法 采用Middlebrook 7H9液体培养基对目的细菌临床分离株进行培养,应用表面增强激光解析-电离飞行时间质谱(SELDI-TOF-MS)技术对7天培养滤液和Middlebrook 7H9液体培养基蛋白质组进行检测,描述性分析其蛋白质组.结果 在Middlebrook 7H9液体培养基与结核分枝杆菌、鸟-胞内分枝杆菌复合群、堪萨斯分枝杆菌、戈登分枝杆菌、瘰疬分枝杆菌7天培养滤液中分别检测到106、164、118、124、109、210种蛋白质/多肽,依据质荷比(m/z)其相对分子量范围分别为1045～3938、982 ～ 9329、1035～4579、1052 ～ 9109、1035 ～ 3940、998 ～ 9329,其表达丰度分别为86 ～ 7838、7～ 210、63 ～ 5730、58 ～ 5559、74～6866、6～190;与Middlebrook 7H9液体培养基蛋白相比,结核分枝杆菌、鸟-胞内分枝杆菌复合群、堪萨斯分枝杆菌、戈登分枝杆菌、瘰疬分枝杆菌7天培养滤液中分别存在157、106、114、99、205种差异蛋白质.结论 结核分枝杆菌、鸟-胞内分枝杆菌复合群、堪萨斯分枝杆菌、戈登分枝杆菌、瘰疠分枝杆菌在繁殖生长早期即存在活跃的蛋白质代谢与分泌活动,而且此5种不同分枝杆菌早期培养滤液中蛋白质种类与表达丰度差异明显.

Objective To find the specific secretory proteins of Mycobacterium tuberculosis resistant to isoniazid. Methods The 7-day and 14-day cultures filtrates of a clinical isolate of isoniazid-resistant Mycobacterium tuberculosis and a clinical isolate of isoniazid-sensitive Mycobacterium tuberculosis growing in Middlebrook 7H9 were captured by weak cation exchange protein chip (WCX2) and detected by surface-enhanced laser desorption/ionization time-of-flight mass spectrometry (SELDI-TOF-MS).Differences were analyzed with descriptive statistics method.Results One hundred sixty four and one hundred seventy five proteins existed in the 7-day and 14-day cultures filtrates of isoniazid-sensitive Myco-bacterium tuberculosis, while 133 and 147 proteins were detected respectively for clinical isolate of isoniazid-resistant Myco-bacterium tuberculosis.Depending on the ratio of mass, their relative molecular masses to charge were 982 ~9328, 997 ~9327, 1050~8903 and 996 ~9369 and the abundance represented relative intensity value calculated by mass spectrometry peak area were 17~2100,21~1943,60~5355 and 41~3267 respectively.Compared with isoniazid-sensitive Mycobacteri-um tuberculosis strain, there were 47 and 58 different proteins in the 7-day and 14-day cultures filtrates of isoniazid-re-sistant Mycobacterium tuberculosis strain.Their relative molecular masses range from 1 083 to 8 812 and from 1 012 to 9327, and levels of abundance were in the range of 76 to 1 811 and 25 to 3 010.Conclusion There are differences of proteins in the culture filtrate during the early and middle of culture courses between isoniazid-resistant and isoniazid-sensitive Myco-bacterium tuberculosis strain.But it remains to be further confirmed whether these different proteins come from secretory pro-tein of Mycobacterium tuberculosis or not.

目的:初步了解结核杆菌体外液培养条件下的早期蛋白质代谢和分泌活动状况.方法:将2株对抗结核一线药物敏感的结核杆菌临床分离菌接种Middlebrook 7H9液体培养基进行培养,并于培养第7天用0.22 μm滤纸过滤培养液制备早期培养滤液,应用与弱阳离子交换蛋白质芯片(WCX2)捕获滤液蛋白行表面增强激光解吸-电离飞行时间质谱(SELDI-TOF-MS)检测,描述性分析比较其蛋白质组.结果:在2株结核杆菌7d培养滤液中分别检测到相对分子质量为982 ～9 329与941～9 331、丰度为7～ 210与11～368(通过质谱峰面积计算相对强度值表示)的164与194种蛋白质；与Middlebrook 7H9液体培养基本身蛋白质组比较,在2株结核杆菌7d培养滤液中分别有153和183种蛋白质新产生(其中10种在2份滤液中均有存在)、92和90种蛋白质消失、10和13种蛋白质丰度变小.结论:在体外液体培养条件下,结核杆菌蛋白质代谢和分泌活动活跃,且不同菌株间有较大差异.

目的本研究主要针对从广东地区500例疑似结核病患者痰液中分离的结核分枝杆菌对异烟肼(INH)、利福平(RFP)、链霉素(SM)耐药情况与其对应的突变基因katG、rpoB、rpsL关系研究。方法采用PCR和DNA测序法对结核分枝杆菌临床分离株katG、rpoB和rpsL基因进行序列分析。结果分离得到耐INH35株、耐RFP27株、耐SM25株。INH耐药株katG基因突变率为71.4%(25/35),RFP耐药株rpoB基因的突变率为81.5%(22/27),SM耐药株rpsL基因突变率为76%(19/25);INH以Ser315Thr突变60%(21/35)最常见,RFP以Ser531Leu突变55.6%(15/27)最常见,SM以Lys43Arg位氨基酸突变72%(18/25)常见。结论 katG、rpoB和rpsL基因突变被证实是广东地区结核分枝杆菌INH、RFP和SM耐药性产生的主要分子机制,PCR-DNA测序法可快速检测结核分枝杆菌INH、RFP和SM耐药性。

目的:对从疑似结核病患者痰液中分离的结核分枝杆菌的rpoB和rpsL基因突变情况进行研究,分别探讨其与耐利福平(RFP)和耐链霉素(SM)之间的关系。方法:采用PCR和DNA测序法对结核分枝杆菌临床分离株和1株卡介苗株(BCG)的rpoB和rpsL基因进行序列分析。结果:分离得到耐利福平27株,耐链霉素25株,经测序分析,所有利福平、链霉素敏感株rpoB、rpsL基因均未发生氨基酸突变;RFP和SM耐药株rpoB和rpsL基因的突变率各为81.5%(22/27)和76%(19/25);耐利福平以Ser531Leu突变最为常见,突变频率为55.6%(15/27),耐链霉素以Lys43Arg位突变最为常见,突变频率为72.0%(18/25)。结论:rpoB和rpsL基因突变分别是结核分枝杆菌RFP和SM耐药性产生的主要分子机制,PCR-DNA测序法可快速检测结核分枝杆菌RFP和SM耐药性。

目的:比较分析菌阳肺结核与菌阴肺结核、肺结核与肺外结核患者的血清蛋白质谱,从蛋白质组学层面了解不同结核病患者群血清蛋白质谱的特征,为结核病诊断标志物的筛选提供指导.方法:应用表面增强激光解吸-电离飞行时间质谱(SELDI-TOF-MS)蛋白质芯片系统,采用弱阳离子交换蛋白质芯片(WCX)对54例菌阳肺结核、45例菌阴肺结核、18例肺外结核患者的血清蛋白质进行检测;应用ClinProTools分析软件依据各蛋白质的质谱峰面积计算相对强度表示该血清蛋白质的丰度,采用t检验(对Anderson-Darling检验P>0.05的正态分布者)或Wilcoxon检验(对Anderson-Darling检验P<0.05的非正态分布者)分析比较茵阳肺结核与茵阴肺结核、肺结核与肺外结核患者各血清蛋白质的平均丰度.结果:(1)在3种不同结核病患者人群中均捕获到102种相对分子质量在1 011.41～9 435.07的不同血清蛋白质,经Anderson-Dading检验,在菌阳肺结核、菌阴肺结核、肺外结核患者人群中分别有31、23和29种血清蛋白质呈正态分布.(2)正态分布的血清蛋白质在不同结核病患者人群间其水平未见显著差异.(3)非正态分布的血清蛋白质:在菌阳肺结核与菌阴肺结核患者人群间其水平未见显著差异;在肺结核与肺外结核患者人群间发现1个差异血清蛋白质(P=0.025 1),其相对分子质量为5 753.76,在肺结核与肺外结核患者人群血清中的平均丰度分别为15.42±4.86和28.29±14.08.结论:不同结核病患者群的血清蛋白质谱具有同质性,为普遍适用性结核病血清学诊断标志物的筛选提供了科学依据.

目的 分析广州地区临床分离的非结核分枝杆菌(NTM)的菌种鉴定结果及耐药情况.方法 用改良罗氏培养基培养法分离680例疑似结核病患者痰液,对确诊为NTM的38例,用传统生化反应进行菌种鉴定,采用以罗氏培养基为基础的绝对浓度法做NTM药物敏感试验.结果 38株NTM按Runyon分类法,检出感染菌种分布为Ⅰ群1株(2.6%),Ⅱ群7株(18.4%),Ⅲ群18株(47.4%),Ⅳ群12株(31.6%);单一耐药有13株(34.2%),且只耐异烟肼;同时耐2种药的比例为10.5%(4/38);同时耐3种药的比例为10.5%(4/38);同时耐4种药耐药率为44.7%(17/38).大多数NTM对常用的抗结核药物呈高度耐药.结论 广州地区NTM感染菌种以鸟-胞内分枝杆菌复合群分离率比例最高,为44.7%(17/38);耐药方式以单一耐异烟肼和同时耐4种药物为主;NTM对常用的抗结核药物呈高度耐药现象.

Objective To evaluate the sensitivity of rapid growing Mycobacterium,including M.abscessus,M.chelonae and M.fortuitum,to various anti-tuberculosis drugs.Methods The sensitivity of mycobacteria to anti-tuberculosis drugs was evaluated using the Absolute Concentrate Method.11 anti-tuberculosis drugs were used in this study.Two different concentrations(μg /mL) were used for each drug.These included Isoniazid(1 /10),Rifampin(50 /250),Streptomyain(10 /100),Ethambutol(5 /50),Amikacin(10 /100),Pasiniazid(1 /10),Capreomycin(100 /1000),Ciprofloxacin(1 /10),Moxifloxacin(1 /10),Levofloxacin(1 /10) and Clarithromycin(1 /10).Results A total of 31 strains of M.abscessus,20 strains of M.chelonae and 9 strains of M.fortuitum were studied.The sensitivity of M.abscessus,M.chelonae and M.fortuitum to clarithromycin were 87.1,75.0 and 77.8,respectively.The sensitivity of M.abscessus,M.chelonae and M.fortuitum to Amikacin were 22.6,55.0 and 33.3,respectively.Moreover,these 3 species of mycobacteria were found to be sensitive to high,but not low,concentration of Ethambutol,Streptomycin and Ciprofloxecin.Conclusion All three species of M.abscessus,M.chelonae and M.fortuitum were found to be sensitive to clarithromycin.

目的对医院内不同实验室或同一实验室不同生化检测系统检测结果进行比对分析和偏差评估,实现医院内不同生化检测系统检测结果一致性。方法参考美国临床实验室标准化委员会(NC-CLS)的EP15-A文件,以Roch Modular P800检测系统(S)为目标检测系统,Roch COBAS 400检测系统(1、2)、CL8000检测系统(3)为实验检测系统,检测患者血清TBIL、ALT、AST含量,进行比对研究和统计学分析,计算偏倚(SE%),实验检测系统比对项目取偏倚最高者与1/4CLIA'88允许误差进行比较,偏倚(SE%)小于等于1/4CLIA'88允许总误差(TBIL、ALT、AST1/4CLIA'88允许总误差均为5%),达到该项目院内检测结果一致性目的。结果①用原校准品校准各检测系统进行比对分析,TBIL、ALT、AST偏倚(SE%)分别为5.6%、19.2%、5.3%②采用目标检测系统定值的新鲜患者混合血清作为临时校准物校准各实验检测系统,第一次比对结果TBIL、ALT、AST偏倚(SE%)分别为5.3%、7.5%、4.7%;第二次比对结果TBIL、ALT、AST偏倚(SE%)分别为4.0%、4.1%、3.6%,实验检测系统比对项目SE%均小于5%,检测结果达院内一致性要求。结论采用目标检测系统定值的患者新鲜混合血清作为临时校准物,校准各实验检测系统的比对分析是实现医院内不同生化检测系统检测结果一致性的有效途径。

目的 对536株耐多药结核分枝杆菌的药敏情况进行分析,从而获得耐多药结核分枝杆菌的耐药趋势,指导临床合理用药.方法 采用WHO推荐的药敏比例法对临床分离结核分枝杆菌进行药物敏感性试验,选择其中的536株耐多药分离株进行耐药情况分析.结果 536株耐多药菌株中,耐HR的共178例(33.2%),耐HRS的共163例(20.4%),耐HRE的共61例(11.4%),耐HRSE的共134例(25.0):不同年度的耐多药患者构成情况有所不同.结论 广州市旧城区结核分枝杆菌的耐多药形势日趋严重,耐HRS菌株不断增加,结核病患者耐多药的性别比例趋向平衡。

目的评估奥蜜佳结核抗体检测试剂的实际应用价值.方法①应用重复性实验、标本稀释试验评价其方法学;②应用敏感性、特异性、阳性预测值、阴性预测值等指标评价其临床应用价值.结果①92例标本3次重复性实验阴阳性结果总体符合率为100%;②20例标本3个稀释度稀释试验示检测线性良好;③832例相关人群标本检测的敏感性、特异性、阳性预测值、阴性预测值分别为:62.6%, 92.2%, 95.4%, 48.6%.结论奥蜜佳结核抗体检测试剂方法学质量保证,并以其高的特异性和中度敏感性对结核病的诊断和鉴别诊断有一定的参考价值,但其敏感性有待进一步提高,操作方法亦需进一步简化.