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1.
In this study two molecular typing methods, a simple double repetitive element PCR-based assay and the standardized restriction fragment length polymorphism (RFLP), were used to confirm cross-contamination in the mycobacteriology laboratory. Clinical specimens from 12 patients, submitted for acid-fast bacilli stain smear and processed for culture in Lowenstein-Jensen on the same day, resulted in positive bacterioscopy (+++) and confluent growth only for one of the patients. The specimens from all the other patients but two were smear-negative and culture-positive, with one or two colonies. None of them had clinical symptoms and radiological findings for active tuberculosis (TB). The suspicion of false-positive cultures arose when a health care worker who had had a PPD skin test conversion, claimed to be healthy and had no TB symptoms, was found to have a positive sputum culture. DRE-PCR demonstrated that all nine cultures typed belonged to one cluster, further confirmed by RFLP. Although DRE-PCR has been found to be poorly reproducible, it has enough discriminatory power to be useful for rapid epidemiological investigation in selected settings.  相似文献   

2.
Abstract The objective of this study was to evaluate the utility of a polymerase chain reaction (PCR) assay in detecting Mycobacterium tuberculosis in bronchoalveolar lavage (BAL) specimens of patients suspected of having active pulmonary tuberculosis (TB) but who were sputum smear-negative. Patients undergoing investigation for suspected pulmonary TB at the University Hospital, Kuala Lumpur, and who were sputum smear-negative underwent fibreoptic bronchoscopy and BAL. One portion of each lavage specimen was submitted for smear examination for acid-fast bacilli and mycobacterial culture and the other portion assayed by PCR for the presence of a 562-base pair DNA segment belonging to the insertion sequence IS986, unique to the M. tuberculosis complex. As controls, lavage specimens from patients with other lung lesions were also similarly tested. The PCR assay gave a positivity rate of 80.9% (55 of 68) compared with 8.8% of smear examination and 7.4% of culture for detecting M. tuberculosis in BAL specimens. The assay was positive in two of 45 BAL specimens from 35 control subjects. The PCR assay was more sensitive than smear and culture in detecting M. tuberculosis in BAL specimens of patients with sputum smear-negative pulmonary TB.  相似文献   

3.
实时荧光PCR技术在不同类型结核病诊断中的应用价值   总被引:1,自引:0,他引:1  
目的 分析比较实时荧光PCR与涂片镜检、培养三种方法在结核杆菌检测的差异.方法 对我院疑似结核病患者的临床标本同时进行实时荧光PCR检测、涂片镜检和培养,将三组检测结果进行比较.结果 215例疑似肺结核患者痰标本同时采用实时荧光PCR、涂片镜检和培养三种方法检测,阳性检出率分别为46.5%、32.6%和44.7%.实时荧光PCR法与涂片镜检法的阳性率间比较有统计学意义,实时荧光PCR法与培养法的阳性率同比较无统计学意义.112例疑似其他类型结核病患者的胸腹水、脑脊液、穿刺液和尿液等非痰标本同时采用上述三种方法检测,阳性率分别为18.8%、3.6%和16.1%.统计学意义同上.结论 实时荧光PCR检测技术在临床上对结核病的诊断和鉴别具有较好的应用价值.  相似文献   

4.
Factors affecting the clinical value of microscopy for acid-fast bacilli   总被引:14,自引:0,他引:14  
In order to assess the clinical value of microscopy for acid-fast bacilli (AFB), the results of 3,207 clinical specimens submitted for mycobacterial smear and culture were analyzed. Mycobacteria grew from 176 (5.5%) of the specimens, 95 (54%) of which were Mycobacterium tuberculosis. Although the overall sensitivity of the smear was low (33%), 65% of respiratory specimens yielding M. tuberculosis had positive AFB smears. Furthermore, 96% of patients with pulmonary tuberculosis from whom more than one specimen was processed had at least a single positive AFB smear. Smear sensitivity correlated well with quantitative growth; 89% of specimens yielding greater than or equal to 50 colonies per slant were smear positive. Specificity of the AFB smear was high; 89% of smear-positive specimens had positive cultures. After the results from culture-negative patients known to have active tuberculosis were eliminated from the analysis, the specificity of a positive smear rose to 98.3%. When the results of all specimens from each patient were considered in toto, the AFB smear had a predictive value of greater than or equal to 96%.  相似文献   

5.
目的 进行环介导等温扩增法(LAMP法)快速检测结核分枝杆菌的临床验证与应用,评估该方法在结核病临床诊断中的效能。方法 选取广东6地市专业结核病防治机构(简称“结防机构”)的肺部不同疾病及健康人员的痰标本2129份,分别采用直接涂片法、罗氏固体培养法及LAMP法检测痰标本中结核分枝杆菌;另广州、汕头两市337例结核病患者的痰标本浓缩处理后进行实时荧光定量(qPCR)及LAMP两种方法检测。 结果 2129例痰标本中,直接涂片法、罗氏固体培养法和LAMP法的阳性检出率分别为13.2%(282/2129)、21.7%(463/2129)和20.3%(432/2129);同罗氏培养法相比,LAMP法灵敏度、特异度、阳性和阴性预测值分别为89.1%(432/485)、95.5%(1570/1644)、85.4%(432/506)和96.7%(1570/1623);337例浓缩处理后的痰标本qPCR阳性率为35.0% (118/337),LAMP阳性率为35.9% (121/337);LAMP法同直接涂片法检测、罗氏固体培养法和qPCR法比较,检测结果实际一致率分别为87.9%(Kappa值=0.612)、96.1%(Kappa值=0.89)和91.4%(Kappa值=0.812)。 结论 LAMP法用于痰标本中结核分枝杆菌检测,其效能同罗氏培养法及qPCR法相当,同直接涂片法相比,LAMP法能够大幅提高阳性检出率,具有很好的临床应用和推广前景。  相似文献   

6.
During intensive bacteriological follow-up of 2123 patients who had received short course chemotherapy regimens in 4 controlled clinical trials in Africa and had not had a bacteriological relapse, 405 isolated positive cultures were obtained from 37 429 sputum specimens in 3 East African laboratories. These cultures might have arisen as a result of clerical error, of transfer of bacilli from positive to negative specimens in the laboratory or from the lesions of the patients. Clerical error in the labelling of specimens or the recording of results did not seem a frequent cause, since isolated positive cultures contained many fewer colonies than cultures from other positive specimens being processed at the same time. Several of evidence suggested that some isolated positive cultures arose from the lesions of patients: a decrease in their incidence occurred in successive time periods after chemotherapy; the number of isolated positives per patient departed significantly from the Poisson distribution; they were more often drug resistant than other cultures processed at the time; positive cultures were obtained less frequently from known autoclaved specimens inserted among the the study specimens than from the study specimens themselves; no association was found between the incidence of isolated positive and of specimens containing numerous viable M. tuberculosis being processed at the same time. Nevertheless some of these cultures probably arose by transfer in the laboratory, since the rates at which transfer were known to occur differed in the 3 laboratories and corresponded to the rates of obtaining isolated positive cultures.  相似文献   

7.
目的 探讨在可疑肺结核病例中能否用两份痰标本涂片法代替三份痰标本涂片法.方法 对1002例可疑肺结核病例的三份痰标本(即时痰、夜间痰、清晨痰)分别进行直接涂片、萋-尼尔逊氏抗酸染色和光学显微镜镜检.结果 即时痰+夜间痰+清晨痰组与夜间痰+清晨痰组的结核阳性率无显著性差异.结论 即时痰不能提高结核阳性率,在可疑肺结核患者中可以用夜间痰+清晨痰两份标本涂片法代替原方案即时痰+夜间痰+清晨痰的三份标本涂片法.  相似文献   

8.
From 1972 to 1974, all sputum specimens and gastric aspirate specimens submitted to the University of Michigan Laboratory for acid-fast smear and culture were studied. Specimens were paired for culture and smear results using the auramine-rhodamine staining technique. Of 1,893 patients, 75 patients without prior antituberculous therapy were found to have either a positive smear or a positive culture of either sputum or gastric material. The data analyzed by patient source revealed the following. (1) Staining sputum with auramine-rhodamine is a clinically reliable technique for detecting pulmonary tuberculosis. It demonstrates a sensitivity of 78 per cent and a relative fraction of false positive smears of only 11 per cent. (2) Staining gastric-aspirated material by the auramine-rhodamine technique is not a clinically reliable method as a routine procedure for the detection of pulmonary tuberculosis, because of a sensitivity of only 58.8 per cent and a relative fraction of false-positive smears of 33 per cent. (3) In the absence of sputum in suspected clinical granulomatous disease, quantified gastric smears may be helpful. In this study, when more than 6 organisms per high power field were found, the patient's sputum or gastric material yielded a pathogenic mycobacterium on culture.  相似文献   

9.
TaqMan聚合酶链反应技术检测结核分支杆菌DNA及其临?…   总被引:23,自引:1,他引:22  
目的 探讨TaqMan聚合酶链反应(TaqMan-PCR)技术在肺结构诊断中的价值。方法 对168例活动性肺结核、57例肺癌患者的痰和外周血及34-例健康对照外周血,同时应用TaqMan-PCR、PCR检测,并与痰涂片法、BACTEC法及改良罗氏培养法结果进行比较。结果 TaqMan-PCR检测痰和外周血总的阳性率分别为53.0%和61.3%,显著高于PCR、痰涂片法、BATCTEC法及改良罗氏培  相似文献   

10.
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.  相似文献   

11.
SETTING: Sputum samples were collected from suspected tuberculosis patients attending out-patient clinics at the Ojha Institute of Chest Diseases, Karachi, Pakistan. OBJECTIVE: To evaluate the performance of the FASTPlaqueTB assay for rapid diagnosis of pulmonary tuberculosis. DESIGN: A comparative study of 584 sputum samples using acid-fast smear microscopy, Lowenstein-Jensen culture and FASTPlaqueTB. RESULTS: A total of 514 samples yielded complete results. Seventy specimens were lost to analysis due to the overgrowth of contaminants. The addition of antimicrobials inhibited growth of gram-positive contaminants, and reduced the overall contamination rate from 18.2% to 7.2%. Mycobacterium tuberculosis was isolated from 175 smear-positive and 70 smear-negative specimens. FASTPlaqueTB detected M. tuberculosis in 81.6% of specimens, with a specificity of 97.7%. The sensitivity and specificity of the assay for smear-positive specimens were respectively 87.4% and 88.2%. For smear-negative specimens, the sensitivity of the assay was 67.1% and the specificity was 98.4%. The combined sensitivity of smear and FASTPlaqueTB for M. tuberculosis was 90%. Test results were available in 48 hours. CONCLUSION: FASTPlaqueTB is a sensitive and specific test for rapid diagnosis of pulmonary tuberculosis in high prevalence areas. The test is sensitive enough to confirm a large number of clinically suspected smear-negative cases and improve case finding.  相似文献   

12.
目的 探讨异种血清抗体检测技术在结核病诊断中的应用价值。方法 采用IgG/IgM抗体试剂盒分别检测102例结核病患者(包括73例肺结核和29例肺外结核)、223例其他肺部疾病患者和100例对照者结核感染情况,以临床诊断为标准评价该方法的敏感度和特异性,同时分别与痰菌培养及痰涂片平行检验的结果作比较,统计学分析采用χ2检验。结果 结核抗体IgG/IgM检测结核病患者的敏感度为74.51%、特异度为91.64%。结核抗体IgG/IgM检测肺结核和肺外结核的敏感度分别为82.19%、55.17%,肺内和肺外结核的敏感度差异有统计学意义(P<0.05),结核抗体lgG/IgM检测结核患者阳性检出率明显高于痰培养法和痰涂片法(P<0.05)。102例结核病患者年龄段分组分析,少年组和老年组检出率分别为58.33%、36%,远低于青年组和中年组的96.15%和89.74%。不同年龄组间进行卡方比较分析显示,P<0.05,差异有统计学意义。425例标本中,共发现8例非结核分枝杆菌,其中6例胞内分枝杆菌, 2例脓肿分枝杆菌,lgG/lgM抗体检测均为阴性。结论 IgG/IgM血清抗体检测肺内、外结核具有快速方便、经济和较高的敏感度,适合用于临床结核筛查。  相似文献   

13.
目的 应用实时荧光PCR快速检测肺结核患儿粪便中Mtb-DNA,并初步评估其临床效果。 方法 收集肺结核住院儿童粪便标本76份,应用实时荧光PCR检测Mtb-DNA,检测结果与20例其他呼吸系统疾病患儿的粪便实时荧光PCR检测Mtb-DNA、76例粪便抗酸杆菌涂片检查以及41例患儿痰标本的涂片和(或)培养、实时荧光PCR检测结果进行比较。 结果 粪便实时荧光PCR检测敏感度达到23.68%(18/76),特异度为100.00%(20/20),肺结核患儿粪便PCR阳性率[23.68%(18/76)]明显高于涂片阳性率[6.58%(5/76)],41例患儿中痰涂片和(或)培养阳性例数(15例)和痰PCR检测阳性例数(18例)高于粪便PCR检测阳性例数(11例)。 结论 实时荧光PCR检测儿童粪便Mtb-DNA,是一种特异度高、比较敏感、非侵入性且相对安全的儿童肺结核快速诊断方法。  相似文献   

14.
A total of 832 respiratory specimens not including the sputum (402 bronchial lavages, 241 bronchial brushing specimens, 136 pumping lavages, 41 pleural effusions, and 12 others) from 462 patients were assayed using the Roche Amplicor Mycobacterium tuberculosis test for amplification and identification of M. tuberculosis, M. avium and M. intracellulare (Amplicor PCR). The results were compared with those obtained using conventional microscopy and cultivation methods. Each patient had little or no sputum and showed an abnormal chest X-ray shadowing of unknown cause. No patients had previously undergone antituberculous therapy. Of the specimens obtained, 24 were both PCR and culture positive, 786 were both PCR and culture negative, 11 were PCR positive and culture negative, and 11 were PCR negative and culture positive. Based on these results, the sensitivity and specificity of Amplicor PCR were determined to be 68.67% and 98.6%, respectively, when compared with culture of respiratory specimens not including the sputum. After correcting for discrepancies due to differences in patient clinical data, the sensitivity of Amplicor PCR was found to be 68.6%, and the specificity to be 99.9%; the corresponding values for culture were 66.7% and 100%, and those for smear were 9.8% and 100%. Thus, Amplicor PCR was shown to possess a similar sensitivity to culture and to be a highly specific technique for the diagnosis of tuberculosis in the respiratory system using non-sputum specimens within hours in patients showing little or no sputum and abnormal chest X-ray shadowing of an indeterminant cause.  相似文献   

15.
It was hypothesised that the time to detect Mycobacterium tuberculosis in liquid culture of sputum from patients with pulmonary tuberculosis may be a better indicator for the duration of respiratory isolation than sputum smear status. Pre-treatment and during-treatment sputum acid-fast bacilli (AFB) smear and culture results were reviewed in 284 patients with pulmonary tuberculosis. The time to detect M. tuberculosis in liquid culture (TTD-TB) was the number of days from inoculation of the Mycobacterial Growth Indicator Tube to culture detection and visualisation of AFB. The median (interquartile range) TTD-TB for smear group 0 (no bacilli seen) was 14 (12-20) days. This value was used as the standard at which release from isolation could be permitted. In smear group 4 (>9 AFB per high-power field (hpf) in sputum specimens before treatment) patients, the TTD-TB exceeded 14 days after a median of 25 days of treatment. The current authors recommend that patients in smear groups 1 and 2 (1-9 AFB per 100 hpf and 1-9 AFB per 10 hpf in sputum specimens before treatment, respectively) receive treatment in respiratory isolation for 7 days, provided the risk of drug resistance is low. Smear group 3 (1-9 AFB per hpf) and 4 patients should receive treatment in respiratory isolation for 14 and 25 days, respectively. These criteria would have reduced the duration of respiratory isolation by 1,516 days in the 143 study participants with sputum smear-positive pulmonary tuberculosis. Provided clinical and radiographical criteria are satisfactory, use of the time to detect Mycobacterium tuberculosis in liquid culture could enable the duration of respiratory isolation to be predicted from the pre-treatment sputum smear grade. The recommendations enable isolation to end well before sputum becomes smear negative, with considerable benefits to patients and healthcare providers.  相似文献   

16.
Abstract
Background:  The misdiagnosis of Mycobacterium tuberculosis infection has many ramifications. These include medical and psychological implications for patients and their families and financial and public health implications for health-care institutions. Microbiology laboratory procedures should minimize the possibility of laboratory cross-contamination of specimens and maximize the ability to recognize a cluster of false-positive cultures. Newer molecular typing methods provide rapid, accurate and effective means of identifying false-positive M. tuberculosis ­cultures.
Aims:  To investigate a cluster of patients with positive M. tuberculosis cultures that were processed in the mycobacteriology laboratory on the same day.
Methods:  Five patients' medical records and radiology results were reviewed to determine whether the cases were epidemiologically linked and whether there was clinical suspicion of tuberculosis. Restriction fragment length polymorphism (DNA fingerprinting) was performed using repetitive elements IS 6110 and pTBN12. Laboratory processing procedures were analysed.
Results:  On the basis of DNA fingerprinting using IS6110, the isolates from all five patients were identical. Molecular typing using pTBN12 was performed on four of the five isolates. All four had identical patterns. There was no epidemiological link between the patients. At least three (and probably four) of the five patients were misdiagnosed with tuberculosis.
Conclusion:  Microbiology laboratories should ensure that appropriate methodologies are in place to avoid cross-contamination of specimens. Clinicians need to critically interpret any positive laboratory result, especially in an unlikely clinical setting. (Intern Med J 2002; 32: 512−519)  相似文献   

17.
目的应用套式PCR-DNA测序方法直接检测痰标本中结核分枝杆菌相关的rpoB基因突变,以期建立一种直接检测分枝杆菌耐利福平的快速方法,并评价其临床应用价值。方法采用套式PCR-DNA测序方法直接检测112例活动性肺结核患者和20例非结核性肺部疾病患者痰标本中结核分枝杆菌rpoB基因突变情况。同份痰标本同时做涂片抗酸染色,罗氏培养及菌型鉴定。结果112例活动性肺结核患者痰标本套式PCR扩增87例呈阳性,产物DNA测序31例有rpoB基因突变,其中分离出耐利福平株的32例痰中29例发生了基因突变,耐药突变率90.6%(29/32),39例菌阴(涂阴培阴)痰中有2例发生突变。分离出对利福平敏感株的37例痰中未发生突变,20例非结核性肺部疾病患者痰标本套式PCR扩增均为阴性,特异性100%。结论套式PCR-DNA测序可望为直接检测临床痰标本中结核分枝杆菌耐利福平的准确、特异、快速的方法。  相似文献   

18.
MTD (GEN-PROBE AMPLIFIED MYCOBACTERIUM TUBERCULOSIS DIRECT TEST) for Mycobacterium tuberculosis, and Amplicor Mycobacterium for Mycobacteria (AMP-M. tb for M. tuberculosis, AMP-M. av for M. avium and AMP-M. in for M. intracellulare) were used for the detection of relevant Mycobacterium. Their sensitivity and specificity were evaluated. Total 244 clinical specimens including 164 sputa were examined by the above two tests. The results were compared with those obtained by the conventional methods. Of 244 samples, number of the M. tuberculosis positive samples by microscopy, cultural test, MTD and AMP-M. tb were 32, 33, 38 and 35, respectively. Among 33 culture positive samples, 25 were MTD positive and 26 were AMP-M. tb positive. Therefore, sensitivity of MTD and AMP-M. tb were 75.8% and 78.8%, and their specificity were 93.8% and 95.7%, respectively. When only sputa were used for the tests as the clinical specimens, both sensitivity of MTD and AMP-M. tb were increased to 94.4%. For MAC, positive samples of M. avium complex by culture, M. avium by AMP-M. av and M. intracellulare by AMP-M. in were 13, 16, and 8, respectively. Sensitivity and specificity of AMP-M. av/M. in were 100% and 95.2%, respectively. Clinical findings of the patients whose MTD tests were positive but negative by culture were reexamined. Three of 9 specimens were also positive in AMP-M. tb. From the records of the isolations of tubercle bacilli or other important pathogens from the other kind of clinical specimens, smear tests and patients' response to tuberculosis chemotherapy, four of 9 specimens were confirmed as true positive, three were suspected as positive, and two other specimens were false positive which might be caused by contamination. From these observations, it could be concluded that MTD and AMP-M. tb are more sensitive than conventional culture method, and MTD is more sensitive than AMP-M. tb but needs more careful treatment to avoid the contamination.  相似文献   

19.
The aim of this study was to assess the value of examining multiple sputum specimens in the diagnosis of pulmonary tuberculosis (PTB). We analyzed sputum smear and culture results of patients diagnosed with culture-proven PTB during 2002. In 1027 patients, the diagnosis was established by detection of Mycobacterium tuberculosis bacilli in sputum samples. The number of sputum specimens submitted to laboratory was one in 634 cases, two in 167 cases, three in 186 cases and more than three in 48 cases. 760 (74%) cases had positive smear examination result. The first sputum smear examination was positive in 82.3% of smear positive cases. Either the first or the second sputum was diagnostic in 94.9% of these cases. Smear examination of third sputum revealed 4.2% additional diagnostic yield. In 863 (84%) cases, culture examination of the first sputum was positive. The second and the third sputum culture examinations revealed additional diagnostic yield of 11% and 4.5%, respectively. Percent 95 of culture-proven cases were diagnosed with the first two sputum cultures. In conclusion the majority of PTB cases can be diagnosed with the examination of two sputum specimens. Three or more sputum specimens submitted obtain a small additional diagnostic yield.  相似文献   

20.
SETTING: A 500-bed government referral institution for patients with tuberculosis and other infectious diseases in Gauteng, South Africa. OBJECTIVES: To assess the usefulness of BACTEC blood cultures over and above that of other microbiological methods for the diagnosis of tuberculosis in patients who are suspected of suffering from tuberculosis. DESIGN: Mycobacterial blood cultures were obtained from patients presenting with symptoms suspicious of tuberculosis and where there was no clinical evidence of other infectious etiologies, and from patients who had failed tuberculosis treatment. RESULTS: Sixteen (22%) of 71 patients included in the study were positive for Mycobacterium tuberculosis on blood culture, while seven (10%) were positive for M. avium complex (MAC). Twelve (75%) of the patients with tuberculosis and positive blood cultures were however also positive for acid-fast bacilli on sputum smears and eight (50%) were initially diagnosed clinically and radiographically as localized pulmonary tuberculosis. Blood cultures positive for mycobacteria were only found among patients with human immunodeficiency virus infection (HIV). CONCLUSIONS: Bacteremia with M. tuberculosis complex was detected in HIV-infected patients with suspected tuberculosis, even in patients presenting with localized pulmonary infection on initial clinical assessment. Among patients with suspected tuberculosis, blood cultures were useful in diagnosing unsuspected MAC disease, but did not add to the diagnostic yield of conventional tests for tuberculosis used routinely, namely sputum microscopy and culture, or occasional biopsy specimens.  相似文献   

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