Rapid Diagnosis Of Mycobacterium Tuberculosis Research Articles

Evaluation of the GeneXpert MTB/RIF Assay for Rapid Diagnosis of Tuberculosis and Detection of Rifampin Resistance in Tuberculous Patients Admitted to Abbassia Chest Hospital

Abstract Background Tuberculosis is considered one of the major infectious diseases especially in the developing countries and a significant diagnostic and therapeutic challenge worldwide. Gene Xpert MTB/RIF assay is a diagnostic test which depends on a real-time polymerase chain reaction for rapid diagnosis of Mycobacterium Tuberculosis and rifampicin resistance in a single test. Aim To evaluate the diagnostic utility of Gene Xpert MTB/RIF assay for rapid diagnosis of pulmonary and extrapulmonary tuberculosis as well as the detection of rifampin resistance in specimens obtained from suspected tuberculous patients in Abbassia Chest Hospital. Patients and Methods Pulmonary and extrapulmonary samples from 77 cases with suspected TB were collected. All retrieved samples were examined with: direct smear microscopy for AFB, culture for Mycobacterium tuberculosis (L.J. media) along with Drug susceptibility testing (D.S.T.) for positive cultures. As well as the gene Xpert MTB/RIF assay and their results were compared to those of both culture and D.S.T. (as gold standard reference methods). Results Gene Xpert MTB/RIF assay showed the highest sensitivity, negative predictive value (NPV) and diagnostic accuracy in detection of pulmonary TB (96.4, 96.4 and 93.1% respectively), with 90%specificity. Xpert MTB/RIF assay showed the lowest sensitivity in diagnosing TB pleural effusion (66.6%) but with the highest specificity and positive predictive value (PPV) (100% for both). As regards diagnosing TB lymphadenitis, gene Xpert MTB/RIF assay showed 75% sensitivity and the lowest specificity (33.3%). Considering rifampin resistance, gene Xpert MTB/RIF assay showed 100% specificity in detection of RIF- Resistance with variable sensitivity according to the type of TB, being the highest in pulmonary TB (90.9% sensitivity). Conclusion Gene Xpert MTB/RIF assay could be used as the first line diagnostic tool for diagnosis of pulmonary TB because of its high diagnostic yield. Yet, it’s not recommended to replace the culture for diagnosing extrapulmonary TB especially in diagnosing tuberculous pleural effusion (pauci-bacillary specimens) which has limited diagnostic utility. Also, the assay showed 100% specificity in diagnosing rifampicin resistance guiding physicians to justifiably starting second line anti-TB treatment.

Burden of pulmonary tuberculosis and challenges related to tuberculosis detection in Ethiopia.

Early and rapid diagnosis of Mycobacterium tuberculosis in clinical specimen is important for the treatment of patients and control of disease transmission to the community. The disease is largely preventable and curable, but without rapid, and correct diagnostic tools for tuberculosis (TB) infection and drug resistance, it is unlikely that we can meet the national TB elimination program in Ethiopia by 2035. Moreover, drug resistant TB is becoming more common and is a great challenge for the successful control and eradication of TB. The need for rapid, accurate and affordable methods for TB management should be considered by policy makers to improve TB detection rate and reduction of TB related deaths in line with the stop TB strategy by 2030 in Ethiopia.

Systematic evaluation of line probe assays for the diagnosis of tuberculosis and drug-resistant tuberculosis

BackgroundLine probe assays (LPAs) are PCR-based assays used for the rapid diagnosis of Mycobacterium tuberculosis (MTB) and drug-resistant tuberculosis (DR-TB). But studies on its performance are insufficient. Thus, in this study, we conducted a systematic review and meta-analysis to evaluate the effect of LPAs in the detection of MTB and drug-resistant TB in comparison with the traditional culture and DST methods. MethodsA systemic literature search was conducted on the Web of Science, Embase, PubMed, the Cochrane Library, Scopus, and OVID databases. All the included studies were classified according to different detecting objects. Sensitivity, specificity, Positive Likely Ratio (PLR), Negative Likely Ratio (NLR), Diagnostic Odds Ratio (DOR), corresponding 95% confidence interval, Area Under Curve (AUC), Deeks’ funnel plot, and Bivariate Boxplot was used to do the evaluation. Results147 studies included 491 datasets, with 182,448 samples, were incorporated into our analysis. The sensitivity (95% CI), specificity (95% CI), PLR, NLR, DOR and AUC for MTB were 0.89 (0.86 to 0.92), 0.94 (0.90 to 0.97), 15.70, 0.11, 139 and 0.96, respectively; for rifampicin-resistant TB were 0.96 (0.95 to 0.97), 0.99 (0.98 to 0.99), 82.9, 0.04, 1994 and 1.00, respectively; for isoniazid-resistant TB were 0.91 (0.89 to 0.93), 0.99 (0.98 to 0.99), 83.4, 0.09, (0.99 to 1.00), 195.7, 0.07, 2783 and 1.00, respectively; for Multi-drug resistant TB (MDR-TB) were 0.93 (0.90 to 0.95), 1.00 (0.99 to 1.00), 195.7, 0.07, 2783 and 1.00, respectively; for extensively drug-resistant TB (XDR-TB) were 0.60 (0.33 to 0.82), 1.00 (0.95 to 1.00), 291.3, 0.4, 726 and 0.95, respectively; for (second-line drug-resistant TB) SLID-TB were 0.83 (0.78 to 0.87), 0.98 (0.97 to 0.99), 44.6, 0.17, 262 and 0.98, respectively. Sensitivity in pre-extensively drug-resistant TB (Pre-XDR-TB) was 0.67, specificity was 0.91. No publication bias existed according to Deeks’ funnel plot. ConclusionHigh diagnosis performance was confirmed in LPAs for the diagnosis of MTB and drug-resistant TB. LPAs might be a good alternative to culture and DST in detecting MTB, RR-TB, INH-TB, XDR-TB, SLID-TB, and MDR-TB. While more studies were still needed to explore the diagnosis performance of LPAs for Pre-XDR TB.

Comparison evaluation between gene Xpert Mtb/Rif and multiplex PCR for rapid diagnosis of mycobacterium tuberculosis.

To compare the efficacy of Gene Xpert mycobacterium tuberculosis-rifampicin and multiplex polymerase chain reacton for the detection of mycobacterium tuberculosis and Rifampicin resistance. The cross-sectional validation study was conducted at the Department of Microbiology, Armed Forces Institute of Pathology, Rawalpindi, Pakistan, from March to October 2018, and comprised mycobacterium tuberculosis-positive rifampicin-resistant and rifampicin-susceptible samples, with the latter acting as negative controls. Gene Xpert mycobacterium tuberculosis-rifampicin assasy and multiplex polymerase chain reacton were applied simultaneously and compared with gold standard mycobacterium growth indicator tube 960. Data was analysed using SPSS 24. Of the 192 samples, 84(44%) were culture-positive rifampicin-resistant and 108(56%) were culture-positive rifampicin-susceptible. Overall, 84(44%) were found positive. Gene Xpert mycobacterium tuberculosis-rifampicin assay detected all 84(100%) rifampicin-resistant samples, while multiplex polymerase chain reacton detected 44(52.3%) such samples. Sensitivity, specificity, positive predictive value and negative predictive value of Gene Xpert were 100% each respectively, while the corresponding values for multiplex polymerase chain reacton were 52%, 100%, 100% and 72% respectively. Molecular detection of mycobacterium tuberculosis and resistance by Gene Xpert and multiplex polymerase chain reacton simultaneously was found to be a rapid and cost-effective method.

Role of GeneXpert in the diagnosis of mycobacterium tuberculosis.

GeneXpert (GX) is a novel, integrated, cartridge-based, nucleic acid amplification test with an established role for rapid diagnosis of Mycobacterium tuberculosis and detection of rifampicin resistance. To evaluate the role of GX in pulmonary and extrapulmonary tuberculosis (TB) cases. A prospective study was conducted in the pulmonary medicine department of a tertiary care hospital after the Ethics Comittee permission. Data of 257 presumptive TB patients was retrieved for GX, acid fast bacilli smear and cul-ture (AFB smear and culture) and drug susceptibility test (DST). Sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV) of GX in diagnosis and determination of rifampicin resistance in pulmonary and extrapulmonary TB cases were calculated and compared with culture and DST results. Our study included 132 pulmonary and 125 extrapulmonary cases. On the basis of clinicoradiological and microbiological correlation, diagnosis of TB was confirmed in 104 pulmonary and 103 extrapulmonary cases. Out of a total of 104 pulmonary TB cases, 73 were rifampicin-sensitive and 31 were rifampicin-resistant cases. 103 extrapulmonary TB patients included 66 rifampicin-sensitive and 37 rifampicin-resistant cases. The sensitivity, specificity, PPV, NPV of GX in diagnosis and detection of rifampicin resistance in pulmonary TB was 95%, 93%, 98%, 84% and 96%, 100%, 100%, 96%, respectively. The sensitivity, specificity, PPV, NPV of GX in diagnosis and detection of rifampicin resistance in extrapulmonary TB cases was 79%, 86%, 96%, 47% and 97%, 95%, 97%, 95%, respectively. GX results are superior to smear microscopy and comparable to culture with shorter turnaround time.We recom-mend using it in routine TB diagnosis as this will expedite the management of patients with presumptive TB.

CapitalBio Mycobacterium real-time polymerase chain reaction detection test: Rapid diagnosis of Mycobacterium tuberculosis and nontuberculous mycobacterial infection

ObjectivesTo evaluate the accuracy of the CapitalBio Mycobacterium real-time polymerase chain reaction (RT-PCR) detection test for pulmonary Mycobacterium tuberculosis (MTB) and nontuberculous mycobacterial (NTM) infection. MethodsThis study analyzed 2,460 samples from patients with suspected pulmonary mycobacterial infection collected between 01 June 2018 and 31 July 2019. It aimed to determine the sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV), and area under the curve (AUC) of the CapitalBio Mycobacterium detection test for MTB and NTM infections, and to evaluate its diagnostic accuracy compared with mycobacterial culture. ResultsThe sensitivity, specificity, PPV, NPV, and AUC of the CapitalBio Mycobacterium detection test for MTB was 83.0%, 79.9%, 80.8%, 82.2%, and 0.81, respectively. This was similar to the diagnostic accuracy of Xpert MTB/RIF for MTB and was significantly higher than that of smear. For pulmonary NTM infection, the sensitivity, specificity, PPV, NPV, and AUC of the test was 82.0%, 99.6%, 94.1%, 98.5%, and 0.91, respectively. The diagnostic accuracy of the CapitalBio Mycobacterium detection test was also significantly higher than that of smear for NTM. ConclusionsThe CapitalBio Mycobacterium detection test had good diagnostic accuracy for MTB and NTM infections. This is of great significance for the differential diagnosis of early pulmonary mycobacterial infection.

Does Xpert\xae MTB/RIF assay give rifampicin resistance results without identified mutation? Review of cases from Addis Ababa, Ethiopia

BackgroundXpert® MTB/RIF assay is currently used in Ethiopia for the rapid diagnosis of Mycobacterium tuberculosis (MTB) and mutations that confer Rifampicin resistance. Rifampicin resistance is determined based on any mutation in the 81 bp of rpoB gene using five overlapping probes represented as Probe A (codons 507–511), Probe B (codons 512–518), Probe C (codons 518–523), Probe D (codons 523–529) and Probe E (codons 529–533). In this review, we assessed the frequency of missed probe types for Rifampicin Resistance results.MethodsData were reviewed from specimens received and tested using Xpert® MTB/RIF assay at Ethiopian National Tuberculosis Reference Laboratory, in Addis Ababa from 15 July 2016 to 31 December 2018 retrospectively. All archived data were reviewed carefully to describe missed probe types and the quantity of DNA in the sample.ResultsA total of 100 specimens were reported as MTB Detected Rifampicin Resistance Detected by Xpert® MTB/RIF assay. More than half (55%) of these results were reported from male patients. The median age was 28.0 years (5 months to 88 years). Majorities (62%) of the cases were detected from sputum. Among the total of 38 extrapulmonary samples, lymph node aspirates were accounted for 50% (19/38). The most common mutations (81.0%) were found in the Probe E region followed by Probe D (10.0%), and Probe B (3.0%). Mutations in Probe A and Probe C regions were not observed. However, six (6.0%) Rifampicin resistance cases were found without any missed probe type. The delta Ct max is ≥4.3. No specimen yielded Rifampicin resistance associated with more than one probe failure or mutation combinations.ConclusionMutations associated with Probe E (codons 529–533) region were identified as the commonest rpoB gene mutations. The Rifampicin resistance results found without any identified missing probe needs further study. The lower DNA amount was observed in extrapulmonary specimens compared with sputum.

Role of Xpert MTB/RIF in Bronchoalveolar lavage fluid of sputum-scarce, suspected Pulmonary TB patients

SettingA prospective observational study conducted in Medical college hospital, in a high-TB- prevalence region of northern Telangana, India. ObjectiveTo know the diagnostic role of Xpert MTB/RIF assay in bronchoalveolar lavage fluid (BALF) in sputum-scarce, suspected pulmonary tuberculosis (PTB) patients. DesignStudy period was between October 2014 and March 2017. Suspected pulmonary tuberculosis patients aged 15 years or more, who were sputum-scarce and conforming to the inclusion criteria were submitted to bronchoscopy. BALF thus obtained was submitted to smear for acid fast bacilli (AFB) and Xpert MTB/RIF assay as index tests along with culture for Mycobacterium tuberculosis complex (MTBC). Culture for M. tuberculosis complex was considered as gold standard for the diagnosis of PTB. The sensitivity, specificity and predictive values were calculated for smear AFB and Xpert MTB/RIF assay. Results56 of the 81 patients were included and evaluated in the final analysis. In 10 of these 56 patients PTB was confirmed by culture positivity. The sensitivity and specificity of Xpert MTB/RIF assay was 90% (9/10,95%CI 59.6- 98.2) and 52.2% (24/46, 95%CI 38. 1-65.9) respectively and that of the smear AFB was 60% (6/10, 95%CI 31.2–83.1) and 67.4% (31/46, 95%CI 53.0–79.1). All the patients considered ‘probable’ PTB (pending culture results), were administered antituberculous treatment and showed complete clinicoradiological improvement on follow up. Three of the 31 Xpert MTB/RIF positive patients were detected as resistance to rifampicin (RR). ConclusionsXpert MTB/RIF assay of BALF in the study cohort provides rapid diagnosis of Mycobacterium tuberculosis, and detection of rifampicin resistance at the very outset, aiding in selection of appropriate ATT regimen. In this context, it can be recommended as the first line investigation. Xpert MTB/RIF assay aided by HRCT Chest and suggestive clinical presentation may be helpful in early institution of ATT especially in smear negative, culture negative cases.

Naked eye detection of the Mycobacterium tuberculosis complex by recombinase polymerase amplification-SYBR green I assays.

Rapid diagnosis of Mycobacterium tuberculosis (Mtb) is key to controlling the spread of tuberculosis, which is a global health concern. In this study, isothermal recombinase polymerase amplification (RPA) was developed to detect specific targets of Mtb, IS6110 and IS1081. Additionally, SYBR Green I was used for endpoint detection of the RPA products by the naked eye. A total of 146 genomic Mtb DNA samples and 24 genomic nontuberculous mycobacteria (NTM) DNA samples were amplified at IS6110 and IS1081 by RPA. After a complete amplification, the RPA amplicons were examined by agarose gel electrophoresis (RPA-AGE) and SYBR Green I (RPA-S) assays. The performance of the RPA assays was evaluated by comparing them to a conventional PCR. The RPA assay demonstrated to have a good capability to differentiate Mtb from NTM with a very short turnaround time at a constant temperature. Compared to conventional PCR, the sensitivities and specificities of RPA-AGE for IS6110 and IS1081 were 100%. The specificity of RPA-S was 100% for both targets; however, its sensitivities for IS6110 and IS1081 were 97.95% and 99.32%, respectively. The limits of detection of IS6110 RPA-AGE and RPA-S were 0.05 and 0.5ng, respectively, while the LODs of IS1081 RPA-AGE and RPA-S were 0.00005 and 0.05ng, respectively. Both RPA assays showed a satisfying diagnostic specificity, with no cross-reaction with other bacteria. A rapid, sensitive, naked eye RPA assay can be integrated into point-of-care diagnosis for Mtb detection, especially in remote areas where laboratory instrument resources are limited.

Evaluation of GeneXpert MTB/RIF and line probe assay for rapid diagnosis of Mycobacterium tuberculosis in Sudanese pulmonary TB patients

Evaluation of GeneXpert MTB/RIF and line probe assay for rapid diagnosis of Mycobacterium tuberculosis in Sudanese pulmonary TB patients

Multicenter evaluation of a real-time loop-mediated isothermal amplification (RealAmp) test for rapid diagnosis of Mycobacterium tuberculosis

The real-time loop-mediated isothermal amplification (RealAmp) test is a novel technology for rapid diagnosis of Mycobacterium tuberculosis (MTB) from sputum samples. This test has not previously been effectively evaluated. In this study, we assessed the performance of the RealAmp test for tuberculosis (TB) detection among TB suspected patients from three provincial hospitals in China. Each eligible TB suspected patients in Shanghai, Guangzhou, and Shandong provided one sputum specimen each, smear microscopy, liquid culture, and the RealAmp test were performed per patient. A total of 1583 TB suspected patients were enrolled in the study, and 1519 were used for the performance analysis. The sensitivity and specificity of the RealAmp test for MTB detection were calculated using liquid culture as reference method. The sensitivity of RealAmp test in smear-negative and culture-positive patients was 60.08%; the sensitivity in smear-positive and culture-positive patients was 97.09%; and the overall sensitivity in culture-positive patients was 74.88%. The specificity of RealAmp test for MTB detection was 86.50%.

Rapid diagnosis of mycobacterium tuberculosis with electrical impedance spectroscopy in suspensions using interdigitated microelectrode

A Mycobacterium tuberculosis (MTB) bacilli are still widely spreading and have to be diagnosed fast and efficiently. Therefore, a new simple and rapid method was proposed to detect MTB by the impedance properties of MTB suspensions using interdigitated microelectrodes. As a result, MTB suspensions in deionized (DI) water with different cell concentrations generated different electrical impedance spectral responses. Whereas MTB suspensions in 0.9 wt. % NaCl solution did not produce any significant differences in the impedance spectra in response to different cell concentrations. In DI water suspensions, the impedance at 1 kHz decreased with increasing cell concentrations. The impedance of MTB suspension in DI water has been discussed; it was found to be resulted from the cell wall charges and release of ions from the cells. There was a linear relationship between the impedance and logarithmic value of the cell concentration in the cell concentration range of 102 to 108 cfu/mL, which can be expressed by the regression equation of Z (kΩ) =–456lnN (cfu/mL) + 9717 with R2 = 0.99. Detection limit was calculated as 104 cfu/mL, which is comparable with many label-free immunosensors for detecting pathogenic bacteria reported in the literature. This work demonstrated that MTB concentration can be determined through measuring the impedance of MTB suspensions in DI water. This new detection mechanism can be an alternative for current impedance methods available for detecting bacterial cells.

Rapid Diagnosis of Mycobacterium Tuberculosis with Electrical Impedance Spectroscopy in Suspensions Using Interdigitated Microelectrode

Rapid Diagnosis of Mycobacterium Tuberculosis with Electrical Impedance Spectroscopy in Suspensions Using Interdigitated Microelectrode

Rapid diagnosis of pediatric mycobacterial lymphadenitis using fine needle aspiration biopsy.

Diagnosis of tuberculosis in children is challenging and fine needle aspiration biopsy (FNAB) is used worldwide in the diagnosis of palpable masses including peripheral lymphadenopathy. Recent studies of the use of nucleic acid amplification such as the Xpert MTB/RIF test on FNAB in adult patients have shown considerable promise. Xpert MTB/RIF allows for the rapid diagnosis of Mycobacterium tuberculosis and identification of rifampicin susceptibility. Studies to date have been predominantly performed in adults. This study aims to determine the accuracy of Xpert MTB/RIF for the detection of M. tuberculosis complex in FNAB from children with clinically suspected mycobacterial lymphadenitis. Prospective hospital-based study of children <13 years referred for FNAB at Tygerberg hospital and Dora Nginza hospital, South Africa, for suspected mycobacterial lymphadenitis. Aspirates were performed and the results of the Xpert MTB/RIF test were compared with liquid (mycobacterial growth indicator tube) culture and cytology. FNABs were collected from 110 children and 38 (35%) cases were excluded. Of the 72 cases included in the study, 32 were positive for M. tuberculosis complex on Xpert MTB/RIF, 36 on cytology and 25 were culture positive for M. tuberculosis complex. Compared with the combined reference standard (cytomorphology suggestive of mycobacterial disease with direct visualization of the organism and/or bacteriological culture), Xpert MTB/RIF identified 32 of 40 cases as positive with a sensitivity and a specificity of 80% and 93.8%, respectively. FNAB and Xpert MTB/RIF enable a rapid diagnosis in pediatric mycobacterial lymphadenitis, expediting appropriate treatment and potentially preventing morbidity and mortality.

Rapid Diagnosis of Mycobacterium tuberculosis with Truenat MTB: A Near-Care Approach

BackgroundControl of the global Tuberculosis (TB) burden is hindered by the lack of a simple and effective diagnostic test that can be utilized in resource-limited settings.MethodsWe evaluated the performance of Truenat MTB™, a chip-based nucleic acid amplification test in the detection of Mycobacterium tuberculosis (MTB) in clinical sputum specimens from 226 patients with suspected pulmonary tuberculosis (TB). The test involved sputum processing using Trueprep-MAG™ (nanoparticle-based protocol run on a battery-operated device) and real-time PCR performed on the Truelab Uno™ analyzer (handheld, battery-operated thermal cycler). Specimens were also examined for presence of MTB using smear microscopy, liquid culture and an in-house nested PCR protocol. Results were assessed in comparison to a composite reference standard (CRS) consisting of smear and culture results, clinical treatment and follow-up, and radiology findings.ResultsBased on the CRS, 191 patients had “Clinical-TB” (Definite and Probable-TB). Of which 154 patients are already on treatment, and 37 were treatment naïve cases. Remaining 35 were confirmed “Non-TB” cases which are treatment naïve cases. The Truenat MTB test was found to have sensitivity and specificity of 91.1% (CI: 86.1–94.7) and 100% (CI: 90.0–100) respectively, in comparison to 90.58% (CI: 85.5–94.3) and 91.43% (CI: 76.9–98.2) respectively for the in-house nested PCR protocol.ConclusionThis preliminary study shows that the Truenat MTB test allows detection of TB in approximately one hour and can be utilized in near-care settings to provide quick and accurate diagnosis.

Comparison of Xpert MTB/RIF with Other Nucleic Acid Technologies for Diagnosing Pulmonary Tuberculosis in a High HIV Prevalence Setting: A Prospective Study

The Xpert MTB/RIF (Cepheid) non-laboratory-based molecular assay has potential to improve the diagnosis of tuberculosis (TB), especially in HIV-infected populations, through increased sensitivity, reduced turnaround time (2 h), and immediate identification of rifampicin (RIF) resistance. In a prospective clinical validation study we compared the performance of Xpert MTB/RIF, MTBDRplus (Hain Lifescience), LightCycler Mycobacterium Detection (LCTB) (Roche), with acid fast bacilli (AFB) smear microscopy and liquid culture on a single sputum specimen. Consecutive adults with suspected TB attending a primary health care clinic in Johannesburg, South Africa, were prospectively enrolled and evaluated for TB according to the guidelines of the National TB Control Programme, including assessment for smear-negative TB by chest X-ray, clinical evaluation, and HIV testing. A single sputum sample underwent routine decontamination, AFB smear microscopy, liquid culture, and phenotypic drug susceptibility testing. Residual sample was batched for molecular testing. For the 311 participants, the HIV prevalence was 70% (n = 215), with 120 (38.5%) culture-positive TB cases. Compared to liquid culture, the sensitivities of all the test methodologies, determined with a limited and potentially underpowered sample size (n = 177), were 59% (47%-71%) for smear microscopy, 76% (64%-85%) for MTBDRplus, 76% (64%-85%) for LCTB, and 86% (76%-93%) for Xpert MTB/RIF, with specificities all >97%. Among HIV+ individuals, the sensitivity of the Xpert MTB/RIF test was 84% (69%-93%), while the other molecular tests had sensitivities reduced by 6%. TB detection among smear-negative, culture-positive samples was 28% (5/18) for MTBDRplus, 22% (4/18) for LCTB, and 61% (11/18) for Xpert MTB/RIF. A few (n = 5) RIF-resistant cases were detected using the phenotypic drug susceptibility testing methodology. Xpert MTB/RIF detected four of these five cases (fifth case not tested) and two additional phenotypically sensitive cases. The Xpert MTB/RIF test has superior performance for rapid diagnosis of Mycobacterium tuberculosis over existing AFB smear microscopy and other molecular methodologies in an HIV- and TB-endemic region. Its place in the clinical diagnostic algorithm in national health programs needs exploration. Please see later in the article for the Editors' Summary.

Efficacy of an in-house polymerase chain reaction assay for rapid diagnosis of Mycobacterium tuberculosis in patients with tubercular lymphadenitis: Comparison with fine needle aspiration cytology and conventional techniques

Tubercular lymphadenitis (TB-L) is the most common manifestation of extrapulmonary tuberculosis. Excisional biopsy with histopathological examination, Ziehl-Neelsen staining (ZNS) and culture and fine needle aspiration (FNA) cytology, although useful in the diagnosis of TB-L, cannot diagnose a substantial proportion of cases. We investigated the role of an in-house polymerase chain reaction (PCR) assay targeting the IS6110 gene from the FNA material in the diagnosis of the disease. The clinical profile of 150 patients with lymphadenopathy was noted and the fine needle aspirate was collected. After cytological processing, ZNS and culture on Lowenstein-Jensen media, mycobacterial DNA was isolated from the residual aspirate material and IS6110 gene PCR was performed. Results of cytology, ZNS, culture and IS6110 gene PCR were compared. There were 49 confirmed patients of TB-L based on laboratory parameters (either culture isolation of Mycobacterium tuberculosis or any two of cytology, ZNS, PCR positive) and clinical response to therapy. Sensitivity and specificity of FNA was 89.8% and 96%, of ZNS was 40.8% and 99%, of culture was 40.8% and 100% and of IS6110 gene PCR test was 100% and 92.1%. IS6110 PCR can be considered a valuable adjunct to cytology, ZNS and culture techniques in the diagnosis of TB-L.

Evaluation of a simplified IS6110 PCR for the rapid diagnosis of Mycobacterium tuberculosis in an area with high tuberculosis incidence

Purpose To assess the diagnostic yield of a simplified IS6110-PCR in an area with high tuberculosis incidence. Methods Pulmonary (218) and extrapulmonary (121) samples were collected from 236 patients including smearpositive leprosy patients. All samples were processed to detect acidfast bacilli by microscopy, culture on solid media and PCR. To remove PCR inhibitors, three washing steps of the decontaminated pellet were included before mycobacterial cell lysis by heat treatment. No detergents, enzymes, or chelating agents were used. From the 339 samples, 34 were selected basing on their large volume and were tested by the commercial kit GenoType Mycobacteria Direct (GTMD) (VER 4, Hain Lifescience, Germany) in addition to the tests cited above. Results The overall sensitivity and specificity of PCR were 93.8 and 98.6% for pulmonary samples, 63.6 and 100% for extrapulmonary samples, respectively. The assay detected MTC in 94.2% of smear positive samples with a positive predictive value of 100%. No inhibition was found among seven samples that were PCR negative but bacteriological confirmed as containing Mycobacterium tuberculosis. No false positive result occurred with samples from leprosy patients. The sensitivities for PCR and GTMD were 81.8 and 75%, respectively. Conclusion PCR could efficiently complement conventional bacteriological tools for the rapid diagnosis of tuberculosis but cannot replace them.

Clinical evaluation of Gen-Probe's amplified mycobacterium tuberculosis direct test for rapid diagnosis of Mycobacterium tuberculosis in Egyptian children at risk for infection.

Abstract Context.—Diagnostic detection of tuberculosis (TB) has improved considerably. Available, standardized, nucleic acid–based amplification techniques have been shown to yield reliable results within 4 to 7 hours of sample processing. Objective.—To study the diagnostic performance of Gen-Probe's technique for direct detection of Mycobacterium tuberculosis in comparison with BACTEC 460 TB culture for both positive and negative Ziehl-Neelsen smears in Egyptian children at risk for TB infection. Design.—We prospectively evaluated 50 children from families with a positive history of TB. All patients were referred from outpatient clinics of the Mansoura University Children's Hospital, Egypt. The children had a positive tuberculin skin test with an induration diameter of more than 10 mm and had scars from a bacille Calmette-Guerin vaccination within the past 2 years. Three consecutive sputum samples were taken from each patient. The samples were examined to detect M tuberculosis by means of the Gen-Probe t...

O477 Evaluation of a new version of the “RT-TB” triplex real-time PCR assay for the rapid diagnosis of Mycobacterium tuberculosis in clinical samples

O477 Evaluation of a new version of the “RT-TB” triplex real-time PCR assay for the rapid diagnosis of Mycobacterium tuberculosis in clinical samples