Diagnostic value of bronchoalveolar lavage and bronchial washing in sputum-scarce or smear-negative cases with suspected pulmonary tuberculosis: a randomized study

ObjectivesBronchoalveolar lavage (BAL) and bronchial washing (BW) are two major methods used to obtain high-quality respiratory specimens from patients with suspected pulmonary tuberculosis (TB) but a sputum-scarce or smear-negative status. We aimed to compare the value of BAL and BW in the diagnosis of TB in such patients.MethodsWe enrolled patients with suspected pulmonary TB but with a sputum-scarce or smear-negative status who were referred for bronchoscopy between October 2013 and January 2016. Participants were randomized into the BAL and BW groups for evaluation. The primary outcome was the diagnostic yield for TB detection. Secondary outcomes included culture positivity, positivity of nucleic acid amplification tests (NAATs) for Mycobacterium tuberculosis and procedure-related complications.ResultsA total of 94 patients were assessed and 91 (43 in the BAL group, 48 in the BW group) were analysed. Twenty-one patients (48.8%) in the BAL group and 30 (62.5%) in the BW group had a final diagnosis of pulmonary TB. The detection rate of M. tuberculosis by culture or NAAT was significantly higher in BAL specimens than in BW specimens (85.7% vs 50.0%, p 0.009). The procedure-related complications were hypoxic events, 2/43 (4.7%) in the BAL group and 5/48 (10.4%) in the BW group; and post-bronchoscopic fever, 3/43 (7.0%) in the BAL group and 4/48 (8.3%) in the BW group.DiscussionAs long as it is tolerable, BAL rather than BW, should be used to obtain specimens for the diagnosis of pulmonary TB in sputum-scarce or smear-negative cases.     

Diagnosis of tuberculosis by trained African giant pouched rats and confounding impact of pathogens and microflora of the respiratory tract

Trained African giant-pouched rats (Cricetomys gambianus) can detect Mycobacterium tuberculosis and show potential for the diagnosis of tuberculosis (TB). However, rats' ability to discriminate between clinical sputum containing other Mycobacterium spp. and nonmycobacterial species of the respiratory tract is unknown. It is also unknown whether nonmycobacterial species produce odor similar to M. tuberculosis and thereby cause the detection of smear-negative sputum. Sputum samples from 289 subjects were analyzed by smear microscopy, culture, and rats. Mycobacterium spp. were isolated on Lowenstein-Jensen medium, and nonmycobacterial species were isolated on four different media. The odor from nonmycobacterial species from smear- and M. tuberculosis culture-negative sputa detected by ≥2 rats ("rat positive") was analyzed by gas chromatography-mass spectrometry and compared to the M. tuberculosis odor. Rats detected 45 of 56 confirmed cases of TB, 4 of 5 suspected cases of TB, and 63 of 228 TB-negative subjects (sensitivity, 80.4%; specificity, 72.4%; accuracy, 73.9%; positive predictive value, 41.7%; negative predictive value, 93.8%). A total of 37 (78.7%) of 47 mycobacterial isolates were M. tuberculosis complex, with 75.7% from rat-positive sputa. Ten isolates were nontuberculous mycobacteria, one was M. intracellulare, one was M. avium subsp. hominissuis, and eight were unidentified. Rat-positive sputa with Moraxella catarrhalis, Streptococcus pneumoniae, Staphylococcus spp., and Enterococcus spp. were associated with TB. Rhodococcus, Nocardia, Streptomyces, Staphylococcus, and Candida spp. from rat-positive sputa did not produce M. tuberculosis-specific volatiles (methyl nicotinate, methyl para-anisate, and ortho-phenylanisole). Prevalence of Mycobacterium-related Nocardia and Rhodococcus in smear-negative sputa did not equal that of smear-negative mycobacteria (44.7%), of which 28.6% were rat positive. These findings and the absence of M. tuberculosis-specific volatiles in nonmycobacterial species indicate that rats can be trained to specifically detect M. tuberculosis.     

Molecular methods in the detection and identification of mycobacterial infections.

Nucleic acid amplification (NAA) tests for direct detection of Mycobacterium tuberculosis complex in respiratory specimens have the potential to provide a more rapid diagnosis of pulmonary tuberculosis (TB) than is currently possible by conventional stain, culture, and identification tests. Currently, 2 NAA tests-enhanced Amplified Mycobacterium Tuberculosis Direct (MTD) Test (Gen-Probe, Inc) and Amplicor Mycobacterium tuberculosis Test (Roche Molecular Systems, Inc)-have been approved by the Food and Drug Administration for testing respiratory specimens that are smear positive for acid-fast bacilli (AFB). This restriction to AFB smear-positive specimens was based on data from the initial clinical trials conducted to evaluate these products that showed low sensitivity (ie, 48%-53%) and less-than-optimal specificity (ie, 96%-99%) in AFB smear-negative specimens. Data from the clinical trial for the enhanced MTD test and from 2 subsequent studies, however, suggest that this version of the MTD test is a reliable tool for rapid diagnosis of pulmonary TB, regardless of the AFB smear result. Both NAA tests have been evaluated for diagnosis of extrapulmonary TB, and results were comparable to the results of tests performed with respiratory specimens. The NAA tests also appear to be reliable for rapid identification of M tuberculosis complex in positive broth cultures of all specimen types except blood. The impact of the NAA tests on patient outcome varies based on the AFB smear result. With smear-positive results, public health and hospital infection control resources are predominantly affected. With smear-negative results, however, the potential for affecting patient outcome is much greater. In patients with smear-negative results, the NAA test can result in earlier diagnosis of TB and subsequent initiation of therapy. Use of these tests also may eliminate the need for invasive diagnostic procedures, which are costly and pose an added risk to the patient, and they may allow earlier discharge of hospitalized patients.     

Prospective clinical evaluation of the serologic tuberculous glycolipid test in combination with the nucleic acid amplification test

We have conducted a prospective controlled multicenter study to evaluate differences in the levels of clinical utility of the tuberculous glycolipid (TBGL) serodiagnostic test and the nucleic acid amplification test in patients with smear-negative active pulmonary tuberculosis (TB). The TBGL test and the PCR test were individually not so useful for the rapid diagnosis of smear-negative active pulmonary TB. However, clinical utility was considerably improved by using the TBGL test and the PCR test in combination, especially in patients with smear-negative and culture-negative active pulmonary TB and in patients with minimally advanced lesions.     

First evaluation of an improved assay for molecular genetic detection of tuberculosis as well as rifampin and isoniazid resistances

The commercially available line probe assay MTBDRplus 2.0 (Hain Lifescience, Nehren, Germany) was evaluated for its ability to detect Mycobacterium tuberculosis complex (MTBC) and mutations conferring resistance to rifampin (RMP) and isoniazid (INH) directly in smear-negative and smear-positive pulmonary clinical specimens under routine laboratory conditions. A total of 348 samples originating from Moldova, a high-incidence country for tuberculosis (TB), were investigated. Two hundred fifty-seven (73.9%) were smear negative, 12 samples were excluded, and 81 (23.3%) were smear positive. Two DNA extraction methods were applied. Compared to culture and clinical data as the reference standard (adapted from Vadwai V et al., J. Clin. Microbiol. 49:2540-2545, 2011), overall sensitivity and specificity were 87.6 and 99.2%, respectively. One hundred four of the 257 smear-negative samples turned out to be culture positive, and 20 were MTBC culture negative but were positive based on clinical symptoms. The combined sensitivity and specificity in the subgroup of smear-negative samples were calculated to be 79.8 and 99.2%, respectively. MTBDRplus 2.0 detected RMP and INH resistance with sensitivity and specificity of 94.3 and 96.0%, respectively. In conclusion, the MTBDRplus 2.0 assay is a rapid and highly sensitive test for the detection of M. tuberculosis strains from smear-positive and -negative clinical specimens and provides additional information on RMP and INH resistance status, which can easily be included in routine laboratory work flow.     

Clinical evaluation of the enhanced Gen-Probe Amplified Mycobacterium Tuberculosis Direct Test for rapid diagnosis of tuberculosis in prison inmates

The reliability of the enhanced Amplified Mycobacterium Tuberculosis Direct Test (E-MTD; Gen-Probe, Inc., San Diego, Calif.) for rapid diagnosis of pulmonary tuberculosis (TB) was evaluated by testing 1, 004 respiratory specimens from 489 Texas prison inmates. Results were compared to those of mycobacterial culture (BACTEC TB 460 and Middlebrook 7H11 biplates), smear for acid-fast bacilli (AFB; auramine O), and clinical course. After chart review, three patients (nine specimens) who were on antituberculosis therapy before the study began were excluded from final analysis. Of the remaining 995 specimens, 21 were AFB smear positive: 13 grew Mycobacterium tuberculosis complex (MTBC), 6 grew nontuberculous mycobacteria, and 2 (from two patients diagnosed with TB and started on therapy after the study began) were culture negative. Twenty-eight specimens (20 patients) were positive for MTBC by culture and E-MTD. Seven specimens (seven patients) were positive by culture alone; three were from patients who had other E-MTD-positive specimens, two were false-positive cultures, and two were false-negative E-MTD results. Eight specimens were positive by E-MTD only; four specimens (four patients) were false-positive E-MTD results, and four specimens were from two patients with earlier E-MTD-positive specimens that grew MTBC. Thus, there were 22 patients with TB (10 smear positive and 12 smear negative). The sensitivity and specificity of the AFB smear for diagnosis of TB, by patient, were 45.5 and 98.9%, respectively. After resolving discrepancies, these same values for E-MTD were 90.9 and 99.1% overall, 100 and 100% for the smear-positive patients, and 83.3 and 99.1% for the smear-negative patients. Excluding the one smear-negative patient whose E-MTD-negative, MTBC culture-positive specimen contained inhibitory substances, the sensitivity of E-MTD was 95.2% overall and 90.9% in smear-negative patients. The specificity and positive predictive value of E-MTD can be improved, without altering other performance characteristics, by modifying the equivocal zone recommended by the manufacturer. These data suggest that E-MTD is a reliable method for rapid diagnosis of pulmonary TB, irrespective of the AFB smear result. Guidelines for the most appropriate use of E-MTD with smear-negative patients are needed.     

Detection of pulmonary and extrapulmonary tuberculosis patients with the 38-kilodalton antigen from Mycobacterium tuberculosis in a rapid membrane-based assay.

A rapid membrane-based serologic assay using the 38-kDa antigen from Mycobacterium tuberculosis for the diagnosis of tuberculosis (TB) was evaluated with 201 patients with pulmonary TB, 67 patients with extrapulmonary TB, 79 Mycobacterium bovis BCG-vaccinated healthy controls, and 77 non-TB respiratory patients. The overall sensitivities, specificities, and positive and negative predictive values were, respectively, 92, 92, 84, and 96% for sputum-positive TB patients; 70, 92, 87, and 79% for sputum-negative TB patients; and 76, 92, 80, and 90% for extrapulmonary-TB patients. Only 2% (1 of 44) of the healthy control BCG-vaccinated subjects gave weak positive signals in the assay, indicating that this rapid serological assay is a valuable aid in clinical diagnosis for both pulmonary and extrapulmonary TB.     

Evaluation of Antigen-Specific Immunoglobulin G Responses in Pulmonary Tuberculosis Patients and Contacts

This study aimed to evaluate the serodiagnostic potential of immunoglobulin G (IgG) responses to Mycobacterium tuberculosis antigens in pulmonary tuberculosis (TB) patients, recent TB contacts with latent TB infection (LTBI), and healthy subjects. Infections were assessed using tuberculin skin tests, QuantiFERON-TB Gold In-Tube tests, drug susceptibility testing, and molecular genotyping of clinical isolates. Serum IgG responses to selective M. tuberculosis antigens, including the 38-kDa and 16-kDa antigens, lipoarabinomannan (LAM), and recombinant early secreted antigen target 6 kDa (ESAT-6) and culture filtrate protein 10 kDa (CFP-10), were determined. We found that the serum IgG responses to all antigens might differentiate between active TB and LTBI, with LAM having the highest diagnostic value (area under the curve [AUC] of 0.7756, P < 0.001). Recurrent TB cases showed significantly higher IgG responses to 38 kDa, CFP-10 (P < 0.01), and LAM (P < 0.05) than new cases, and male patients had higher levels of antigen-specific IgG than females (P < 0.05). Conversely, drug resistance and patient body mass index did not affect IgG responses (P > 0.05). LAM-specific IgG responses differentiated between acid-fast bacillus (AFB) smear-positive and -negative patients (P < 0.01), whereas antigen-specific IgG responses did not vary with the M. tuberculosis genotype (P > 0.05). Significantly higher IgG responses to 38 kDa and 16 kDa were observed in AFB smear-negative patients than in controls. These results suggest that assessment of serum IgG responses to selective purified M. tuberculosis antigens may help improve the diagnosis of active TB, particularly for sputum smear-negative patients or recurrent cases, and these may also help to differentiate between active TB and LTBI.     

Use of multiepitope polyproteins in serodiagnosis of active tuberculosis

Screening of genomic expression libraries from Mycobacterium tuberculosis with sera from tuberculosis (TB) patients or rabbit antiserum to M. tuberculosis led to the identification of novel antigens capable of detecting specific antibodies to M. tuberculosis. Three antigens, Mtb11 (also known as CFP-10), Mtb8, and Mtb48, were tested together with the previously reported 38-kDa protein, in an enzyme-linked immunosorbent assay (ELISA) to detect antibodies in TB patients. These four proteins were also produced as a genetically fused polyprotein, which was tested with two additional antigens, DPEP (also known as MPT32) and Mtb81. Sera from individuals with pulmonary and extrapulmonary TB, human immunodeficiency virus (HIV)-TB coinfections, and purified protein derivative (PPD)-positive and PPD-negative status with no evidence of disease were tested. In samples from HIV-negative individuals, the ELISA detected antibodies in >80% of smear-positive individuals and >60% smear-negative individuals, with a specificity of approximately 98%. For this group, smears detected 81.6% but a combination of smear and ELISA had a sensitivity of approximately 93%. The antigen combination detected a significant number of HIV-TB coinfections as well as antibodies in patients with extrapulmonary infections. Improved reactivity in the HIV-TB group was observed by including the antigen Mtb81 that was identified by proteomics. The data indicate that the use of multiple antigens, some of which are in a single polyprotein, can be used to facilitate the development of a highly sensitive test for M. tuberculosis antibody detection.     

Molecular and immunological characterization of Mycobacterium tuberculosis CFP-10, an immunodiagnostic antigen missing in Mycobacterium bovis BCG

In order to identify antigens that may be used in the serodiagnosis of active tuberculosis (TB), we screened a Mycobacterium tuberculosis genomic expression library with a pool of sera from patients diagnosed with active pulmonary TB. The sera used lacked reactivity with a recombinant form of the M. tuberculosis 38-kDa antigen (r38kDa), and the goal was to identify antigens that might complement r38kDa in a serodiagnostic assay. Utilizing this strategy, we identified a gene, previously designated lhp, which encodes a 100-amino-acid protein referred to as culture filtrate protein 10 (CFP-10). The lhp gene is located directly upstream of esat-6, within a region missing in M. bovis BCG. Immunoblot analysis demonstrated that CFP-10 is present in M. tuberculosis CFP, indicating that it is likely a secreted or shed antigen. Purified recombinant CFP-10 (rCFP-10) was shown to be capable of detecting specific antibody in a percentage of TB patients that lack reactivity with r38kDa, most notably in smear-negative cases, where sensitivity was increased from 21% for r38kDa alone to 40% with the inclusion of rCFP-10. In smear-positive patient sera, sensitivity was increased from 49% for r38kDa alone to 58% with the inclusion of rCFP-10. In addition, rCFP-10 was shown to be a potent T-cell antigen, eliciting proliferative responses and gamma interferon production from peripheral blood mononuclear cells in 70% of purified protein derivative-positive individuals without evident disease. The responses to this antigen argue for the inclusion of rCFP-10 in a polyvalent serodiagnostic test for detection of active TB infection. rCFP-10 could also contribute to the development of a recombinant T-cell diagnostic test capable of detecting exposure to M. tuberculosis.     

Potential Role of Immunodiagnosis for Pulmonary Tuberculosis Using Induced Sputum Cells

PurposeTo evaluate the diagnostic utility and predictors for determinate results of an enzyme-linked immunospot assay using induced sputum cells (IS ELISPOT) for a rapid diagnosis of pulmonary tuberculosis (TB).ResultsA total of 43 subjects, including 25 with TB (TB group) and 18 with non-TB disease (non-TB group) were enrolled. Results of IS ELISPOT were determinate in only 17/43 (39%) subjects, but all of determinate results were consistent with the final diagnosis. Of the 43 sputum samples, 11 (26%) were inadequate to perform IS ELISPOT. Of 32 adequate sputum samples, the proportion of determinate results was significantly higher in the TB group (75%, 15/20) than in the non-TB group (17%, 2/12) (p=0.002). The status of active TB was a unique predictor but smear positivity was not a significant predictor for determinate results. In addition, sensitivity of IS ELISPOT (75%, 9/12) in smear negative TB was higher than that of TB-polymerase chain reaction (25%, 3/12).ConclusionIS ELISPOT showed relatively high diagnostic value and accuracy in the TB group, independent of smear positivity. IS ELISPOT may provide additional diagnostic yield for microbiological tools in the rapid diagnosis of smear-negative TB.     

Comparison of the MB/BacT and BACTEC 460 TB systems for recovery of mycobacteria from various clinical specimens

A total of 1,830 specimens (75.7% respiratory and 24.3% nonrespiratory) were cultured in parallel with the MB/BacT and BACTEC 460 TB systems and on Lowenstein-Jensen (LJ) medium. Mycobacteria were identified from 173 (6.5%) specimens. The most common species recovered were Mycobacterium tuberculosis complex (65. 9%), Mycobacterium avium complex (22.5%), and Mycobacterium chelonae (9.2%). The recovery rates by individual systems were 96.5, 99.4, and 95.9% for MB/BacT, BACTEC 460 TB, and LJ medium, respectively, for all mycobacteria; the recovery rates were 99.1, 100, and 98.2%, respectively, for M. tuberculosis complex alone. The difference among the recovery rates for all mycobacteria and those for individual species was not significant. The BACTEC 460 TB system detected M. tuberculosis isolates more rapidly than the MB/BacT system (8 versus 11.8 days for smear-positive specimens [P < 0.01] and 18 versus 21 days for smear-negative specimens [P < 0.05]), whereas the MB/BacT system more rapidly detected the nontuberculous mycobacteria (17.1 versus 12.7 days [P < 0.01]). These results indicate that the nonradiometric MB/BacT system is a rapid, sensitive, and efficient method for the recovery of M. tuberculosis and nontuberculous mycobacteria from both pulmonary and extrapulmonary clinical specimens.     

Use of Transrenal DNA for the Diagnosis of Extrapulmonary Tuberculosis in Children: a Case of Tubercular Otitis Media

The diagnosis of tuberculosis (TB) is difficult in children, especially for smear-negative pulmonary and extrapulmonary TB, which are common at this age. We report an 11-year-old girl with TB otitis media with negative smear microscopy and Xpert MTB/RIF but positive Mycobacterium tuberculosis-specific transrenal DNA (Tr-MTB-DNA) test results and culture for M. tuberculosis.     

Use of Multiepitope Polyproteins in Serodiagnosis of Active Tuberculosis

Screening of genomic expression libraries from Mycobacterium tuberculosis with sera from tuberculosis (TB) patients or rabbit antiserum to M. tuberculosis led to the identification of novel antigens capable of detecting specific antibodies to M. tuberculosis. Three antigens, Mtb11 (also known as CFP-10), Mtb8, and Mtb48, were tested together with the previously reported 38-kDa protein, in an enzyme-linked immunosorbent assay (ELISA) to detect antibodies in TB patients. These four proteins were also produced as a genetically fused polyprotein, which was tested with two additional antigens, DPEP (also known as MPT32) and Mtb81. Sera from individuals with pulmonary and extrapulmonary TB, human immunodeficiency virus (HIV)-TB coinfections, and purified protein derivative (PPD)-positive and PPD-negative status with no evidence of disease were tested. In samples from HIV-negative individuals, the ELISA detected antibodies in >80% of smear-positive individuals and >60% smear-negative individuals, with a specificity of ~98%. For this group, smears detected 81.6% but a combination of smear and ELISA had a sensitivity of ~93%. The antigen combination detected a significant number of HIV-TB coinfections as well as antibodies in patients with extrapulmonary infections. Improved reactivity in the HIV-TB group was observed by including the antigen Mtb81 that was identified by proteomics. The data indicate that the use of multiple antigens, some of which are in a single polyprotein, can be used to facilitate the development of a highly sensitive test for M. tuberculosis antibody detection.     

Assessing the serodiagnostic potential of 35 Mycobacterium tuberculosis proteins and identification of four novel serological antigens

Improved diagnostic reagents are needed for the detection of Mycobacterium tuberculosis infections, and the development of a serodiagnostic test would complement presently available diagnostic methods. The aim of the present study was to identify novel serological targets for use for the future serodiagnosis of tuberculosis (TB). We cloned and expressed 35 M. tuberculosis proteins as recombinant proteins in Escherichia coli and analyzed their serodiagnostic potentials. By a two-step selection process, four superior seroantigens, TB9.7, TB15.3, TB16.3, and TB51, were identified, none of which has been described before. The four novel antigens were tested with panels of sera from smear-positive and smear-negative TB patients from areas both where TB is endemic and where TB is not endemic, with recognition frequencies ranging from 31 to 93% and with a specificity of at least 97%. The single most potent antigen was TB16.3, which had a sensitivity of 48 to 55% with samples from Danish resident TB patients and a sensitivity of 88 to 98% with samples from African TB patients. Importantly, the TB16.3 and the TB9.7 antigens were recognized by more than 85% of the samples from TB patients coinfected with human immunodeficiency virus, a patient group for which it is in general difficult to detect M. tuberculosis-specific antibodies.     

Clinical evaluation of the polymerase chain reaction for the rapid diagnosis of tuberculosis

AIMS: Use of the polymerase chain reaction for the detection of Mycobacterium tuberculosis (TB PCR) as a basis for making clinical decisions on the initiation of antituberculosis treatment was studied. METHODS: A retrospective study involving a cohort of 155 patients being investigated for tuberculosis in an infectious disease consultation service was undertaken. TB PCR was performed on pulmonary and extrapulmonary specimens from these patients. The sensitivity of TB PCR was analysed. RESULTS: Of the 155 patients, 144 fitted the clinical diagnosis of tuberculosis, and 112 of them were culture positive for M tuberculosis. Sixty (58.3%) patients with clinical features suggestive of tuberculosis received antituberculosis treatment based on positive TB PCR alone. Of 224 clinical specimens (138 pulmonary and 86 extrapulmonary) sent for TB PCR, 148 (99 pulmonary and 49 extrapulmonary) were positive in 117 patients. Of the 690 clinical specimens sent for culture, 279 were positive for M tuberculosis in 112 patients. The diagnostic sensitivity of TB PCR was 75.9% (85 of 112) and 81.3% (117 of 144) in patients with culture confirmed and clinically diagnosed tuberculosis, respectively. Using culture as the gold standard, the overall sensitivity of TB PCR was 78.3%, and for pulmonary and extrapulmonary specimens it was 82.3% and 72.0%, respectively. CONCLUSIONS: TB PCR is a rapid and reliable test in the diagnosis and management of tuberculosis.     

Field evaluation of a rapid immunochromatographic test for tuberculosis

Rapid diagnostic tests for tuberculosis (TB) are needed to facilitate early treatment of TB and prevention of Mycobacterium tuberculosis transmission. The ICT Tuberculosis test is a rapid, card-based immunochromatographic test for detection of antibodies directed against M. tuberculosis antigens. The objective of the study was to evaluate the performance of the ICT Tuberculosis test for the diagnosis of active pulmonary TB (PTB) with whole blood, plasma, and serum from patients suspected of having PTB and from asymptomatic controls in a setting with a high prevalence of PTB. Seventy patients suspected of having PTB (and who were later confirmed to have or not to have PTB by use of M. tuberculosis culture as the "gold standard") and 42 controls were studied. Twenty-one controls were neither vaccinated with Mycobacterium bovis bacillus Calmette-Guérin (BCG) nor tuberculin skin test (TST) positive (group A controls), and 21 controls were TST positive and/or had previously been vaccinated with BCG (group B controls). Study subjects were drawn from one hospital and one primary health care unit in Rio de Janeiro City, Brazil. One version of the test (ICT-1) was evaluated by using whole blood, plasma, and serum samples. Sera obtained for this study were frozen and later tested with a manufacturer-modified version of the test (ICT-2). Among the patients suspected of having PTB, the sensitivities of the ICT-1 with whole blood, serum, and plasma were 83, 65, and 70%, respectively, and the specificities were 46, 67, and 56%, respectively. Among the group A controls, the specificities of ICT-1 with the three specimen types were 95, 100, and 95%, respectively. Among the group B controls, the specificities of ICT-1 with the three specimen types were 71, 86, and 86%, respectively. Among the patients suspected of having PTB, the sensitivity of ICT-2 was 70% and the specificity was 65%. Among the group A controls, the specificity of ICT-2 was 95%, and among the group B controls, the specificity of ICT-2 was 81%. With a 29% observed prevalence of PTB among patients suspected of having PTB, the positive predictive values of the ICT tests ranged from 39 to 50% and the negative predictive values ranged from 82 to 87%. The ICT Tuberculosis tests were not sufficiently predictive to warrant their widespread use as routine diagnostic tests for PTB in this setting. However, further evaluation of these tests in specific epidemiologic settings may be warranted.     

Detection of Mycobacterium tuberculosis by EasyNAT Diagnostic Kit in Sputum Samples from Tanzania

The EasyNAT assay was evaluated for the detection of tuberculosis in sputum smears from presumptive pulmonary tuberculosis (TB) patients in an African high-TB and high-HIV setting. The sensitivity of the EasyNAT assay was 66.7%, and the specificity and positive predictive value were 100% for the culture-positive patients. The sensitivity was only 10% in the smear-negative and culture-positive patients.     

Levels of Circulating Immune Complexes Containing Mycobacterium Tuberculosis-specific Antigens in Pulmonary Tuberculosis and Sarcoidosis Patients

The present study was conducted to understand the aetiological link between tuberculosis (TB) and sarcoidosis. Sera from smear-positive TB subjects (n = 24), smear-negative TB subjects (n = 24), sarcoidosis patients (n = 24) and healthy controls (n = 24) were collected and circulating immune complexes were isolated. Sandwich ELISA was performed for detecting four highly specific mycobacterial regions of difference (RD) proteins (early secretory antigenic target 6 [ESAT6], 10 KDa culture filtrate protein [CFP10], 21 KDa CFP [CFP21] and mycobacterial protein from species TB [MPT 64]). Sensitivity and specificity was calculated, and receiver operating characteristic plots were plotted. Non-parametric Mann–Whitney U-test was used to calculate statistical significance. Seventy per cent of sarcoidosis patients showed the presence of immune complexes of mycobacterial RD proteins similar to that observed in the sera of smear-negative TB patients as opposed to antibody-based detection assay based on these RD proteins. Thus, immunoassays based on specific mycobacterial RD proteins also need to be developed and validated carefully to differentiate TB and sarcoidosis, a close mimic of smear-negative tuberculosis.     

Multicenter Evaluation of Anyplex Plus MTB/NTM MDR-TB Assay for Rapid Detection of Mycobacterium tuberculosis Complex and Multidrug-Resistant Isolates in Pulmonary and Extrapulmonary Specimens

The rapid diagnosis of tuberculosis (TB) and the detection of drug-resistant Mycobacterium tuberculosis strains are critical for successful public health interventions. Therefore, TB diagnosis requires the availability of diagnostic tools that allow the rapid detection of M. tuberculosis and drug resistance in clinical samples. Here, we performed a multicenter study to evaluate the performance of the Seegene Anyplex MTB/NTM MDR-TB assay, a new molecular method based on a multiplex real-time PCR system, for detection of Mycobacterium tuberculosis complex (MTBC), nontuberculous mycobacteria (NTM), and genetic determinants of drug resistance. In total, the results for 755 samples (534 pulmonary and 221 extrapulmonary samples) were compared with the results of smears and cultures. For pulmonary specimens, the sensitivities of the Anyplex assay and acid-fast bacillus smear testing were 86.4% and 75.0%, respectively, and the specificities were 99% and 99.4%. For extrapulmonary specimens, the sensitivities of the Anyplex assay and acid-fast bacillus smear testing were 83.3% and 50.0%, respectively, and the specificities of both were 100%. The negative and positive predictive values of the Anyplex assay for pulmonary specimens were 97% and 100%, respectively, and those for extrapulmonary specimens were 84.6% and 100%. The sensitivities of the Anyplex assay for detecting isoniazid resistance in MTBC strains from pulmonary and extrapulmonary specimens were 83.3% and 50%, respectively, while the specificities were 100% for both specimen types. These results demonstrate that the Anyplex MTB/NTM MDR-TB assay is an efficient and rapid method for the diagnosis of pulmonary and extrapulmonary TB and the detection of isoniazid resistance.