Evaluation of Two Line Probe Assays for Rapid Detection of Mycobacterium tuberculosis,Tuberculosis (TB) Drug Resistance,and Non-TB Mycobacteria in HIV-Infected Individuals with Suspected TB

Limited performance data from line probe assays (LPAs), nucleic acid tests used for the rapid diagnosis of tuberculosis (TB), nontuberculosis mycobacteria (NTM), and Mycobacterium tuberculosis drug resistance are available for HIV-infected individuals, in whom paucibacillary TB is common. In this study, the strategy of testing sputum with GenoType MTBDRplus (MTBDR-Plus) and GenoType Direct LPA (Direct LPA) was compared to a gold standard of one mycobacterial growth indicator tube (MGIT) liquid culture. HIV-positive (HIV⁺) individuals with suspected TB from southern Africa and South America with <7 days of TB treatment had 1 sputum specimen tested with Direct LPA, MTBDR-Plus LPA, smear microscopy, MGIT, biochemical identification of mycobacterial species, and culture-based drug-susceptibility testing (DST). Of 639 participants, 59.3% were MGIT M. tuberculosis culture positive, of which 276 (72.8%) were acid-fast bacillus (AFB) smear positive. MTBDR-Plus had a sensitivity of 81.0% and a specificity of 100%, with sensitivities of 44.1% in AFB smear-negative versus 94.6% in AFB smear-positive specimens. For specimens that were positive for M. tuberculosis by MTBDR-Plus, the sensitivity and specificity for rifampin resistance were 91.7% and 96.6%, respectively, and for isoniazid (INH) they were 70.6% and 99.1%. The Direct LPA had a sensitivity of 88.4% and a specificity of 94.6% for M. tuberculosis detection, with a sensitivity of 72.5% in smear-negative specimens. Ten of 639 MGIT cultures grew Mycobacterium avium complex or Mycobacterium kansasii, half of which were detected by Direct LPA. Both LPA assays performed well in specimens from HIV-infected individuals, including in AFB smear-negative specimens, with 72.5% sensitivity for M. tuberculosis identification with the Direct LPA and 44.1% sensitivity with MTBDR-Plus. LPAs have a continued role for use in settings where rapid identification of INH resistance and clinically relevant NTM are priorities.     

Comparison of the Automated Mycobacteria Growth Indicator Tube System (BACTEC 960/MGIT) with Löwenstein-Jensen medium for recovery of mycobacteria from clinical specimens

We examined whether the BACTEC/Mycobacteria Growth Indicator Tube (MGIT) system alone could supplant the use of a supplemental L?wenstein-Jensen (LJ) slant for routine recovery of Mycobacterium species from clinical specimens. A total of 6,062 specimens were included in the study. Of these, 273 specimens were positive for 278 mycobacterial isolates while 15 specimens were smear positive but culture negative using both media. Further analysis showed that 143 (51.4%) of the 278 total isolates were recovered from both the MGIT and LJ media. An additional 106 isolates (38.1%) were recovered from the MGIT only, while 29 (10.4%) isolates grew only on the LJ slant. The overall sensitivities of the MGIT and LJ media were 86.5% and 59.7%, respectively, for the recovery of mycobacteria from clinical materials. This study shows that although the MGIT system demonstrates better sensitivity for the recovery of mycobacteria from clinical specimens, both media types are necessary to maximize the sensitivity of detection.     

Comparison of mycobacteria growth indicator tube with BACTEC 460 for detection and recovery of mycobacteria from clinical specimens.

We compared the Mycobacteria Growth Indicator Tube (MGIT) system with the BACTEC 460 (B460) and Lowenstein Jensen (LJ) systems for the recovery of mycobacteria (acid-fast bacteria [AFB]) from 1,441 clinical specimens. Excluding 13 isolates of Mycobacterium gordonae, 178 significant AFB isolates were recovered from 113 patients. Isolates (119) of the Mycobacterium avium complex (MAC) accounted for 67% of all isolates, while isolates (30) of the Mycobacterium tuberculosis complex (MTB) accounted for 17% of isolates. The MGIT system recovered 98 (82%) MAC and 27 (90%) MTB isolates, while the B460 system recovered 101 (85%) MAC and 28 (93%) MTB isolates and the LJ system recovered 91 (76%) MAC and 25 (83%) MTB isolates. Overall, the MGIT system recovered 152 isolates of AFB (85.4% sensitivity), and the B460 and LJ systems recovered 151 (84.8% sensitivity) and 137 (76.9% sensitivity) AFB isolates, respectively. The recoveries of AFB with combinations of media were as follows: MGIT + LJ, 93.2%; B460 + LJ, 92.1%; and MGIT + B460, 96.6%. Although the sensitivity of MGIT was equivalent to that of B460, MGIT required a longer incubation (median, 11 days) than did B460 (median, 8 days) to become positive (P < 0.05).     

EVALUATION OF THE BACTEC MGIT 960 TB SYSTEM FOR RECOVERY AND IDENTIFICATION OF MYCOBACTERIUM TUBERCULOSIS COMPLEX IN A HIGH THROUGH PUT TERTIARY CARE CENTRE

Aim: To evaluate the performance of an automated BACTEC MGIT 960, a non-radioactive, non-invasive liquid culture system for cultivation of M. tuberculosis complex in terms of recovery rate and time. Materials and Methods: From March 2005 to December 2007, 14,597 specimens were processed using the MGIT 960 system and the results were compared with conventional L.J medium. We standardised ρ-nitro benzoic acid (PNBA) assay on MGIT 960 TB system for identification of M. tuberculosis complex and evaluated its usefulness by comparing the results with an in-house molecular assay and sequencing. Results and Discussion: Of the total 6143 (42%) isolates positive for M. tuberculosis complex, 6015 (41%) were positive by MGIT 960 TB system. In contrast, 3526 (24%) M. tuberculosis complex isolates grew on the conventional L.J medium. The mean turn around time for mycobacterial growth in smear-positive specimens was nine days for MGIT 960, and 38 days for L.J. medium whereas in smear negative specimens it was 16 days by MGIT vs. 48 days by L.J. Conclusion: MGIT 960 system with PNBA assay for identification of M. tuberculosis complex is a rapid and useful method in laboratories processing a large number of specimens.     

A comparative study for the detection of Mycobacteria by BACTEC MGIT 960, Lowenstein Jensen media and direct AFB smear examination

PURPOSE: To compare BACTEC MGIT 960 (M960) with conventional culture on Lowenstein Jensen (LJ) media and direct acid fast bacilli (AFB) smear examination for the detection of Mycobacteria in clinical samples obtained from suspected cases of pulmonary and extra pulmonary tuberculosis (TB). METHODS: A total of 500 samples were processed for direct AFB smear examination, and culture on M960 and LJ media. RESULTS: Two hundred fifty-eight out of 500 (51.6%) isolates of Mycobacteria were obtained by combined use of the two culture methods. Two hundred and fifty-three (50.6%) were positive in culture by M960 and LJ media and 28% (140/500) by direct AFB smear examination. The positivity rate of M960 system alone was 34.10% (88/258) and of LJ alone was 1.93% (5/258). Average time to detect growth (TTD) was 9.66 days by M960 and 28.81 days by LJ. CONCLUSIONS: M960 system is a rapid and sensitive method for early diagnosis of pulmonary and extrapulmonary TB. But for maximum recovery of Mycobacteria , a combination of both M960 and LJ media should be used.     

Comparison of the BACTEC MGIT 960 and BACTEC 460TB systems for detection of mycobacteria in clinical specimens

The reliability of the Mycobacteria Growth Indicator Tube (MGIT) 960 system for rapid detection of mycobacteria in clinical specimens was evaluated and compared to the radiometric method (BACTEC 460TB) and to mycobacterial culture on Lowenstein-Jensen (LJ) medium. Clinical specimens (n = 590) were tested without selection. A total of 121 (20.5%) isolates of mycobacteria were recovered; 98 (81.0%) of them were recovered with the BACTEC 460TB system, 86 (71.1%) were recovered with the BACTEC MGIT 960 system, and 55 (45.5%) were recovered with LJ medium (MGIT 960 versus BACTEC 640TB, p >0.05; MGIT 960 or BACTEC 460TB versus LJ, p <0.001). The mean time to detection (TTD) was 18 da for BACTEC 460 TB, and 13 da for BACTEC MGIT 960. The mean time to detection in each system, based upon data where both systems were culture positive, was significantly different (16.6 da for BACTEC 460TB and 13 da for BACTEC MGIT 960, p<0.001). The contamination rate of the BACTEC MGIT 960 system was 13.2%, which was intermediate between the BACTEC 460TB system (11.7%) and the LJ medium (14.7%). These data indicate that the fully automated MGIT 960 system is an accurate, non-radiometric alternative to the BACTEC 460TB method for rapid detection of mycobacteria in a clinical setting.     

Comparison of solid and liquid culture media with the polymerase chain reaction for detection ofMycobacterium tuberculosis in clinical samples

The aim of this study was to determine the sensitivity of different methods — two commercial polymerase chain reaction (PCR) kits (a protocol of nested PCR and a protocol of amplification of the IS6110 insertion element), the radiometric Bactec system, the Septi-Chek AFB culture system, and culture in Löwenstein-Jensen (LJ) solid medium — for the detection ofMycobacterium tuberculosis. One hundred clinical samples from 51 patients with culture-positive tuberculosis (81 specimens) and 19 controls (19 specimens) were used. Eighty-nine percent of the samples were smear negative. In the 81 specimens obtained from patients with tuberculosis, the frequency of positivity was 66.6% for nested PCR, 63% for culture in liquid media, 38.3% for the IS6110 assay, and 28.4% for culture on LJ medium. In 18 samples obtained by invasive procedures in patients with tuberculosis, mycobacterial DNA was detected by nested PCR in 83.3% (including all samples positive by culture on liquid media), by culture in liquid media in 77.7%, by culture on LJ medium in 27.7%, and by the IS6110 assay in 11.1%. No false-positive results were obtained from the negative control specimens with any of the techniques tested. The sensitivity of the reamplification protocol appears to be superior to that of the IS6110 assay and similar to that of the Bactec system.     

Use of Colorimetric Culture Methods for Detection of Mycobacterium tuberculosis Complex Isolates from Sputum Samples in Resource-Limited Settings

Despite recent advances, tuberculosis (TB) diagnosis remains imperfect in resource-limited settings due to its complexity and costs, poor sensitivity of available tests, or long times to reporting. We present a report on the use of colorimetric methods, based on the detection of mycobacterial growth using colorimetric indicators, for the detection of Mycobacterium tuberculosis in sputum specimens. We evaluated the nitrate reductase assay (NRA), a modified NRA using para-nitrobenzoic acid (PNB) (NRAp), and the resazurin tube assay using PNB (RETAp) to differentiate tuberculous and nontuberculous mycobacteria. The performances were assessed at days 18 and 28 using mycobacterium growth indicator tube (MGIT) and Löwenstein-Jensen (LJ) medium culture methods as the reference standards. We enrolled 690 adults with suspected pulmonary tuberculosis from a regional referral hospital in Uganda between March 2010 and June 2011. At day 18, the sensitivities and specificities were 84.6% and 90.0% for the NRA, 84.1% and 92.6% for the NRAp, and 71.2% and 99.3% for the RETAp, respectively. At day 28, the sensitivity of the RETAp increased to 82.6%. Among smear-negative patients with suspected TB, sensitivities at day 28 were 64.7% for the NRA, 61.3% for the NRAp, and 50% for the RETAp. Contamination rates were found to be 5.4% for the NRA and 6.7% for the RETAp, compared with 22.1% for LJ medium culture and 20.4% for MGIT culture. The median times to positivity were 10, 7, and 25 days for colorimetric methods, MGIT culture, and LJ medium culture,respectively. Whereas the low specificity of the NRA/NRAp precludes it from being used for TB diagnosis, the RETAp might provide an alternative to LJ medium culture to decrease the time to culture results in resource-poor settings.     

Detection of 123 bp fragment of insertion element IS6110 Mycobacterium tuberculosis for diagnosis of extrapulmonary tuberculosis

Purpose: Extrapulmonary tuberculosis (EPTB) is emerging problem in developing and developed countries. The diagnosis of EPTB in its different clinical presentations remains a true challenge. IS6110-based polymerase chain reaction (PCR) is used for rapid identification and positivity rate of the Mycobacterium tuberculosis complex in clinical isolates of different sites of EPTB. The present study was carried out to study the prevalence of M. tuberculosis complex in clinical isolates of EPTB at tertiary care centres in Lucknow. Materials and Methods: Seven hundred fifty-six specimens were collected from the suspected cases of EPTB which were processed for Mycobacteria by Ziehl Neelson (ZN) staining and BACTEC culture. All the specimens were also processed for IS6110-based PCR amplification with primers targeting 123 bp fragment of insertion element IS6110 of the M. tuberculosis complex. Results: Of these 756 specimens, 71(9.3%) were positive for acid fast bacilli (AFB) by ZN staining, 227(30.1%) were positive for mycobacteria by BACTEC culture and IS6110 PCR were positive for M. tuberculosis complex in 165 (20.7%) isolates. We found a significant difference in sensitivities of different tests (P<0.05). Conclusions: This study reveals the positivity of M. tuberculosis complex in clinical isolates of EPTB case in tertiary care hospitals in Northern India. 72.7% of M. tuberculosis complex was confirmed by IS6110-PCR in culture isolates from different sites of EPTB. The high prevalence of the M. tuberculosis complex was seen in lymph node aspirate and synovial fluid. However, utility of PCR may play a potentially significant role in strengthening the diagnosis of EPTB especially targeting IS6110.     

Use of the real-time PCR assay in conjunction with MagNA Pure for the detection of mycobacterial DNA from fixed specimens.

Tuberculosis in immunocompromised patients is often caused by Mycobacterial species other than Mycobacterium tuberculosis. Thus, detection of and differentiation between M. tuberculosis and nontuberculosis species is necessary for diagnosis of disease in these patients. Furthermore, when tissue changes show granulomatous inflammation, quick confirmation testing for mycobacterial infection is needed for conclusive diagnosis. The aim of this study was to validate the utility of a real-time polymerase chain reaction (PCR) assay in conjunction with the MagNA Pure LC automated extraction system for the detection of mycobacterial DNA from formalin-fixed, paraffin-embedded specimens. A total of 46 archived, paraffin-embedded, fixed specimens showing granulomatous inflammation were studied for mycobacterial infection by real-time PCR. Bacterial DNA was extracted and isolated using the MagNA Pure extraction system. Real-time PCR was performed on the LightCycler using the Artus Real Art Mycob Diff ASR kit from Qiagen. Thirteen of the 46 patient specimens were positive for mycobacterial infection by acid-fast bacilli (AFB) stain. Of the13 reported positive by AFB stain, 12 where positive by real-time PCR. All 13 specimens reported positive by AFB were sent for culture confirmation. Eleven of 13 were returned positive by culture. Specimens reported as negative by culture and positive by real-time PCR were confirmed positive by a second PCR method from another reference laboratory. We believe that these studies are beneficial in the differential diagnosis of mycobacterial infection from fixed tissue specimens where tuberculosis might not have been clinically initially suspected and when specimens are not suitable for microbiologic examination.     

Real-time PCR assay using fine-needle aspirates and tissue biopsy specimens for rapid diagnosis of mycobacterial lymphadenitis in children

A real-time PCR assay was developed to diagnose and identify the causative agents of suspected mycobacterial lymphadenitis. Primers and probes for the real-time PCR were designed on the basis of the internal transcribed spacer sequence, enabling the recognition of the genus Mycobacterium and the species Mycobacterium avium and M. tuberculosis. The detection limit for the assay was established at 1,100 CFU/ml of pus, and the specificity tests showed no false-positive reaction with other mycobacterial species and other pathogens causing lymphadenitis. From 67 children with suspected mycobacterial lymphadenitis based on a positive mycobacterial skin test, 102 samples (58 fine-needle aspirates [FNA] and 44 tissue specimens) were obtained. The real-time PCR assay detected a mycobacterial infection in 48 patients (71.6%), whereas auramine staining and culturing were positive for 31 (46.3%) and 28 (41.8%) of the patients. The addition of the real-time PCR assay to conventional diagnostic tests resulted in the recognition of 13 more patients with mycobacterial disease. These results indicate that the real-time PCR is more sensitive than conventional staining and culturing techniques (P = 0.006). The M. avium-specific real-time PCR was positive for 38 patients, and the M. tuberculosis-specific real-time PCR was positive for 1 patient. Analysis of 27 patients from whom FNA and tissue biopsy specimens were collected revealed significantly more positive real-time PCR results for FNA than for tissue biopsy specimens (P = 0.003). Samples from an age-matched control group of 50 patients with PCR-proven cat scratch disease were all found to be negative by the real-time PCR. We conclude that this real-time PCR assay with a sensitivity of 72% for patients with lymphadenitis and a specificity of 100% for the detection of atypical mycobacteria can provide excellent support for clinical decision making in children with lymphadenitis.     

Rapid culture diagnosis of tuberculous lymphadenitis from a tertiary care centre in an endemic nation: Potential and pitfalls

In spite of low sensitivity and specificity, standard diagnostic algorithm recommends fine needle aspiration cytology (FNAC) and direct microscopic screening for acid-fast bacilli (AFB) for the routine diagnosis of tuberculous lymphadenopathy (LNTB). In this study, the diagnostic utility of liquid broth based automated culture (BacT/ALERT 3D) technique was assessed in comparison with conventional techniques in 89 clinically suspected tubercular lymphadenitis patients. 60% (n = 53) were positive by FNAC and 38.4% (n = 34) demonstrated AFB in smear examination. BacT/ALERT yielded isolation in 43.1% (n = 38) aspirates, confirming tubercular aetiology. We also found six paediatric culture-positive cases which showed negative outcome by both FNAC and smear. Thus, we conclude that culture by BacT/ALERT, may be used for faster yield of Mycobacteria in LNTB, especially in children. Additionally, this could also be used as a platform for further differentiation of Mycobacterium tuberculosis from non-tuberculous mycobacteria (NTM) infection and for testing of anti-tubercular chemotherapeutic agents whenever drug resistance is suspected.     

Diagnostic potential of IS6110, 38kDa, 65kDa and 85B sequence-based polymerase chain reaction in the diagnosis of Mycobacterium tuberculosis in clinical samples

Purpose: The correlation between the presence of specific gene sequence of M. tuberculosis and specific diagnosis of clinical tuberculosis is not known. This study compared the results of polymerase chain reaction (PCR) amplification of M. tuberculosis specific DNA sequences (IS6110, 65kDa, 38kDa and mRNA coding for 85 B protein) from different clinical samples of pulmonary and extrapulmonary tuberculosis. Methods: One hundred and seventy-two clinical samples from suspected tuberculosis patients were tested for smear examination, culture (LJ and rapid BACTEC 460 TB system) and PCR. PCR was performed with specific primers for the targets: IS6110, 65kDa, 38kDa and 85B. Results: Each PCR test was found to have a much higher positivity than conventional test and BACTEC culture (P 0.05). Smear positive samples (56) and the samples (36) showing positive results by conventional methods (smear and LJ medium culture) and BACTEC were found to be positive by all PCR protocols. No significant difference was found between the four PCR protocols (P >0.05). The primer specific for amplifying the 123bp IS6110 fragment gave the highest positivity (83%), followed by 65kDa, 38kDa and 85B RT-PCR in descending order. Conclusions: These data suggest that the presence of IS6110 correlates more closely with the diagnosis of clinical tuberculosis than that of 65kDa, 38kDa and 85B     

Diagnostic Accuracy of Xpert MTB/RIF Assay in Extrapulmonary Tuberculosis

Introduction: The WHO endorsed Xpert Mycobacterium tuberculosis/rifampicin (MTB/RIF) assay, has been evaluated for pulmonary TB in a number of studies but very few have investigated it for extrapulmonary specimens. The present study evaluates the performance of Xpert MTB/RIF assay in the diagnosis of extrapulmonary TB (EPTB). Aim and Objectives: The aim of the study is to determine sensitivity and specificity of Xpert MTB/RIF assay for diagnosis of EPTB and RIF resistance in comparison to culture on Lowenstein–Jensen (LJ) medium and proportion method (PM), respectively. Materials and Methods: A total of 738 specimens from clinically suspected cases of EPTB were subjected to Ziehl–Neelsen staining, Xpert MTB/RIF assay and culture on LJ medium. PM was done on MTB isolates. Results: The sensitivity, specificity of Xpert MTB/RIF assay for diagnosis of EPTB were 84.91% (95% confidence interval [CI] 72.41%–93.25%) and 86.72% (95% CI 83.94%–89.17%) and for RIF resistance detection were 60.00% (95% CI 32.29%–83.66%) and 94.74% (95% CI 73.97%–99.87%), respectively. Among culture-positive cases, the sensitivity of Xpert MTB/RIF assay was 94.12% in smear positive and 80.56% in smear-negative cases. Xpert MTB/RIF showed maximum sensitivity of MTB detection from lymph node specimens (100% [95% CI 54.07%–100.00%]) and other body fluids (100% [95% CI 15.81%–100.00%]). Conclusion: The present study establishes Xpert MTB/RIF assay as a promising tool in the rapid diagnosis of EPTB and detection of RIF resistance.     

Comparison of the Septi-Chek AFB and BACTEC systems and conventional culture for recovery of mycobacteria.

The performance of the Septi-Chek AFB system was compared with that of the BACTEC radiometric system and that of Lowenstein-Jensen agar slants (LJ) for detection of mycobacteria in clinical specimens. A total of 642 specimens were cultured; 61 (9.5%) yielded mycobacteria. Mycobacterium tuberculosis (34 isolates) and Mycobacterium avium complex (25 isolates) were the predominant species isolated. Of the 61 culture-positive specimens, 30 were smear positive and 31 were smear negative. Overall, 95% of the positive specimens were detected by Septi-Chek and BACTEC (100% of M. tuberculosis isolates) and 75% by LJ (82% of M. tuberculosis isolates). The mean times to detection were 15 days for BACTEC, 23 days for Septi-Chek, and 27 days for LJ. Of the 30 smear-positive specimens, 100% were recovered by Septi-Chek and BACTEC and 90% were recovered by LJ. Of the 31 smear-negative specimens, 90% were detected by Septi-Chek and BACTEC and 61% were detected by LJ. The Septi-Chek and BACTEC systems are superior to the conventional (LJ) mycobacterial culture method. Although Septi-Chek requires more time for the detection of mycobacteria than BACTEC, it is comparable in terms of overall recovery.     

Application of PCR from the fine needle aspirates for the diagnosis of cervical tuberculous lymphadenitis.

Tuberculosis remains a major public health problem worldwide. A definitive and accurate diagnosis of tuberculosis in cervical lymphadenopathy is important because satisfactory results can be achieved with chemotherapy alone, obviating surgery. Recently, fine needle aspiration cytology (FNAC) has provided an alternative and easy procedure for collection of material for cytomorphologic and bacteriologic examination. But the detection rate for M. tuberculosis from the aspirate material is still low with Ziehl-Neelson stain and even with culture. The authors therefore performed polymerase chain reaction (PCR) for mycobacterial DNA sequences in 31 cases of cytodiagnosis of tuberculous lymphadenitis and compared conventional bacteriologic methods. Ziehl-Neelson staining for acid-fast bacilli (AFB) was positive in 3 cases (10%) in direct smears, and the cultures for M. tuberculosis were positive in 6 cases (19%). In 19 (61%) among 31 samples, mycobacterial DNA fragments were detected, using the PCR method. With combined conventional and PCR method, the rate of detection was increased to 68 percent high. In conclusion, PCR is the most sensitive technique in the demonstration of M. tuberculosis in patient with clinically suspected as tuberculosis, who have AFB stain or culture negative cytology. Combined conventional and PCR methods as well as cytologic findings are of further help in the detection and characterization of M. tuberculosis.     

Bacterial Lymphadenitis at a Major Referral Hospital in France from 2008 to 2012

Lymph node enlargement is a common medical problem, and in a large number of patients, the causes of lymphadenopathy remain undiagnosed. We report a thorough microbiological analysis of 1,688 lymph node biopsy specimens collected in our bartonellosis reference center. We studied lymph node biopsy samples from patients with suspected regional infectious lymph node enlargement from January 2008 to December 2012. To evaluate a useful strategy for the diagnosis of infectious lymphadenitis, specimens were cultured and subjected to molecular assays. Histologic analysis was done when possible. A total of 642 (38%) biopsy specimens were infected with a bacterial agent, and quantitative PCR (qPCR) was significantly better than 16S rRNA gene PCR (rrs) for the detection of Bartonella henselae (P = 0.05), Mycobacterium tuberculosis (P = 0.05), and Mycobacterium avium (P = 0.007). Molecular assays were significantly better than bacterial cultures for the diagnosis of Francisella tularensis (P = 0.017) but were less effective for detecting M. tuberculosis (P = 0.004) and M. avium (P = 0.001). Histologic analysis was done for 412 lymph nodes, and 20% of these were compatible with an infectious lymphadenitis, whereas a neoplasm was found in 29% of these lymph nodes. M. tuberculosis was detected significantly more in female than in male patients (P = 0.01), and patients with cat scratch disease (CSD) were younger than patients with M. tuberculosis, Tropheryma whipplei, and F. tularensis. Negative rrs PCR does not exclude the diagnosis of infectious lymphadenitis. Histologic analysis of lymph node biopsy specimens is critical, as a diagnosis of infectious lymphadenitis does not preclude other concurrent diseases.     

Comparison of the mycobacteria growth indicator tube with MB redox, Löwenstein-Jensen, and Middlebrook 7H11 media for recovery of mycobacteria in clinical specimens

The rate of recovery and the mean time to detection of mycobacteria in clinical specimens were evaluated with two nonradiometric broth-based systems, the Mycobacteria Growth Indicator Tube (MGIT) and MB Redox systems. The data obtained for each system were compared with each other and with those obtained with the L?wenstein-Jensen (LJ) and Middlebrook 7H11 reference media. A total of 117 mycobacterial isolates (Mycobacterium tuberculosis, n = 112; nontuberculous mycobacteria, n = 5) were detected in 486 clinical specimens. The recovery rates for M. tuberculosis were 91 of 112 (81.3%) isolates with MGIT and 81 of 112 (72.3%) isolates with MB Redox. The combination of MGIT plus MB Redox recovered 104 of the 112 (92.9%) M. tuberculosis isolates. MGIT plus LJ plus Middlebrook 7H11 recovered 106 of the 112 (94.6%) isolates, MB Redox plus LJ plus Middlebrook 7H11 recovered 99 of the 112 (88.4%) isolates, and LJ plus Middlebrook 7H11 recovered 84 of the 112 (75. 0%) isolates. The mean time to detection of M. tuberculosis in smear-positive specimens was 7.2 days with MGIT, 6.9 days with MB Redox, 20.4 days with LJ, and 17.6 days with Middlebrook 7H11. The mean time to detection of M. tuberculosis in smear-negative specimens was 19.1 days with MGIT, 15.5 days with MB Redox, 25.8 days with LJ, and 21.6 days with Middlebrook 7H11. The contamination rates were 4.4, 3.8, 2.1, and 2.7% for MGIT, MB Redox, LJ, and Middlebrook 7H11, respectively. In conclusion, MGIT and MB Redox can be viable tools in the routine mycobacteriology laboratory.     

Improvement of tuberculosis diagnosis by the Mycobacteria Growth Indicator Tube (MGIT) in a developing country laboratory

In order to improve tuberculosis diagnosis in a developing country (Senegal), we evaluated a new liquid-based medium and nonradioactive system, Mycobacteria Growth Indicator Tube (MGIT), with individual clinical specimens collected in Dakar. The main purpose was to compare the time to detection and the rate of recovery of Mycobacterium tuberculosis complex and to determine its importance with respect to Lowenstein-Jensen (LJ), a liquid-based-medium for isolation of M. tuberculosis complex. 531 specimens were processed with Mycoprep kit containing NaOH-N-acetyl L-cystein and inoculated on both LJ and MGIT and incubated at 37 degrees C for 60 days. For each medium, the recovery rate and the time to detection were recorded. Among the 531 specimens, of which 121 smears were positive, 32.5% (173/531) grew the M. tuberculosis complex. Of these, 103 were smear positive (S+) and 70 smear negative (S-). LJ recovered 54.9% (95/173) and MGIT recovered 91.9% (159/173). Disagreements were observed with 92 isolates, LJ failed to recover 78 while MGIT failed to recover 14. The overall mean time to detection was 20.1 days for LJ and 10.5 days for MGIT. MGIT has shown a better sensitivity in isolation with significant reduction in reporting culture for M. tuberculosis complex. As a simple and a nonradiometric system, it could be used in conjunction with egg-based media in developing countries laboratories.     

Comparison of the conventional diagnostic modalities, bactec culture and polymerase chain reaction test for diagnosis of tuberculosis

Purpose: To evaluate the performance of 65 kDa antigen based PCR assay in clinical samples obtained from pulmonary and extrapulmonary cases of tuberculosis. Methods: One hundred and fifty six samples were processed for detection of Mycobacterium tuberculosis by ZN smear examination, LJ medium culture, BACTEC radiometric culture and PCR tests. Results: A significant difference was seen in the sensitivities of different tests, the figures being 74.4% for PCR test, 33.79% for ZN smear examination, 48.9% for LJ culture and 55.8% for BACTEC culture (P<0.05). However, there was no significant difference (P>0.05) as far as specificity of different tests was concerned. PCR test sensitivity in pulmonary and extrapulmonary clinical samples were 72.7% and 75.9% respectively and found to be significantly higher (P<0.05) when compared with those of other tests. The mean detection time for M.tuberculosis was 24.03 days by LJ medium culture, 12.89 days by BACTEC culture and less than one day by PCR test. Conclusions: PCR is a rapid and sensitive method for the early diagnosis of pulmonary and extrapulmonary tuberculosis.