Evaluation of GeneXpert MTB/RIF and BACTEC-MGIT 960 for the detection of tuberculosis among pneumoconiosis-associated tuberculosis patients / 中华劳动卫生职业病杂志

Objective@#To evaluate the performence of GeneXpert MTB/RIF and BACTEC-MGIT 960 on detecting Mycobacterium tuberculosis and rifampicin resistance for pneumoconiosis-associated tuberculosis patients.@*Methods@#The recruited 133 suspected active pneumoconiosis-associated tuberculosis hospitalized cases, morning sputum samples were collected to do modified L-J culture, conventional proportion method drug susceptibility test, GeneXpert MTB/RIF and BACTEC-MGIT 960. Analyze the sensitivity and specificity of the 133 sputum from patients, the positive rates of patients with tuberculosis in GeneXpert MTB/RIF test, BACTEC-MGIT 960 and modified L-J culture were 37.59%, 34.59% and 30.08% respectively. There was no significant difference among the three tests respectively (P>0.05) . According to the modified L-J culture, the sensitivity of GeneXpert MTB/RIF and BACTEC-MGIT 960 in detecting tuberculosis were 92.5% and 95.0% respectively, and specificity in rifampicin resistance were 86.0% and 91.4% respectively. There was no significant difference between GeneXpert MTB/RIF and BACTEC-MGIT 960 (P>0.05) . According to conventional proportion method drug susceptibility test, the sensitivity of GeneXpert MTB/RIF and BACTEC-MGIT 960 in detecting rifampicin resistance were 90.0% and 100%, and specificity were 92.6% and 96.4%. There was no significant difference between GeneXpert MTB/RIF and BACTEC-MGIT 960 (P>0.05) .@*Conclusion@#The GeneXpert MTB/RIF has good performence of detecting tuberculosis and rifampicin resistance. It has good application value among pneumoconiosis-associated tuberculosis patients.

Rapid culture-based detection of living mycobacteria using microchannel electrical impedance spectroscopy (m-EIS)

BACKGROUND: Multiple techniques exist for detecting Mycobacteria, each having its own advantages and drawbacks. Among them, automated culture-based systems like the BACTEC-MGIT™ are popular because they are inexpensive, reliable and highly accurate. However, they have a relatively long "time-to-detection" (TTD). Hence, a method that retains the reliability and low-cost of the MGIT system, while reducing TTD would be highly desirable. METHODS: Living bacterial cells possess a membrane potential, on account of which they store charge when subjected to an AC-field. This charge storage (bulk capacitance) can be estimated using impedance measurements at multiple frequencies. An increase in the number of living cells during culture is reflected in an increase in bulk capacitance, and this forms the basis of our detection. M. bovis BCG and M. smegmatis suspensions with differing initial loads are cultured in MGIT media supplemented with OADC and Middlebrook 7H9 media respectively, electrical "scans" taken at regular intervals and the bulk capacitance estimated from the scans. Bulk capacitance estimates at later time-points are statistically compared to the suspension's baseline value. A statistically significant increase is assumed to indicate the presence of proliferating mycobacteria. RESULTS: Our TTDs were 60 and 36 h for M. bovis BCG and 20 and 9 h for M. smegmatis with initial loads of 1000 CFU/ml and 100,000 CFU/ml respectively. The corresponding TTDs for the commercial BACTEC MGIT 960 system were 131 and 84.6 h for M. bovis BCG and 41.7 and 12 h for M smegmatis, respectively. CONCLUSION: Our culture-based detection method using multi-frequency impedance measurements is capable of detecting mycobacteria faster than current commercial systems.

Evaluation of anti-tubercular activity of linolenic acid and conjugated-linoleic acid as effective inhibitors against Mycobacterium tuberculosis

OBJECTIVE@#To evaluate a new pharmacological activity/effect of linolenic acid (α- and γ-form) and conjugated-linoleic acid (CLA) causing antibacterial activity against Mycobacterium tuberculosis (Mtb).@*METHODS@#The anti-Mtb activity/effect of linolenic acid and CLA were determined using different anti-Mtb indicator methods such as resazurin microtiter assay (REMA) and MGIT 960 system assay. The Mtb was incubated with various concentrations (12.5-200 μg/mL) of the compounds and anti-Mtb first-line drugs for 5 d in the REMA, and for 3 wk in MGIT 960 system assay.@*RESULTS@#Linolenic acid and CLA obviously indicated their anti-Mtb activity/effect by strongly inhibiting the growth/proliferation of Mtb in a dose-dependent manner in the REMA and the MGIT 960 system assay. Interestingly, linolenic acid and CLA consistently induced anti-Mtb activity/effect by effectively inhibiting the growth/proliferation of Mtb in MGIT 960 system for 21 d with a single treatment, and their minimum inhibitory concentrations were measured as 200 μg/mL respectively.@*CONCLUSIONS@#These results demonstrate that linolenic acid and CLA not only have effective anti-Mtb activity/properties, but also induce the selective-anti-Mtb effects by strongly inhibiting and blocking the growth/proliferation of Mtb through a new pharmacological activity/action. Therefore, this study provides novel perspectives for the effective use of them and the potential that can be used as potent anti-Mtb candidate drugs, as well as suggests the advantage of reducing the cost and/or time for developing a new/substantive drug by effectively repurposing the existing drugs or compounds as one of new strategies for the global challenge of tuberculosis.

The anti-tubercular activity of Melia azedarach L. and Lobelia chinensis Lour. and their potential as effective anti-Mycobacterium tuberculosis candidate agents

Objective: To evaluate the anti-mycobacterial activity of Melia azedarach L. (M. azedarach) and Lobelia chinensis Lour. (L. chinensis) extracts against the growth of Mycobacterium tuberculosis (M. tuberculosis). Methods: The anti-M. tuberculosis activity of M. azedarach and L. chinensis extracts were evaluated using different indicator methods such as resazurin microtiter assay (REMA) and mycobacteria growth indicator tube (MGIT) 960 system assay. The M. tuberculosis was incubated with various concentrations (50–800 mg/mL) of the ex-tracts for 5 days in the REMA, and for 4 weeks in MGIT 960 system assay. Results: M. azedarach and L. chinensis extracts showed their anti-M. tuberculosis ac-tivity by strongly inhibiting the growth of M. tuberculosis in a concentration-dependent manner in the REMA and the MGIT 960 system assay. Particularly, the methanol extract of M. azedarach and n-hexane extract of L. chinensis consistently exhibited their effects by effectively inhibiting the growth of M. tuberculosis in MGIT 960 system for 4 weeks with a single-treatment, indicating higher anti-M. tuberculosis activity than other extracts, and their minimum inhibitory concentrations were measured as 400 mg/mL and 800 mg/mL, respectively. Conclusions: These results demonstrate that M. azedarach and L. chinensis extracts not only have unique anti-M. tuberculosis activity, but also induce the selective anti-M. tuberculosis effects by consistently inhibiting or blocking the growth of M. tuberculosis through a new pharmacological action. Therefore, this study suggests the potential of them as effective candidate agents of next-generation for developing a new anti-tuberculosis drug, as well as the advantage for utilizing traditional medicinal plants as one of effective strategies against tuberculosis.

Evaluation of anti-tubercular activity of linolenic acid and conjugated-linoleic acid as effective inhibitors against Mycobacterium tuberculosis

Objective: To evaluate a new pharmacological activity/effect of linolenic acid (α- and γ-form) and conjugated-linoleic acid (CLA) causing antibacterial activity against Mycobacterium tuberculosis (Mtb). Methods: The anti-Mtb activity/effect of linolenic acid and CLA were determined using different anti-Mtb indicator methods such as resazurin microtiter assay (REMA) and MGIT 960 system assay. The Mtb was incubated with various concentrations (12.5-200 μg/mL) of the compounds and anti-Mtb first-line drugs for 5 d in the REMA, and for 3 wk in MGIT 960 system assay. Results: Linolenic acid and CLA obviously indicated their anti-Mtb activity/effect by strongly inhibiting the growth/proliferation of Mtb in a dose-dependent manner in the REMA and the MGIT 960 system assay. Interestingly, linolenic acid and CLA consistently induced anti-Mtb activity/effect by effectively inhibiting the growth/proliferation of Mtb in MGIT 960 system for 21 d with a single treatment, and their minimum inhibitory concentrations were measured as 200 μg/mL respectively. Conclusions: These results demonstrate that linolenic acid and CLA not only have effective anti-Mtb activity/properties, but also induce the selective-anti-Mtb effects by strongly inhibiting and blocking the growth/proliferation of Mtb through a new pharmacological activity/action. Therefore, this study provides novel perspectives for the effective use of them and the potential that can be used as potent anti-Mtb candidate drugs, as well as suggests the advantage of reducing the cost and/or time for developing a new/substantive drug by effectively repurposing the existing drugs or compounds as one of new strategies for the global challenge of tuberculosis.

The anti-tubercular activity of Melia azedarach L. and Lobelia chinensis Lour. and their potential as effective anti-Mycobacterium tuberculosis candidate agents

Objective To evaluate the anti-mycobacterial activity of Melia azedarach L. (M. azedarach) and Lobelia chinensis Lour. (L. chinensis) extracts against the growth of Mycobacterium tuberculosis (M. tuberculosis). Methods The anti-M. tuberculosis activity of M. azedarach and L. chinensis extracts were evaluated using different indicator methods such as resazurin microtiter assay (REMA) and mycobacteria growth indicator tube (MGIT) 960 system assay. The M. tuberculosis was incubated with various concentrations (50–800 μg/mL) of the extracts for 5 days in the REMA, and for 4 weeks in MGIT 960 system assay. Results M. azedarach and L. chinensis extracts showed their anti-M. tuberculosis activity by strongly inhibiting the growth of M. tuberculosis in a concentration-dependent manner in the REMA and the MGIT 960 system assay. Particularly, the methanol extract of M. azedarach and n-hexane extract of L. chinensis consistently exhibited their effects by effectively inhibiting the growth of M. tuberculosis in MGIT 960 system for 4 weeks with a single-treatment, indicating higher anti-M. tuberculosis activity than other extracts, and their minimum inhibitory concentrations were measured as 400 μg/mL and 800 μg/mL, respectively. Conclusions These results demonstrate that M. azedarach and L. chinensis extracts not only have unique anti-M. tuberculosis activity, but also induce the selective anti-M. tuberculosis effects by consistently inhibiting or blocking the growth of M. tuberculosis through a new pharmacological action. Therefore, this study suggests the potential of them as effective candidate agents of next-generation for developing a new anti-tuberculosis drug, as well as the advantage for utilizing traditional medicinal plants as one of effective strategies against tuberculosis.

Effectiveness of Tuberculosis Smear Microscopy in Laboratory Diagnosis of Mycobacterium tuberculosis in Resource Constrained Settings.

Aim: To determine the correlation of accuracy of direct smear microscopy compared with BACTEC MGIT 960. Design: The study prospectively compare direct smear microscopy with BACTEC MGIT 960 using the reference standard, Lowenstein Jensen culture. Place and Duration: The study was conducted in Zankli Medical Centre, Abuja, between November 2004 and July 2005. Methodology: 340 suspected patients for Mycobacterium tuberculosis referred from direct observation therapy clinics located in six different government owned health facilities were referred to our facility. These patients; male (192) and female (148) were between the age of 10 and 64 years old. Three sputa samples were collected over two consecutive days and direct smear microscopy and culture were performed on these samples. Results: When compared with the reference standard, BACTEC MGIT 960 has a sensitivity and specificity of 100.0% and 56.4% respectively, and a negative predictive value of 100.0%; indicating the proportion of AFB negative participants were actually not infected with M. tuberculosis when tested with BACTEC MGIT 960. The sensitivity of direct microscopy was significantly lower than BACTEC MGIT 960 (84.9% versus 100%, p<0.001) and the specificity was significantly higher (96.6% versus 56.4%, p<0.001). Conclusions: For the purpose of effectiveness of tuberculosis program in developing countries, direct smear microscopy may still be relevant in the diagnosis of Mycobacterium tuberculosis.

Detection of Mycobacterium in fine-needle aspiration biopsy specimens from children with tuberculous lymphadenitis / 中华微生物学和免疫学杂志

Objective To improve the diagnosis of tuberculosis ( TB) by analyzing Mycobacterium infection in fine-needle aspiration biopsy specimens from children with tuberculous lymphadenitis .Methods Fine-needle aspiration biopsy was performed on 269 children with tuberculous lymphadenitis diagnosed by Shanghai Public Health Clinical Center from January 2011 to September 2013 .The needle aspiration biopsy specimens were processed for acid-fast bacillus (AFB) smear test, mycobacterial culture and Mycobacterium identification ( p-nitrobenzoic acid inhibition test ) .Results Cytological diagnosis of tuberculous lymphade-nitis was made for 269 patients.The positive results by AFB smear test were detected in 63.19% of 269 specimens (n=170) and 40.15%(n=108) specimens were positive in mycobacterial culture .The differ-ence between the two tests were significant (P<0.01).The positive rate of Mycobacterium detected by using BACTEC MGIT 960 automated system and L?wenstein-Jensen culture method were 38 .66% ( n=104 ) and 28.99%(n=78), respectively, showing the significant difference between two tests (P<0.05).AFB smear test in combination with mycobacterial culture could precisely diagnose 70.63% of tuberculous lym-phadenitis in children.Of the 108 clinical isolates, 105 strains (97.2%) were Mycobacterium tuberculosis complex and the rest were non-tuberculous Mycobacterium strains (2.8%).Conclusion The positive rate by AFB smear test was significantly increased in fine needle aspiration biopsy specimens after a series of treatments including sample digestion , centrifugation and precipitation , but the positive rate of mycobacterial culture was reduced .Diagnostic accuracy could be significantly improved by using BACTEC MGIT 960 sys-tem.Mycobacterium tuberculosis complex was the predominant pathogenic bacterium in children with tubercu-lous lymphadenitis .

Antibacterial Activity of Medicinal Aqueous Plant Extracts against Mycobacterium tuberculosis

Tuberculosis (TB) remains a serious health problem in many regions of the world, and the development of resistance to antibiotics by this microbe created the need for new drugs to replace those which have lost effectiveness. This study assesses the medicinal anti-Mycobacterium tuberculosis properties of natural products obtained from plants collected from Eastern Libya. In this study aqueous extracts of nine different plants were assayed for their Mycobacterium tuberculosis inhibitory activity using the BACTEC MGIT960 susceptibility test method. The aqueous extracts of Ceratonia siliqua L, Helichrysum stoechas (L.) Moench and Thymus algeriensis did not show any activity against M.tuberculosis in different concentrations. The aqueous extract of Marrubium vulgare L. from Syria showed high activity against M. tuberculosis. Marrubium alysson L., Marrubium vulgare L., Pistacia lentiscus L, Quercus coccifera L, Thymus capitatus (L.) Hoffm. & Link, showed varying degrees of activity against M. tuberculosis. The results of this study show that aqueous extracts from six different medicinal plants have different effects against M. tuberculosis in vitro.

Comparison of MGIT 960 & pyrazinamidase activity assay for pyrazinamide susceptibility testing of Mycobacterium tuberculosis.

Background & objectives: Pyrazinamide is an important front line antimycobacterial drug, which is also being used in the treatment of multi drug resistant tuberculosis along with second line drugs in DOTS plus programme. Conventional testing of pyrazinamide on solid medium is difficult as it is active at acidic pH. Therefore, there is a need for a rapid and simple method for susceptibility testing of pyrazinamide. This study was carried out to compare pyrazinamide susceptibility testing by MGIT 960 and two rapid pyrazinamidase activity tests. Methods: Pyrazinamide susceptibility was tested in 136 clinical isolates of Mycobacterium tuberculosis by MGIT 960 and pyrazinamidase activity was tested by classical Wayne’s method and modified PZase agar method. Results: There was 88.9 per cent concordance between MGIT 960 and classical Wayne’s method and 93.38 per cent with modified method for pyrazinamidase activity. Using MGIT 960 results as gold standard the sensitivity and specificity of Wayne’s method was 88.15 and 90 per cent respectively and that of modified method was 89.4 and 98.3 per cent. Interpretation & conclusions: Our study demonstrates that the modified pyrazinamidase activity test can be used as a screening test to detect resistance to pyrazinamide specially in resource limited settings but confirmation of susceptibility should be done by standard methods like MGIT 960.

Evaluation of the bactec MGIT 960 TB system for recovery and identification of Mycobacterium tuberculosis complex in a high volume 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 r-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.

Evaluation of Combination of BACTEC Mycobacteria Growth Indicator Tube 960 System and Ogawa Media for Mycobacterial Culture / 대한진단검사의학회지

BACKGROUND: The combined use of liquid media and solid media is recommended for mycobacterial culture. We evaluated diagnostic performance of combination of BACTEC Mycobacteria Growth Indicator Tube (MGIT; Becton Dickinson, USA) and 2% Ogawa media (Korean Institute of Tuberculosis, Korea) for recovery of mycobacteria. METHODS: In September 2007, 1,764 specimens from 1,059 patients were cultured with MGIT and Ogawa. Acid fast bacilli (AFB) smear was fluorochrome-stained. The isolates were identified into Mycobacterium tuberculosis (MTB) and nontuberculous mycobacteria (NTM) with PCR using Seeplex TB Detection Kit (Seegene, Korea). Recovery rate, time to detection (TTD), contamination rate, mixed growth rate and species distribution were analyzed. RESULTS: Two hundred thirty-five specimens (13.3%) from 165 patients (15.6%) were positive for mycobacterial culture. Recovery rates of mycobacteria from the group using both media, MGIT only, and Ogawa only were 13.3%, 12.1%, and 7.8%, respectively. While MGIT recovered 98.9% of MTB and 79.7% of NTM, Ogawa recovered 65.9% of MTB and 54.1% of NTM. TTDs of total mycobacteria/MTB/NTM in MGIT and Ogawa were 10.6/11.4/9.7 days and 31/29/33 days, respectively. MGIT TTDs of total mycobacteria/MTB/NTM from AFB-positive specimens were significantly shorter than those of AFB-negative specimens; 8.2/9.5/4.4 days vs 11.6/12.7/10.7 days. Contamination and mixed growth rate of MGIT were 9.6% and 3.7%. Primary culture of Ogawa recovered 1 MTB and 1 NTM among the 170 MGIT-contaminated specimens and 38 mycobacteria among 66 specimens that showed mixed cultures of MGIT. CONCLUSIONS: MGIT warrants sensitive and rapid isolation of mycobacteria. However, the combination of MGIT and Ogawa is more desirable to recover mycobacteria in the case of contaminations or mixed cultures.

Evaluation of MGIT 960 System for Recovery of Mycobacteria from Body Fluids / 대한임상미생물학회지

BACKGROUND: In this study, we evaluated the BACTEC MGIT 960 system (Becton Dickinson Microbiology Systems, Sparks, Md, USA), which is fully automated, noninvasive and nonradiometric fluorescent indicator broth detection system, for the growth and detection of mycobacteria with body fluid specimens. METHODS: Total of 1,891 body fluid specimens were included (pleural fluid 752, ascitic fluid 629, cerebrospinal fluid 214, joint fluid 79, peritozol 54, others 163). Specimens were inoculated into MGIT and solid media (3% ogawa, Japan). Polymerase chain reaction was performed for the discrimination of Mycobacterium tuberculosis from Mycobacterium other than tuberculosis (MOTT). RESULTS: A total of 62 isolates of mycobacteria were recovered from all culture system. With MGIT system, 56 isolates were recovered, compared with solid system recovered 33 isolates. 29 isolates were recovered with MGIT only and 6 isolates recovered with solid media only. Among 62 isolates recovered, 11 isolates were positive in acid fast stain. 10 isolates were recovered with MGIT. One isolate was recovered with solid system. 51 isolates were negative in acid fast stain. Among this, 46 isolates were recovered with MGIT. The mean detection time was 14.2 days with MGIT system, and 38.2 days with solid media. Contamination rate for each system with body fluid specimens were 4.1% for MGIT and 1.7% for solid media. CONCLUSION: In body fluid, the MGIT system has the advantages of improved detection rate and rapid recovery than solid media to recover mycobacteria.

Evaluation of the BACTEC MGIT 960 system for the recovery of mycobacteria / 대한임상병리학회지

BACKGROUND: We evaluated the BACTEC MGIT 960 system(Becton Dickinson Microbiology Systems, Sparks, Md, USA), which is a fully automated, noninvasive and nonradiometric fluorescent indicator broth detection system for the growth and detection of mycobacteria with a capacity to incubate and continuously monitor 7 ml MGIT(Mycobacterium growth indicator tube) culture tube. METHODS: We studied 1,690 specimens(1,258 respiratory and 432 non-respiratory specimens). Processed specimens with 2% NaOH-NALC(final NaOH concentration: 1%) were inoculated into MGIT and solid media(3% Ogawa, Japan). Polymerase chain reaction was performed for the discrimination of Mycobacterium tuberculosis complex from Mycobacterium other than tuberculosis(MOTT). RESULTS: From all culture system, a total of 181 isolates of mycobacteria were recovered. The greatest number of isolates of mycobacteria was recovered with MGIT 960 system(174, 96.1%), followed by solid media(73, 40.3%). 108 isolates(59.7%) were recovered with MGIT only and 7(3.9%) recovered with solid media only( P value < 0.0001). From a total of 1,617 AFB smear negative specimens, 101(6.3%) were recovered with MGIT 960 system and 34(2.1%) recovered with solid media. The mean times to detection were average 11.1 days for MGIT 960 system and 31.6 days for solid media(P value < 0.0001). Contamination rates for each system were 14.9% for MGIT 960 and 2.6% for solid media. CONCLUSIONS: The newly introduced MGIT 960 system is easy to use and has advantages of high detection rate and rapid recovery than solid media, but there are some problems such as high contamination rate and high cost in the management of this liquid system.

Detection of Mycobacterium tuberculosis using BACTEC Mycobacteria Growth Indicator Tube(MGIT) 960 system: Comparison with BACTEC 460 TB system and Ogawa Media / 대한임상병리학회지

BACKGROUND: BACTEC MGIT 960 system(Becton Dickinson, USA; MGIT 960) is a fully automated, noninvasive culture system for mycobacteria, which has been regarded as a sensitive and least labor-intensive method. This study was purposed to evaluate the performance of MGIT 960 compared to BACTEC 460 TB radiometric system(Becton Dickinson, USA; BACTEC 460) and Ogawa media. METHODS: A total of 1,067 clinical specimens submitted from April to June in 1999 was cultured for acid fast bacilli(AFB). All specimens were digested, decontaminated by the 6% sodium hydroxide(final concentration of 1.5%) and 0.5% N-acetyl-L-cysteine method. All specimens were inoculated into three kinds of media: a MGIT, a BACTEC 12B, and an Ogawa medium. The AFB recovered from cultures were identified to M. tuberculosis complex and MOTT by NAP test. RESULTS: Of 106 isolates of M. tuberculosis recovered from all culture systems, 101(95.3 %) were detected in the MGIT 960, 95(89.6%) in the BACTEC 460 and 76(71.7%) on Ogawa media. MGIT 960 plus Ogawa media detected 104(98.1%) isolates and BACTEC 460 plus Ogawa media recovered 96(90.6%) isolates. The mean time required for detection of M. tuberculosis was 12.7+/-5.8 days with MGIT 960, 16.2+/-7.7 days with BACTEC 460, and 22.8+/-9.5 days with Ogawa media. The contamination rate were 5.1% for MGIT 960, 2.7% for BACTEC 460, and 6.7% for Ogawa media. CONCLUSIONS: MGIT 960 is a sensitive and rapid method to isolate M. tuberculosis.