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This study aimed to assess the possible causes of discordant results between Xpert MTB/RIF (Xpert) and Bactec MGIT 960 Culture System (MGIT960) regarding rifampicin (RIF) susceptibility in Mycobacterium tuberculosis. Patients with previous RIF-resistant tuberculosis who were admitted to Wenzhou Central Hospital from January 2020 to December 2022 were enrolled. The isolates obtained from these patients were subjected to RIF susceptibility tests using Xpert and MGIT960, and the minimum inhibitory concentration (MIC) of RIF was determined by the MYCOTB MIC plate test. Additionally, molecular docking and molecular dynamics (MD) simulations were performed to evaluate the binding efficacy of rpoB and RIF based on rpoB mutations detected in the isolates with discordant RIF susceptibility results. A total of 28 isolates with discordant RIF susceptibility test results were detected, 15 of them were RIF susceptible with MICs ≤ 0.5 µg/mL. Twelve out of 15 isolates contained borderline RIF resistance-associated mutations [L430P (n = 6), H445N (n = 6)], 1 isolate had D435Y and Q429H double mutation, and the remaining 2 isolates had a silent (Q432Q) mutation. Compared with the affinity of RIF toward the wild type (WT) (-45.83 kcal/mol) by MD, its affinity toward L452P (-55.52 kcal/mol), D435Y (-47.39 kcal/mol), L430P (approximately -69.72 kcal/mol), H445N (-49.53 kcal/mol), and Q429H (-55.67 kcal/mol) increased. Borderline RIF resistance-associated mutations were the main cause for the discordant RIF susceptibility results between Xpert and MGIT960, and the mechanisms of the resistance need further investigated.IMPORTANCEThis study is aimed at assessing discordant results between Xpert MTB/RIF (Xpert) assay and Bactec MGIT 960 Culture System (MGIT960) regarding the detection of rifampicin (RIF)-resistant Mycobacterium tuberculosis isolates in Wenzhou, China. The discordant results of RIF between these two assays were mainly caused by borderline RIF resistance-associated mutations, subsequently by silent mutations of rpoB. Borderline RIF resistance- associated mutations detected in our study were demonstrated to not be affected by the affinity of rpoB and RIF by molecular dynamics, and the mechanism of resistance was needed to be clarified. For the discordant results of RIF by Xpert and MGIT960 that occurred, rpoB DNA sequencing was recommended to investigate its association with resistance to RIF.
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Proteínas de Bactérias , Testes de Sensibilidade Microbiana , Mutação , Mycobacterium tuberculosis , Rifampina , Tuberculose Resistente a Múltiplos Medicamentos , Mycobacterium tuberculosis/efeitos dos fármacos , Mycobacterium tuberculosis/genética , Mycobacterium tuberculosis/isolamento & purificação , Rifampina/farmacologia , Humanos , China , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Tuberculose Resistente a Múltiplos Medicamentos/microbiologia , Antituberculosos/farmacologia , RNA Polimerases Dirigidas por DNA/genética , Farmacorresistência Bacteriana/genética , Simulação de Acoplamento MolecularRESUMO
Nocardia spp. is an aerobic Gram-positive bacillus responsible for nocardiosis. Herein, we performed a retrospective study to evaluate the performance of BACTEC MGIT 960 system, in comparison with smear microscopy and blood agar plate (BAP) culture, to recover Nocardia from different clinical specimens. Furthermore, the inhibitory effect of antibiotics contained in MGIT 960 tube on Nocardia was also evaluated. The sensitivities for Nocardia recovery using smear microscopy, BAP culture, and MGIT 960 were 39.4% (54/137), 46.1% (99/215), and 81.3% (156/192), respectively. N. farcinica was the most detected species (60.4%, 136/225). In MGIT 960-recovered Nocardia strains, N. farcinica accounted for 76.9%. Furthermore, trimethoprim in MGIT 960 tube inhibited less N. farcinica growth than that of other Nocardia species, partially explaining why MGIT 960 recovered more N. farcinica from sputa. The current study demonstrated that MGIT 960 could recover Nocardia strains from heavily-contaminated samples if its components and antibiotics are redesigned.
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Nocardiose , Nocardia , Humanos , Técnicas Bacteriológicas , Nocardia/genética , Estudos Retrospectivos , Meios de Cultura , Ágar , Antibacterianos/farmacologia , Nocardiose/diagnóstico , Nocardiose/tratamento farmacológicoRESUMO
The fast and accurate detection of susceptibility in drugs is a major challenge for a successful tuberculosis (TB) control programme. This study evaluated the performance of WHO-endorsed rapid diagnostic tools, such as BACTEC MGIT 960 SIRE (MGIT SIRE), GenoType MTBDRplus (MTBDRplus) and Xpert MTB/RIF (Xpert), for detecting susceptibility to first-line anti-TB drugs among pulmonary TB patients in Bangladesh. A total of 825 sputum samples with results from drug susceptibility testing (DST) against first-line anti-TB drugs in the MGIT SIRE, MTBDRplus and Xpert assays were evaluated and compared with the gold standard proportion susceptibility method of the Lowenstein-Jensen (LJ) medium. The overall sensitivities of MGIT SIRE were 97.6%, 90.0%, 61.3% and 44.9%, while specificities were 89.9%, 94.5%, 91.3% and 92.2% for detection of susceptibility to isoniazid (INH), rifampicin (RIF), streptomycin (STR) and ethambutol (EMB), respectively. For MTBDRplus, the sensitivities were 88.0% and 88.7%, and the specificities were 97.4% and 97.8% for the detection of susceptibility to INH and RIF, respectively. Xpert demonstrated a sensitivity and specificity of 94.8% and 99.5%, respectively, for the detection of RIF susceptibility. All tests performed significantly better in retreated TB patients compared with primary TB cases. For detection of RIF and INH susceptibility, all three assays showed almost perfect agreement with the LJ method, although MGIT SIRE exhibited low agreement for STR and EMB. Considering the high performance, shorter turnaround time and ease of use, molecular-based approaches Xpert and MTBDRplus can be widely implemented throughout the country for the rapid detection of drug-resistant TB.
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Heteroresistance is defined as the coexistence of both susceptible and resistant bacteria in a bacterial population. Previously published data show that it may occur in 9 to 57% of Mycobacterium tuberculosis isolates for various drugs. Pyrazinamide (PZA) is an important first-line drug used for treatment of both drug-susceptible and PZA-susceptible multidrug-resistant TB. Clinical PZA resistance is defined as a proportion of resistant bacteria in the isolate exceeding 10%, when the drug is no longer considered clinically effective. The ability of traditional drug susceptibility testing techniques to detect PZA heteroresistance has not yet been evaluated. The aim of this study was to compare the capacity of Bactec MGIT 960, Wayne's test, and whole-genome sequencing (WGS) to detect PZA-resistant subpopulations in bacterial suspensions prepared with different proportions of mutant strains. Both Bactec MGIT 960 and WGS were able to detect the critical level of 10% PZA heteroresistance, whereas Wayne's test failed to do so, with the latter falsely reporting highly resistant samples as PZA susceptible. Failure to detect drug-resistant subpopulations may lead to inadvertently weak treatment regimens if ineffective drugs are included, with the risk of treatment failure with the selective growth of resistant subpopulations. We need clinical awareness of heteroresistance as well as evaluation of new diagnostic tools for their capacity to detect heteroresistance in TB.
Assuntos
Mycobacterium tuberculosis , Tuberculose Resistente a Múltiplos Medicamentos , Amidoidrolases/genética , Antituberculosos/farmacologia , Antituberculosos/uso terapêutico , Farmacorresistência Bacteriana/genética , Humanos , Testes de Sensibilidade Microbiana , Mutação , Mycobacterium tuberculosis/genética , Pirazinamida/farmacologia , Tuberculose Resistente a Múltiplos Medicamentos/tratamento farmacológicoRESUMO
BACKGROUND: Tuberculosis is a devastating and a deadly disease despite the novel advances in its diagnostic tools and drug therapy. Drug resistant Mycobacterium contributes a great share to tuberculosis mortality. Status of drug resistance and patients' awareness toward the disease is unknown in northeastern Ethiopia. Thus, the aim of this study was to determine the phenotypic and genotypic drug sensitivity patterns and associated factors in Oromia Special Zone and Dessie Town, northeastern Ethiopia. METHODS: In a cross-sectional study, 384 smear positive tuberculosis cases were recruited and Löwenstein-Jensen culture was done. The performance of GenoTypic MTBDRplus assay using the conventional BACTEC MGIT 960 as a "gold standard" was determined. Drug resistant strains were identified using spoligotyping. Pearson Chi-square test was used to determine the association of drug sensitivity test and tuberculosis type, lineages, dominant strains and clustering of the isolates. RESULTS: The 384 smear positive Mycobacterium samples were cultured on LJ media of which 29.2% (112/384) as culture positive. A fair agreement was found between MTBDRplus assay and the conventional MGIT test in detecting the Mycobacterium tuberculosis with sensitivity, specificity, positive and negative predictive value of 94.2, 30.2, 68.4 and 76.5%, respectively. Among LJ culture positive samples 95 of them gave valid result for MTBDRplus assay and 16.8% (16/95) as drug resistant. Similarly, MGIT subculture was made for the 112 isolates and 69 of them gave positive result with 15.9% (11/69) as drug resistant. Cohen's kappa value showed almost a perfect agreement between the two testing methods in detecting rifampicin (sensitivity 100% and specificity 98.3%) and multi-drug resistance (sensitivity 83.3% and specificity 100%). Spoligotyping identified 76.5% (13/17) of the drug resistant isolates as Euro-American and family 33 as the predominant family. Significant association was observed between drug resistant isolates and the dominant strains (χ2: 34.861; p = 0.040) of the Mycobacterium. CONCLUSION: Higher magnitude of drug resistance was found in the study area. The GenoTypic MDRTBplus assay had an acceptable drug sensitivity testing performance.
Assuntos
Mycobacterium tuberculosis/genética , Tuberculose/diagnóstico , Adolescente , Adulto , Idoso , Antituberculosos/farmacologia , Antituberculosos/uso terapêutico , Estudos Transversais , Farmacorresistência Bacteriana/efeitos dos fármacos , Farmacorresistência Bacteriana/genética , Etiópia , Feminino , Genótipo , Humanos , Isoniazida/farmacologia , Isoniazida/uso terapêutico , Masculino , Testes de Sensibilidade Microbiana , Pessoa de Meia-Idade , Mycobacterium tuberculosis/isolamento & purificação , Fenótipo , Kit de Reagentes para Diagnóstico , Rifampina/farmacologia , Rifampina/uso terapêutico , Tuberculose/tratamento farmacológico , Adulto JovemRESUMO
BACKGROUND: Since the implementation of the Xpert MTB/RIF in Sao Paulo, Brazil, numerous Mycobacterium tuberculosis isolates presenting "rifampicin-resistant genotype with rifampicin-susceptible phenotype" were observed. OBJECTIVE: To evaluate the prevalence, rpoB mutations and transmission of M. tuberculosis resistant to rifampicin on Xpert MTB/RIF but susceptible on BACTEC MGIT system, in Sao Paulo state. METHODS: Patients' isolates with this pattern of rifampicin discordance, collected from 2014 to 2017, had their rpoB predominant rifampicin-resistance-determining region sequenced and were genotyped by IS6110 restriction fragment-length polymorphism. FINDINGS: The prevalence of rifampicin-discordant M. tuberculosis with genotypic resistance was 55.1% (156/283). Among the sequenced and genotyped isolates, 75.5% (111/147) were in clusters, largely associated with the type of rpoB mutation. Most isolates (98.6%; 72/73) harbouring the predominant mutation, His445Asn, were pooled into the two largest clusters, SP2ga (42/72; 58.3%) and SP5o (12/72; 16.7%). Ranking second, isolates carrying the silent mutation Phe433Phe were mostly (92.3%; 24/26) gathered into four groups of the family SP25. CONCLUSION: These findings suggest that this unusual high rifampicin discrepancy proportion was greatly influenced by few actively circulating clusters. Further studies on many of the rpoB mutations identified in our setting are needed to elucidate their association with phenotypic rifampicin resistance.
Assuntos
Proteínas de Bactérias/genética , Farmacorresistência Bacteriana/genética , Epidemias/estatística & dados numéricos , Mutação , Mycobacterium tuberculosis/genética , Rifampina/farmacologia , Tuberculose Resistente a Múltiplos Medicamentos/transmissão , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Antibióticos Antituberculose/farmacologia , Brasil/epidemiologia , Estudos Transversais , RNA Polimerases Dirigidas por DNA/genética , Feminino , Genótipo , Humanos , Masculino , Pessoa de Meia-Idade , Mycobacterium tuberculosis/efeitos dos fármacos , Mycobacterium tuberculosis/isolamento & purificação , Fenótipo , Tuberculose Resistente a Múltiplos Medicamentos/tratamento farmacológico , Tuberculose Resistente a Múltiplos Medicamentos/microbiologia , Adulto JovemRESUMO
The results of the comparative testing of the susceptibility of M. tuberculosis clinical strains to isoniazid, streptomycin, rifampicin and ethambutol using the TB test kit, developed in SCRAMB, (Obolensk) and the absolute concentrations method; the TB test kit and the BACTEC MGIT 960 automated system are presented in the study. A total of 629 and 220 strains, respectively, were tested. A high degree of agreement of the results was shown: 89.1-98.6% for isoniazid, 96.2-98.0% for rifampicin, 91.5-98.2% for streptomycin and 89.1-95.9% for ethambutol. The smallest number of discrepancies in the results was obtained when comparing the TB test kit and BACTEC MGIT 960. The discrepant results analysis was performed by the proportion method, PCR sequencing, or re-testing on new lots of the TB test kit and Lowenstein-Jensen medium with anti-tuberculosis drugs, after which the sensitivity, the specificity and the efficiency of the TB test kit have exceeded 95 % for all anti-tuberculosis drugs. The turnaround time with the TB test kit (median 9.25-9.9 days, ranged from 8 to13 days) was significantly shorter than that with the absolute concentration method (median 21-23 days, ranged from 20 to 28 days) and is commensurate with the turnaround time with BACTEC MGIT 960 (average 7.2 days, ranged from 5 to 12 days). The TB test kit is easy to use, does not require expensive equipment and special staff training.
Assuntos
Antituberculosos/farmacologia , Testes de Sensibilidade Microbiana , Mycobacterium tuberculosis/efeitos dos fármacos , Kit de Reagentes para Diagnóstico , Etambutol , Humanos , Isoniazida , Rifampina , EstreptomicinaRESUMO
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.
Assuntos
Farmacorresistência Bacteriana , Pneumoconiose/complicações , Pneumoconiose/microbiologia , Tuberculose/diagnóstico , Humanos , Mycobacterium tuberculosis , Rifampina , Sensibilidade e Especificidade , EscarroRESUMO
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.
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We evaluated the microscopic and macroscopic characteristics of mycobacteria growth indicator tube (MGIT) cultures for the presumptive identification of the Mycobacterium tuberculosis complex (MTBC) and assessed the reliability of this strategy for correctly directing isolates to drug susceptibility testing (DST) or species identification. A total of 1526 isolates of mycobacteria received at the Instituto Adolfo Lutz were prospectively subjected to presumptive identification by the observation of growth characteristics along with cord formation detection via microscopy. The presumptive identification showed a sensitivity, specificity and accuracy of 98.8, 92.5 and 97.9â%, respectively. Macroscopic analysis of MTBC isolates that would have been erroneously classified as non-tuberculous mycobacteria based solely on microscopic morphology enabled us to direct them rapidly to DST, representing a substantial gain to patients. In conclusion, the growth characteristics of mycobacteria in MGIT, when considered along with cord formation, increased the reliability of the presumptive identification, which has a great impact on the laboratory budget and turnaround times.
Assuntos
Técnicas de Cultura de Células/métodos , Infecções por Mycobacterium/diagnóstico , Mycobacterium tuberculosis/classificação , Micobactérias não Tuberculosas/classificação , Antígenos de Bactérias/genética , Antígenos de Bactérias/metabolismo , Antituberculosos/farmacologia , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Chaperonina 60/genética , Chaperonina 60/metabolismo , Imunoensaio , Testes de Sensibilidade Microbiana , Infecções por Mycobacterium/tratamento farmacológico , Infecções por Mycobacterium/microbiologia , Mycobacterium tuberculosis/efeitos dos fármacos , Mycobacterium tuberculosis/crescimento & desenvolvimento , Mycobacterium tuberculosis/isolamento & purificação , Micobactérias não Tuberculosas/efeitos dos fármacos , Micobactérias não Tuberculosas/crescimento & desenvolvimento , Micobactérias não Tuberculosas/isolamento & purificação , Estudos Prospectivos , Reprodutibilidade dos Testes , Sensibilidade e Especificidade , Especificidade da EspécieRESUMO
BACKGROUND: Several nucleic acid amplification techniques (IS6110, 16S rRNA, and 85B mRNA) were developed for the rapid, direct detection of Mycobacterium tuberculosis. We aimed to assess the diagnostic performance of 85B mRNA-based RT-qPCR by comparing with the real-time PCR COBAS TaqMan MTB Kit while using the BACTEC MGIT 960 method as the gold standard. METHODS: 60 patients with confirmed pulmonary TB and 60 individuals without TB were included as the study and control groups, respectively. Sputum specimens were cultured using LJ and BACTEC MGIT 960 systems. Extracted DNA was used for COBAS PCR in a CONAS TaqMan 48 analyzer. 85B mRNA detection was performed by RT-qPCR. RESULTS: The sensitivity, specificity, positive predictive value, negative predictive value, and accuracy of COBAS TaqMan MTB Test were detected as 93.3%, 83.3%, 84.8%, 92.6%, and 88.3%, respectively. The same diagnostic parameters of RT-qPCR were: 98.3%, 95.0%, 95.2%, 98.3%, and 96.7%, respectively. According to the binary logistic regression analysis, RT-qPCR (OR: 19,924, p<0.001) was identified as the more optimal test. CONCLUSION: RT-qPCR targeting the 85B gene of M. tuberculosis seems to be a more useful and rapid technique than DNA-based methods for detecting live M. tuberculosis bacilli from sputum specimens.
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Aciltransferases/genética , Antígenos de Bactérias/genética , Proteínas de Bactérias/genética , Técnicas de Diagnóstico Molecular/métodos , RNA Mensageiro/análise , Reação em Cadeia da Polimerase em Tempo Real/métodos , Escarro/microbiologia , Tuberculose Pulmonar/diagnóstico , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Valor Preditivo dos Testes , RNA Mensageiro/genética , Sensibilidade e Especificidade , Adulto JovemRESUMO
Pyrazinamide forms a core part of treatment for all types of tuberculosis (TB) in Zambia. Due to challenges associated with pyrazinamide testing, little information is available to indicate the frequency of resistance to this drug in Zambia. To determine the frequency of pyrazinamide (PZA) resistance and its correlation with mutation in pncA in Mycobacterium tuberculosis isolated from patients in Lusaka, Zambia, BACTEC MGIT M960 was used for phenotypic PZA susceptibility testing while sequencing was used to determine resistance-conferring mutations in the pncA. Of the 131 isolates analyzed, 32 were phenotypically resistant to PZA. Among multidrug-resistant (MDR) M. tuberculosis isolates, the frequency of PZA resistance was 21 of 35 (58.3%). And 27 of 32 PZA resistant isolates had mutations in the pncA that seem to confer resistance. With BACTEC MGIT 960 as the reference standard, gene sequencing showed 84.4% sensitivity and 100% specificity. Nine new mutations were identified and the single nucleotide substitution T104G and C195T were the most frequent mutations. However, they were observed in both susceptible and resistant strains and indicating that they are non-resistance conferring mutations. This study has demonstrated that PZA susceptibility testing is necessary especially in patients suffering from MDR-TB as approximately half of the patients have PZA resistant TB. Similar studies will have to be carried out in other provinces to get an accurate estimate of PZA resistance in Zambia. Mutations in pncA were the major mechanism of PZA resistance with no involvement of rpsA and panD genes. However, the presence of mutations among phenotypically PZA susceptible M. tuberculosis isolates makes it challenging to independently use genotyping method for the determination of PZA resistance.
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Amidoidrolases/genética , Antituberculosos/uso terapêutico , Farmacorresistência Bacteriana Múltipla/genética , Mutação , Mycobacterium tuberculosis/genética , Pirazinamida/uso terapêutico , Tuberculose Resistente a Múltiplos Medicamentos/microbiologia , Tuberculose Pulmonar/microbiologia , Análise Mutacional de DNA , Genótipo , Humanos , Testes de Sensibilidade Microbiana , Taxa de Mutação , Mycobacterium tuberculosis/efeitos dos fármacos , Mycobacterium tuberculosis/isolamento & purificação , Fenótipo , Tuberculose Resistente a Múltiplos Medicamentos/diagnóstico , Tuberculose Resistente a Múltiplos Medicamentos/tratamento farmacológico , Tuberculose Pulmonar/diagnóstico , Tuberculose Pulmonar/tratamento farmacológico , ZâmbiaAssuntos
Antituberculosos/farmacologia , Erros de Diagnóstico/prevenção & controle , Farmacorresistência Bacteriana/efeitos dos fármacos , Testes de Sensibilidade Microbiana/métodos , Mycobacterium tuberculosis/efeitos dos fármacos , Pirazinamida/farmacologia , Amidoidrolases/genética , Farmacorresistência Bacteriana/genética , Humanos , Mutação , Mycobacterium tuberculosis/classificação , Mycobacterium tuberculosis/genética , Tuberculose/diagnóstico , Tuberculose/microbiologiaRESUMO
Drug resistance in tuberculosis is a major threat to public health and control of the disease worldwide. Given the need of a rapid and accurate detection of Mycobacterium tuberculosis resistance to second-line drugs, this study evaluated the performance of the BACTEC MGIT 960 for second-line, drug susceptibility testing in comparison with the resazurin microtiter assay (REMA), in order to implement the automated methodology in the diagnostic routine of a reference laboratory. Drug susceptibility testing (DST) for second-line drugs of 151 MDR M. tuberculosis clinical isolates was performed by both BACTEC MGIT 960 and REMA, and a panel of 26 M. tuberculosis reference isolates from a proficiency test was tested by the BACTEC MGIT 960. DST for second-line drugs by the BACTEC MGIT 960 system was more rapid, highly reproducible and showed 100% of proficiency. After these results, this methodology was successfully implemented in our diagnostic routine for all MDR-TB patients.
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Antituberculosos/farmacologia , Testes de Sensibilidade Microbiana/métodos , Mycobacterium tuberculosis/efeitos dos fármacos , Oxazinas/metabolismo , Xantenos/metabolismo , Automação Laboratorial , Humanos , Testes de Sensibilidade Microbiana/instrumentação , Tuberculose Resistente a Múltiplos Medicamentos/microbiologiaRESUMO
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.
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Técnicas Bacteriológicas/métodos , Espectroscopia Dielétrica , Mycobacterium/crescimento & desenvolvimento , Mycobacterium/isolamento & purificação , Meios de Cultura , Humanos , Mycobacterium/classificação , Reprodutibilidade dos Testes , Fatores de TempoRESUMO
BACKGROUND: The pattern of Mycobacterium tuberculosis susceptibility to first line drugs and multidrug resistance in Al-Madinah Al-Munawarah, a seasonally overcrowded are during Hajj and Omrah, is not well studied. OBJECTIVE: This study aimed to investigate anti-tuberculosis drug resistance and its distribution among new cases in Al-Madinah Al-Monawarah. METHODS: Study subjects included 622 patients with first time confirmed TB referred to the central tuberculosis laboratory in Al-Madinah between January 2012 and December 2014. RESULTS: Out of the 622 isolates, 99 (15.9%) were Mycobacteria Other Than Tuberculosis (MOTTS) and 25 (4.0%), three of which (12%) were children under five years of age, revealed multidrug resistance (MDR). Monoresistance to isoniazid (H) was (1.8%), to rifampin (R) was (1.4%), to streptomycin (S) was (1.9 %) to ethambutol (E) was (1.1 %) and to pyrazinamide (Z) was (2.1%). CONCLUSION: Being among the new cases, multidrug resistant tuberculosis (MDR TB) is supposed to be caused by strains which are originally multidrug resistant. Neither nationality nor gender was found to be associated with MDR TB. Since 12% of MDR cases were among children, a probability of primary infection with MDR strains is to be considered. Moreover, mass gathering during Hajj and Omrah seasons does not seem to increase the burden of MDR in the region. However, further investigation is needed to molecularly characterize MDR isolates and their phylogenetics and geographical origin.
Assuntos
Efeitos Psicossociais da Doença , Mycobacterium tuberculosis/isolamento & purificação , Tuberculose Resistente a Múltiplos Medicamentos/epidemiologia , Tuberculose Resistente a Múltiplos Medicamentos/microbiologia , Adolescente , Adulto , Antituberculosos/farmacologia , Antituberculosos/uso terapêutico , Criança , Feminino , Humanos , Islamismo , Isoniazida/farmacologia , Isoniazida/uso terapêutico , Masculino , Testes de Sensibilidade Microbiana , Mycobacterium tuberculosis/efeitos dos fármacos , Mycobacterium tuberculosis/genética , Filogenia , Rifampina/farmacologia , Rifampina/uso terapêutico , Arábia Saudita/epidemiologia , Estreptomicina/farmacologia , Estreptomicina/uso terapêutico , Viagem , Tuberculose Resistente a Múltiplos Medicamentos/diagnóstico , Tuberculose Resistente a Múltiplos Medicamentos/tratamento farmacológico , Adulto JovemRESUMO
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.
Assuntos
Humanos , Técnicas Bacteriológicas/métodos , Espectroscopia Dielétrica , Mycobacterium/isolamento & purificação , Mycobacterium/crescimento & desenvolvimento , Fatores de Tempo , Reprodutibilidade dos Testes , Meios de Cultura , Mycobacterium/classificaçãoRESUMO
Objective A retrospective study comparing the diagnostic performance of the BACTEC™ MGIT™ 960 system (M960 system; BD Worldwide, Franklin Lakes, NJ, USA) with Löwenstein-Jensen (L-J) culture to detect mycobacteria in sputum specimens from patients with suspected pulmonary tuberculosis (TB). Methods Sputum samples were analysed for the presence of acid-fast bacilli (AFB). Samples were inoculated into the M960 system and L-J culture. Positive cultures were examined for the presence of AFB. Results The M960 method detected significantly more positive samples than L-J culture (818/1676 [48.8%] vs 692/1676 [41.3%]). Using L-J culture as reference, the sensitivity, specificity, positive predictive and negative predictive values of the M960 system were 91.0%, 76.1%, 77.0% and 92.2%, respectively. The time-to-detection of mycobacteria was 11.78 ± 5.16 days for M960 and 24.17 ± 8.73 days for L-J. Conclusions The M960 system had better diagnostic capability than L-J culture. Clinical value may be maximized by combining results from both methods.
Assuntos
Mycobacterium tuberculosis/isolamento & purificação , Escarro/microbiologia , Tuberculose Pulmonar/diagnóstico , China , Meios de Cultura , Humanos , Estudos Retrospectivos , Tuberculose Pulmonar/microbiologiaRESUMO
BACKGROUND/PURPOSE: The emergence of resistance to anti-tuberculosis (TB) drugs has become an obstacle to effective TB control. Thus, there is an urgent need to identify patients and initiate adequate treatment for drug-resistant cases in a timely manner. The BACTEC MGIT 960 system is well known for its rapid culturing time, and is in widespread use in Taiwan. In this study, we evaluated the possibility of replacing the traditional indirect agar proportion method with a modified direct agar proportion method (MDAPM), as a technique for rapid testing the drug susceptibility of Mycobacterium tuberculosis without additional cost. METHODS: In this study, 432 positive MGIT 960 samples that were identified as M. tuberculosis complex using the MeDiPro M. tuberculosis Antigen Rapid Test or the Cobas Amplicor MTB test were evaluated. Each sample was tested separately by the MDAPM and indirect agar proportion method, between July 2008 and December 2008, to compare the consistency and total turnaround time. RESULTS: Four first-line anti-TB drugs-rifampin, isoniazid, ethambutol, and streptomycin-were tested. For the MDAPM and indirect agar proportion method, the respective consistencies for each drug were 99.31%, 98.38%, 98.38%, and 97.22%. Our results also indicated that the MDAPM leads to an average saving in working time of 2 weeks, compared with the traditional indirect agar proportion method. CONCLUSION: In addition to having the potential to shorten turnaround time without compromising diagnostic quality, the MDAPM also provides a more efficient and cost-effective procedure. This modified procedure presents potential benefits for TB diagnosis in laboratories already equipped with the MGIT 960 system.
Assuntos
Antituberculosos/farmacologia , Meios de Cultura/química , Testes de Sensibilidade Microbiana/métodos , Mycobacterium tuberculosis/efeitos dos fármacos , Tuberculose/microbiologia , Análise Custo-Benefício , Humanos , Taiwan , Fatores de TempoRESUMO
Tubercular liver abscess is generally secondary to some other primary foci in the body, most notably pulmonary and gastrointestinal system. To find primary tubercular liver abscess is rare, with prevalence of 0.34% in patients with hepatic tuberculosis. Abscess tracking into abdominal wall from spinal and para spinal tuberculosis is known, however primary liver tuberculosis rupturing into anterior abdominal wall has been reported only twice in literature. We report a case of 43-year-old female with direct invasion of the anterior abdominal wall from an isolated tubercular parenchymal liver abscess, caused by Mycobacterium tuberculosis complex, diagnosed primarily on smear for Acid Fast Bacilli (AFB), imaging and isolated by culture and BACTEC MGIT 960 KIT. We discuss here the diagnostic dilemma, management and outcome of primary tubercular liver parenchymal abscess with direct invasion into anterior abdominal wall.