ABSTRACT
Mycobacterium abscessus is one of the most common and pathogenic nontuberculous mycobacteria (NTM) isolated in clinical laboratories. It consists of three subspecies: M. abscessus subsp. abscessus, M. abscessus subsp. bolletii, and M. abscessus subsp. massiliense. Due to their different antibiotic susceptibility pattern, a rapid and accurate identification method is necessary for their differentiation. Although matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) has proven useful for NTM identification, the differentiation of M. abscessus subspecies is challenging. In this study, a collection of 325 clinical isolates of M. abscessus was used for MALDI-TOF MS analysis and for the development of machine learning predictive models based on MALDI-TOF MS protein spectra. Overall, using a random forest model with several confidence criteria (samples by triplicate and similarity values >60%), a total of 96.5% of isolates were correctly identified at the subspecies level. Moreover, an improved model with Spanish isolates was able to identify 88.9% of strains collected in other countries. In addition, differences in culture media, colony morphology, and geographic origin of the strains were evaluated, showing that the latter had an impact on the protein spectra. Finally, after studying all protein peaks previously reported for this species, two novel peaks with potential for subspecies differentiation were found. Therefore, machine learning methodology has proven to be a promising approach for rapid and accurate identification of subspecies of M. abscessus using MALDI-TOF MS.
Subject(s)
Mycobacterium Infections, Nontuberculous , Mycobacterium abscessus , Mycobacterium , Humans , Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization/methods , Nontuberculous Mycobacteria , Mycobacterium Infections, Nontuberculous/diagnosis , Mycobacterium Infections, Nontuberculous/microbiologyABSTRACT
Diagnosis of latent tuberculosis infection (LTBI) is considered key in the control of tuberculosis. Interferon gamma (IFN-γ) release assays, such as the QuantiFERON-TB Gold Plus test (QFT-Plus), are now widely implemented for the in vitro diagnosis of LTBI. To date, the detection and quantification of IFN-γ has been mostly performed with semiautomated enzyme-linked immunosorbent assays (ELISAs), but several limitations currently exist. The study aims to evaluate the chemiluminescence immunoassay (CLIA) analyzer Liaison XL compared to ELISA for the performance of the QFT-Plus test. Between February and April 2020, 333 heparin blood samples from 323 adult patients were collected at a tertiary teaching hospital in Barcelona, Spain. Overall, the CLIA analyzer Liaison XL performed well for the detection of IFN-γ compared to the ELISA method, demonstrating substantial agreement (κ, 0.872) and great correlation between assays (r, >0.950). CLIA produced significantly higher values of IFN-γ IU per milliliter than the ELISA (P = 0.004 for the TB1 tube and P = 0.010 for the TB2 tube). Many discrepant cases (8/15, 53.3%) corresponded to indeterminate results with ELISA (NIL-corrected mitogen value of <0.5 IU/ml), which, when analyzed with the CLIA analyzer Liaison XL, reverted to interpretable results. In conclusion, this analysis suggests that CLIA presents a greater sensitivity for the identification of LTBI, especially among immunocompromised patients. Furthermore, the analytical variability reported between both ELISA and CLIA methods, especially around the standardized 0.35-IU/ml positivity threshold, suggests the need to refine the interpretative algorithm.
Subject(s)
Latent Tuberculosis , Mycobacterium tuberculosis , Tuberculosis , Adult , Humans , Incidence , Interferon-gamma Release Tests , Latent Tuberculosis/diagnosis , Luminescence , Spain , Tuberculin Test , Tuberculosis/diagnosisABSTRACT
Identification of mycobacteria by matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) requires not only a good protein extraction protocol but also an adequate cutoff score in order to provide reliable results. The aim of this study was to assess the cutoff scores proposed by the MALDI-TOF MS system for mycobacterial identification. A total of 693 clinical isolates from a liquid medium and 760 from a solid medium were analyzed, encompassing 67 different species of nontuberculous mycobacteria (NTM). MALDI-TOF MS identified 558 (80.5%) isolates from the liquid medium and 712 (93.7%) isolates from the solid medium with scores of ≥1.60. Among these, four (0.7%) misidentifications were obtained from the liquid medium and four (0.5%) from the solid medium. With regard to species diversity, MALDI-TOF MS successfully identified 64 (95.5%) different species, while PCR-reverse hybridization (GenoType Mycobacterium CM and AS assays) identified 24 (35.8%) different species. With MALDI-TOF MS scores of ≥2, all isolates were correctly identified, and with scores in the range from 1.60 to 1.99, most isolates were correctly identified, except for Mycobacterium angelicum, M. parascrofulaceum, M. peregrinum, M. porcinum, and M. gastri In conclusion, MALDI-TOF MS is a useful method for identifying a large diversity of NTM species. A score threshold of 1.60 proved useful for identifying almost all the isolates tested; only a few species required a higher score (≥2.00) to obtain a valid definitive identification.
Subject(s)
Mycobacterium Infections, Nontuberculous , Mycobacterium , Humans , Mycobacterium/genetics , Mycobacterium Infections, Nontuberculous/diagnosis , Nontuberculous Mycobacteria/genetics , Spectrometry, Mass, Matrix-Assisted Laser Desorption-IonizationABSTRACT
Tuberculosis (TB) remains a major health problem worldwide. Control of TB requires rapid, accurate diagnosis of active disease. However, extrapulmonary TB is very difficult to diagnose because the clinical specimens have very low bacterial loads. Several molecular methods involving direct detection of the Mycobacterium tuberculosis complex (MTBC) have emerged in recent years. Real-time PCR amplification simultaneously combines the amplification and detection of candidate sequences by using fluorescent probes (mainly TaqMan or Molecular Beacons) in automated systems. The multiplex real-time PCR-short assay is performed using locked nucleic acid (LNA) probes (length, 8 to 9 nucleotides) in combination with CodUNG to detect multiple pathogens in clinical samples. In this study, we evaluated the performance of this novel multiplex assay for detecting the MTBC in comparison with that of the conventional culture-based method. The multiplex real-time PCR-shortTUB assay targets two genes, whiB3 (redox-responsive transcriptional regulator) and pstS1 (phosphate-specific transporter), yielding limits of detection (LOD) of 10 copies and 100 copies, respectively, and amplification efficiencies of 92% and 99.7%, respectively. A total of 94 extrapulmonary samples and pulmonary samples with low mycobacterial loads (all smear negative; 75 MTBC culture positive) were analyzed using the test, yielding an overall sensitivity of 88% and a specificity of 95%. For pleural fluid and tissues/biopsy specimens, the sensitivity was 83% and 85%, respectively. In summary, this technique could be implemented in routine clinical microbiology testing to reduce the overall turnaround time for MTBC detection and may therefore be a useful tool for the diagnosis of extrapulmonary tuberculosis and diagnosis using pulmonary samples with low mycobacterial loads.
Subject(s)
Bacterial Load/methods , Lung/microbiology , Multiplex Polymerase Chain Reaction/standards , Mycobacterium tuberculosis/genetics , Tuberculosis/microbiology , ATP-Binding Cassette Transporters/genetics , Bacterial Proteins/genetics , Humans , Limit of Detection , Multiplex Polymerase Chain Reaction/methods , Mycobacterium tuberculosis/growth & development , Oligonucleotides/genetics , Pleural Effusion/microbiology , Sensitivity and SpecificityABSTRACT
Background: Screening strategies based on interferon-γ release assays in tuberculosis contact tracing may reduce the need for preventive therapy without increasing subsequent active disease. Methods: We conducted an open-label, randomized trial to test the noninferiority of a 2-step strategy with the tuberculin skin test (TST) followed by QuantiFERON-TB Gold In-Tube (QFT-GIT) as a confirmatory test (the TST/QFT arm) to the standard TST-alone strategy (TST arm) for targeting preventive therapy in household contacts of patients with tuberculosis. Participants were followed for 24 months after randomization. The primary endpoint was the development of tuberculosis, with a noninferiority margin of 1.5 percentage points. Results: A total of 871 contacts were randomized. Four contacts in the TST arm and 2 in the TST/QFT arm developed tuberculosis. In the modified intention-to-treat analysis, this accounted for 0.99% in the TST arm and 0.51% in the TST/QFT arm (-0.48% difference; 97.5% confidence interval [CI], -1.86% to 0.90%); in the per-protocol analysis, the corresponding rates were 1.67% and 0.82% in the TST and TST/QFT arms, respectively (-0.85% difference; 97.5% CI, -3.14% to 1.43%). Of the 792 contacts analyzed, 65.3% in the TST arm and 42.2% in the TST/QFT arm were diagnosed with tuberculosis infection (23.1% difference; 95% CI, 16.4% to 30.0%). Conclusions: In low-incidence settings, screening household contacts with the TST and using QFT-GIT as a confirmatory test is not inferior to TST-alone for preventing active tuberculosis, allowing a safe reduction of preventive treatments. Clinical Trials Registration: NCT01223534.
Subject(s)
Contact Tracing , Interferon-gamma Release Tests/standards , Latent Tuberculosis/diagnosis , Reagent Kits, Diagnostic/standards , Tuberculin Test/standards , Adult , Cost-Benefit Analysis , Family Characteristics , Female , Humans , Incidence , Male , Mass Screening/methods , Middle Aged , Preventive Health Services/methodsABSTRACT
In Spain, systematic reporting of pulmonary infections with nontuberculous mycobacteria is not mandatory. Therefore, to determine trends, we retrospectively identified cases for January 1994-December 2014 in Catalonia. Over the 21 years, prevalence increased and was associated with being male. Mycobacterium avium complex and M. abscessus prevalence increased; M. kansasii prevalence decreased.
Subject(s)
Mycobacterium Infections, Nontuberculous/epidemiology , Mycobacterium Infections, Nontuberculous/microbiology , Nontuberculous Mycobacteria , Tuberculosis, Pulmonary/epidemiology , Tuberculosis, Pulmonary/microbiology , Adolescent , Adult , Aged , Aged, 80 and over , Female , History, 20th Century , History, 21st Century , Humans , Male , Middle Aged , Mycobacterium Infections, Nontuberculous/history , Population Surveillance , Prevalence , Spain/epidemiology , Tuberculosis, Pulmonary/history , Young AdultABSTRACT
The rapid detection of Mycobacterium tuberculosis complex (MTUBC) in clinical samples is essential for successful treatment. New techniques such as real-time PCR have been developed in order to facilitate rapid diagnosis, but their sensitivity is low in extrapulmonary specimens, due to the low bacillary load in such samples. A next-generation assay has recently been developed to try to overcome this limitation. The aim of this study was to analyze the effectiveness of the Xpert MTB/RIF Ultra (GX-Ultra) for the detection of MTUBC DNA in 108 smear-negative extrapulmonary specimens that were MTUBC culture positive. In addition, 40 extrapulmonary culture-negative samples and 20 samples with nontuberculous mycobacteria were tested to evaluate the specificity of the assay. All samples were collected between May 1999 and May 2017. The GX-Ultra detected DNA of MTUBC in 82 extrapulmonary specimens that were MTUBC culture positive (75.9% sensitivity; 95% confidence interval [CI], 66.6 to 83.4%). The assay was negative for all clinical specimens that were MTUBC culture negative and the samples with nontuberculous mycobacteria (100% specificity). Furthermore, two (1.8%) samples presented mutations related to rifampin resistance. The highest sensitivity was obtained in samples of lymph nodes (94.1%) and nonsterile fluids (93.7%), followed by tissue specimens (86.6%), stool material (80%), abscess aspirates (64.7%), and sterile fluids (60.5%). Pleural fluids, one of the least optimal samples for detecting DNA of MTUBC, were GX-Ultra positive in 10/21 (47.6%) of cases. In summary, GX-Ultra showed excellent specificity and high sensitivity in paubacillary specimens, making it a useful tool for rapid diagnosis of extrapulmonary tuberculosis.
Subject(s)
Diagnostic Tests, Routine/methods , Molecular Diagnostic Techniques/methods , Mycobacterium tuberculosis/isolation & purification , Tuberculosis/diagnosis , Adult , Antibiotics, Antitubercular/pharmacology , Drug Resistance, Bacterial/genetics , Humans , Mycobacterium tuberculosis/genetics , Nontuberculous Mycobacteria/genetics , Nontuberculous Mycobacteria/isolation & purification , Rifampin/pharmacology , Sensitivity and Specificity , Time Factors , Tuberculosis/microbiology , Tuberculosis/pathologyABSTRACT
Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) has proved to be a useful diagnostic method for identifying conventional bacteria. In the case of mycobacteria, a good protein extraction protocol is essential in order to obtain reliable identification results. To date, no such protocol has been definitively established. The aim of this study was to compare the manufacturer's recommended protein extraction protocol (protocol A) with two novel protocols (protocols B and C), which apply different freezing temperatures and mechanical disruption times using an automatic tissue homogenizer. A total of 302 clinical isolates, comprising 41 nontuberculous mycobacteria (NTM) species, were grown in parallel on solid and liquid media and analyzed: 174 isolates were slow-growing mycobacteria (SGM) and 128 isolates were rapid-growing mycobacteria (RGM). Overall, MALDI-TOF MS identified a higher number of NTM isolates from solid than from liquid media, especially with protocol C (83.4 and 68.2%, respectively; P < 0.05). From solid media, this protein extraction method identified 57.9 and 3.9% more isolates than protocols A (P < 0.001) and B (P < 0.05), respectively. In the case of liquid media, protocol C identified 49.7 and 6.3% more isolates than protocols A and B, respectively (P < 0.001). With regard to the growth rate, MALDI-TOF MS identified more RGM isolates than SGM isolates in all of the protocols studied. In conclusion, the application of freezing and automatic tissue homogenizer improved protein extraction of NTM and boosted identification rates. Consequently, MALDI-TOF MS, which is a cheap and simple method, could be a helpful tool for identifying NTM species in clinical laboratories.
Subject(s)
Bacterial Proteins/isolation & purification , Freezing , Nontuberculous Mycobacteria/isolation & purification , Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization/methods , Stress, Mechanical , Bacterial Proteins/chemistry , Culture Media/chemistry , Humans , Mycobacterium Infections, Nontuberculous/diagnosis , Mycobacterium Infections, Nontuberculous/microbiology , Nontuberculous Mycobacteria/chemistry , Nontuberculous Mycobacteria/growth & development , Research Design , Specimen Handling/methods , Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization/economicsABSTRACT
Mycobacteria are a large group of microorganisms, multiple species of which are major causes of morbidity and mortality, such as tuberculosis and leprosy. At present, the emergence and spread of multidrug-resistant strains of Mycobacterium tuberculosis complex are one of the most serious health problems worldwide. Furthermore, in contrast to M. tuberculosis and Mycobacterium leprae, non-tuberculous mycobacteria (NTM) are more frequently isolated and, in many cases, treatment is based on drug susceptibility testing. This article is a review of the different methods to determine the in vitro drug susceptibility of M. tuberculosis complex and the most relevant NTM isolates. The molecular techniques currently used for rapid detection of resistance of clinical specimens are also analysed.
Subject(s)
Anti-Bacterial Agents/pharmacology , Drug Resistance, Microbial , Microbial Sensitivity Tests/methods , Mycobacterium/drug effects , Antitubercular Agents/pharmacology , Drug Resistance, Multiple, Bacterial , Humans , Molecular Diagnostic Techniques , Mycobacterium/classification , Mycobacterium Infections/microbiology , Nontuberculous Mycobacteria/drug effects , Species SpecificityABSTRACT
To date, more than 170 species of mycobacteria have been described, of which more than one third may be pathogenic to humans, representing a significant workload for microbiology laboratories. These species must be identified in clinical practice, which has long been a major problem due to the shortcomings of conventional (phenotypic) methods and the limitations and complexity of modern methods largely based on molecular biology techniques. The aim of this review was to briefly describe different aspects related to the use of MALDI-TOF (matrix-assisted laser desorption ionization time-of-flight) mass spectrometry (MS) for the identification of mycobacteria. Several difficulties are encountered with the use of this methodology in these microorganisms mainly due to the high pathogenicity of some mycobacteria and the peculiar structure of their cell wall, requiring inactivation and special protein extraction protocols. We also analysed other relevant aspects such as culture media, the reference methods employed (gold standard) in the final identification of the different species, the cut-off used to accept data as valid, and the databases of the different mass spectrometry systems available. MS has revolutionized diagnosis in modern microbiology; however, specific improvements are needed to consolidate the use of this technology in mycobacteriology.
Subject(s)
Bacteriological Techniques/methods , Mycobacterium Infections/diagnosis , Mycobacterium/isolation & purification , Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization/methods , Bacterial Proteins/analysis , Bacterial Proteins/isolation & purification , Bacterial Typing Techniques/methods , Bacteriological Techniques/instrumentation , Culture Media , Equipment Design , Genotype , Humans , Mycobacterium/chemistry , Mycobacterium/genetics , Mycobacterium Infections/microbiology , Phenotype , Reference Standards , Specimen Handling , Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization/instrumentationABSTRACT
We evaluated the performance of the molecular lab-on-chip-based VerePLEX Biosystem for detection of multidrug-resistant tuberculosis (MDR-TB), obtaining a diagnostic accuracy of more than 97.8% compared to sequencing and MTBDRplus assay for Mycobacterium tuberculosis complex and rifampin and isoniazid resistance detection on clinical isolates and smear-positive specimens. The speed, user-friendly interface, and versatility make it suitable for routine laboratory use.
Subject(s)
Lab-On-A-Chip Devices , Molecular Diagnostic Techniques/methods , Tuberculosis, Multidrug-Resistant/diagnosis , Antitubercular Agents/pharmacology , Drug Resistance, Multiple, Bacterial , Humans , Isoniazid/pharmacology , Mycobacterium tuberculosis/drug effects , Rifampin/pharmacologyABSTRACT
OBJECTIVES: Ethambutol resistance has mostly been related to mutations in the embB gene. The objective of the present study was to characterize the embB gene in a collection of ethambutol-resistant and ethambutol-susceptible isolates of Mycobacterium tuberculosis complex (MTBC) from Barcelona, and to develop a DNA microarray for the rapid detection of embB mutations in our area. METHODS: Fifty-three ethambutol-resistant and 702 ethambutol-susceptible isolates of MTBC were sequenced in internal 982-1495 bp fragments of the embB gene. In addition, a low-cost, low-density array was designed to include the embB codons identified as being most frequently mutated in our area (LD-EMB array). RESULTS: The global prevalence of embB mutations found among the ethambutol-resistant isolates was 77.4% (41/53). Substitutions in embB306 were the most common [53.7% (22/41)], followed by substitutions in embB406 [26.8% (11/41)]. The presence of mutations in embB406 was related to higher levels of ethambutol resistance and to multidrug resistance. Among unrelated isolates (from 24-locus MIRU-VNTR genotyping), the percentage of embB-mutated isolates was 72.9% (27/37)--59.3% (16/27) in embB306 and 25.9% (7/27) in embB406. None of the ethambutol-susceptible isolates studied showed a mutation in codon 306 or 406. The LD-EMB array showed 100% sensitivity and specificity in identifying the main embB substitutions in our area. CONCLUSIONS: Mutations at codons 306 and 406 of embB have a relevant role in resistance to ethambutol in our area. The LD-EMB array developed in this study would appear to be a good molecular test for rapid detection of ethambutol resistance.
Subject(s)
Antitubercular Agents/pharmacology , Drug Resistance, Bacterial , Ethambutol/pharmacology , Mutation, Missense , Mycobacterium tuberculosis/genetics , Pentosyltransferases/genetics , Codon , Genotype , Humans , Mycobacterium tuberculosis/isolation & purification , Oligonucleotide Array Sequence Analysis , Sequence Analysis, DNA , SpainABSTRACT
OBJECTIVE: For non-tuberculous mycobacteria (NTM), minimum inhibitory concentration (MIC) distributions of wild-type isolates have not been systematically evaluated despite their importance for establishing antimicrobial susceptibility testing (AST) breakpoints. METHODS: We gathered MIC distributions for drugs used against the Mycobacterium avium complex (MAC) and Mycobacterium abscessus (MAB) obtained by commercial broth microdilution (SLOMYCOI and RAPMYCOI) from 12 laboratories. Epidemiological cut-off values (ECOFFs) and tentative ECOFFs (TECOFFs) were determined by EUCAST methodology including quality control (QC) strains. RESULTS: The clarithromycin ECOFF was 16 mg/L for M. avium (n = 1271) whereas TECOFFs were 8 mg/L for M. intracellulare (n = 415) and 1 mg/L for MAB (n = 1014) confirmed by analysing MAB subspecies without inducible macrolide resistance (n = 235). For amikacin, the ECOFFs were 64 mg/L for MAC and MAB. For moxifloxacin, the WT spanned >8 mg/L for both MAC and MAB. For linezolid, the ECOFF and TECOFF were 64 mg/L for M. avium and M. intracellulare, respectively. Current CLSI breakpoints for amikacin (16 mg/L), moxifloxacin (1 mg/L) and linezolid (8 mg/L) divided the corresponding WT distributions. For QC M. avium and M. peregrinum, ≥95% of MIC values were well within recommended QC ranges. CONCLUSION: As a first step towards clinical breakpoints for NTM, (T)ECOFFs were defined for several antimicrobials against MAC and MAB. Broad wild-type MIC distributions indicate a need for further method refinement which is now under development within the EUCAST subcommittee for anti-mycobacterial drug susceptibility testing. In addition, we showed that several CLSI NTM breakpoints are not consistent in relation to the (T)ECOFFs.
Subject(s)
Mycobacterium Infections, Nontuberculous , Mycobacterium abscessus , Mycobacterium avium-intracellulare Infection , Mycobacterium tuberculosis , Humans , Mycobacterium avium Complex , Anti-Bacterial Agents/pharmacology , Anti-Bacterial Agents/therapeutic use , Nontuberculous Mycobacteria , Amikacin/pharmacology , Moxifloxacin/pharmacology , Linezolid/pharmacology , Mycobacterium avium-intracellulare Infection/microbiology , Microbial Sensitivity Tests , Drug Resistance, Bacterial , Macrolides/pharmacology , Mycobacterium Infections, Nontuberculous/drug therapy , Mycobacterium aviumABSTRACT
BACKGROUND: Interferon-y Release Assays (IGRA) reversions have been reported in different clinical scenarios for the diagnosis of tuberculosis (TB) infection. This study aimed to determine the rate of QuantiFERON-TB Gold Plus (QFT-Plus) reversions during contact investigation as a potential strategy to reduce the number of preventive treatments. METHODS: Prospective, multicentre cohort study of immunocompetent adult contacts of patients with pulmonary TB tested with QFT-Plus. Contacts with an initial positive QFT-Plus (QFT-i) underwent a second test within 4 weeks (QFT-1), and if negative, underwent a repeat test 4 weeks later (QFT-2). Based on the QFT-2 result, we classified cases as sustained reversion if they remained negative and as temporary reversion if they turned positive. RESULTS: We included 415 contacts, of whom 96 (23.1%) had an initial positive test (QFT-i). Following this, 10 had negative QFT-1 results and 4 (4.2%) of these persisted with a negative result in the QFT-2 (sustained reversions). All four sustained reversions occurred in contacts with IFN-γ concentrations between ≥0.35 and ≤0.99 IUâ¢mL-1 in one or both QFT-i tubes. CONCLUSION: In this study, TB contact investigations rarely reveal QFT-Plus reversion. These results do not support retesting cases with an initial positive result to reduce the number of preventive treatments.
Subject(s)
Latent Tuberculosis , Tuberculosis, Pulmonary , Tuberculosis , Adult , Humans , Cohort Studies , Prospective Studies , Tuberculosis/diagnosis , Tuberculosis, Pulmonary/diagnosisABSTRACT
Early extrapulmonary tuberculosis (EPTB) diagnosis is particularly difficult. Among 108 smear-negative extrapulmonary samples showing a positive culture for Mycobacterium tuberculosis complex (43 body fluids and 65 nonliquid specimens), 63 (58.3%) were positive with the Xpert MTB/RIF assay (GX). GX sensitivity was quite low for samples from sterile locations (especially for pleural fluids: 26.9%) but high for some nonliquid samples, like abscess aspirates (76.5%). In summary, GX may be a useful tool to be considered for EPTB diagnosis.
Subject(s)
Mycobacterium tuberculosis/isolation & purification , Real-Time Polymerase Chain Reaction/methods , Tuberculosis/diagnosis , Tuberculosis/microbiology , Humans , Mycobacterium tuberculosis/genetics , Sensitivity and SpecificityABSTRACT
Several molecular systems for direct detection of Mycobacterium tuberculosis complex (MTBC) have recently been developed. The GenoQuick MTB assay (GQ-MTB) used in this study detected 82 of the 96 (85.4%) samples with MTBC, including 50 of 64 (78.1%) samples with negative acid-fast bacillus smears. Fifteen samples containing nontuberculous mycobacteria were also studied: 13 were GQ-MTB negative, one was positive, and one was indeterminate. GQ-MTB showed good effectiveness for the direct detection of MTBC from clinical samples.
Subject(s)
Bacteriological Techniques/methods , Molecular Diagnostic Techniques/methods , Mycobacterium tuberculosis/isolation & purification , Tuberculosis/diagnosis , Tuberculosis/microbiology , Humans , Mycobacterium tuberculosis/genetics , Sensitivity and SpecificityABSTRACT
Tuberculosis is an opportunistic infection with high morbidity and mortality in solid organ transplant patients. The reasons for this high morbidity and mortality lie mostly in diagnostic difficulties, which cause delays in starting treatment, and associated pharmaceutical toxicity. There are still major issues and difficulties in managing tuberculosis in solid organ transplant patients. These include problems due to interactions between antituberculosis and immunosuppressant drugs, the high risk of toxicity of antituberculosis drugs (particularly in liver transplant patients) and the absence of clear indications for the treatment of latent tuberculous infection. This article updates current understanding of tuberculosis in solid organ transplant patients.
Subject(s)
Organ Transplantation/adverse effects , Tuberculosis/etiology , Humans , Latent Tuberculosis/etiology , Tuberculosis/diagnosis , Tuberculosis/drug therapyABSTRACT
The ability of MALDI-TOF for the identification of nontuberculous mycobacteria (NTM) has improved recently thanks to updated databases and optimized protein extraction procedures. Few multicentre studies on the reproducibility of MALDI-TOF have been performed so far, none on mycobacteria. The aim of this study was to evaluate the reproducibility of MALDI-TOF for the identification of NTM in 15 laboratories in 9 European countries. A total of 98 NTM clinical isolates were grown on Löwenstein-Jensen. Biomass was collected in tubes with water and ethanol, anonymized and sent out to the 15 participating laboratories. Isolates were identified using MALDI Biotyper (Bruker Daltonics). Up to 1330 MALDI-TOF identifications were collected in the study. A score ≥ 1.6 was obtained for 100% of isolates in 5 laboratories (68.2-98.6% in the other). Species-level identification provided by MALDI-TOF was 100% correct in 8 centres and 100% correct to complex-level in 12 laboratories. In most cases, the misidentifications obtained were associated with closely related species. The variability observed for a few isolates could be due to variations in the protein extraction procedure or to MALDI-TOF system status in each centre. In conclusion, MALDI-TOF showed to be a highly reproducible method and suitable for its implementation for NTM identification.
Subject(s)
Nontuberculous Mycobacteria/isolation & purification , Humans , Nontuberculous Mycobacteria/classification , Reproducibility of Results , Species Specificity , Spectrometry, Mass, Matrix-Assisted Laser Desorption-IonizationABSTRACT
Sixty-four of 85 (75.3%) smear-negative respiratory (n = 78) and nonrespiratory (n = 7) samples with positive cultures of Mycobacterium tuberculosis complex (MTC) were detected by the GeneXpert system using the Xpert MTB/RIF assay (GX). In addition, GX found rpoB mutations in all six of the rifampin-resistant strains detected. The test was negative in 20 culture-negative and 20 nontuberculous culture-positive samples (100% specificity). GX offers high potential for the diagnosis of tuberculosis due to its capacity for direct detection of MTC, its rapidity, and its simplicity.