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1.
Int Arch Occup Environ Health ; 95(8): 1785-1796, 2022 10.
Artículo en Inglés | MEDLINE | ID: mdl-35551477

RESUMEN

PURPOSE: Exposures related to beryllium (Be) are an enduring concern among workers in the nuclear weapons and other high-tech industries, calling for regular and rigorous biological monitoring. Conventional biomonitoring of Be in urine is not informative of cumulative exposure nor health outcomes. Biomarkers of exposure to Be based on non-invasive biomonitoring could help refine disease risk assessment. In a cohort of workers with Be exposure, we employed blood plasma extracellular vesicles (EVs) to discover novel biomarkers of exposure to Be. METHODS: EVs were isolated from plasma using size-exclusion chromatography and subjected to mass spectrometry-based proteomics. A protein-based classifier was developed using LASSO regression and validated by ELISA. RESULTS: We discovered a dual biomarker signature comprising zymogen granule protein 16B and putative protein FAM10A4 that differentiated between Be-exposed and -unexposed subjects. ELISA-based quantification of the biomarkers in an independent cohort of samples confirmed higher expression of the signature in the Be-exposed group, displaying high predictive accuracy (AUROC = 0.919). Furthermore, the biomarkers efficiently discriminated high- and low-exposure groups (AUROC = 0.749). CONCLUSIONS: This is the first report of EV biomarkers associated with Be exposure and exposure levels. The biomarkers could be implemented in resource-limited settings for Be exposure assessment.


Asunto(s)
Berilio , Vesículas Extracelulares , Berilio/metabolismo , Biomarcadores , Vesículas Extracelulares/química , Vesículas Extracelulares/metabolismo , Humanos , Espectrometría de Masas , Proteómica/métodos
2.
Emerg Infect Dis ; 25(3): 473-481, 2019 03.
Artículo en Inglés | MEDLINE | ID: mdl-30789130

RESUMEN

Attention to environmental sources of Mycobacterium avium complex (MAC) infection is a vital component of disease prevention and control. We investigated MAC colonization of household plumbing in suburban Philadelphia, Pennsylvania, USA. We used variable-number tandem-repeat genotyping and whole-genome sequencing with core genome single-nucleotide variant analysis to compare M. avium from household plumbing biofilms with M. avium isolates from patient respiratory specimens. M. avium was recovered from 30 (81.1%) of 37 households, including 19 (90.5%) of 21 M. avium patient households. For 11 (52.4%) of 21 patients with M. avium disease, isolates recovered from their respiratory and household samples were of the same genotype. Within the same community, 18 (85.7%) of 21 M. avium respiratory isolates genotypically matched household plumbing isolates. Six predominant genotypes were recovered across multiple households and respiratory specimens. M. avium colonizing municipal water and household plumbing may be a substantial source of MAC pulmonary infection.


Asunto(s)
Microbiología Ambiental , Infección por Mycobacterium avium-intracellulare/epidemiología , Infección por Mycobacterium avium-intracellulare/microbiología , Mycobacterium avium/clasificación , Microbiología del Agua , Adulto , Anciano , Anciano de 80 o más Años , Femenino , Genotipo , Historia del Siglo XXI , Humanos , Masculino , Persona de Mediana Edad , Repeticiones de Minisatélite , Tipificación de Secuencias Multilocus , Mycobacterium avium/genética , Mycobacterium avium/aislamiento & purificación , Complejo Mycobacterium avium/clasificación , Complejo Mycobacterium avium/genética , Complejo Mycobacterium avium/aislamiento & purificación , Infección por Mycobacterium avium-intracellulare/historia , Philadelphia/epidemiología , Filogenia , Vigilancia en Salud Pública , Secuenciación Completa del Genoma
3.
J Clin Microbiol ; 57(2)2019 02.
Artículo en Inglés | MEDLINE | ID: mdl-30429252

RESUMEN

We characterize three respiratory isolates of the recently described species Mycobacterium talmoniae recovered in Texas, Louisiana, and Massachusetts, including the first case of disease in a patient with underlying cystic fibrosis. The three isolates had a 100% match to M. talmoniae NE-TNMC-100812T by complete 16S rRNA, rpoB region V, and hsp65 gene sequencing. Core genomic comparisons between one isolate and the type strain revealed an average nucleotide identity of 99.8%. The isolates were susceptible to clarithromycin, amikacin, and rifabutin, while resistance was observed for tetracyclines, ciprofloxacin, and linezolid. M. talmoniae should be added to the list of potential pulmonary pathogens, including in the setting of cystic fibrosis.


Asunto(s)
Bronquiectasia/complicaciones , Fibrosis Quística/complicaciones , Enfermedades Pulmonares Fúngicas/diagnóstico , Infecciones por Mycobacterium/diagnóstico , Mycobacterium/clasificación , Mycobacterium/aislamiento & purificación , Filogenia , Anciano , Antibacterianos/farmacología , Proteínas Bacterianas/genética , Chaperonina 60/genética , Niño , Análisis por Conglomerados , ADN Bacteriano/química , ADN Bacteriano/genética , ADN Ribosómico/química , ADN Ribosómico/genética , ARN Polimerasas Dirigidas por ADN/genética , Femenino , Humanos , Louisiana , Enfermedades Pulmonares Fúngicas/microbiología , Masculino , Massachusetts , Pruebas de Sensibilidad Microbiana , Mycobacterium/efectos de los fármacos , Mycobacterium/genética , Infecciones por Mycobacterium/microbiología , ARN Ribosómico 16S/genética , Análisis de Secuencia de ADN , Texas
4.
J Clin Microbiol ; 56(8)2018 08.
Artículo en Inglés | MEDLINE | ID: mdl-29875193

RESUMEN

The accuracy and robustness of the Vitek MS v3.0 matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry (MS) system was evaluated by identifying mycobacteria from automated liquid-medium systems using patient samples. This is the first report to demonstrate that proteins within the liquid medium, its supplements, and decontamination reagents for nonsterile patient samples do not generate misidentification or false-positive results by use of the Vitek MS v3.0 system. Prior to testing with patient samples, a seeded-culture study was conducted to challenge the accuracy of the Vitek MS system at identifying mycobacteria from liquid medium by mimicking a clinical workflow. Seventy-seven Mycobacterium strains representing 21 species, seeded in simulated sputum, were decontaminated, inoculated into BacT/Alert MP liquid culture medium, incubated until positivity, and identified using the Vitek MS system. A total of 383 liquid cultures were tested, of which 379 (99%) were identified correctly to the species/complex/group level, 4 (1%) gave a "no-identification" result, and no misidentifications were observed. Following the simulated-sputum study, a total of 73 smear-positive liquid-medium cultures detected using BD BBL MGIT and VersaTREK Myco liquid media were identified by the Vitek MS system. Sixty-four cultures (87.7%) were correctly identified to the species/complex/group level; 7 (9.6%) resulted in no identification; and 2 (2.7%) were misidentified at the species level. These results indicate that the Vitek MS v3.0 system is an accurate tool for routine diagnostics of Mycobacterium species isolated from liquid cultures.


Asunto(s)
Técnicas Bacteriológicas/métodos , Infecciones por Mycobacterium no Tuberculosas/diagnóstico , Mycobacterium/clasificación , Mycobacterium/aislamiento & purificación , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción , Técnicas Bacteriológicas/instrumentación , Medios de Cultivo , Pruebas Diagnósticas de Rutina , Humanos , Mycobacterium/química , Esputo/microbiología
5.
J Clin Microbiol ; 56(6)2018 06.
Artículo en Inglés | MEDLINE | ID: mdl-29643203

RESUMEN

This multicenter study was designed to assess the accuracy and reproducibility of the Vitek MS v3.0 matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry system for identification of Mycobacterium and Nocardia species compared to DNA sequencing. A total of 963 clinical isolates representing 51 taxa were evaluated. In all, 663 isolates were correctly identified to the species level (69%), with another 231 (24%) correctly identified to the complex or group level. Fifty-five isolates (6%) could not be identified despite repeat testing. All of the tuberculous mycobacteria (45/45; 100%) and most of the nontuberculous mycobacteria (569/606; 94%) were correctly identified at least to the group or complex level. However, not all species or subspecies within the M. tuberculosis, M. abscessus, and M. avium complexes and within the M. fortuitum and M. mucogenicum groups could be differentiated. Among the 312 Nocardia isolates tested, 236 (76%) were correctly identified to the species level, with an additional 44 (14%) correctly identified to the complex level. Species within the N. nova and N. transvalensis complexes could not always be differentiated. Eleven percent of the isolates (103/963) underwent repeat testing in order to get a final result. Identification of a representative set of Mycobacterium and Nocardia species was highly reproducible, with 297 of 300 (99%) replicates correctly identified using multiple kit lots, instruments, analysts, and sites. These findings demonstrate that the system is robust and has utility for the routine identification of mycobacteria and Nocardia in clinical practice.


Asunto(s)
Mycobacterium tuberculosis/aislamiento & purificación , Nocardia/aislamiento & purificación , Micobacterias no Tuberculosas/aislamiento & purificación , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción/instrumentación , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción/métodos , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Mycobacterium tuberculosis/genética , Nocardia/genética , Nocardiosis/diagnóstico , Nocardiosis/microbiología , ARN Ribosómico 16S/genética , Juego de Reactivos para Diagnóstico , Reproducibilidad de los Resultados , Tuberculosis/diagnóstico , Tuberculosis/microbiología
6.
Int J Syst Evol Microbiol ; 68(11): 3557-3562, 2018 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-30204586

RESUMEN

Two mycobacterial strains with close similarity to the Mycobacterium tuberculosis complex (MTBC) were isolated from cutaneous lesions of patients in the USA and Italy. At the phenotypic level, similarities to the MTBC included slow growth rate, rough morphotype of the unpigmented colonies and nearly identical high-performance liquid chromatography profiles of mycolic acids. In contrast to the MTBC, the strains were niacin- and nitrate-negative, and catalase-positive both at 68 °C and in semi-quantitative tests. The clinical isolates were more closely related to M. tuberculosis than to any other known mycobacterium and scored positive with commercial DNA probes (Hologic AccuProbe M. tuberculosis). Both average nucleotide identity and genome-to-genome distance suggested the strains are different from the MTBC. Therefore, given the distinguishing phenotypic and genomic-scale differences, we submit that the strains belong to a new species we have named Mycobacteriumdecipiens with type strain TBL 1200985T (=ATCC TSD-117T=DSM 105360T).


Asunto(s)
Infecciones por Mycobacterium/microbiología , Mycobacterium/clasificación , Filogenia , Tuberculosis Cutánea/microbiología , Técnicas de Tipificación Bacteriana , Composición de Base , ADN Bacteriano/genética , Humanos , Italia , Mycobacterium/genética , Mycobacterium/aislamiento & purificación , Mycobacterium tuberculosis , Ácidos Micólicos/química , ARN Ribosómico 16S/genética , Análisis de Secuencia de ADN , Estados Unidos
7.
Clin Infect Dis ; 64(7): 902-911, 2017 Apr 01.
Artículo en Inglés | MEDLINE | ID: mdl-28077517

RESUMEN

BACKGROUND: Nontuberculous mycobacteria (NTM) commonly colonize municipal water supplies and cause healthcare-associated outbreaks. We investigated a biphasic outbreak of Mycobacterium abscessus at a tertiary care hospital. METHODS: Case patients had recent hospital exposure and laboratory-confirmed colonization or infection with M. abscessus from January 2013 through December 2015. We conducted a multidisciplinary epidemiologic, field, and laboratory investigation. RESULTS: The incidence rate of M. abscessus increased from 0.7 cases per 10000 patient-days during the baseline period (January 2013-July 2013) to 3.0 cases per 10000 patient-days during phase 1 of the outbreak (August 2013-May 2014) (incidence rate ratio, 4.6 [95% confidence interval, 2.3-8.8]; P < .001). Thirty-six of 71 (51%) phase 1 cases were lung transplant patients with positive respiratory cultures. We eliminated tap water exposure to the aerodigestive tract among high-risk patients, and the incidence rate decreased to baseline. Twelve of 24 (50%) phase 2 (December 2014-June 2015) cases occurred in cardiac surgery patients with invasive infections. Phase 2 resolved after we implemented an intensified disinfection protocol and used sterile water for heater-cooler units of cardiopulmonary bypass machines. Molecular fingerprinting of clinical isolates identified 2 clonal strains of M. abscessus; 1 clone was isolated from water sources at a new hospital addition. We made several water engineering interventions to improve water flow and increase disinfectant levels. CONCLUSIONS: We investigated and mitigated a 2-phase clonal outbreak of M. abscessus linked to hospital tap water. Healthcare facilities with endemic NTM should consider similar tap water avoidance and engineering strategies to decrease risk of NTM infection.


Asunto(s)
Infección Hospitalaria , Brotes de Enfermedades , Infecciones por Mycobacterium no Tuberculosas/epidemiología , Infecciones por Mycobacterium no Tuberculosas/microbiología , Mycobacterium abscessus/clasificación , Mycobacterium abscessus/genética , Anciano , Femenino , Genes Bacterianos , Hospitales , Humanos , Incidencia , Masculino , Persona de Mediana Edad , Tipificación de Secuencias Multilocus , Infecciones por Mycobacterium no Tuberculosas/diagnóstico , Infecciones por Mycobacterium no Tuberculosas/etiología , Factores de Riesgo
8.
J Clin Microbiol ; 55(2): 574-584, 2017 02.
Artículo en Inglés | MEDLINE | ID: mdl-27927925

RESUMEN

Bedaquiline (BDQ), a diarylquinoline antibiotic that targets ATP synthase, is effective for the treatment of Mycobacterium tuberculosis infections that no longer respond to conventional drugs. While investigating the off-label use of BDQ as salvage therapy, seven of 13 patients with Mycobacterium intracellulare lung disease had an initial microbiological response and then relapsed. Whole-genome comparison of pretreatment and relapse isolates of M. intracellulare uncovered mutations in a previously uncharacterized locus, mmpT5 Preliminary analysis suggested similarities between mmpT5 and the mmpR5 locus, which is associated with low-level BDQ resistance in M. tuberculosis Both genes encode transcriptional regulators and are adjacent to orthologs of the mmpS5-mmpL5 drug efflux operon. However, MmpT5 belongs to the TetR superfamily, whereas MmpR5 is a MarR family protein. Targeted sequencing uncovered nonsynonymous mmpT5 mutations in isolates from all seven relapse cases, including two pretreatment isolates. In contrast, only two relapse patient isolates had nonsynonymous changes in ATP synthase subunit c (atpE), the primary target of BDQ. Susceptibility testing indicated that mmpT5 mutations are associated with modest 2- to 8-fold increases in MICs for BDQ and clofazimine, whereas one atpE mutant exhibited a 50-fold increase in MIC for BDQ. Bedaquiline shows potential for the treatment of M. intracellulare lung disease, but optimization of treatment regimens is required to prevent the emergence of mmpT5 variants and microbiological relapse.


Asunto(s)
Antituberculosos/uso terapéutico , Diarilquinolinas/uso terapéutico , Farmacorresistencia Bacteriana , Mutación Missense , Complejo Mycobacterium avium/genética , Factores de Transcripción/genética , Tuberculosis Pulmonar/tratamiento farmacológico , Anciano , Femenino , Genoma Bacteriano , Humanos , Masculino , Pruebas de Sensibilidad Microbiana , Persona de Mediana Edad , Complejo Mycobacterium avium/aislamiento & purificación , Recurrencia , Análisis de Secuencia de ADN , Tuberculosis Pulmonar/microbiología
9.
Int J Syst Evol Microbiol ; 67(11): 4345-4351, 2017 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-28984546

RESUMEN

A previously undescribed, rapidly growing, scotochromogenic species of the genus Mycobacterium (represented by strains PB739T and GK) was isolated from two clinical sources - the sputum of a 76-year-old patient with severe chronic obstructive pulmonary disease, history of tuberculosis exposure and Mycobacterium avium complex isolated years prior; and the blood of a 15-year-old male with B-cell acute lymphoblastic leukaemia status post bone marrow transplant. The isolates grew as dark orange colonies at 25-37 °C after 5 days, sharing features in common with other closely related species. Analysis of the complete 16S rRNA gene sequence (1492 bp) of strain PB739T demonstrated that the isolate shared 98.8 % relatedness with Mycobacterium wolinskyi. Partial 429 bp hsp65 and 744 bp rpoB region V sequence analyses revealed that the sequences of the novel isolate shared 94.8 and 92.1 % similarity with those of Mycobacterium neoaurum and Mycobacterium aurum, respectively. Biochemical profiling, antimicrobial susceptibility testing, HPLC/gas-liquid chromatography analyses and multilocus sequence typing support the taxonomic status of these isolates (PB739T and GK) as representatives of a novel species. Both isolates were susceptible to the Clinical and Laboratory Standards Institute recommended antimicrobials for susceptibility testing of rapidly growing mycobacteria including amikacin, ciprofloxacin, moxifloxacin, doxycycline/minocycline, imipenem, linezolid, clarithromycin and trimethropin/sulfamethoxazole. Both isolates PB739T and GK showed intermediate susceptibility to cefoxitin. We propose the name Mycobacterium grossiae sp. nov. for this novel species and have deposited the type strain in the DSMZ and CIP culture collections. The type strain is PB739T (=DSM 104744T=CIP 111318T).


Asunto(s)
Infecciones por Mycobacterium/microbiología , Mycobacterium/clasificación , Filogenia , Adolescente , Anciano , Técnicas de Tipificación Bacteriana , Cultivo de Sangre , ADN Bacteriano/genética , Humanos , Masculino , Tipificación de Secuencias Multilocus , Mycobacterium/genética , Mycobacterium/aislamiento & purificación , Infecciones por Mycobacterium/sangre , Pigmentación , ARN Ribosómico 16S/genética , Análisis de Secuencia de ADN , Esputo/microbiología
10.
J Clin Microbiol ; 54(5): 1340-51, 2016 05.
Artículo en Inglés | MEDLINE | ID: mdl-26962085

RESUMEN

Mycobacterium terrae complex has been recognized as a cause of tenosynovitis, with M. terrae and Mycobacterium nonchromogenicum reported as the primary etiologic pathogens. The molecular taxonomy of the M. terrae complex causing tenosynovitis has not been established despite approximately 50 previously reported cases. We evaluated 26 isolates of the M. terrae complex associated with tenosynovitis or osteomyelitis recovered between 1984 and 2014 from 13 states, including 5 isolates reported in 1991 as M. nonchromogenicum by nonmolecular methods. The isolates belonged to three validated species, one new proposed species, and two novel related strains. The majority of isolates (20/26, or 77%) belonged to two recently described species: Mycobacterium arupense (10 isolates, or 38%) and Mycobacterium heraklionense (10 isolates, or 38%). Three isolates (12%) had 100% sequence identity to each other by 16S rRNA and 99.3 to 100% identity by rpoB gene region V sequencing and represent a previously undescribed species within the M. terrae complex. There were no isolates of M. terrae or M. nonchromogenicum, including among the five isolates reported in 1991. The 26 isolates were susceptible to clarithromycin (100%), rifabutin (100%), ethambutol (92%), and sulfamethoxazole or trimethoprim-sulfamethoxazole (70%). The current study suggests that M. arupense, M. heraklionense, and a newly proposed species ("M. virginiense" sp. nov.; proposed type strain MO-233 [DSM 100883, CIP 110918]) within the M. terrae complex are the major causes of tenosynovitis and osteomyelitis in the United States, with little change over 20 years. Species identification within this complex requires sequencing methods.


Asunto(s)
Infecciones por Mycobacterium no Tuberculosas/microbiología , Mycobacterium/clasificación , Mycobacterium/aislamiento & purificación , Osteomielitis/microbiología , Tenosinovitis/microbiología , Adulto , Anciano , Antituberculosos/farmacología , ADN Ribosómico/química , ADN Ribosómico/genética , ARN Polimerasas Dirigidas por ADN , Femenino , Humanos , Masculino , Pruebas de Sensibilidad Microbiana , Persona de Mediana Edad , Mycobacterium/efectos de los fármacos , Mycobacterium/genética , ARN Ribosómico 16S/genética , Análisis de Secuencia de ADN , Estados Unidos , Adulto Joven
11.
J Clin Microbiol ; 54(4): 891-901, 2016 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-26739155

RESUMEN

"Mycobacterium aviumsubsp.hominissuis" is an important cause of pulmonary disease. It is acquired from environmental sources, but there is no methodology for large population studies. We evaluated the potential of variable-number tandem-repeat (VNTR) analysis. Clinical and household biofilmM. aviumisolates underwent molecular identification. Testing for IS901was done to separateM. aviumsubsp.aviumfromM. aviumsubsp.hominissuis VNTR types were defined using VNTR loci, and subtyping was performed using 3'hsp65and internal transcribed spacer (ITS) sequencing. Forty-nine VNTR types and eight subtypes ofM. aviumsubsp.hominissuis(IS901negative) were identified among 416 isolates ofM. aviumfrom 121 patients and 80 biofilm sites. Of those types, 67% were found only among patient isolates, 11% only among household water isolates, and 23% among both. Of 13 VNTR types that included ≥4 patients, the majority (61.5%) represented geographic clustering (same city). Most VNTR types with multiple patients belonged to the same 3'hsp65sequence code (sequevar). A total of 44 isolates belonging to fourM. aviumsubsp.hominissuisVNTR types (8%), including three with the rare Mav-F ITS sequence and 0/8 subspecies, produced amplicons with IS901PCR primers. By sequencing, all 44 amplicons were not IS901but ISMav6, which was recently observed in Japan but had not been previously described among U.S. isolates. VNTR analysis ofM. aviumsubsp.hominissuisisolates is easier and faster than pulsed-field gel electrophoresis. Seven VNTR loci separated 417 isolates into 49 types. No isolates ofM. aviumsubsp.aviumwere identified. The distributions of the VNTR copy numbers, the allelic diversity, and the low prevalence of ISMav6 differed from the findings for respiratory isolates reported from Japan.


Asunto(s)
Biopelículas , Repeticiones de Minisatélite , Tipificación Molecular/métodos , Mycobacterium avium/clasificación , Mycobacterium avium/aislamiento & purificación , Tuberculosis/microbiología , Microbiología del Agua , Proteínas Bacterianas/genética , Chaperonina 60/genética , Análisis por Conglomerados , Elementos Transponibles de ADN , ADN Espaciador Ribosómico/genética , Composición Familiar , Genotipo , Humanos , Japón , Epidemiología Molecular/métodos , Mycobacterium avium/genética , Filogeografía , Reacción en Cadena de la Polimerasa/métodos
12.
J Clin Microbiol ; 53(11): 3686-90, 2015 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-26292312

RESUMEN

Nocardia thailandica is a rare pathogen related to Nocardia asteroides, Nocardia neocaledoniensis, and Nocardia caishijiensis that, since its original description in 2004, has only been reported to cause wound and ocular infections in humans. We report a case of pulmonary nocardiosis caused by Nocardia thailandica in a 66-year-old solid organ transplant patient from Connecticut, which was identified at the molecular taxonomic level by secA1 analysis, 16S rRNA gene sequencing, and matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS). To our knowledge, this is the first reported case of N. thailandica in the United States and the first report of pulmonary infection by this pathogen in the literature.


Asunto(s)
Huésped Inmunocomprometido/inmunología , Enfermedades Pulmonares/diagnóstico , Nocardiosis/diagnóstico , Nocardiosis/inmunología , Nocardia/aislamiento & purificación , Adenosina Trifosfatasas/genética , Anciano , Proteínas Bacterianas/genética , Secuencia de Bases , Connecticut , ADN Bacteriano/genética , ADN Ribosómico/genética , Trasplante de Corazón/efectos adversos , Humanos , Terapia de Inmunosupresión/efectos adversos , Enfermedades Pulmonares/inmunología , Enfermedades Pulmonares/microbiología , Masculino , Proteínas de Transporte de Membrana/genética , Datos de Secuencia Molecular , Nocardiosis/microbiología , ARN Ribosómico 16S/genética , Canales de Translocación SEC , Proteína SecA , Análisis de Secuencia de ADN , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción
13.
J Clin Microbiol ; 53(4): 1211-5, 2015 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-25653399

RESUMEN

The erm(41) gene confers inducible macrolide resistance in Mycobacterium abscessus subsp. abscessus, calling into question the usefulness of macrolides for treating M. abscessus subsp. abscessus infections. With an extended incubation (14 days), isolates with MICs of ≥8 µg/ml are considered macrolide resistant by current CLSI guidelines. Our goals were to determine the incidence of macrolide susceptibility in U.S. isolates, the validity of currently accepted MIC breakpoints, and the erm(41) sequences associated with susceptibility. Of 349 isolates (excluding those with 23S rRNA gene mutations), 85 (24%) had clarithromycin MICs of ≤8 µg/ml. Sequencing of the erm(41) genes from these isolates, as well as from isolates with MICs of ≥16 µg/ml, including ATCC 19977T, revealed 10 sequevars. The sequence in ATCC 19977T was designated sequevar (type) 1; most macrolide-resistant isolates were of this type. Seven sequevars contained isolates with MICs of >16 µg/ml. The T28C substitution in erm(41), previously associated with macrolide susceptibility, was identified in 62 isolates (18%) comprising three sequevars, with MICs of ≤2 (80%), 4 (10%), and 8 (10%) µg/ml. No other nucleotide substitution was associated with macrolide susceptibility. We recommend that clarithromycin susceptibility breakpoints for M. abscessus subsp. abscessus be changed from ≤2 to ≤4 µg/ml and that isolates with an MIC of 8 µg/ml have repeat MIC testing or erm sequencing performed. Our studies suggest that macrolides are useful for treating approximately 20% of U.S. isolates of M. abscessus subsp. abscessus. Sequencing of the erm gene of M. abscessus subsp. abscessus will predict inducible macrolide susceptibility.


Asunto(s)
Antibacterianos/farmacología , Claritromicina/farmacología , Farmacorresistencia Bacteriana , Metiltransferasas/genética , Mycobacterium/efectos de los fármacos , Mycobacterium/enzimología , Genotipo , Humanos , Macrólidos/farmacología , Pruebas de Sensibilidad Microbiana , Datos de Secuencia Molecular , Proteínas Mutantes/genética , Mycobacterium/genética , Análisis de Secuencia de ADN , Estados Unidos
14.
J Clin Microbiol ; 53(3): 875-8, 2015 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-25568437

RESUMEN

Macrolide resistance has been linked to the presence of a functional erythromycin ribosomal methylase (erm) gene in most species of pathogenic rapidly growing mycobacteria (RGM). For these Mycobacterium isolates, extended incubation in clarithromycin is necessary to determine macrolide susceptibility. In contrast, the absence of a detectable erm gene in isolates of M. chelonae, M. senegalense, and M. peregrinum and a nonfunctional erm gene in M. abscessus subsp. massiliense and 15% to 20% of M. abscessus subsp. abscessus isolates renders these species intrinsically macrolide susceptible. Not all RGM species have been screened for the presence of an erm gene, including the Mycobacterium mucogenicum group (M. mucogenicum, M. phocaicum, and M. aubagnense) and Mycobacterium immunogenum. A total of 356 isolates of these two pathogenic RGM taxa from two reference laboratories (A.R.U.P. Reference Laboratories and the Mycobacteria/Nocardia Laboratory at the University of Texas Health Science Center at Tyler) underwent clarithromycin susceptibility testing with readings at 3 to 5 days and 14 days. Only 13 of the 356 isolates had resistant clarithromycin MICs at initial extended MIC readings, and repeat values on all available isolates were ≤2 µg/ml. These studies suggest that these two additional RGM groups do not harbor functional erm genes and, like M. chelonae, do not require extended clarithromycin susceptibility testing. We propose to the Clinical Laboratory and Standards Institute that isolates belonging to these above-mentioned six rapidly growing mycobacterial groups based on molecular identification with no known functional erm genes undergo only 3 to 5 days of susceptibility testing (to exclude mutational resistance).


Asunto(s)
Antibacterianos/farmacología , Claritromicina/farmacología , Metiltransferasas/genética , Micobacterias no Tuberculosas/efectos de los fármacos , Micobacterias no Tuberculosas/genética , Humanos , Macrólidos/farmacología , Pruebas de Sensibilidad Microbiana , Infecciones por Mycobacterium/microbiología , Micobacterias no Tuberculosas/aislamiento & purificación , Estudios Retrospectivos , Factores de Tiempo
16.
J Clin Microbiol ; 52(6): 2265-9, 2014 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-24740075

RESUMEN

Mycobacterium canariasense is a recently described late-pigmenting, rapidly growing mycobacterium linked to bacteremia in patients with underlying malignant diseases. We report a case of M. canariasense infection in a patient from Massachusetts with underlying diffuse B cell lymphoma, which was identified both by multilocus sequence typing and matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS). To our knowledge, this is the first description after its original identification in Spain and the first report of this opportunistic pathogen in the Americas.


Asunto(s)
Bacteriemia/diagnóstico , Infecciones Relacionadas con Catéteres/diagnóstico , Infecciones por Mycobacterium no Tuberculosas/diagnóstico , Mycobacterium/aislamiento & purificación , Adulto , Bacteriemia/microbiología , Bacteriemia/patología , Infecciones Relacionadas con Catéteres/microbiología , Infecciones Relacionadas con Catéteres/patología , Humanos , Linfoma de Células B Grandes Difuso/complicaciones , Masculino , Massachusetts , Datos de Secuencia Molecular , Tipificación de Secuencias Multilocus , Mycobacterium/química , Mycobacterium/clasificación , Mycobacterium/genética , Infecciones por Mycobacterium no Tuberculosas/microbiología , Infecciones por Mycobacterium no Tuberculosas/patología , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción
17.
J Clin Microbiol ; 52(12): 4414-8, 2014 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-25253791
18.
Microbiology (Reading) ; 159(Pt 11): 2323-2332, 2013 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-24014661

RESUMEN

Nearly half of US clinical isolates of the emerging pathogen Mycobacterium abscessus were reported to exhibit smeared DNA during PFGE. This DNA degradation (Dnd) phenotype results from DNA phosphorothioation, a sulfur modification found in other bacteria and conferred by dnd genes located on mobile elements. Putative dnd genes are located on a 19.6 kbp genomic island (GI) in the M. abscessus type strain ATCC 19977. We confirmed that ATCC 19977(T) is Dnd-positive by PFGE and we developed a PCR assay to predict Dnd phenotype. Dnd-positive strains generated an amplicon from dndC whereas Dnd-negative strains generated a bridge amplicon that spanned the GI insertion site, indicating they lacked the entire 'Dnd-GI'. Comparative analyses of sequences from the bridge amplicon with ATCC 19977(T) revealed the Dnd-GI is flanked by 22 bp repeats in M. abscessus sensu stricto and inserted downstream of a tRNA-Ala gene and between inverted repeats. Regions flanking the Dnd-GI were highly conserved within the M. abscessus complex. Bioinformatics studies suggest the Dnd-GI inserted independently into a strain of Mycobacterium massiliense and that other species of mycobacteria also have dnd genes, supporting reports that the Dnd phenotype is common among actinomycetes. Within the M. abscessus complex, Dnd-positive clinical isolates were primarily M. abscessus sensu stricto, and tandem repeat typing indicated these isolates were highly related, confirming previous PFGE studies and revealing a widespread family of strains with significance in human disease.


Asunto(s)
Cromosomas Bacterianos , ADN Bacteriano/metabolismo , Islas Genómicas , Mycobacterium/genética , Mycobacterium/metabolismo , Tionucleótidos/metabolismo , ADN Bacteriano/química , ADN Bacteriano/genética , Electroforesis en Gel de Campo Pulsado , Orden Génico , Datos de Secuencia Molecular , Reacción en Cadena de la Polimerasa , Recombinación Genética , Análisis de Secuencia de ADN
19.
J Clin Microbiol ; 51(2): 409-16, 2013 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-23175249

RESUMEN

Strain comparison is important to population genetics and to evaluate relapses in patients with Mycobacterium avium complex (MAC) lung disease, but the "gold standard" of pulsed-field gel electrophoresis (PFGE) is time-consuming and complex. We used variable-number tandem repeats (VNTR) for fingerprinting of respiratory isolates of M. intracellulare from patients with underlying bronchiectasis, to establish a nonsequence-based database for population analysis. Different genotypes identified by PFGE underwent species identification using a 16S rRNA gene multiplex PCR. Genotypes of M. intracellulare were confirmed by internal transcribed spacer 1 (ITS1) sequencing and characterized using seven VNTR primers. The pattern of VNTR amplicon sizes and repeat number defined each specific VNTR type. Forty-two VNTR types were identified among 84 genotypes. PFGE revealed most isolates with the same VNTR type to be clonal or exhibit similar grouping of bands. Repetitive sequence-based PCR (rep-PCR) showed minimal pattern diversity between VNTR types compared to PFGE. Fingerprinting of relapse isolates from 31 treated patients using VNTR combined with 16S multiplex PCR unambiguously and reliably distinguished different genotypes from the same patient, with results comparable to those of PFGE. VNTR for strain comparison is easier and faster than PFGE, is as accurate as PFGE, and does not require sequencing. Starting with a collection of 167 M. intracellulare isolates, VNTR distinguished M. intracellulare into 42 clonal groups. Comparison of isolates from different geographic areas, habitats, and clinical settings is now possible.


Asunto(s)
Genotipo , Repeticiones de Minisatélite , Complejo Mycobacterium avium/clasificación , Complejo Mycobacterium avium/genética , Infección por Mycobacterium avium-intracellulare/diagnóstico , Alelos , Técnicas de Tipificación Bacteriana , Bases de Datos de Ácidos Nucleicos , Humanos , Datos de Secuencia Molecular , Tipificación de Secuencias Multilocus , ARN Ribosómico 16S
20.
J Clin Microbiol ; 51(6): 1747-52, 2013 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-23536397

RESUMEN

Recent studies have shown that respiratory isolates from pulmonary disease patients and household water/biofilm isolates of Mycobacterium avium could be matched by DNA fingerprinting. To determine if this is true for Mycobacterium intracellulare, household water sources for 36 patients with Mycobacterium avium complex (MAC) lung disease were evaluated. MAC household water isolates from three published studies that included 37 additional MAC respiratory disease patients were also evaluated. Species identification was done initially using nonsequencing methods with confirmation by internal transcribed spacer (ITS) and/or partial 16S rRNA gene sequencing. M. intracellulare was identified by nonsequencing methods in 54 respiratory cultures and 41 household water/biofilm samples. By ITS sequencing, 49 (90.7%) respiratory isolates were M. intracellulare and 4 (7.4%) were Mycobacterium chimaera. In contrast, 30 (73%) household water samples were M. chimaera, 8 (20%) were other MAC X species (i.e., isolates positive with a MAC probe but negative with species-specific M. avium and M. intracellulare probes), and 3 (7%) were M. avium; none were M. intracellulare. In comparison, M. avium was recovered from 141 water/biofilm samples. These results indicate that M. intracellulare lung disease in the United States is acquired from environmental sources other than household water. Nonsequencing methods for identification of nontuberculous mycobacteria (including those of the MAC) might fail to distinguish closely related species (such as M. intracellulare and M. chimaera). This is the first report of M. chimaera recovery from household water. The study underscores the importance of taxonomy and distinguishing the many species and subspecies of the MAC.


Asunto(s)
Complejo Mycobacterium avium/clasificación , Complejo Mycobacterium avium/aislamiento & purificación , Tuberculosis Pulmonar/microbiología , Microbiología del Agua , Biopelículas , ADN Bacteriano/química , ADN Bacteriano/genética , ADN Ribosómico/química , ADN Ribosómico/genética , ADN Espaciador Ribosómico/química , ADN Espaciador Ribosómico/genética , Composición Familiar , Humanos , ARN Ribosómico 16S/genética , Análisis de Secuencia de ADN
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