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Molecular-typing can help in unraveling epidemiological scenarios and improvement for disease control strategies. A literature review of Mycobacterium tuberculosis transmission in Brazil through genotyping on 56 studies published from 1996-2019 was performed. The clustering rate for mycobacterial interspersed repetitive units - variable tandem repeats (MIRU-VNTR) of 1,613 isolates were: 73%, 33% and 28% based on 12, 15 and 24-loci, respectively; while for RFLP-IS6110 were: 84% among prison population in Rio de Janeiro, 69% among multidrug-resistant isolates in Rio Grande do Sul, and 56.2% in general population in São Paulo. These findings could improve tuberculosis (TB) surveillance and set up a solid basis to build a database of Mycobacterium genomes.

https://doi.org/10.1590/0074-02760200517 744 downloads

Evaluation of drug susceptibility profile of Mycobacterium tuberculosis Lineage 1 from Brazil based on whole genome sequencing and phenotypic methods
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Guimarães, Arthur Emil dos Santos

; Sharma, Abhinav

; Furlaneto, Ismari Perini

; Rutaihwa, Liliana

; Cardoso, Jedson Ferreira

; da Conceição, Marília Lima

; Spinassé, Lizânia Borges

; Machado, Edson

; Lopes, Maria Luiza

; Duarte, Rafael Silva

; Gagneux, Sebastien

; Suffys, Philip Noel

; Lima, Karla Valéria Batista

; Conceição, Emilyn Costa

Memórias do Instituto Oswaldo Cruz

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2020, Volume 115 elocation e200520

Resumo: En | Texto: En | PDF: En

BACKGROUND The evaluation of procedures for drug susceptibility prediction of Mycobacterium tuberculosis based on genomic data against the conventional reference method test based on culture is realistic considering the scenario of growing number of tools proposals based on whole-genome sequences (WGS). OBJECTIVES This study aimed to evaluate drug susceptibility testing (DST) outcome based on WGS tools and the phenotypic methods performed on isolates of M. tuberculosis Lineage 1 from the state of Pará, Brazil, generally associated with low levels of drug resistance. METHODOLOGY Culture based DST was performed using the Proportion Method in Löwenstein-Jensen medium on 71 isolates that had been submitted to WGS. We analysed the seven main genome sequence-based tools for resistance and lineage prediction applied to M. tuberculosis and for comparison evaluation we have used the Kappa concordance test. FINDINGS When comparing the WGS-based tools against the DST, we observed the highest level of agreement using TB-profiler. Among the tools, TB-profiler, KvarQ and Mykrobe were those which identified the largest number of TB-MDR cases. Comparing the four most sensitive tools regarding resistance prediction, agreement was observed for 43 genomes. MAIN CONCLUSIONS Drug resistance profiling using next-generation sequencing offers rapid assessment of resistance-associated mutations, therefore facilitating rapid access to effective treatment.

https://doi.org/10.1590/0074-02760200520 427 downloads

Rapid detection of Mycobacterium tuberculosis in children using blood and urine specimens
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Costa-Lima, Juliana Figueirêdo da

; Pimentel, Lílian Maria Lapa Montenegro

; Santos, Fabiana Cristina Fulco

; Salazar, Marcela Pereira

; Duarte, Rafael Silva

; Mello, Fernanda Carvalho de Queiroz

; Schindler, Haiana Charifker

Revista da Sociedade Brasileira de Medicina Tropical

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2020, Volume 53 elocation e20200051

Resumo: En | Texto: En | PDF: En

Abstract INTRODUCTION: Laboratory and clinical features of childhood tuberculosis (TB) are non-specific and establishing an accurate diagnosis remains a challenge. This study evaluated a Single tube nested-PCR (STNPCR) to detect genomic DNA of Mycobacterium tuberculosis complex in blood and urine. METHODS: Biological samples were obtained from children (<15 years old) with clinical suspicion of pulmonary and extrapulmonary TB at public hospitals in Recife-Pernambuco, Brazil. Cultures yielded negative results in a majority of childhood TB cases, which are generally paucibacillary. A set of clinical, epidemiological, radiological, and laboratory criteria with evident clinical improvement after anti-TB treatment were frequently used to define childhood TB cases. RESULTS: Ninety children with clinical suspicion were enrolled in this study (44 with TB and 46 without TB). The pulmonary TB group had 20 confirmed cases and 46 negative controls, while the extrapulmonary TB group had 24 confirmed cases. The STNPCR showed sensitivities to pulmonary and extrapulmonary TB of 47.4% and 52.2% (blood) and 38.8% and 20% (urine), respectively. Considering the low performance of STNPCR on separate samples, we decided to perform a combined analysis (parallel sensitivity analysis) of the results from blood and urine samples. The parallel sensitivity increased to 65% in blood and 62.5% in urine. The specificity in both samples ranged from 93.5-97.8%. CONCLUSIONS: Although STNPCR showed moderate sensitivity, the specificity is high; therefore, the test can be used as an auxiliary tool to diagnose TB in children. It is a rapid test that demonstrated better performance than other diagnostic tests in paucibacillary samples as it does in childhood tuberculosis.

https://doi.org/10.1590/0037-8682-0051-2020 474 downloads

Newly sequenced genomes of four Bacillus Calmette Guerin vaccines
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Sisco, Maria Carolina

; Silva, Marlei Gomés

; Lopez, Beatriz

; Arguelles, Claudia

; Mendonça-Lima, Leila

; de Waard, Jacobus H

; Duarte, Rafael Silva

; Suffys, Philip Noel

Memórias do Instituto Oswaldo Cruz

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2020, Volume 115 elocation e190401

Resumo: En | Texto: En | PDF: En

Bacillus Calmette Guerin (BCG) vaccines comprise a family of related strains. Whole genome sequencing has allowed the better characterisation of the differences between many of the BCG vaccines. As sequencing technologies improve, updating of publicly available sequence data becomes common practice. We hereby announce the draft genome of four commonly used BCG vaccines in Brazil, Argentina and Venezuela.

https://doi.org/10.1590/0074-02760190401 669 downloads

Resistance profile of strains of Mycobacterium fortuitum isolated from clinical specimens
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Santos, Debora Ribeiro de Souza

; Lourenço, Maria Cristina Silva

; Coelho, Fábrice Santana

; Mello, Fernanda Carvalho Queiroz

; Duarte, Rafael Silva

Jornal Brasileiro de Pneumologia

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Ago 2016, Volume 42 Nº 4 Páginas 299 - 301

Resumo: Pt En | Texto: Pt En | PDF: Pt | PDF: En

https://doi.org/10.1590/s1806-37562016000000073 2981 downloads

Rapid detection and differentiation of mycobacterial species using a multiplex PCR system
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Lima, Andrea Santos

; Duarte, Rafael Silva

; Montenegro, Lilian Maria Lapa

; Schindler, Haiana Charifker

Revista da Sociedade Brasileira de Medicina Tropical

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Jul 2013, Volume 46 Nº 4 Páginas 447 - 452

Resumo: En | Texto: En | PDF: En

Introduction The early diagnosis of mycobacterial infections is a critical step for initiating treatment and curing the patient. Molecular analytical methods have led to considerable improvements in the speed and accuracy of mycobacteria detection. Methods The purpose of this study was to evaluate a multiplex polymerase chain reaction system using mycobacterial strains as an auxiliary tool in the differential diagnosis of tuberculosis and diseases caused by nontuberculous mycobacteria (NTM) Results Forty mycobacterial strains isolated from pulmonary and extrapulmonary origin specimens from 37 patients diagnosed with tuberculosis were processed. Using phenotypic and biochemical characteristics of the 40 mycobacteria isolated in LJ medium, 57.5% (n=23) were characterized as the Mycobacterium tuberculosis complex (MTBC) and 20% (n=8) as nontuberculous mycobacteria (NTM), with 22.5% (n=9) of the results being inconclusive. When the results of the phenotypic and biochemical tests in 30 strains of mycobacteria were compared with the results of the multiplex PCR, there was 100% concordance in the identification of the MTBC and NTM species, respectively. A total of 32.5% (n=13) of the samples in multiplex PCR exhibited a molecular pattern consistent with NTM, thus disagreeing with the final diagnosis from the attending physician. Conclusions Multiplex PCR can be used as a differential method for determining TB infections caused by NTM a valuable tool in reducing the time necessary to make clinical diagnoses and begin treatment. It is also useful for identifying species that were previously not identifiable using conventional biochemical and phenotypic techniques.

https://doi.org/10.1590/0037-8682-0097-2013 5312 downloads Citado 1 vez em SciELO

Rapid tests for the detection of the Mycobacterium abscessus subsp. bolletii strain responsible for an epidemic of surgical-site infections in Brazil
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Matsumoto, Cristianne Kayoko

; Chimara, Erica

; Ramos, Jesus Pais

; Campos, Carlos Eduardo Dias

; Caldas, Paulo Cesar de Souza

; Lima, Karla Valeria Batista

; Lopes, Maria Luiza

; Duarte, Rafael Silva

; Leão, Sylvia Cardoso

Memórias do Instituto Oswaldo Cruz

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Dez 2012, Volume 107 Nº 8 Páginas 969 - 977

Resumo: En | Texto: En | PDF: En

A single strain of Mycobacterium abscessus subsp. bolletii, characterised by a particular rpoB sequevar and two highly related pulsed field gel electrophoresis patterns has been responsible for a nationwide outbreak of surgical infections in Brazil since 2004. In this study, we developed molecular tests based on polymerase chain reaction restriction-enzyme analysis (PRA) and sequencing for the rapid identification of this strain. Sequences of 15 DNA regions conserved in mycobacteria were retrieved from GenBank or sequenced and analysed in silico. Single nucleotide polymorphisms specific to the epidemic strain and located in enzyme recognition sites were detected in rpoB, the 3' region of the 16S rDNA and gyrB. The three tests that were developed, i.e., PRA-rpoB, PRA-16S and gyrB sequence analysis, showed 100%, 100% and 92.31% sensitivity and 93.06%, 90.28% and 100% specificity, respectively, for the discrimination of the surgical strain from other M. abscessus subsp. bolletii isolates, including 116 isolates from 95 patients, one environmental isolate and two type strains. The results of the three tests were stable, as shown by results obtained for different isolates from the same patient. In conclusion, due to the clinical and epidemiological importance of this strain, these tests could be implemented in reference laboratories for the rapid preliminary diagnosis and epidemiological surveillance of this epidemic strain.

2577 downloads Citado 1 vez em SciELO

Molecular characterization of the first fluoroquinolone resistant strains of Streptococcus agalactiae isolated in Brazil
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Barros, Rosana Rocha

; Kegele, Fabíola Cristina Oliveira

; Paula, Geraldo Renato de

; Brito, Monique Araújo de

; Duarte, Rafael Silva

Brazilian Journal of Infectious Diseases

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Out 2012, Volume 16 Nº 5 Páginas 476 - 478

Resumo: En | Texto: En | PDF: En

Three isolates of Streptococcus agalactiae, recovered from residents of the metropolitan area of Rio de Janeiro with significant bacteriuria, were found to be resistant to levofloxacin. Determination of the minimal inhibitory concentration (MIC) confirmed one isolate as intermediate and two as resistant to levofloxacin. No reduction in levofloxacin MIC was observed with reserpine, indicating that resistance was not caused by an efflux mechanism. Typical point mutations were observed in the quinolone resistance determinant region of gyrA and parC. Other point mutations in parC generated novel altered codons: Ser80→Pro in the intermediate resistance isolate, and Gly128→Asp in a resistant isolate. Through molecular modeling, it was possible to observe that these novel substitutions might not play a role in resistance, since these amino acids were not involved in the antibiotic binding site. Pulsed field gel electrophoresis profiles revealed a non-clonal trend among these isolates. This is the first report of genetic characterization of levofloxacin-resistant S. agalactiae strains in Brazil.

https://doi.org/10.1016/j.bjid.2012.05.003 1549 downloads

Molecular identification and typing of Mycobacterium massiliense isolated from postsurgical infections in Brazil
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Monego, Fernanda

; Duarte, Rafael Silva

; Nakatani, Sueli Massumi

; Araújo, Wildo Navegantes

; Riediger, Irina Nastassja

; Brockelt, Sonia

; Souza, Verena

; Cataldo, Jamyra Iglesias

; Dias, Rubens Clayton da Silva

; Biondo, Alexander Welker

Brazilian Journal of Infectious Diseases

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Out 2011, Volume 15 Nº 5 Páginas 436 - 441

Resumo: En | Texto: En | PDF: En

OBJECTIVE: One hundred thirty-one cases of postsurgical infections were reported in Southern Region of Brazil between August 2007 and January 2008. Thirty-nine (29.8%) cases were studied; this report describes epidemiological findings, species identification, antimicrobial susceptibility and clonal diversity of rapidly growing mycobacteria isolated in this outbreak. METHODS: All 39 isolates were analyzed by Ziehl-Nielsen stained smear, bacterial culture and submitted to rpoB partial gene sequencing for identification. The isolates were also evaluated for their susceptibility to amikacin, cefoxitin, clarithromycin, ciprofloxacin, doxycycline, tobramycin and sulfamethoxazole. RESULTS: Thirty-six isolates out of the confirmed cases were identified as Mycobacterium massilienseand the remaining three were identified as Mycobacterium abscessus, Mycobacterium chelonae and Mycobacterium fortuitum. All M. massiliense isolates were susceptible to amikacin (MIC90 = 8 µg/mL) and clarithromycin (MIC90 = 0.25 µg/mL) but resistant to cefoxitin, ciprofloxacin, doxycycline, tobramycin and sulfamethoxazole. Molecular analysis by pulsed-field gel electrophoresis clustered all 36 M. massiliense isolates and showed the same pattern (BRA 100) observed in three other outbreaks previously reported in Brazil. CONCLUSIONS: These findings suggest a common source of infection for all patients and reinforce the hypotheses of spread of M. massiliense BRA100 in Brazilian hospital surgical environment in recent years.

3134 downloads Citado 1 vez em SciELO

10.

Identificação de Mycobacterium bovis em cepas micobacterianas isoladas espécimes clínicos humanos em um complexo hospitalar na cidade do Rio de Janeiro
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Sobral, Luciana Fonseca

; Duarte, Rafael Silva

; Vieira, Gisele Betzler de Oliveira

; Silva, Marlei Gomes da

; Boechat, Neio

; Fonseca, Leila de Souza

Jornal Brasileiro de Pneumologia

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Out 2011, Volume 37 Nº 5 Páginas 664 - 668

Resumo: Pt En | Texto: Pt En | PDF: Pt | PDF: En

Entre 2005 e 2006, 8.121 espécimes clínicos enviados ao Laboratório de Micobactérias do Hospital Universitário Clementino Fraga Filho/Instituto de Doenças do Tórax, no Rio de Janeiro, RJ, foram inoculados em meio Löwenstein-Jensen contendo glicerol e piruvato. Desses espécimes, 79 isolados de micobactérias tiveram crescimento somente em meio com piruvato, sendo selecionados para a identificação presuntiva de Mycobacterium bovis. Esses isolados foram submetidos à identificação por testes bioquímicos, amplificação por PCR com primers específicos (Rv0577 e Rv1510) e teste de suscetibilidade à pirazinamida. Todas as cepas apresentaram padrões fenotípicos e genotípicos de M. tuberculosis, não sendo detectado M. bovis.

In 2005 and 2006, 8,121 clinical specimens submitted to the Mycobacteriology Laboratory of the Clementino Fraga Filho University Hospital/Thoracic Diseases Institute, in the city of Rio de Janeiro, Brazil, were inoculated on Löwenstein-Jensen medium containing glycerol and pyruvate. There were 79 mycobacteria isolates that presented growth only on pyruvate-containing medium, and those isolates were selected for the presumptive identification of Mycobacterium bovis. The selected isolates were screened with biochemical tests, PCR amplification (with the specific primers Rv0577 and Rv1510), and pyrazinamide susceptibility tests. All of the strains isolated showed specific phenotypical and genotypical patterns characteristic of M. tuberculosis, and no M. bovis strains were detected.

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11.

Mycobacterium massiliense BRA100 strain recovered from postsurgical infections: resistance to high concentrations of glutaraldehyde and alternative solutions for high level disinfection
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Lorena, Nádia Suely de Oliveira

; Pitombo, Marcos Bettini

; Côrtes, Patrícia Barbur

; Maya, Maria Cristina Araújo

; Silva, Marlei Gomes da

; Carvalho, Ana Carolina da Silva

; Coelho, Fábrice Santana

; Miyazaki, Neide Hiromi Tokumaru

; Marques, Elizabeth Andrade

; Chebabo, Alberto

; Freitas, Andréa D'Ávila

; Lupi, Otília

; Duarte, Rafael Silva

Acta Cirúrgica Brasileira

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Out 2010, Volume 25 Nº 5 Páginas 455 - 459

Resumo: Pt En | Texto: Pt En | PDF: Pt | PDF: En

OBJETIVO: Avaliar a concentração mínima inibitória (CMI) de GTA frente a M. massiliense e a susceptibilidade a produtos alternativos para desinfecção de alto nível (DAN). MÉTODOS: Cepas de M. massiliense de origem clínica e de referência foram incluídas no estudo. As culturas ativadas foram submetidas a testes qualitativos com diluições de GTA (de 1,5% a 8%) e com soluções comerciais de ortoftaldeído (OPA) ou ácido peracético (PA), utilizando os tempos de exposição recomendados pela Agência Nacional de Vigilância Sanitária para DAN. RESULTADOS: Todas as cepas de referência e M. massiliense não-BRA100, obtida de escarro, foram susceptíveis às concentrações de GTA, e soluções de OPA e PA. As cepas de M. massiliense BRA100 apresentaram CMI de 8% para GTA e foram susceptíveis a OPA e PA. CONCLUSÃO: M. massiliense BRA100 é resistente a altas concentrações de GTA (até 7%), o que demonstra que esse composto não é eficaz, e deve ser substituído por OPA ou PA nos processos de DAN.

PURPOSE: To evaluate the minimum inhibitory concentration (MIC) of GTA against these microorganisms and alternative disinfectants for high-level disinfection (HLD). METHODS: Reference mycobacteria and clinical M. massiliense strains were included in this study. Active cultures were submitted to susceptibility qualitative tests with GTA dilutions (ranging from 1.5% to 8%), and commercial orthophthaldehyde (OPA) and peracetic acid (PA) - based solutions, during the period of exposure as recommended by National Agency of Sanitary Surveillance for HLD. RESULTS: All reference and M. massiliense non-BRA100 strains, recovered from sputum, were susceptible to any GTA concentration, OPA and PA solutions. M. massiliense BRA100 strains presented MIC of 8% GTA and were susceptible to OPA and PA. CONCLUSION: M. massiliense BRA100 strain is resistant to high GTA concentrations (up to 7%), which proves that this product is non-effective against specific rapidly growing mycobacteria and should be substituted by OPA or PA - based solutions for HLD.

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12.

Infecções por micobactérias de crescimento rápido resistentes a desinfetantes: uma problemática nacional?
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Pitombo, Marcos Bettini

; Lupi, Otília

; Duarte, Rafael Silva

Revista Brasileira de Ginecologia e Obstetrícia

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Nov 2009, Volume 31 Nº 11 Páginas 529 - 533

Resumo: Pt En | Texto: Pt En | PDF: Pt | PDF: En

Rapidly growing mycobacteria (RGM) are opportunistic microorganisms and widely distributed into aqueous environment and soil. Human RGM infections are usually associated with contaminated solutions or medical instruments used during invasive procedures. RGM postsurgical infections have recently emerged in Brazil and have caused national alert, considering the risk factors and epidemiological aspects. This study aimed at analysing the main factors linked to the recent RGM outbreaks, with focus on the national epidemic of Mycobacterium massiliense infections related to the BRA100 strains resistant to 2% glutaraldehyde commercial solutions commonly used for preoperative high-level disinfection. Based on previous studies and laboratorial results of assays and colaborations, it has been observed that the cases have been associated with videolaparoscopy for different applications and elective esthetic procedures, such as lipoaspiration and mammary prosthesis implant. Furthermore, outbreaks between 2004 and 2008 and the epidemic in Rio de Janeiro state may be considered particular Brazilian events. Although there are a few epidemiological published studies, some hypotheses based on common aspects related to most national nosocomial occurrences are possible, such as lack of protocols for cleaning and high-level disinfection, use of 2% glutaraldehyde as high-level disinfectant for surgical instruments, and dissemination of M. massiliense BRA100 by unknown mechanisms.

9248 downloads Citado 2 vezes em SciELO

13.

Infecção por micobactérias de crescimento rápido após procedimentos videocirúrgicos - a hipótese do glutaraldeído
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Lorena, Nádia Suely de Oliveira

; Duarte, Rafael Silva

; Pitombo, Marcos Bettini

Revista do Colégio Brasileiro de Cirurgiões

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Jul 2009, Volume 36 Nº 3 Páginas 266 - 267

Resumo: Pt En | Texto: Pt En | PDF: Pt | PDF: En

Between August 2006 and February 2007, in the state of Rio de Janeiro, Brazil, a massive outbreak of RGM infections after video laparoscopy was mainly associated to the recently described Mycobacterium massiliense species. All confirmed and probable cases reports described the use of high-level disinfection of medical devices by using 2% glutaraldehyde (2% GA) for 30 min before the surgical procedures. We investigated the susceptibility of the M. massiliense isolates recovered during the outbreak to high-level disinfection after 30 min, 1h, 6h and 10h of exposure to the commercial disinfectants. Reference strains for official mycobactericidal tests such as Mycobacterium abscessus, Mycobacterium bovis, Mycobacterium chelonae, Mycobacterium neoaurum and Mycobacterium smegmatis were included as controls. Although all the reference strains were eliminated in 30 min of exposure to 2% GA, we observed the recovery of all M. massiliense clinical isolates even after 10h of exposure. This study suggests that failures in high-level disinfection and the high tolerance of these M. massiliense clinical strains to the 2% GA were strongly associated to the magnitude of the outbreak.

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