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Spoligotyping, genotyping, and spatial distribution of Mycobacterium bovis in cattle in the state of Bahia, Brazil
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Santos, Evelin S.V.

; Alzamora Filho, Fernando

; Santos, Bruno R.

; Silva, Alana V.

; Gomes, Harrison M.

; Suffys, Philip N.

; Costa, Joselito N.

Pesquisa Veterinária Brasileira

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2021, Volume 41 elocation e06729

Abstract: Pt En | Text: Pt En | PDF: Pt | PDF: En

RESUMO: O Mycobacterium bovis é o responsável pela tuberculose bovina e bubalina, doença zoonótica importante e com distribuição global. O conhecimento da distribuição e a identificação precisa dessa enfermidade, incluindo diagnósticos mais avançados como o spoligotyping, permite escolher as melhores estratégias de combate ao avanço da doença. O presente trabalho objetivou investigar a presença de micobactérias, genotipar suas estirpes e avaliar a distribuição espacial dos casos de tuberculose a partir de lesões sugestivas nas carcaças de bovinos e bubalinos inspecionadas em frigoríficos sob regime de inspeção oficial no estado da Bahia. Foram investigados 453.417 animais dentre os quais 31 (0,007%) foram suspeitos de doença e provenientes de 17 municípios. Após o crescimento em meio de cultura, 95% foram categorizados como bacilos álcool-ácido resistentes (BAAR). Todos os isolados foram submetidos à spoligotyping e 95% foram confirmados M. bovis (SB0120, SB0121, SB0852, SB0828, SB0295, SB0881, SB1648, SB6119, SB0140, SB1055). A cepa SB0120 foi a mais prevalente e este perfil vem sendo descrito na literatura com casos de tuberculose humana por M. bovis ressaltando o potencial zoonótico deste perfil. Este estudo também identificou cepas nunca relatadas no estado da Bahia, destacando um padrão distinto de outras partes do Brasil, além da existência de infecções mistas. Permitiu ainda relatar linhagens nunca antes descritas no estado com destaque para um padrão novo no Brasil (SB6119 e SB0852 respectivamente). Um perfil inédito identificado foi identificado e inserido no banco de dados internacional (Mbovis.org), nomeado SB2715.

ABSTRACT: Mycobacterium bovis is responsible for bovine and buffalo tuberculosis, an important zoonotic disease with global distribution. The knowledge of the distribution and the precise identification of this disease, including advanced diagnoses such as spoligotyping, allows choosing the best strategies to fight the disease’s progress. The present work aimed to investigate mycobacteria’s presence, genotype their strains, and evaluate tuberculosis cases’ spatial distribution from suggestive lesions in carcasses of bovine and buffalo inspected in slaughterhouses under an official inspection regime in the state of Bahia, Brazil. The study investigated 453,417 animals. Among these, 31 (0.007%) from 17 municipalities were suspected of tuberculosis. Among the culture medium growth, 95% of these were categorized as alcohol-acid resistant bacilli (BAAR). All isolates were subjected to spoligotyping and 95% were confirmed as M. bovis (SB0120, SB0121, SB0852, SB0828, SB0295, SB0881, SB1648, SB6119, SB0140, SB1055). The strain SB0120 was the most prevalent, and this profile has been described in cases of human tuberculosis by M. bovis, highlighting the zoonotic potential of this profile. This study also identified strains never reported in Bahia, highlighting a distinctive pattern from other parts of Brazil, besides mixed infections. Besides, to identify strains never before described in the state, highlighting a distinctive pattern in Brazil (SB6119 and SB0852, respectively). An unpublished profile was identified and inserted in the international database (Mbovis.org), named SB2715.

https://doi.org/10.1590/1678-5150-pvb-6729 197 downloads

Whole genome sequence of Mycobacterium kansasii isolates of the genotype 1 from Brazilian patients with pulmonary disease demonstrates considerable heterogeneity
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Machado, Edson

; Vasconcellos, Sidra Ezidio Gonçalves

; Cerdeira, Camillo

; Gomes, Lia Lima

; Junqueira, Ricardo

; Carvalho, Luciana Distasio de

; Ramos, Jesus Pais

; Redner, Paulo

; Campos, Carlos Eduardo Dias

; Caldas, Paulo Cesar de Souza

; Gomes, Ana Paula Chaves Sobral

; Goldenberg, Telma

; Montes, Fatima Fandinho

; Mello, Fernanda Carvalho de Queiroz

; Mussi, Vinicius de Oliveira

; Lasunskaia, Elena

; Soolingen, Dick van

; Miranda, Antonio Basílio de

; Rigouts, Leen

; Jong, Bouke C de

; Meehan, Conor J

; Catanho, Marcos

; Suffys, Philip N

Memórias do Instituto Oswaldo Cruz

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2018, Volume 113 N. 9 elocation e180085

Abstract: En | Text: En | PDF: En

Mycobacterium kansasii is an opportunistic pathogen and one of the most commonly encountered species in individuals with lung disease. We here report the complete genome sequence of 12 clinical isolates of M. kansasii from patients with pulmonary disease in Brazil.

https://doi.org/10.1590/0074-02760180085 1130 downloads

Direct detection of Mycobacterium tuberculosis complex in bovine and bubaline tissues through nested-PCR
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Araújo, Cristina P.

; Osório, Ana Luiza A.R.

; Jorge, Klaudia S.G.

; Ramos, Carlos A.N.

; Souza Filho, Antonio F.

; Vidal, Carlos E.S.

; Vargas, Agueda P.C.

; Roxo, Eliana

; Rocha, Adalgiza S.

; Suffys, Philip N.

; Fonseca Júnior, Antônio A.

; Silva, Marcio R.

; Barbosa Neto, José D.

; Cerqueira, Valíria D.

; Araújo, Flábio R.

Brazilian Journal of Microbiology

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jun 2014, Volume 45 N. 2 Pages 633 - 640

Abstract: En | Text: En | PDF: En

Post-mortem bacterial culture and specific biochemical tests are currently performed to characterize the etiologic agent of bovine tuberculosis. Cultures take up to 90 days to develop. A diagnosis by molecular tests such as PCR can provide fast and reliable results while significantly decreasing the time of confirmation. In the present study, a nested-PCR system, targeting rv2807, with conventional PCR followed by real-time PCR, was developed to detect Mycobacterium tuberculosis complex (MTC) organisms directly from bovine and bubaline tissue homogenates. The sensitivity and specificity of the reactions were assessed with DNA samples extracted from tuberculous and non-tuberculous mycobacteria, as well as other Actinomycetales species and DNA samples extracted directly from bovine and bubaline tissue homogenates. Regarding the analytical sensitivity, DNA of the M. bovis AN5 strain was detected up to 1.5 pg by nested-PCR, whereas DNA of M. tuberculosis H37Rv strain was detected up to 6.1 pg. The nested-PCR system showed 100% analytical specificity for MTC when tested with DNA of reference strains of non-tuberculous mycobacteria and closely-related Actinomycetales. A clinical sensitivity level of 76.7% was detected with tissues samples positive for MTC by means of the culture and conventional PCR. A clinical specificity of 100% was detected with DNA from tissue samples of cattle with negative results in the comparative intradermal tuberculin test. These cattle exhibited no visible lesions and were negative in the culture for MTC. The use of the nested-PCR assay to detect M. tuberculosis complex in tissue homogenates provided a rapid diagnosis of bovine and bubaline tuberculosis.

2540 downloads

Múltiplas estirpes de isolados de Mycobacteriumbovis identificados por tipagem molecular em bovinos abatidos em matadouros-frigoríficos
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Alzamora Filho, Fernando

; Vasconcellos, Sidra E.G.

; Gomes, Harrison M.

; Cavalcante, Miriam P.

; Suffys, Philip N.

; Costa, Joselito N.

Pesquisa Veterinária Brasileira

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feb 2014, Volume 34 N. 2 Pages 103 - 108

Abstract: Pt En | Text: Pt En | PDF: Pt | PDF: En

O objetivo do presente trabalho foi utilizar métodos bacteriológicos e moleculares para a identificação do Mycobacteriumbovis em lesões observadas em carcaças de bovinos durante a inspeção postmortem de rotina em matadouros-frigoríficos com serviço de inspeção oficial. Foi acompanhado o abate e a inspeção de 825.394 bovinos, sadios ao exame ante mortem pelo serviço de inspeção oficial em dez matadouros-frigoríficos do estado da Bahia. Carcaça de 180 bovinos apresentaram lesões sugestivas de tuberculose e por outras linfadenites. No isolamento bacteriano, 25 amostras apresentaram crescimento disgônico de colônias de coloração creme-amareladas em meio de cultura Stonebrink-Leslie. Desses isolados, 14 foram identificados como M. bovis PCR multiplex e pela técnica do spoligotyping foram discriminados oito diferentes espoligotipos do M. bovis, sendo sete descritos na literatura e um novo spoligotipo sem descrição anterior. O espoligotipo majoritário foi o SB0121, com cinco amostras, sendo descrito no Brasil e em outros países, seguidos por dois clusters, SB295 e SB1055, com dois isolados cada. O espoligotipo SB1145 e SB1648 foram referidos apenas no Brasil e Dinamarca, respectivamente. O espoligotipo SB140 já foi encontrado no Brasil, Argentina, Uruguai e Paraguai. Estes resultados demonstram que os espoligotipos obtidos são compartilhados, até o momento, entre estados brasileiros e entre países da América Latina e Europa. Sendo assim, a discriminação molecular de isolados de M. bovis através do Spoligotyping constitui-se numa ferramenta para estudos epidemiológicos da tuberculose bovina no Estado da Bahia.

The aim of this study was to use bacteriological and molecular methods to identify Mycobacteriumbovis in lesions observed in cattle carcasses during routine post-mortem inspection in slaughterhouses with official inspection service. It was accompanied the slaughter and inspection of 825,394 cattle, healthy ante mortem examination by the official inspection service in ten slaughterhouses in the state of Bahia. Carcasses of 180 cattle presented lesions suggestive of tuberculosis and other lymphadenitis. In bacterial isolation, 25 samples showed dysgonic growth of colonies of creamy-yellow in medium-Stonebrink Leslie. From these isolates, 14 were identified as M. bovis and the multiplex PCR technique spoligotyping was discriminated against eight different spoligotypes of M. bovis, seven previously described in the literature and a new spoligotypes without former description. The major spoligotypes was SB0121, with five samples which has been described in Brazil and other countries, followed by two clusters, SB295 and SB1055, with two isolates each. The SB1145 and SB1648 spoligotypes were reported only in Brazil and Denmark, respectively. The spoligotypes SB140 has been found in Brazil, Argentina, Uruguay and Paraguay. These results demonstrate that the spoligotypes obtained are shared, so far, among Brazilian states and among Latin America and Europe. Thus, molecular discrimination of isolates of M. bovis by Spoligotyping constitutes a tool for epidemiological studies of bovine tuberculosis in the state of Bahia.

2713 downloads

Mycobacterium tuberculosis strains of the Beijing genotype are rarely observed in tuberculosis patients in South America
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Ritacco, Viviana

; López, Beatriz

; Cafrune, Patricia I

; Ferrazoli, Lucilaine

; Suffys, Philip N

; Candia, Norma

; Vásquez, Lucy

; Realpe, Teresa

; Fernández, Jorge

; Lima, Karla V

; Zurita, Jeannete

; Robledo, Jaime

; Rossetti, Maria L

; Kritski, Afranio L

; Telles, Maria A

; Palomino, Juan C

; Heersma, Herre

; van Soolingen, Dick

; Kremer, Kristin

; Barrera, Lucía

Memórias do Instituto Oswaldo Cruz

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aug 2008, Volume 103 N. 5 Pages 489 - 492

Abstract: En | Text: En | PDF: En

The frequency of the Beijing genotype of Mycobacterium tuberculosis as a cause of tuberculosis (TB) in South America was determined by analyzing genotypes of strains isolated from patients that had been diagnosed with the disease between 1997 and 2003 in seven countries of the subcontinent. In total, 19 of the 1,202 (1.6%) TB cases carried Beijing isolates, including 11 of the 185 patients from Peru (5.9%), five of the 512 patients from Argentina (1.0%), two of the 252 Brazilian cases (0.8%), one of the 166 patients from Paraguay (0.6%) and none of the samples obtained from Chile (35), Colombia (36) and Ecuador (16). Except for two patients that were East Asian immigrants, all cases with Beijing strains were native South Americans. No association was found between carrying a strain with the Beijing genotype and having drug or multi-drug resistant disease. Our data show that presently transmission of M. tuberculosis strains of the Beijing genotype is not frequent in Latin America. In addition, the lack of association of drug resistant TB and infection with M. tuberculosis of the Beijing genotype observed presently demands efforts to define better the contribution of the virulence and lack of response to treatment to the growing spread of Beijing strains observed in other parts of the world.

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Typing of Mycobacterium tuberculosis strains isolated in Community Health Centers of Rio de Janeiro City, Brazil
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Oelemann, Maraníbia C

; Fontes, Amanda NB

; Pereira, Marcia A da Silva

; Bravin, Yolanda

; Silva, Guida

; Degrave, Wim

; Carvalho, Anna Cristina

; Brito, Rossana C

; Kritski, Afrânio L

; Suffys, Philip N

Memórias do Instituto Oswaldo Cruz

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jun 2007, Volume 102 N. 4 Pages 455 - 462

Abstract: En | Text: En | PDF: En

Fingerprinting of Mycobacterium tuberculosis strains from tuberculosis (TB) patients attended in Community Health Centers (CHCs) of Rio de Janeiro was performed to verify possible risk factors for TB transmission. A prospective community-based study was performed during the period of July 1996 to December 1996 by collecting sputum samples of 489 patients in 11 different CHCs in four different planning areas (APs) of the city. Bacteriological, clinical, and epidemiological information was collected and M. tuberculosis genotypes defined after restriction fragment length polymorphism (IS6110-RFLP) and double repetitive element (DRE) fingerprinting of RFLP-clustered cases. Risk factors for TB transmission were looked for using three levels of cluster stringency. Among 349 (71%) positive cultures obtained, IS6110-RFLP typing could be performed on strains from 153 different patients. When using identity of RFLP patterns as cluster definition, 49 (32%) of the strains belonged to a cluster and none of the clinical or epidemiologic characteristics was associated with higher clustering levels. However, higher clustering level was observed in the AP including the central region of the city when compared to others. This strongly suggests that more recent transmission occurs in that area and this may be related with higher incidence of TB and HIV in this region.

1846 downloads

Drug susceptibility of Brazilian strains of Mycobacterium bovis using traditional and molecular techniques
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Parreiras, Patrícia M

; Lobato, Francisco CF

; Alencar, Andréa P

; Figueiredo, Telma de

; Gomes, Harrison M

; Boéchat, Neio

; Lage, Andrey P

; Assis, Ronnie A

; Pereira, Márcia AS

; Souza, Patrícia R

; Mota, Pedro MPC

; Suffys, Philip N

Memórias do Instituto Oswaldo Cruz

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nov 2004, Volume 99 N. 7 Pages 749 - 752

Abstract: En | Text: En | PDF: En

Transmission of Mycobacterium bovis from cattle to humans has been reported and can cause tuberculosis (Tb) and a problem in certain risk populations. Therefore, knowledge of resistance of M. bovis towards antibiotics used for therapy of human Tb could help avoiding cure delay and treatment cost increase when dealing with drug resistant organisms. We therefore evaluated the susceptibility of M. bovis isolates towards streptomycin, isoniazide, rifampicin, ethambutol, and ethionamide, the first line antibiotics for human Tb. Therefore, 185 clinical samples from cattle with clinical signs of tuberculosis were processed and submitted to culturing and bacterial isolates to identification and drug susceptibility testing using the proportion method. Among 89 mycobacterial strains, 65 were identified as M. bovis and none were resistant to any of the antibiotics used. Confirmation of present results by future studies, enrolling a large number of isolates and designed to properly represent Brazilian regions, may favor the idea of using isoniazide preventive therapy as part of a Tb control strategy in special situations. Also, nucleic acids from bacterial isolates were submitted to rifoligotyping, a recently described reverse hybridization assay for detection of mutations causing resistance towards rifampicin. Concordance between the conventional and the molecular test was 100%, demonstrating the use of such methodology for rapid evaluation of drug susceptibility in M. bovis.

2353 downloads

The Changing Face of the Epidemiology of Tuberculosis due to Molecular Strain Typing: A Review
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Suffys, Philip N

; Araujo, Marcelo E Ivens de

; Degrave, Wim M

Memórias do Instituto Oswaldo Cruz

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may 1997, Volume 92 N. 3 Pages 297 - 316

Abstract: En | Text: En | PDF: En

About one third of the world population is infected with tubercle bacilli, causing eight million new cases of tuberculosis (TB) and three million deaths each year. After years of lack of interest in the disease, World Health Organization recently declared TB a global emergency and it is clear that there is need for more efficient national TB programs and newly defined research priorities. A more complete epidemiology of tuberculosis will lead to a better identification of index cases and to a more efficient treatment of the disease. Recently, new molecular tools became available for the identification of strains of Mycobacterium tuberculosis (M. tuberculosis), allowing a better recognition of transmission routes of defined strains. Both a standardized restriction-fragment-length-polymorphism-based methodology for epidemiological studies on a large scale and deoxyribonucleic acids (DNA) amplification-based methods that allow rapid detection of outbreaks with multidrug-resistant (MDR) strains, often characterized by high mortality rates, have been developed. This review comments on the existing methods of DNA-based recognition of M. tuberculosis strains and their peculiarities. It also summarizes literature data on the application of molecular fingerprinting for detection of outbreaks of M. tuberculosis, for identification of index cases, for study of interaction between TB and infection with the human immunodeficiency virus, for analysis of the behavior of MDR strains, for a better understanding of risk factors for transmission of TB within communities and for population-based studies of TB transmission within and between countries

2455 downloads

Use of PCR for the determination of the frequency of the deltaF508 mutation in Brasilian cistic fibrosis patients
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Miranda, Antonio B. de

; LLerena Junior, Juan

; Dallalana, Ludma T.

; Moura-Neto, Rodrigo S.

; Suffys, Philip N.

; Degrave, Wim M.

Memórias do Instituto Oswaldo Cruz

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jun 1993, Volume 88 N. 2 Pages 309 - 312

Abstract: En | Text: En | PDF: En

The [Delta]F508 mutation in the cystic fibrosis (CF) gene was studied in a population of 18 Brazilian CF patients and their 17 families by use of PCR and differential hybridization with oligonucleotides. In a total of 34 chromosomes considered, 12 (35%) carried the F508 deletion, a frequency much lower than that reported in most other populations. As a consequence, CF in Brazil would be predominantly caused by mutations different from the F508 deletion

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