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Vol. 11. Issue 6.
Pages 533-556 (November - December 2005)
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Vol. 11. Issue 6.
Pages 533-556 (November - December 2005)
Original Article
Open Access
Spoligtyping e polimorfismo do gene pncA: Um cenário em duas etapas para o diagnóstico de Mycobacterium bovis em Portugal
Spoligotyping and pncA polymorphisme: A two step scenario for Mycobacterium bovis diagnosis in Portugal
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Suzana David
,
Corresponding author
suzanadavid@ihmt.unl.pt

Correspondência/Correspondence to: Suzana David, UEI de Micobacteriologia, Instituto de Higiene e Medicina Tropical. Rua da Junqueira, 96; 1349-008 Lisboa, Portugal Tel.: +351-21-3652600; Fax: +351-21-3632105.
, Clara Portugal**, Abílio Antunes***, Ana Calado*, Ângela Cardoso*, Vanessa Barros*, Luísa Sancho**, José Germano de Sousa**
* Unidade de Ensino e Investigação (UEI) de Micobacteriologia, Instituto de Higiene e Medicina Tropical (IHMT)/Universidade Nova de Lisboa (UNL), Lisboa/ Microbacteriology Teaching and Research Unit (TRU), Hygiene and Tropical Medicine Institute (HTMI)/ Nova University Lisbon (NUL), Lisbon.
** Serviço de Patologia Clínica, Hospital Fernando Fonseca, Amadora/Clinical Pathology Department, Fernando Fonseca Hospital, Amadora.
*** UEI de Clínica e Doenças Tropicais/IHMT/UNL, Lisboa/TRU of the Tropical Diseases Clinic, HTMI, NUL, Lisbon.
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Resumo

O diagnóstico diferencial do Mycobacterium bovis é importante no controlo da transmissão para a população humana e também no tratamento da doença, visto que o M. bovis é naturalmente resistente à pirazinamida. Foram analisados onze isolados clínicos do Hospital Fernando Fonseca, com um padrão de Spoligotype indicativo de M. bovis pela ausência dos espaçadores 39-43, mas que também contaram com a ausência do espaçador 38. Para a identificação destas estirpes procedeu-se à análise fenotípica da resistência à pirazinamida e estudo dos polimorfismos dos genes pncA e gyrB. Pelo estudo do polimorfismo do pncA, verificámos que as estirpes analisadas não continham a mutação específica do M. bovis. Em relação aos polimorfismos do gyrB, utilizando o kit GenoType® MTBC, as estirpes foram identificadas como pertencentes ao grupo M. tuberculosis, M. africanum subtipo II e M. canetti. O presente trabalho de investigação permitiu-nos definir novos genótipos sobre os quais deverão apoiar-se estudos bacteriológicos futuros. Entre estes foi desde já considerado importante o estudo do polimorfismo do gene pncA, devido às implicações de utilidade prática e imediata para o clínico. Avaliar a transmissão e definir grupos de risco é um objectivo para o qual se considera importante o apoio dos serviços veterinários.

Rev Port Pneumol 2005; XI (6): 533-556

Palavras-chave:
Complexo Mycobacterium tuberculosis
Mycobacterium bovis
Spoligotyping
kit GenoType® MTBC
pncA
gyrB
pirazinamida
Abstract

The differential diagnosis of Mycobacterium bovis is important in the control of transmission to the human population and also for treatment since M. bovis is naturally resistant to pyrazinamide. Eleven clinical isolates from the Fernando Fonseca Hospital with Spoligotypes indicative of M. bovis, through the absence of spacers 39-43 but that also counted with the absence of spacer 38, were analyzed. For the identification of these strains, the phenotypic analysis of pyrazinamide resistance and study of the polymorphisms of the pncA and gyrB genes were carried out. The study of the pncA polymorphism revealed that the strains analyzed did not contain the M. bovis specific mutation. In relation the gyrB polymorphisms, using the GenoType® MTBC kit, the strains were identified as belonging to the group M. tuberculosis, M. africanum subtipo II e M. canetti. The present investigation enabled us to define new genotypes on which future bacteriological studies should be based. Amongst these the study of the pncA polymorphism was considered important due to the immediate practical implications for the clinician. Evaluating transmission and defining groups of risk is an objective for which support from the veterinary services is considered relevant.

Rev Port Pneumol 2005; XI (6): 533-556

Key-words:
Mycobacterium tuberculosis complex
Mycobacterium bovis
Spoligotyping
kit GenoType® MTBC
pncA
gyrB
pyrazinamide
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Bibliografia/Bibliography
[1.]
C.J. Darbon, J.M. Grange.
HIV/AIDS and its implications for the control of animal tuberculosis.
Br Vet J, 149 (1993), pp. 405-417
[2.]
J. Blazquez, L.E. de Los Monteros, S. Samper, C. Martin, A. Guerrero, J. Cobo, J.D.A. Van Embden, F. Baquero, E. Gomez-Mampaso.
Genetic characterization of multidrug-resistant Mycobacterium bovis strains from a hospital outbreak involving human immunodeficiency virus-positive patients.
J Clin Microbiol, 35 (1997), pp. 1390-1393
[3.]
A. Riviero, M. Márquez, J. Santos, A. Pinedo, M.-A. Sánchez, A. Esteve, S. Samper, C. Martin.
High rate of tuberculosis reinfection during a nosocomial outbreak of multidrug-resistant tuberculosis caused by Mycobacterium bovis strain B.
Clin Infect Dis, 32 (2001), pp. 159-161
[4.]
E.A. Fanning.
Mycobacterium bovis infection in animals and humans.
Clinical tuberculosis, pp. 351-364
[5.]
A.L. Gibson, G. Hewinson, T. Goodchild, B. Watt, A. Story, J. Inwald, F.A. Drobniewski.
Molecular epidemiology of disease to Mycobacterium bovis in humans in the United Kingdom.
J Clin Microbiol, 42 (2004), pp. 431-434
[6.]
J.R. Dalovisio, M. Setter, S. Mikota-Wells.
Rhinoceros rhinorrhea: cause of an outbreak of infection due to airborn Mycobacterium bovis in zookeepers.
Clin Infect Dis, 15 (1992), pp. 598-600
[7.]
T. Kubica, S. Rüsch-Gerdes, S. Niemann.
Mycobacterium bovis subsp. caprae caused one-third of human M. bovis-associated tuberculosis cases reported in Germany between 1999 and 2001.
J Clin Microbiol, 41 (2003), pp. 3070-3077
[8.]
M.P.U. Sales, G.M. Taylor, S. Hughes, M. Yates, G. Hewinson, D.B. Young, R.J. Shaw.
Genetic diversity among Mycobacterium bovis isolates: a preliminary study of strains from animal and human sources.
J Clin Microbiol, 39 (2001), pp. 4558-4562
[9.]
M. Gutiérrez, S. Samper, M.S. Jiménez, J.D.A. Van Embden, J.F. Marín, C. Martín.
Identification by spoligotyping of a caprine genotype in Mycobacterium bovis strains causing human tuberculosis.
J Clin Microbiol, 35 (1997), pp. 3328-3330
[10.]
A. Aranaz, E. Liébana, A. Mateos, L. Dominguez, D. Vidal, M. Domingo, O. Gonzolez, E.F. Rodriguez-Ferri, A.E. Bunschoten, J.D.A. Van Embden, D. Cousins.
Spacer oligonucleotide typing of Mycobacterium bovis strains from cattle and other animals: a tool for studying epidemiology of tuberculosis.
J Clin Microbiol, 34 (1996), pp. 2734-2740
[11.]
P.M.A. Groenen, A.E. Bunschoten, D. Van Soolingen, J.D.A. Van Embden.
Nature of DNA polymorphism in the direct repeat cluster of Mycobacterium tuberculosis; Application for strain differentiation by a novel method.
Mol Microbiol, 10 (1993), pp. 1057-1065
[12.]
D. Van Soolingen.
Molecular epidemiology of tuberculosis and other mycobacterial infections: main methodologies and achievements.
J Intern Med, 249 (2001), pp. 1-26
[13.]
E. Richter, M. Weizenegger, S. Rüsch-Gerdes, S. Niemann.
Evaluation of genotype MTBC assay for differenciation of clinical Mycobacterium tuberculosis complex isolates.
J Clin Microbiol, 41 (2003), pp. 2672-2675
[14.]
S. Niemann, D. Harmsen, S. Rüsch-Gerdes, E. Richter.
Differentiation of clinical Mycobacterium tuberculosis complex isolates by gyrB DNA sequence polymorphism analysis.
J Clin Microbiol, 38 (2000), pp. 3231-3234
[15.]
H. David, L. Brum, E. Prieto.
Manual de micobacteriologia em saúde pública: princípios e métodos.
Instituto de Higiene e Medicina Tropical, (1994),
[16.]
C.H. Collins, M.D. Yates, J.M. Grange.
A study of bovine strains of Mycobacterium tuberculosis isolated from humans in South-East England, 1977-1979.
Tubercle, 62 (1981), pp. 117-121
[17.]
S. Niemann, D. Harmsen, S. Rüsch-Gerdes.
Differenciation among members of the Mycobacterium tuberculosis complex by molecular and biochemical features: evidence for two pyrazinamide-susceptible subtypes of M. bovis.
J Clin Microbiol, 38 (2000), pp. 152-157
[18.]
S. Sreevatsan, X. Pan, Y. Zhang, B.N. Kreiswirth, J.M. Musser.
Mutations associated with pyrazinamide resistance in pncA of Mycobacterium tuberculosis complex organismes.
Antimicrobiol Agents Chemother, 41 (1997), pp. 636-640
[19.]
L.E. de Los Monteros, C. Galan, M. Gutierrez, S. Samper, J.F. Garcia-Martin, C. Martin, L. Dominguez, L. De Rafael, F. Baquero, E. Gomez-Mampaso, J. Blazquez.
Allele-specific PCR method based on pncA and oxyR sequences for distinguishing Mycobacterium bovis from Mycobacterium tuberculosis intraspecific M. bovis pncA polymorphism.
J Clin Microbiol, 36 (1998), pp. 239-242
[20.]
A. Scorpio, D. Collins, D. Whipple, D. Cave, J. Bates, Y. Zhang.
Rapid differentiation of bovine and human tubercle bacilli based on a characteristic mutation in the bovine pyrazinamidase gene.
J Clin Microbiol, 35 (1997), pp. 106-110
[21.]
S. David, C. Portugal, A. Antunes, A. Cardoso, A. Calado, V. Barros, L. Sancho.
Identificação molecular pelo método de Spoligotyping de estirpes do complexo Mycobacterium tuberculosis isoladas no Hospital Fernando Fonseca.
Rev Port Pneumologia, X (2004), pp. 195-204
[24.]
E.A. Talbot, D.L. William, R. Frothingham.
PCR identification of Mycobacterium bovis BCG.
J Clin Microbiol, 35 (1997), pp. 566-569
[25.]
M. Salfinger, L.B. Reller, F.M. Kafader.
Abstr. 90th Annu Meet Am Soc Microbiol, 12 (1990), pp. 38
[26.]
H.L. David, T.M. Jahan, A. Jumin, J. Grandry, E.H. Lehman.
Numerical taxonomy analysis of Mycobacterium africanum.
Int. J. Systematic Bacteriol, (1978), pp. 467-472
[27.]
C. Viana-Niero, C. Gutierrez, C. Sola, I. Filliol, F. Boulahbal, V. Vincent, N. Rastogi.
Genetic diversity of Mycobacterium africanum clinical isolates based on IS6110-restriction fragment length polymorphism analysis, spoligotyping, and variable number of tandem DNA repeats.
J Clin Microbiol, 39 (2001), pp. 57-65
[28.]
M.C. Gutierrez, J.C. Galan, J. Blazquez, E. Bouvet, V. Vincent.
Molecular markers demonstrate that the first described multidrug-resistant Mycobacterium bovis outbreak was due to Mycobacterium tuberculosis.
J Clin Microbiol, 37 (1999), pp. 971-975
Copyright © 2005. Sociedade Portuguesa de Pneumologia/SPP
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