@2024 Afarand., IRAN
ISSN: 2252-0805 The Horizon of Medical Sciences 2013;19(1):1-6
ISSN: 2252-0805 The Horizon of Medical Sciences 2013;19(1):1-6
Gene Polymorphism of BCG Vaccine Strains Using in Iran
ARTICLE INFO
Article Type
Original ResearchAuthors
Fallah F. (1)Goudarzi H. (2)
Doustdar F. (2)
Zahraei S.M. (3)
Mohammadzadeh A.R. (*)
(*) Department of Microbiology, Faculty of Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran
(1) “Department of Microbiology, Faculty of Medicine”, & “Pediatric Infections Research Center”, Shahid Beheshti University of Medical Sciences, Tehran, Iran
(2) Department of Microbiology, Faculty of Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran
(3) Contagious Diseases Management Center, Ministry of Health, Care & Medical Education, Tehran, Iran
Correspondence
Address: Microbiology Department, Medical Faculty, Shahid Beheshti University of Medical Sciences, Koodakyar Street, Daneshgah Boulevard, Velenjak, Tehran, Iran.Phone: +985357225027
Fax: +985357223914
alm13604@gmail.com
Article History
Received: September 19, 2012Accepted: March 4, 2013
ePublished: March 10, 2013
ABSTRACT
Aims
The Bacille Calmette-Guérin (BCG) is the only vaccine available against tuberculosis. The BCG strain used in Iran as the vaccine is not specified. The purpose of the present study was to confirm and make a qualitative control for BCG vaccine in Iran and to determine the genetic polymorphism of the given strain.
Materials & Methods Twenty eight lyophilized BCG vaccines (Pasteur Institute, Iran) with different batch numbers were collected over a year (2011-12) and transferred under the temperature condition of 4°C to the laboratory. Three strains of vaccine were selected randomly out of each batch number. The DNA was extracted using CTAB method. DNA samples were kept at -20°C. Of four primers, RD14 and DU1 genes were used for PCR. The sequence of RD16 region in BCG strains was determined using DNA analyzer ABI3730XL using Chromas and Mega4 software. VNTR typing was done for six genic positions.
Findings All strains were positive in terms of their culture on the medium. In all BCG strains under study, DU1 duplication in the bacteria’s chromosome was observed. None of strains had RD14 gene. All RD16 regions had similar sequence with RD16 region in the genome of Pasture1173p2 strain. The number and size of repeated units for six gene positions including MIRU4, MIRU24, MIRU26, MIRU40, VNTR1895 and VNTR11b in 28 strains of different BCG in the vaccine samples were respectively (2×77)+122, (2×54)+393, (5×51)+285, (2×54)+354, (4×57)+281 and (3×69)+67.
Conclusion BCG strain using in Iran is the very Pasteur 1173p2 and may be used as vaccine.
Materials & Methods Twenty eight lyophilized BCG vaccines (Pasteur Institute, Iran) with different batch numbers were collected over a year (2011-12) and transferred under the temperature condition of 4°C to the laboratory. Three strains of vaccine were selected randomly out of each batch number. The DNA was extracted using CTAB method. DNA samples were kept at -20°C. Of four primers, RD14 and DU1 genes were used for PCR. The sequence of RD16 region in BCG strains was determined using DNA analyzer ABI3730XL using Chromas and Mega4 software. VNTR typing was done for six genic positions.
Findings All strains were positive in terms of their culture on the medium. In all BCG strains under study, DU1 duplication in the bacteria’s chromosome was observed. None of strains had RD14 gene. All RD16 regions had similar sequence with RD16 region in the genome of Pasture1173p2 strain. The number and size of repeated units for six gene positions including MIRU4, MIRU24, MIRU26, MIRU40, VNTR1895 and VNTR11b in 28 strains of different BCG in the vaccine samples were respectively (2×77)+122, (2×54)+393, (5×51)+285, (2×54)+354, (4×57)+281 and (3×69)+67.
Conclusion BCG strain using in Iran is the very Pasteur 1173p2 and may be used as vaccine.
CITATION LINKS
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[2]Herr H, Morales A. History of Bacillus calmette-guerin and bladder cancer: An immunotherapy success story. J Urol. 2008;179:53-6.
[3] Behr MA. Comparative genomics of BCG vaccines. Tuberculosis. 2001;81(1/2):165-8.
[4] Pym A, Brodin P, Brosch R, Huerre M, Cole S. Loss of RD1 contributed to the attenuation of the live tuberculosis vaccines Mycobacterium bovis BCG and Mycobacterium microti. Mol Microbiol. 2002;46(3):709-17.
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[7] Brosch R, Gordon S, Garnier T, Eiglmeier K, Frigui W, Valenti P, et al. Genome plasticity of BCG and impact on vaccine efficacy. Proc Natl Acad Sci. 2007;104(13):5596- 601.
[8] Knezevic I, Corbel MJ. WHO discussion on the improvement of the quality control of BCG vaccines. Pasteur Institute, Paris, France, 7 June 2005 (Meeting report). Vaccine. 2006;24(18):3874-7.
[9] Magdalena J, Supply P, Locht C. Specific differentiation between Mycobacterium bovis BCG and virulent strains of the Mycobacterium tuberculosis complex. J Clin Microbiol. 1998;36(9):2471-6.
[10] Roring S, Scott AN, Hewinson RG, Neill SD, Skuce RA. Evaluation of Variable Number Tandem Repeat (VNTR) loci in molecular typing of Mycobacterium bovis isolates from Ireland. Vet Microbiol. 2004;101(1):65-73.
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[13] Markey K, Hoa MM, Choudhurya B, Sekib M, Juc L, Castello-Brancod L RR, et al. Report of an international collaborative study to evaluate the suitability of multiplex PCR as an identity assay for different sub-strains of BCG vaccine. Vaccine. 2010;28:6964-9.
[14] Okazaki T, Ebihara S, Takahashi H, Asada M, Sato A, Seki M, et al. Multiplex PCR-identified cutaneous tuberculosis evoked by Mycobacterium bovis BCG vaccination in a healthy baby. J Clin Microbiol. 2005;43(1):523-5.
[15]Kim SH, Kim SY, Eun BW, Yoo WJ, Park KU, Choi EH, et al. BCG osteomyelitis caused by the BCG Tokyo strain and confirmed by molecular method. Vaccine. 2008;26:4379-81.
[16]Talbot E, Williams D, Frothingham R. PCR identification of Mycobacterium bovis BCG. J Clin Microbiol. 1997;35(3):566-9.
[17] Allix C, Supply P, Fauville-Dufaux M. Utility of fast mycobacterial interspersed repetitive unit-variable number tandem repeat genotyping in clinical mycobacteriological analysis. Clin Infect Dis. 2004;39(6):783-9.
[18] Ritz N, Curtis N. Mapping the global use of different BCG vaccine strains. Tuberculosis. 2009;89:248-51.
[19] Ausubel F, Brent R, Kingston RE, Moore DD, Seidman JG, Smith JA, et al. Short protocols in molecular biology. 5th ed. New York: John Wiley and Sons; 2002.
[20] Somerville W, Thibert L, Schwartzman K, Behr MA. Extraction of Mycobacterium tuberculosis DNA: A question of containment. J Clin Microbiol. 2005;43(6):2996-7.
[21] Supply P, Allix C, Lesjean S, Cardoso-Oelemann M, Rusch-Gerdes S, Willery E. Proposal for standardization of optimized mycobacterial interspersed repetitive unitvariable- number tandem repeat typing of Mycobacterium tuberculosis. J Clin Microbiol. 2006;44(12):4498-510.
[22] World Health Organization. Global TB database. Geneva: World Health Organization; 2012. Available from: http://www.who.int/tb/country/global_tb_database/en/;2012
[23] Khazaei HA, Rezaei N, Bagheri GhR, Dankoub MA, Shahryari Kh, Tahai A. Epidemiology of tuberculosis in the southeastern Iran. Eur J Epidemiol. 2005;20:879-83.
[24]Liu J, Tran V, Leung A, Alexander D, Zhu B. BCG vaccines: Their mechanisms of attenuation and impact on safety and protective efficacy. Hum Vac. 2009;5(2):70-8.
[25] World Health Organization. BCG vaccine. WHO position paper. Wkly Epidemiol Rec. 2004;79:27-38.
[26] Brosch R, Gordon S, Buchrieser C, Pym A, Garnier T, Cole S. Comparative genomics uncovers large tandem chromosomal duplications in Mycobacterium bovis BCG Pasteur. Yeast. 2000;17(2):111-23.
[27]Cowan LS, Mosher L, Diem L, Massey JP, Crawford JT. Variable-number tandem repeat typing of Mycobacterium tuberculosis isolates with low copy numbers of IS6110 by using mycobacterial interspersed repetitive units. J Clin Microbiol. 2002;40(5):1592-602.
[28] Duarte E, Domingos M, Amado A, Cunha M, Botelho A. MIRU-VNTR typing ads discriminatory value to groups of Mycobacterium bovis and Mycobacterium caprae strains defined by spoligotyping. Vet Microbiol. 2010;143(2- 4):299-306.
[29] Stefanova T, Chouchkova M, Hinds J, Butcher P, Inwald J, Dale J, et al. Genetic composition of Mycobacterium bovis BCG substrain Sofia. J Clin Microbiol. 2003;41(11):5349.
[2]Herr H, Morales A. History of Bacillus calmette-guerin and bladder cancer: An immunotherapy success story. J Urol. 2008;179:53-6.
[3] Behr MA. Comparative genomics of BCG vaccines. Tuberculosis. 2001;81(1/2):165-8.
[4] Pym A, Brodin P, Brosch R, Huerre M, Cole S. Loss of RD1 contributed to the attenuation of the live tuberculosis vaccines Mycobacterium bovis BCG and Mycobacterium microti. Mol Microbiol. 2002;46(3):709-17.
[5] Bedwell J, Kairo SK, Behr MA, Bygraves JA. Identification of substrains of BCG vaccine using multiplex PCR. Vaccine. 2001;19:2146-51.
[6] Behr MA, Wilson MA, Gill WP, Salamon H, Schoolnik GK, Rane S, et al. Comparative genomics of BCG vaccines by whole-genome DNA microarray. Science. 1999;284:1520-3.
[7] Brosch R, Gordon S, Garnier T, Eiglmeier K, Frigui W, Valenti P, et al. Genome plasticity of BCG and impact on vaccine efficacy. Proc Natl Acad Sci. 2007;104(13):5596- 601.
[8] Knezevic I, Corbel MJ. WHO discussion on the improvement of the quality control of BCG vaccines. Pasteur Institute, Paris, France, 7 June 2005 (Meeting report). Vaccine. 2006;24(18):3874-7.
[9] Magdalena J, Supply P, Locht C. Specific differentiation between Mycobacterium bovis BCG and virulent strains of the Mycobacterium tuberculosis complex. J Clin Microbiol. 1998;36(9):2471-6.
[10] Roring S, Scott AN, Hewinson RG, Neill SD, Skuce RA. Evaluation of Variable Number Tandem Repeat (VNTR) loci in molecular typing of Mycobacterium bovis isolates from Ireland. Vet Microbiol. 2004;101(1):65-73.
[11] European Pharmacopoeia Commission. BCG vaccine, freeze dried. European pharmacopoeia. 6th ed. Strasbourg: Council of Europe; 2008.
[12] Donikian R, Gheorghiu M, Jablokova TB. Requirements for dried BCG vaccine. WHO Tech Rep Ser. 1987:60-92
[13] Markey K, Hoa MM, Choudhurya B, Sekib M, Juc L, Castello-Brancod L RR, et al. Report of an international collaborative study to evaluate the suitability of multiplex PCR as an identity assay for different sub-strains of BCG vaccine. Vaccine. 2010;28:6964-9.
[14] Okazaki T, Ebihara S, Takahashi H, Asada M, Sato A, Seki M, et al. Multiplex PCR-identified cutaneous tuberculosis evoked by Mycobacterium bovis BCG vaccination in a healthy baby. J Clin Microbiol. 2005;43(1):523-5.
[15]Kim SH, Kim SY, Eun BW, Yoo WJ, Park KU, Choi EH, et al. BCG osteomyelitis caused by the BCG Tokyo strain and confirmed by molecular method. Vaccine. 2008;26:4379-81.
[16]Talbot E, Williams D, Frothingham R. PCR identification of Mycobacterium bovis BCG. J Clin Microbiol. 1997;35(3):566-9.
[17] Allix C, Supply P, Fauville-Dufaux M. Utility of fast mycobacterial interspersed repetitive unit-variable number tandem repeat genotyping in clinical mycobacteriological analysis. Clin Infect Dis. 2004;39(6):783-9.
[18] Ritz N, Curtis N. Mapping the global use of different BCG vaccine strains. Tuberculosis. 2009;89:248-51.
[19] Ausubel F, Brent R, Kingston RE, Moore DD, Seidman JG, Smith JA, et al. Short protocols in molecular biology. 5th ed. New York: John Wiley and Sons; 2002.
[20] Somerville W, Thibert L, Schwartzman K, Behr MA. Extraction of Mycobacterium tuberculosis DNA: A question of containment. J Clin Microbiol. 2005;43(6):2996-7.
[21] Supply P, Allix C, Lesjean S, Cardoso-Oelemann M, Rusch-Gerdes S, Willery E. Proposal for standardization of optimized mycobacterial interspersed repetitive unitvariable- number tandem repeat typing of Mycobacterium tuberculosis. J Clin Microbiol. 2006;44(12):4498-510.
[22] World Health Organization. Global TB database. Geneva: World Health Organization; 2012. Available from: http://www.who.int/tb/country/global_tb_database/en/;2012
[23] Khazaei HA, Rezaei N, Bagheri GhR, Dankoub MA, Shahryari Kh, Tahai A. Epidemiology of tuberculosis in the southeastern Iran. Eur J Epidemiol. 2005;20:879-83.
[24]Liu J, Tran V, Leung A, Alexander D, Zhu B. BCG vaccines: Their mechanisms of attenuation and impact on safety and protective efficacy. Hum Vac. 2009;5(2):70-8.
[25] World Health Organization. BCG vaccine. WHO position paper. Wkly Epidemiol Rec. 2004;79:27-38.
[26] Brosch R, Gordon S, Buchrieser C, Pym A, Garnier T, Cole S. Comparative genomics uncovers large tandem chromosomal duplications in Mycobacterium bovis BCG Pasteur. Yeast. 2000;17(2):111-23.
[27]Cowan LS, Mosher L, Diem L, Massey JP, Crawford JT. Variable-number tandem repeat typing of Mycobacterium tuberculosis isolates with low copy numbers of IS6110 by using mycobacterial interspersed repetitive units. J Clin Microbiol. 2002;40(5):1592-602.
[28] Duarte E, Domingos M, Amado A, Cunha M, Botelho A. MIRU-VNTR typing ads discriminatory value to groups of Mycobacterium bovis and Mycobacterium caprae strains defined by spoligotyping. Vet Microbiol. 2010;143(2- 4):299-306.
[29] Stefanova T, Chouchkova M, Hinds J, Butcher P, Inwald J, Dale J, et al. Genetic composition of Mycobacterium bovis BCG substrain Sofia. J Clin Microbiol. 2003;41(11):5349.