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Draft Genome Sequence of a Mycobacterium africanum Clinical Isolate from Antioquia, Colombia U. A. Hurtado,a,b J. S. Solano,a,b A. Rodriguez,a,b J. Robledo,a,c F. Rouzauda,d Corporación para Investigaciones Biológicas (CIB), Medellín, Colombiaa; Universidad Nacional de Colombia, Medellín, Colombiab; Universidad Pontificia Bolivariana (UPB), Medellín, Colombiac; Equal Opportunity Life Sciences (EQUOLS), Rockville, Maryland, USAd
Mycobacterium africanum is a member of the Mycobacterium tuberculosis complex. Most commonly found in West African countries, it has scarcely been described in South America. Here, we report the first genome sequence of a Colombian M. africanum clinical isolate. It is composed of 4,493,502 bp, with 4,069 genes. Received 18 April 2016 Accepted 21 April 2016 Published 2 June 2016 Citation Hurtado UA, Solano JS, Rodriguez A, Robledo J, Rouzaud F. 2016. Draft genome sequence of a Mycobacterium africanum clinical isolate from Antioquia, Colombia. Genome Announc 4(3):e00486-16. doi:10.1128/genomeA.00486-16. Copyright © 2016 Hurtado et al. This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International license. Address correspondence to J. Robledo, [email protected].
M
ycobacterium africanum is the causative pathogen of nearly half of the pulmonary tuberculosis cases in West Africa (1, 2). It has been characterized in 1968 in Senegal as an intermediate strain between Mycobacterium tuberculosis and Mycobacterium bovis (3). It consists of 2 phylogenetically distinct lineages within the M. tuberculosis complex, known as West African 1 and West African 2. The symptoms of infection caused by M. africanum are similar to those caused by its M. tuberculosis complex counterparts, and it is likely spread by aerosol transmission as well (4). Human tuberculosis caused by M. africanum has been reported in Africa, Australia, Europe, North America, and South America (3– 6). However, no M. africanum isolate has been reported in Colombia to date. Twenty-four-locus mycobacterial interspersed repetitiveunit–variable-number tandem-repeat (MIRU-VNTR) analysis was used to confirm that isolate UT307 belongs to the M. africanum lineage (7–9). An IS6110-based restriction fragment length polymorphism (RFLP) analysis revealed the presence of 4 copies of the insertion element. Spoligotyping showed the absence of spacers 7 to 9, 22 to 24, 35, 36, and 39. Phenotypic susceptibility tests were performed using the Bactec MGIT 960 system. It was determined that UT307 is susceptible to streptomycin, isoniazid, rifampin, and ethambutol. Genomic DNA was obtained from isolate UT307 using the cetyltrimethylammonium bromide (CTAB) method (10). Wholegenome sequencing was performed at the BioFrontiers Institute Next-Generation Sequencing Facility (University of ColoradoBoulder), using an Illumina HiSeq 2000 platform with 100-bp reads and to about 228⫻ coverage. De novo assembly was carried out using the SOAPdenovo2 assembler (11), revealing a sequence length of 4,493,502 bp, with 65.1% G⫹C content. The N50 and mean contig lengths were 195,659 bp and 19,278 bp, respectively. The functional and structural annotations were completed with the Prokka software, which uses several sources of evidence, including the TIGRFAMs and Pfam protein family databases (12). The genome contains 4,018 coding sequences (CDSs) and 45 tRNAs. Large-sequence polymorphisms (LSPs) were evaluated,
May/June 2016 Volume 4 Issue 3 e00486-16
and we identified partial deletions of RD7, RD9, and RD10, and complete deletions of RD8 and RD702. Single-nucleotide polymorphisms (SNPs) Rv2427828GC and Rv378404GA (13) were identified, validating the assignment of isolate UT307 to the West African 2 lineage. The whole-genome sequencing of UT307 confirms the circulation of M. africanum in Colombia. This first report will serve as a baseline further comparative genome analysis of M. africanum in Colombia and in the Americas. Nucleotide sequence accession numbers. The genome sequence of the M. africanum isolate UT307 has been deposited at DDBJ/EMBL/GenBank under the accession no. CP014617. The version described in this paper is the first version, CP014617.1. ACKNOWLEDGMENT We thank the “Centro Colombiano de Investigación en Tuberculosis CCITB,” which performed initial identification and genotyping of the M. africanum isolate.
FUNDING INFORMATION This work, including the efforts of Uriel Hurtado Páez, Juan Sebastian Solano, Hector Alejandro Rodriguez Cabal, and Francois Rouzaud, was funded by Departamento Administrativo de Ciencia, Tecnología e Innovación (COLCIENCIAS) (657057636375).
REFERENCES 1. De Jong BC, Antonio M, Gagneux S. 2010. Mycobacterium africanum— review of an important cause of human tuberculosis in West Africa. PLoS Negl Trop Dis 4:e744. http://dx.doi.org/10.1371/journal.pntd.0000744. 2. Haas W, Brezel G, Amthor B, Schilke K, Krommes G, Rusch-Gerdes S, Sticht-Groh V, Bremer H. 1997. Comparison of DNA fingerprint patterns of isolates of Mycobacterium africanum from East and West Africa. J Clin Microbiol 35:663– 666. 3. Castets M, Boisvert H, Grumbach F, Brunel M, Rist N. 1968. Tuberculosis bacilli of the African type: preliminary note. Rev Tuberc Pneumol (Paris) 32:179 –184. (In French.) 4. Desmond E, Ahmed A, Probert W, Ely J, Jang Y, Sanders C, Lin S, Flood J. 2004. Mycobacterium africanum cases, California. Emerg Infect Dis 10:5. 5. Grange JM, Yates MD. 1989. Incidence and nature of human tuberculosis
Genome Announcements
genomea.asm.org 1
Hurtado et al.
6.
7. 8.
9.
10.
due to Mycobacterium africanum in South-East England: 1977-87. Epidemiol Infect 103:127–132. http://dx.doi.org/10.1017/S0950268800030429. Lumb R, Bastian I, Carter R, Jelfs P, Keehner T, Sievers A. Tuberculosis in Australia: bacteriologically confirmed cases and drug resistance, 2010. A report of the Australian Mycobacterium Reference Laboratory Network. Commun Dis Intell Q Rep 37:E40 –E46. Mostowy S, Onipede A, Gagneux S, Niemann S, Kremer K, Desmond EP, Kato-Maeda M, Behr M. 2004. Genomic analysis distinguishes Mycobacterium africanum. J Clin Microbiol 42:3594 –3599. Van Ingen J, Rahim Z, Mulder A, Boeree M, Simeone R, Brosch R, Van Soolingen D. 2012. Characterization of Mycobacterium orygis as M. tuberculosis complex subspecies. Emerg Infect Dis 18:653– 655. http:// dx.doi.org/10.3201/eid1804.110888. Weniger T, Krawczyk J, Weniger T, Krawczyk J, Supply P, Niemann S, Harmsen D. 2010. MIRU-VNTRplus: a Web tool for polyphasic genotyping of Mycobacterium tuberculosis complex bacteria. Nucleic Acids Res 38:W326 –W331. http://dx.doi.org/10.1093/nar/gkq351. Belisle JT, Sonnenberg MG. 1998. Isolation of genomic DNA from
2 genomea.asm.org
mycobacteria, p 31– 44. In Parish T, Stoker NG (ed.), Methods in molecular biology, vol 101. Mycobacteria protocols. Humana Press, Totowa, NJ. 11. Ruibang L, Binghang L, Yinlong X, Zhenyu L, Weihua H, Jianying Y, Guangzhu H, Yanxiang C, Qi P, Yunjie L, Jingbo T, Gengxiong W, Hao Z, Yujian S, Yong L, Chang Y, Bo W, Yao L, Changlei H, David WC, Siu-Ming Y, Shaoliang P, Zhu X, Guangming L, Xiangke L, Yingrui L, Huanming Y, Jian W, Tak-Wah L, Jun W. 2012. SOAPdenovo2: an empirically improved memory-efficient short-read de novo assembler. GigaScience 1:18. http://dx.doi.org/10.1186/2047-217X-1-18. 12. Seemann T. 2014. Prokka: rapid prokaryotic genome annotation. Bioinformatics 30:2068 –2069. http://dx.doi.org/10.1093/ bioinformatics/btu153. 13. Stucki D, Malla B, Hostettler S1, Huna T, Feldmann J, Yeboah-Manu D, Borrell S, Fenner L, Comas I, Coscollá M, Gagneux S. 2012. Two new rapid SNP-typing methods for classifying Mycobacterium tuberculosis complex into the main phylogenetic lineages. PLoS One 7:e41253. http:// dx.doi.org/10.1371/journal.pone.0041253.
Genome Announcements
May/June 2016 Volume 4 Issue 3 e00486-16
Draft Genome Sequence of a Mycobacterium africanum Clinical Isolate from Antioquia, Colombia U. A. Hurtado,a,b J. S. Solano,a,b A. Rodriguez,a,b J. Robledo,a,c F. Rouzauda,d Corporación para Investigaciones Biológicas (CIB), Medellín, Colombiaa; Universidad Nacional de Colombia, Medellín, Colombiab; Universidad Pontificia Bolivariana (UPB), Medellín, Colombiac; Equal Opportunity Life Sciences (EQUOLS), Rockville, Maryland, USAd
Mycobacterium africanum is a member of the Mycobacterium tuberculosis complex. Most commonly found in West African countries, it has scarcely been described in South America. Here, we report the first genome sequence of a Colombian M. africanum clinical isolate. It is composed of 4,493,502 bp, with 4,069 genes. Received 18 April 2016 Accepted 21 April 2016 Published 2 June 2016 Citation Hurtado UA, Solano JS, Rodriguez A, Robledo J, Rouzaud F. 2016. Draft genome sequence of a Mycobacterium africanum clinical isolate from Antioquia, Colombia. Genome Announc 4(3):e00486-16. doi:10.1128/genomeA.00486-16. Copyright © 2016 Hurtado et al. This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International license. Address correspondence to J. Robledo, [email protected].
M
ycobacterium africanum is the causative pathogen of nearly half of the pulmonary tuberculosis cases in West Africa (1, 2). It has been characterized in 1968 in Senegal as an intermediate strain between Mycobacterium tuberculosis and Mycobacterium bovis (3). It consists of 2 phylogenetically distinct lineages within the M. tuberculosis complex, known as West African 1 and West African 2. The symptoms of infection caused by M. africanum are similar to those caused by its M. tuberculosis complex counterparts, and it is likely spread by aerosol transmission as well (4). Human tuberculosis caused by M. africanum has been reported in Africa, Australia, Europe, North America, and South America (3– 6). However, no M. africanum isolate has been reported in Colombia to date. Twenty-four-locus mycobacterial interspersed repetitiveunit–variable-number tandem-repeat (MIRU-VNTR) analysis was used to confirm that isolate UT307 belongs to the M. africanum lineage (7–9). An IS6110-based restriction fragment length polymorphism (RFLP) analysis revealed the presence of 4 copies of the insertion element. Spoligotyping showed the absence of spacers 7 to 9, 22 to 24, 35, 36, and 39. Phenotypic susceptibility tests were performed using the Bactec MGIT 960 system. It was determined that UT307 is susceptible to streptomycin, isoniazid, rifampin, and ethambutol. Genomic DNA was obtained from isolate UT307 using the cetyltrimethylammonium bromide (CTAB) method (10). Wholegenome sequencing was performed at the BioFrontiers Institute Next-Generation Sequencing Facility (University of ColoradoBoulder), using an Illumina HiSeq 2000 platform with 100-bp reads and to about 228⫻ coverage. De novo assembly was carried out using the SOAPdenovo2 assembler (11), revealing a sequence length of 4,493,502 bp, with 65.1% G⫹C content. The N50 and mean contig lengths were 195,659 bp and 19,278 bp, respectively. The functional and structural annotations were completed with the Prokka software, which uses several sources of evidence, including the TIGRFAMs and Pfam protein family databases (12). The genome contains 4,018 coding sequences (CDSs) and 45 tRNAs. Large-sequence polymorphisms (LSPs) were evaluated,
May/June 2016 Volume 4 Issue 3 e00486-16
and we identified partial deletions of RD7, RD9, and RD10, and complete deletions of RD8 and RD702. Single-nucleotide polymorphisms (SNPs) Rv2427828GC and Rv378404GA (13) were identified, validating the assignment of isolate UT307 to the West African 2 lineage. The whole-genome sequencing of UT307 confirms the circulation of M. africanum in Colombia. This first report will serve as a baseline further comparative genome analysis of M. africanum in Colombia and in the Americas. Nucleotide sequence accession numbers. The genome sequence of the M. africanum isolate UT307 has been deposited at DDBJ/EMBL/GenBank under the accession no. CP014617. The version described in this paper is the first version, CP014617.1. ACKNOWLEDGMENT We thank the “Centro Colombiano de Investigación en Tuberculosis CCITB,” which performed initial identification and genotyping of the M. africanum isolate.
FUNDING INFORMATION This work, including the efforts of Uriel Hurtado Páez, Juan Sebastian Solano, Hector Alejandro Rodriguez Cabal, and Francois Rouzaud, was funded by Departamento Administrativo de Ciencia, Tecnología e Innovación (COLCIENCIAS) (657057636375).
REFERENCES 1. De Jong BC, Antonio M, Gagneux S. 2010. Mycobacterium africanum— review of an important cause of human tuberculosis in West Africa. PLoS Negl Trop Dis 4:e744. http://dx.doi.org/10.1371/journal.pntd.0000744. 2. Haas W, Brezel G, Amthor B, Schilke K, Krommes G, Rusch-Gerdes S, Sticht-Groh V, Bremer H. 1997. Comparison of DNA fingerprint patterns of isolates of Mycobacterium africanum from East and West Africa. J Clin Microbiol 35:663– 666. 3. Castets M, Boisvert H, Grumbach F, Brunel M, Rist N. 1968. Tuberculosis bacilli of the African type: preliminary note. Rev Tuberc Pneumol (Paris) 32:179 –184. (In French.) 4. Desmond E, Ahmed A, Probert W, Ely J, Jang Y, Sanders C, Lin S, Flood J. 2004. Mycobacterium africanum cases, California. Emerg Infect Dis 10:5. 5. Grange JM, Yates MD. 1989. Incidence and nature of human tuberculosis
Genome Announcements
genomea.asm.org 1
Hurtado et al.
6.
7. 8.
9.
10.
due to Mycobacterium africanum in South-East England: 1977-87. Epidemiol Infect 103:127–132. http://dx.doi.org/10.1017/S0950268800030429. Lumb R, Bastian I, Carter R, Jelfs P, Keehner T, Sievers A. Tuberculosis in Australia: bacteriologically confirmed cases and drug resistance, 2010. A report of the Australian Mycobacterium Reference Laboratory Network. Commun Dis Intell Q Rep 37:E40 –E46. Mostowy S, Onipede A, Gagneux S, Niemann S, Kremer K, Desmond EP, Kato-Maeda M, Behr M. 2004. Genomic analysis distinguishes Mycobacterium africanum. J Clin Microbiol 42:3594 –3599. Van Ingen J, Rahim Z, Mulder A, Boeree M, Simeone R, Brosch R, Van Soolingen D. 2012. Characterization of Mycobacterium orygis as M. tuberculosis complex subspecies. Emerg Infect Dis 18:653– 655. http:// dx.doi.org/10.3201/eid1804.110888. Weniger T, Krawczyk J, Weniger T, Krawczyk J, Supply P, Niemann S, Harmsen D. 2010. MIRU-VNTRplus: a Web tool for polyphasic genotyping of Mycobacterium tuberculosis complex bacteria. Nucleic Acids Res 38:W326 –W331. http://dx.doi.org/10.1093/nar/gkq351. Belisle JT, Sonnenberg MG. 1998. Isolation of genomic DNA from
2 genomea.asm.org
mycobacteria, p 31– 44. In Parish T, Stoker NG (ed.), Methods in molecular biology, vol 101. Mycobacteria protocols. Humana Press, Totowa, NJ. 11. Ruibang L, Binghang L, Yinlong X, Zhenyu L, Weihua H, Jianying Y, Guangzhu H, Yanxiang C, Qi P, Yunjie L, Jingbo T, Gengxiong W, Hao Z, Yujian S, Yong L, Chang Y, Bo W, Yao L, Changlei H, David WC, Siu-Ming Y, Shaoliang P, Zhu X, Guangming L, Xiangke L, Yingrui L, Huanming Y, Jian W, Tak-Wah L, Jun W. 2012. SOAPdenovo2: an empirically improved memory-efficient short-read de novo assembler. GigaScience 1:18. http://dx.doi.org/10.1186/2047-217X-1-18. 12. Seemann T. 2014. Prokka: rapid prokaryotic genome annotation. Bioinformatics 30:2068 –2069. http://dx.doi.org/10.1093/ bioinformatics/btu153. 13. Stucki D, Malla B, Hostettler S1, Huna T, Feldmann J, Yeboah-Manu D, Borrell S, Fenner L, Comas I, Coscollá M, Gagneux S. 2012. Two new rapid SNP-typing methods for classifying Mycobacterium tuberculosis complex into the main phylogenetic lineages. PLoS One 7:e41253. http:// dx.doi.org/10.1371/journal.pone.0041253.
Genome Announcements
May/June 2016 Volume 4 Issue 3 e00486-16
