danДоклады Национальной академии наук БеларусиDoklady of the National Academy of Sciences of Belarus1561-83232524-2431The Republican Unitary Enterprise Publishing House "Belaruskaya Navuka"10.29235/1561-8323-2023-67-3-231-241dan-1133Research ArticleМЕДИЦИНАMEDICINEХарактеристика генома Mycobacterium tuberculosis генотипа Beijing кластера 100-32 с пре-широкой лекарственной устойчивостьюCharacteristic of the Mycobacterium tuberculosis genome of Beijing genotype of cluster 100-32 displaying pre-extended drug resistancehttps://orcid.org/0000-0003-1846-6100СлизеньВ. В.SlizenV. V.

Слизень Вероника Вячеславовна – канд. мед. наук, доцент, докторант

Долгиновский тракт, 157, 220080, Минск, Республика Беларусь

Slizen Veronika V. – Ph. D. (Medicine), Associate Professor, Doctoral Candidate

157, Dolginovski tract, 220080, Minsk, Republic of Belarus

veronal@tut.byОхремчукА. Э.AkhremchukA. E.

Oхремчук Артур Эдуардович – науч. сотрудник

ул. Купревича, 2, 220114, Минск, Республика Беларусь

Akhremchuk Artur E. – Researcher

2, Kuprevich Str., 220114, Minsk, Republic of Belarus

okhrem4uartur@gmail.comСурковаЛ. К.SurkovaL. K.

Суркова Лариса Константиновна – д-р мед. наук, профессор, заведующий отделом

Долгиновский тракт, 157, 220080, Минск, Республика Беларусь

Surkova Larisa K. – D. Sc. (Medicine), Professor, Head of the Department

157, Dolginovski tract, 220080, Minsk, Republic of Belarus

niipulm@tut.byhttps://orcid.org/0000-0001-5683-2322ГуревичГ. Л.GurevichG. L.

Гуревич Геннадий Львович – член-корреспондент, д-р мед. наук, профессор, директор

Долгиновский тракт, 157, 220080, Минск, Республика Беларусь

Gurevich Gennady L. – Corresponding Member, D. Sc. (Medicine), Professor, Director

157, Dolginovski tract, 220080, Minsk, Republic of Belarus

niipulm@tut.byhttps://orcid.org/0000-0002-4524-3730ТитовЛ. П.TitovL. P.

Титов Леонид Петрович – академик, д-р мед. наук, профессор, заведующий лабораторией

ул. Филимонова, 23, 220114, Минск, Республика Беларусь

Titov Leonid P. – Academician, D. Sc. (Medicine), Professor, Head of Laboratory

23, Filimonov Str., 220114, Minsk, Republic of Belarus

leonidtitov@tut.byРеспубликанский научно-практический центр пульмонологии и фтизиатрииRepublican Scientific and Practical Center of Pulmonology and PhthisiatryИнститут микробиологии Национальной академии наук БеларусиInstitute of Microbiology of the National Academy of Sciences of BelarusРеспубликанский научно-практический центр эпидемиологии и микробиологииRepublican Research and Practical Center for Epidemiology and Microbiology202306072023673231241Copyright © Слизень В.В., Охремчук А.Э., Суркова Л.К., Гуревич Г.Л., Титов Л.П., 20232023Слизень В.В., Охремчук А.Э., Суркова Л.К., Гуревич Г.Л., Титов Л.П.Slizen V.V., Akhremchuk A.E., Surkova L.K., Gurevich G.L., Titov L.P.Данная работа распространяется под лицензией Creative Commons Attribution 4.0.This work is licensed under a Creative Commons Attribution 4.0 License.https://doklady.belnauka.by/jour/article/view/1133

Проведено полногеномное секвенирование штамма М. tuberculosis 11502 (база биообразцов NCBI, код доступа SAMN17832565), отнесенного, на основании структуры MIRU-VNTR локусов (n = 24), к генотипу Beijing подтипу B0/W148 кластеру 100-32 и проявлявшего пре-широкую лекарственную устойчивость. Штамм М. tuberculosis 11502 был резистентен к изониазиду, рифампицину, этамбутолу, левофлоксацину, этионамиду, что коррелировало с присутствием в резистоме мутаций в соответствующих генах: к рифампицину – мутации в генах rpoB (p.S450L), rpoC (p.I491T), к изониазиду – в промоторе гена fabG1 (g.-8T>C), промоторе katG (p.S315T), к этионамиду – в ethA (делеция Т в положении 4 335 027 (gatgc-gagc)); к фторхинолонам – в гене gyrA (p.D94G); к этамбутолу – в embB (p.M306I), к стрептомицину – в rpsL (p.K43R). Геном М. tuberculosis 11502 (код доступа GenBank NCBI – CP070338) содержит 4 420 561 пару оснований, 4 104 гена, 4 053 кодирующие последовательности (кодирующие белки – 3 874) и отличается от референтного штамма М. tuberculosis H37Rv присутствием 2 055 мутаций, при этом зарегистрирован незначительный дрейф мутаций в сторону накопления G+C, что свидетельствует о важности поддержания высокого содержания G+C в геноме микобактерий. Геном М. tuberculosis 11502 имеет большее количество мутаций в сравнении с ранее секвенированным штаммом М. tuberculosis 4860 (код доступа в GenBank, NCBI: CP053092.1), относящимся к генотипу LAM (2055 против 1577 мутаций), что может быть результатом более длительной или более активной циркуляции М. tuberculosis 11502, или существования биологических особенностей, обеспечивающих промутагенный эффект.

A whole genome sequencing was performed of strain M. tuberculosis 11502 (NCBI biosamples database, access code SAMN17832565) that was assigned to the Beijing genotype subtype B0/W148 of cluster 100-32, based on the MIRU- VNTR loci (n = 24) structure, a nd t hat exhibited pre-extended d rug resistance. M. tuberculosis 11502 was resistant to isoniazid, rifampicin, ethambutol, levofloxacin, and ethionamide, which correlated with the presence of mutations in the genes: resistance to isoniazid – the mutations in the fabG1 promoter (p.-8T>C), the katG promoter (p.S315T), to ethionamide – the mutations in ethA (deletion of T at position 4 335 027 (gatgc-gagc)); to fluoroquinolones – in the gyrA gene (p.D94G); to ethambutol – in the embB gene (p.M306I); to streptomycin – in the rpsL gene (p.K43R). M. tuberculosis 11502 genome (Gen- Bank NCBI access code – CP070338) contained 4 420 561 base pairs, 4 104 genes, 4 053 CDSs (coding proteins – 3 874) and differed from reference strain M. tuberculosis H37Rv by the presence of 2 055 mutations. A slight drift of mutations towards the G+C accumulation was revealed, which indicates the importance of maintaining a high G+C content in the Mycobacterium spp.genome Strain M. tuberculosis 11502 has a higher number of mutations in comparison to previously sequenced M. tuberculosis 4860 (GenBank Access Code, NCBI: CP053092) belonging to the LAM genotype (2055 vs. 1577 mutations), which may be a consequence of a longer circulation of M. tuberculosis 11502, or some biological features providing the promutagenic effect.

геном микобактерий туберкулезагенотип Beijing подтип B0/W148кластер 100-32пре-широкая лекарственная устойчивостьMycobacterium tuberculosis genomegenotype Beijing subtype B0/W148 cluster B100-32pre-extended drug resistanceReferencesAllué-Guardia, A. Evolution of drug-resistant Mycobacterium tuberculosis strains and their adaptation to the human lung environment / А. Allué-Guardia, J. I. García, J. B. Torrelles // Front. Microbiol. – 2021. – Vol. 12. – Art. 612675. https://doi.org/10.3389/fmicb.2021.612675Allué-Guardia A., García J. I., Torrelles J. B. Evolution of drug-resistant Mycobacterium tuberculosis strains and their adaptation to the human lung environment. Frontiers in Microbiology, 2021, vol. 12, art. 612675. https://doi.org/10.3389/fmicb.2021.612675Deciphering the biology of Mycobacterium tuberculosis from the complete genome sequence / S. T. Cole [et al.] // Nature. – 1998. – Vol. 393. – P. 537–544. https://doi.org/10.1038/31159Cole S., Brosch R., Parkhill J., Garnier T., Churcher C., Harris D., Gordon S. V. [et al.]. Deciphering the biology of Mycobacterium tuberculosis from the complete genome sequence. Nature, 1998, vol. 393, no. 6685, pp. 537–544. https://doi.org/10.1038/31159Biofilm formation in the lung contributes to virulence and drug tolerance of Mycobacterium tuberculosis / P. Chakraborty [et al.] // Nat. Commun. – 2021. – Vol. 12, N 1. – Art. 1606. https://doi.org/10.1038/s41467-021-21748-6Chakraborty P., Bajeli S., Kaushal D., Radotra B. D., Kumar A. Biofilm formation in the lung contributes to virulence and drug tolerance of Mycobacterium tuberculosis. Nature Communications, 2021, vol. 12, no. 1, art. 1606. https://doi.org/10.1038/s41467-021-21748-6Evolution and diversity of clonal bacteria: the paradigm of Mycobacterium tuberculosis / T. Dos Vultos [et al.] // PloS One. – 2008. – Vol. 3, N 2. – Art. e1538. https://doi.org/10.1371/journal.pone.0001538Dos Vultos T., Mestre O., Rauzier J., Golec M., Rastogi N., Rasolofo V., Tonjum T., Sola C., Matic I., Gicquel B. Evolution and diversity of clonal bacteria: the paradigm of Mycobacterium tuberculosis. PloS One, 2008, vol. 3, no. 2, art. e1538. https://doi.org/10.1371/journal.pone.0001538Rapid detection of Mycobacterium tuberculosis Beijing genotype strains by real-time PCR / D. Hillemann [et al.] // J. Clin. Microbiol. – 2006. – Vol. 44, N 2. – P. 302–306. https://doi.org/10.1128/jcm.44.2.302-306.2006Hillemann D., Warren R., Kubica T., Rüsch-Gerdes S., Niemann S. Rapid detection of Mycobacterium tuberculosis Beijing genotype strains by real-time PCR. Journal of Clinical Microbiology, 2006, vol. 44, no. 2, pp. 302–306. https://doi.org/10.1128/jcm.44.2.302-306.2006Russian “successful” clone B0/W148 of Mycobacterium tuberculosis Beijing genotype: a multiplex PCR assay for rapid detection and global screening / I. Mokrousov [et al.] // J. Clin. Microbiol. – 2012. – Vol. 50, N 11. – P. 3757–3759. https://doi.org/10.1128/jcm.02001-12Mokrousov I., Narvskaya O., Vyazovaya A., Otten T., Jiao W.-W., Gomes L. L., Suffys P. N., Shen A.-D., Vishnevsky B. Russian “successful” clone B0/W148 of Mycobacterium tuberculosis Beijing genotype: a multiplex PCR assay for rapid detection and global screening. Journal of Clinical Microbiology, 2012, vol. 50, no. 11, pp. 3757–3759. https://doi.org/10.1128/jcm.02001-12JSpeciesWS: a web server for prokaryotic species circumscription based on pairwise genome comparison / M. Richter [et al.] // Bioinformatics. – 2016. – Vol. 32, N 6. – P. 929–931. https://doi.org/10.1093/bioinformatics/btv681Richter M., Rosselló-Móra R., Glöckner F. O., Peplies J. JSpeciesWS: a web server for prokaryotic species circumscription based on pairwise genome comparison. Bioinformatics, 2016, vol. 32, no. 6, pp. 929–931. https://doi.org/10.1093/bioinformatics/btv681Hershberg, R. Evidence that mutation is universally biased towards AT in bacteria / R. Hershberg, D. Petrov // PLoS Genetics. – 2010. – Vol. 6, N 9. – Art. e1001115. https://doi.org/10.1371/journal.pgen.1001115Hershberg R., Petrov D. A. Evidence that mutation is universally biased towards AT in bacteria. PLoS Genetics, 2010, vol. 6, no. 9, art. e1001115. https://doi.org/10.1371/journal.pgen.1001115Seward, E. A. Dietary nitrogen alters codon bias and genome composition in parasitic microorganisms / E. A. Seward, S. Kelly // Genome Biology. – 2016. – Vol. 17, N 1. – Art. 226. https://doi.org/10.1186/s13059-016-1087-9Seward E. A., Kelly S. Dietary nitrogen alters codon bias and genome composition in parasitic microorganisms. Genome Biology, 2016, vol. 17, no. 1, art. 226. https://doi.org/10.1186/s13059-016-1087-9Restricted structural gene polymorphism in the Mycobacterium tuberculosis complex indicates evolutionarily recent global dissemination / S. Sreevatsan [et al.] // Proceedings of the National Academy of Sciences. – 1997. – Vol. 94, N 18. – P. 9869–9874. https://doi.org/10.1073/pnas.94.18.9869Sreevatsan S., Pan X. I., Stockbauer K. E., Connell N. D., Kreiswirth B. N., Whittam T. S., Musser J. M. Restricted structural gene polymorphism in the Mycobacterium tuberculosis complex indicates evolutionarily recent global dissemination. Proceedings of the National Academy of Sciences, 1997, vol. 94, no. 18, pp. 9869–9874. https://doi.org/10.1073/pnas.94.18.9869Relationship between Mycobacterium tuberculosis genotype and the clinical phenotype of pulmonary and meningeal tuberculosis / G. Thwaites [et al.] // J. Clin. Microbiol. – 2008. – Vol. 46, N 4. – P. 1363–1368. https://doi.org/10.1128/jcm.02180-07Thwaites G., Caws M., Chau T. T. H., D’Sa A., Lan N. T. N., Huyen M. N. T., Gagneux S. Relationship between Mycobacterium tuberculosis genotype and the clinical phenotype of pulmonary and meningeal tuberculosis. Journal of Clinical Microbiology, 2008, vol. 46, no. 4, pp. 1363–1368. https://doi.org/10.1128/jcm.02180-07Tuberculous spondylitis in Russia and prominent role of multidrug-resistant clone Mycobacterium tuberculosis Beijing B0/W148 / A. Vyazovaya [et al.] // Antimicrob. Agents Chemother. – 2015. – Vol. 59, N 4. – P. 2349–2357. https://doi.org/10.1128/aac.04221-14Vyazovaya A., Mokrousov I., Solovieva N., Mushkin A., Manicheva O., Vishnevsky B., Zhuravlev V., Narvskaya O. Tuberculous spondylitis in Russia and prominent role of multidrug-resistant clone Mycobacterium tuberculosis Beijing B0/ W148. Antimicrobial Agents and Chemotherapy, 2015, vol. 59, no. 4, pp. 2349–2357. https://doi.org/10.1128/aac.04221-14A marked difference in pathogenesis and immune response induced by different Mycobacterium tuberculosis genotypes / B. Lopez [et al.] // Clinical Experimental Immunology. – 2003. – Vol. 133, N 1. – P. 30–37. https://doi.org/10.1046/j.1365-2249.2003.02171.xLopez B., Aguilar D., Orozco H., Burger M., Espitia C., Ritacco V., Barrera L., Kremer K., Hernandez-Pando R., Huygen K., Van Soolingen D. A marked difference in pathogenesis and immune response induced by different Mycobacterium tuberculosis genotypes. Clinical and Experimental Immunology, 2003, vol. 133, no. 1, pp. 30–37. https://doi.org/10.1046/j.1365-2249.2003.02171.xEmergence of multidrug-resistant Mycobacterium tuberculosis of the Beijing lineage in Portugal and Guinea-Bissau: a snapshot of moving clones by whole-genome sequencing / J. Perdigão [et al.] // Emerging Microbes Infections. – 2020. – Vol. 9, N 1. – P. 1342–1353. https://doi.org/10.1080/22221751.2020.1774425Perdigão J., Silva C., Maltez F., Machado D., Miranda A., Couto I., Rabna P. [et al.]. Emergence of multidrug-resistant Mycobacterium tuberculosis of the Beijing lineage in Portugal and Guinea-Bissau: a snapshot of moving clones by whole-genome sequencing. Emerging Microbes and Infections, 2020, vol. 9, no. 1, pp. 1342–1353. https://doi.org/10.1080/22221751.2020.1774425Population structure of drug-resistant Mycobacterium tuberculosis in Central Asia / A. Engström [et al.] // BMC Infectious Diseases. – 2019. – Vol. 19, N 1. – Art. 908. https://doi.org/10.1186/s12879-019-4480-7Engström A., Antonenka U., Kadyrov A., Kalmambetova G., Kranzer K., Merker M., Kabirov O. [et al.]. Population structure of drug-resistant Mycobacterium tuberculosis in Central Asia. BMC Infectious Diseases, 2019, vol. 19, no. 1, art. 908. https://doi.org/10.1186/s12879-019-4480-7

The authors declare that there are no conflicts of interest present.