Issue 4

Journal for Veterinary Medicine, Biotechnology and Biosafety

Volume 4, Issue 4, December 2018, Pages 5–11

ISSN 2411-3174 (print version) ISSN 2411-0388 (online version)

PHYLOGENETIC ANALYSIS OF UKRAINIAN BACILLUS ANTHRACIS STRAINS

Biloivan O. V. 1, Stegniy B. T. 1, Arefiev V. L. 1, Solodiankin O. S. 1, Gerilovych A. P. 1, Duerr A. 2, Schwarz J. 2, Grass G. 2, Napnenko O. O. 3, Deryabin O. M. 3

1 National Scientific Center ‘Institute of Experimental and Clinical Veterinary Medicine’, Kharkiv, Ukraine, e-mail: silverscreen91@gmail.com

2 Bundeswehr Institute of Microbiology, Munich, Germany

3 State Scientific Control Institute of Biotechnology and Strains of Microorganisms, Kyiv, Ukraine

Download PDF (print version)

Citation for print version: Biloivan, O. V., Stegniy, B. T., Arefiev, V. L., Solodiankin, O. S., Gerilovych, A. P., Duerr, A., Schwarz, J., Grass, G., Napnenko, O. O. and Deryabin, O. M. (2018) ‘Phylogenetic analysis of Ukrainian Bacillus anthracis strains’, Journal for Veterinary Medicine, Biotechnology and Biosafety, 4(4), pp. 5–11.

Download PDF (online version)

Citation for online version: Biloivan, O. V., Stegniy, B. T., Arefiev, V. L., Solodiankin, O. S., Gerilovych, A. P., Duerr, A., Schwarz, J., Grass, G., Napnenko, O. O. and Deryabin, O. M. (2018) ‘Phylogenetic analysis of Ukrainian Bacillus anthracis strains’, Journal for Veterinary Medicine, Biotechnology and Biosafety. [Online] 4(4), pp. 5–11. Available at: http://jvmbbs.kharkov.ua/archive/2018/volume4/issue4/oJVMBBS_2018044_005-011.pdf

Summary. In many countries anthrax is a common zoonotic disease which poses a serious threat to human and animal health. Sporadic cases of anthrax occur each year both among farm animals and humans in Ukraine. The cutaneous form of anthrax is the most widespread in Ukraine. The capability of Bacillus anthracis spores to remain viable in soil for decades, as well as the possibility to use this pathogen as biological terror agent make effective diagnostic and research capabilities extremely important. This comprises molecular methods including state-of-the-art methods for accurate genotyping of B. anthracis strains. A total of 12 B. anthracis DNA samples from a Ukrainian strain collection were studied by qPCR to confirm chromosomal and plasmid markers. To characterize regional and global phylogeographic relationships of these strains, canonical Single Nucleotide Polymorphism analysis (canSNP) and Multiple-Locus Variable-number of tandem repeat Analysis (MLVA‑25) were conducted. B. anthracis chromosomal DNA-markers (dhp61 and gyrA) as well as those of the pXO1 plasmid could be detected in all 12 DNA samples. However, only 5 out of 12 tested strains contained the pXO2 plasmid-marker. All pXO2 positive strains group into the A.Br.008/009 SNP-clade, which belongs to the major ‘A’ branch of B. anthracis. MLVA‑25 analysis suggested that Ukrainian B. anthracis genotypes are related to strains from Southern Europe (in particular, to Bulgarian, Greek, and Italian isolates). In contrast, the pXO2‑ negative strains might be related to the Russian vaccine strain STI as they grouped to A.Br.008/011 canSNP group. The infrequent occurrence of anthrax in the country of Ukraine is likely caused by a heterogeneous population of B. anthracis. This population is phylogenetically composed of at least two different canSNP groups of the world-wide dominating A‑branch of the pathogen. While one group might stem from environmental recovery of live vaccine strains used in Ukraine (or the former Soviet Union in the past) the other one, the A.Br.008/009 group, could be enzootic as indicated by the presence of related strains in countries of southeastern Europe in relatively close geographical vicinity to Ukraine.

Keywords: anthrax, genotyping, PCR, HRM, canonical SNP, MLVA, Ukraine

References:

Antwerpen, M. H., Zimmermann, P., Bewley, K., Frangoulidis, D. and Meyer, H. (2008) ‘Real-time PCR system targeting a chromosomal marker specific for Bacillus anthracis’, Molecular and Cellular Probes, 22(5–6), pp. 313–315. http://dx.doi.org/10.1016/j.mcp.2008.06.001

Antwerpen, M., Ilin, D., Georgieva, E., Meyer, H., Savov, E. and Frangoulidis, D. (2011) ‘MLVA and SNP analysis identified a unique genetic cluster in Bulgarian Bacillus anthracis strains’, European Journal of Clinical Microbiology and Infectious Diseases, 30(7), pp. 923–930. http://dx.doi.org/10.1007/s10096-011-1177-2

Beyer, W., Bellan, S., Eberle, G., Ganz, H. H., Getz, W. M., Haumacher, R., Hilss, K. A., Kilian, W., Lazak, J., Turner, W. C. and Turnbull, P. C. B. (2012) ‘Distribution and molecular evolution of Bacillus anthracis genotypes in Namibia’, PLoS Neglected Tropical Diseases, 6(3), p. e1534. http://dx.doi.org/10.1371/journal.pntd.0001534

Bezymennyi, M., Bagamian, K. H., Barro, A., Skrypnyk, A., Skrypnyk, V. and Blackburn, J. K. (2014) ‘Spatio-temporal patterns of livestock Anthrax in Ukraine during the past century (1913–2012)’, Applied Geography, 54, pp. 129–138. http://dx.doi.org/10.1016/j.apgeog.2014.07.016

Birdsell, D. N., Pearson, T., Price, E. P., Hornstra, H. M., Nera, R. D., Stone, N., Gruendike, J., Kaufman, E. L., Pettus, A. H., Hurbon, A. N., Buchhagen, J. L., Harms, N. J., Chanturia, G., Gyuranecz, M., Wagner, D. M. and Keim, P. S. (2012) ‘Melt analysis of Mismatch Amplification Mutation Assays (Melt-MAMA): a functional study of a cost-effective SNP genotyping assay in bacterial models’, PLoS ONE, 7(3), p. e32866. http://dx.doi.org/10.1371/journal.pone.0032866

Bobyliova, O. O. and Mukharska, L. M. (2002) ‘The epidemic situation concerning especially dangerous infections in Ukraine in the last decade’ [Epidemichna sytuatsiia z osoblyvo nebezpechnykh infektsii v Ukraini za ostannie desiatylittia], Infectious Diseases [Infektsiini khvoroby], 1, pp. 5–12. [in Ukrainian]

Bobyliova, O. O., Mukharska, L. M., Nekrasova, A. S. and Nesterenko, L. P. (2001) ‘Anthrax in Ukraine. Epidemiological analysis for 55 years (1946–2001)’ [Sybirka v Ukraini. Epidemiolohichnyi analiz za 55 rokiv (1946–2001)], Current Infections [Suchasni infektsii], 3, pp. 5–9. [in Ukrainian]

Derzelle, S. (2015) ‘Single-nucleotide polymorphism discrimination using high-resolution melting analysis for the genotyping of Bacillus anthracis’, in Cunha, M. V. and Inácio, J. (eds) Veterinary Infection Biology: Molecular Diagnostics and High-Throughput Strategies. Methods in Molecular Biology, 1247. New York, NY: Humana Press, pp. 361–371. http://dx.doi.org/10.1007/978-1-4939-2004-4_26

Derzelle, S., Laroche, S., Le Flèche, P., Hauck, Y., Thierry, S., Vergnaud, G. and Madani, N. (2011) ‘Characterization of genetic diversity of Bacillus anthracis in France by using high-resolution melting assays and Multilocus Variable-number tandem-repeat Analysis’, Journal of Clinical Microbiology, 49(12), pp. 4286–4292. http://dx.doi.org/10.1128/JCM.05439-11

Gierczyński, R., Kałużewski, S., Rakin, A., Jagielski, M., Zasada, A., Jakubczak, A., Borkowska-Opacka, B. and Rastawicki, W. (2004) ‘Intriguing diversity of Bacillus anthracis in eastern Poland — the molecular echoes of the past outbreaks’, FEMS Microbiology Letters, 239(2), pp. 235–240. http://dx.doi.org/10.1016/j.femsle.2004.08.038

Hoffmaster, A. R., Fitzgerald, C. C., Ribot, E., Mayer, L. W. and Popovic, T. (2002) ‘Molecular subtyping of Bacillus anthracis and the 2001 bioterrorism-associated Anthrax outbreak, United States’, Emerging Infectious Diseases, 8(10), pp. 1111–1116. http://dx.doi.org/10.3201/eid0810.020394

Keim, P., Kalif, A., Schupp, J., Hill, K., Travis, S. E., Richmond, K., Adair, D. M., Hugh-Jones, M., Kuske, C. R. and Jackson, P. (1997) ‘Molecular evolution and diversity in Bacillus anthracis as detected by amplified fragment length polymorphism markers’, Journal of Bacteriology, 179(3), pp. 818–824. http://dx.doi.org/10.1128/jb.179.3.818-824.1997

Keim, P., Price, L. B., Klevytska, A. M., Smith, K. L., Schupp, J. M., Okinaka, R., Jackson, P. J. and Hugh-Jones, M. E. (2000) ‘Multiple-Locus Variable-number tandem repeat Analysis reveals genetic relationships within Bacillus anthracis’, Journal of Bacteriology, 182(10), pp. 2928–2936. http://dx.doi.org/10.1128/JB.182.10.2928-2936.2000

Keim, P., Van Ert, M. N., Pearson, T., Vogler, A. J., Huynh, L. Y. and Wagner, D. M. (2004) ‘Anthrax molecular epidemiology and forensics: using the appropriate marker for different evolutionary scales’, Infection, Genetics and Evolution, 4(3), pp. 205–213. http://dx.doi.org/10.1016/j.meegid.2004.02.005

Korotich, A. S. and Pogrebnyak, L. I. (1976) Anthrax [Sibirskaya yazva]. Kiev: Urozhay. [in Russian]

Le Flèche, P., Hauck, Y., Onteniente, L., Prieur, A., Denoeud, F., Ramisse, V., Sylvestre, P., Benson, G., Ramisse, F. and Vergnaud, G. (2001) ‘A tandem repeats database for bacterial genomes: application to the genotyping of Yersinia pestis and Bacillus anthracis’, BMC Microbiology, 1, p. 2. http://dx.doi.org/10.1186/1471-2180-1-2

Leski, T. A., Caswell, C. C., Pawlowski, M., Klinke, D. J., Bujnicki, J. M., Hart, S. J. and Lukomski, S. (2009) ‘Identification and classification of bcl genes and proteins of Bacillus cereus group organisms and their application in Bacillus anthracis detection and fingerprinting’, Applied and Environmental Microbiology, 75(22), pp. 7163–7172. http://dx.doi.org/10.1128/AEM.01069-09

Lista, F., Faggioni, G., Valjevac, S., Ciammaruconi, A., Vaissaire, J., Le Doujet, C., Gorgé, O., De Santis, R., Carattoli, A., Ciervo, A., Fasanella, A., Orsini, F., D’Amelio, R., Pourcel, C., Cassone, A. and Vergnaud, G. (2006) ‘Genotyping of Bacillus anthracis strains based on automated capillary 25-loci Multiple Locus Variable-number tandem repeats Analysis’, BMC Microbiology, 6(1), p. 33. http://dx.doi.org/10.1186/1471-2180-6-33

Maly, V. P., Lyadova, T. I., Bryatko, N. V., Volhina, S. I. and Morozov, V. Z. (2013) ‘Anthrax in Ukraine’ [Sybirka v Ukraini], Infectious Diseases [Infektsiini khvoroby], 3, pp. 107–110. [in Ukrainian]

Martin J. W., Christopher G. W., Eitzen E. M. (2007) ‘Chapter 1. History of biological weapons: from poisoned darts to intentional epidemics’, in Dembek Z. F. (ed.) Medical Aspects of Chemical and Biological Warfare. Falls Church, Virginia; Washington, D. C.: Office of the Surgeon General; Borden Institute, pp. 1–20. Available at: http://purl.access.gpo.gov/GPO/LPS101470

Pearson, T., Busch, J. D., Ravel, J., Read, T. D., Rhoton, S. D., U’Ren, J. M., Simonson, T. S., Kachur, S. M., Leadem, R. R., Cardon, M. L., Van Ert, M. N., Huynh, L. Y., Fraser, C. M. and Keim, P. (2004) ‘Phylogenetic discovery bias in Bacillus anthracis using Single-Nucleotide Polymorphisms from whole-genome sequencing’, Proceedings of the National Academy of Sciences, 101(37), pp. 13536–13541. http://dx.doi.org/10.1073/pnas.0403844101

Pearson, T., Okinaka, R. T., Foster, J. T. and Keim, P. (2009) ‘Phylogenetic understanding of clonal populations in an era of whole genome sequencing’, Infection, Genetics and Evolution, 9(5), pp. 1010–1019. http://dx.doi.org/10.1016/j.meegid.2009.05.014

Purcell, B. K., Worsham, P. L. and Freidlander, A. M. (2007) ‘Chapter 4. Anthrax’, in Dembek Z. F. (ed.) Medical Aspects of Chemical and Biological Warfare. Falls Church, Virginia; Washington, D. C.: Office of the Surgeon General; Borden Institute, pp. 69–90. Available at: http://purl.access.gpo.gov/GPO/LPS101470

Rume, F. I., Ahsan, C. R., Biswas, P. K., Yasmin, M., Braun, P., Walter, M. C., Antwerpen, M., Grass, G. and Hanczaruk, M. (2016) ‘Unexpected genomic relationships between Bacillus anthracis strains from Bangladesh and Central Europe’, Infection, Genetics and Evolution, 45, pp. 66–74. http://dx.doi.org/10.1016/j.meegid.2016.08.017

Smith, K. L., DeVos, V., Bryden, H., Price, L. B., Hugh-Jones, M. E. and Keim, P. (2000) ‘Bacillus anthracis diversity in Kruger National Park’, Journal of Clinical Microbiology, 38(10), pp. 3780–3784. Available at: https://jcm.asm.org/content/38/10/3780

Thierry, S., Tourterel, C., Le Flèche, P., Derzelle, S., Dekhil, N., Mendy, C., Colaneri, C., Vergnaud, G. and Madani, N. (2014) ‘Genotyping of French Bacillus anthracis strains based on 31-loci Multi Locus VNTR Analysis: epidemiology, marker evaluation, and update of the internet genotype database’, PLoS ONE, 9(6), p. e95131. http://dx.doi.org/10.1371/journal.pone.0095131

Van Ert, M. N., Easterday, W. R., Huynh, L. Y., Okinaka, R. T., Hugh-Jones, M. E., Ravel, J., Zanecki, S. R., Pearson, T., Simonson, T. S., U’Ren, J. M., Kachur, S. M., Leadem-Dougherty, R. R., Rhoton, S. D., Zinser, G., Farlow, J., Coker, P. R., Smith, K. L., Wang, B., Kenefic, L. J., Fraser-Liggett, C. M., Wagner, D. M. and Keim, P. (2007) ‘Global genetic population structure of Bacillus anthracis’, PLoS ONE, 2(5), p. e461. http://dx.doi.org/10.1371/journal.pone.0000461

Zaviriuha, H. A., Yanenko, U. M. and Zaviriuha, A. I. (2015) ‘Prevention of Anthrax epizootic appearance in uncontrolled risk areas using pathogens exotoxins’ [Poperedzhennia vynyknennia epizootii shchodo sybirky v nekontrolovanykh zonakh ryzyku iz zastosuvanniam ekzotoksyniv patohennykh mikroorhanizmiv], Scientific Herald of the National University of Life and Environmental Sciences of Ukraine [Naukovyi visnyk Natsionalnoho universytetu bioresursiv i pryrodokorystuvannia Ukrainy], 227, pp. 87–94. Available at: https://elibrary.ru/item.asp?id=25065083. [in Ukrainian]