Skip to main content

Advertisement

Springer Nature Link
Log in

Molecular detection of Mycobacterium tuberculosis complex in the 8th century skeletal remains from the territory of Slovakia

  • Published:
Biologia Aims and scope Submit manuscript

Abstract

DNA was extracted using a Silica Bead Extraction kit from bone samples taken from a Slavonic-Avar individual found at the archaeological site of Cífer-Pác. The analysed skeletal remains from the grave number 62/79 belong to a young adult male (20–30 years at death) and are dated to the 8th–9th century anno Domini. The isolated ancient DNA (aDNA) was amplified by a targeted PCR with a primer pair designed to recognize the Mycobacterium tuberculosis complex insertion sequence IS6110. The aim of this molecular approach was to test and optimize a methodology for aDNA M. tuberculosis complex extraction from bone samples with osteological evidence of tuberculosis. Despite of the currently biased authenticity of the mentioned fragment, in this case study we prove that macroscopic evidence for tuberculosis additionally supported by a positive result of molecular testing can be considered authentic enough to be the proof of a tuberculosis infection caused by MTBC (Mycobacterium tuberculosis complex) bacteria when additional skeletal trauma and changes potentially caused by MOTT (mycobacteria other than tuberculosis) bacteria can be excluded. Positivity was confirmed in all of the three samples (thoracic vertebrae, lumbar vertebrae and right femur). Our results confirmed the diagnosis of tuberculosis of the spine and right hip joint. This is the first molecular evidence for the occurrence of tuberculosis on the territory of Slovakia. Through this bio-molecular approach we wish to provide a basis for aDNA examinations on other skeletal collections and provide epidemiological data concerning historical populations living on the territory of Slovakia.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Subscribe and save

Springer+
from $39.99 /Month
  • Starting from 10 chapters or articles per month
  • Access and download chapters and articles from more than 300k books and 2,500 journals
  • Cancel anytime
View plans

Buy Now

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

Explore related subjects

Discover the latest articles and news from researchers in related subjects, suggested using machine learning.

Abbreviations

AD:

anno Domini

aDNA:

ancient DNA

BC:

before Christ

MOTT:

mycobacteria other than tuberculosis

MTBC:

Mycobacterium tuberculosis complex

NTM:

non-tuberculous mycobacteria

TBC:

tuberculosis

References

  • Aranaz A., Cousins D., Mateos A. & Domínguez L. 2003. Elevation of Mycobacterium tuberculosis subsp. caprae to species rank as Mycobacterium caprae comb. nov., sp. nov. Int. J. Syst. Evol. Microbiol. 53: 1785–1789.

    Article  CAS  Google Scholar 

  • Aufderheide A.C. & Rodríguez-Martin C. 1998. The Cambridge Encyclopedia of Human Paleopathology. Cambridge University Press, Cambridge, 481 pp.

    Google Scholar 

  • Baron H., Hummel S. & Herrmann B. 1996. Mycobacterium tuberculosis complex DNA in ancient human bones. J. Archaeol. Sci. 23: 667–671.

    Article  Google Scholar 

  • Benuš R., Baldovic M. & Masnicová S. 2002. A probable case of tuberculosis in a Slavonic-Avar population from Cífer-Pác (West Slovakia). Bull. Slov. Antropol. Spol. SAV 5: 1–9 (In Slovak).

    Google Scholar 

  • Bos K.I., Harkins K.M., Herbig A., Coscolla M., Weber N., Comas I., Forrest S.A., Bryant J.M., Harris S.R., Schuenemann V.J., Campbell T.J., Majander K., Wilbur A.K., Guichon R.A., Wolfe Steadman D.L., Cook D.C., Niemann S., Behr M.A., Zumarraga M., Bastida R., Huson D., Nieselt K., Young D., Parkhill J., Buikstra J.E., Gagneux S., Stone A.C. Krause J. 2014. Pre-Columbian mycobacterial genomes reveal seals as a source of New World human tuberculosis. Nature 514: 494–497.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Burger J., Hummel S., Herrmann B. & Henke W. 1999. DNA preservation: a microsatellite-DNA study on ancient skeletal remains. Electrophoresis 20: 1722–1728.

    Article  CAS  PubMed  Google Scholar 

  • Canci A., Minozzi S. & Borgognini Tarli S.M. 1996. New evidence of tuberculous spondylitis from Neolithic Liguria (Italy). Int. J. Osteoarchaeol. 6: 497–501.

    Article  Google Scholar 

  • Comas I., Coscolla M., Luo T., Borrell S., Holt K.E., Kato-Maeda M., Parkhill J., Malla B., Berg S., Thwaites G., Yeboah-Manu D., Bothamley G., Mei J., Wei L., Bentley S., Harris S.R., Niemann S., Diel R., Aseffa A., Gao Q., Young D. & Gagneux S. 2013. Out-of-Africa migration and Neolithic coexpansion of Mycobacterium tuberculosis with modern humans. Nat. Genet. 45: 1176–1182.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Coros A., DeConno E. & Derbyshire K.M. 2008. IS6110, a Mycobacterium tuberculosis complex-specific insertion sequence, is also present in the genome of Mycobacterium smegmatis, suggestive of lateral gene transfer among mycobacterial species. J. Clin. Microbiol. 190: 3408–3410.

    CAS  Google Scholar 

  • Eisenach K.D., Cave M.D., Bates J.H. & Crawford J.T. 1990. Polymerase chain reaction amplification of a repetitive DNA sequence specific for Mycobacterium tuberculosis. J. Infect. Dis. 161: 977–981.

    Article  CAS  PubMed  Google Scholar 

  • Elsayed S. & Read R. 2006. Mycobacterium haemophylum osteomyelitis: case report and review of the literature. BMC Infect. Dis. 6: 70.

    Article  PubMed  PubMed Central  Google Scholar 

  • Fairhurst R.M., Kubak B.M., Pegues D.A., Moriguchi J.D., Han K.F., Haley J.C. & Kobashigawa J.A. 2002. Mycobacterium haemophilum infections in heart transplant recipients: case report and review of the literature. Am. J. Transplant. 5: 476–479.

    Article  Google Scholar 

  • Gutiérrez M., Samper S., Jiménez M. S., van Embden J.D.A., Marin J.F. & Martín C. 1997. Identification by spoligotyping of a caprine genotype in Mycobacterium bovis strains causing human tuberculosis. J. Clin. Microbiol. 35: 3328–3330.

    PubMed  PubMed Central  Google Scholar 

  • Haynes S., Searle J.B., Bretman A. & Dobney K.M. 2002. Bone preservation and ancient DNA: the application of screening methods for predicting DNA survival. J. Archaeol. Sci. 29: 585–592.

    Article  Google Scholar 

  • Hirsch R., Miller S.M., Kazi S., Cate T.R. & Reveille J.D. 1996. Human immunodeficiency virus-associated atypical mycobacterial skeletal infections. Semin. Arthritis Reum. 25: 347–356.

    Article  CAS  Google Scholar 

  • Hofreiter M., Serre D., Poinar H.N., Kuch M. & Pääbo S. 2001. Ancient DNAU. Nat. Rev. Genet. 2: 353–359.

    Article  CAS  PubMed  Google Scholar 

  • Kemp B.M., Malhi R.S., McDonough J., Bolnick D.A., Eshleman J.A., Rickards O., Martinez-Labarga C., Johnson J.R., Lorenz J.G., Dixon E.J., Fifield T.E., Heaton T.H., Worl R. & Smith D.G. 2007. Genetic analysis of early holocene skeletal remains from Alaska and its implications for the settlement of the Americas. Am. J. Phys. Anthropol. 132: 605–621.

    Article  PubMed  Google Scholar 

  • Kiehn T.E. & White M. 1994. Mycobacterium haemophylum: an emerging pathogen. Eur. J. Clin. Microbiol. 11: 925–931.

    Article  Google Scholar 

  • Kiers A., Klarenbeek A., Mendelts B., Van Soolingen D. & Koëter G. 2008. Transmission of Mycobacterium pinnipedii to humans in a zoo with marine mammals. Int. J. Tuberc. Lung Dis. 12: 1469–1473.

    CAS  PubMed  Google Scholar 

  • Masson M., Bereczki Z., Molnár E., Donoghue H.E., Minnikin D.E., Lee O.Y., Wu H.H., Besra G.S., Bull I.D. & Pálfi G. 2015. 7000 year-old tuberculosis cases from Hungary — Osteological and biomolecular evidence. Tuberculosis (Edinb) 95 (Suppl. 1): S13–S17.

    Article  Google Scholar 

  • Müller R., Roberts C.A. & Brown T.A. 2014. Biomolecular identification of ancient Mycobacterium tuberculosis complex DNA in human remains from Britain and continental Europe. Am. J. Phys. Anthropol. 153: 178–189.

    Article  PubMed  Google Scholar 

  • Müller R., Roberts C.A. & Brown T. 2014. Genotyping of ancient Mycobacterium tuberculosis strains reveals historic genetic diversity. Proc. Biol. Sci. 281: 20133236.

  • Müller R., Roberts C.A. & Brown T.A. 2015. Complications in the study of ancient tuberculosis: non-specificity of IS6110 PCRs. Sci. Technol. Archaeol. Res. 1: 1–8.

    Google Scholar 

  • Nicklisch N., Maixner F., Ganslmeier R., Friederich S., Dresely V., Meller H., Zink A. & Alt K. 2012. Rib lesions in skeletons from early Neolithic sites in central Germany: on the trail of tuberculosis at the onset of agriculture. Am. J. Phys. Anthropol. 149: 391–404.

    Article  PubMed  Google Scholar 

  • Ortner D.J. 2003. Methods used in the analysis of skeletal lesions, pp. 45–55. In: Identification of Pathological Conditions in Human Skeletal Remains. Academic Press, San Diego.

    Chapter  Google Scholar 

  • Pálfi G., Maixner F., Maczel M., Molnár E., Pósa A., Kristóf L. A., Marcsik A., Balázs J., Masson M., Paja L., Palkó A., Szentgyörgyi R., Nerlich A., Zink A. & Dutour O. 2015. Unusual spinal tuberculosis in an Avar Age skeleton (Csongrád-Felgyȍ, Ürmös-tanya, Hungary): a morphological and biomolecular study. Tuberculosis (Edinb) 95 (Suppl. 1): S29–S34.

    Article  Google Scholar 

  • Pruvost M., Schwarz R., Bessa Correia V., Champlot S., Braguier S., Morel N., Fernanez-Jalvo Y., Grange T. & Geigl E.M. 2007. Freshly excavated fossil bones are best for amplification of ancient DNA. Proc. Natl. Acad. Sci. USA 104: 739–744.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Resnick D. & Niwayama G. 1988. Diagnosis of Bone and Joint Disorders. Saunders, Philadelphia, 4944 pp.

    Google Scholar 

  • Resnick D. & Niwayama G. 1995. Osteomyelitis, septic arthritis, and soft tissue infection: organisms, pp. 2419–2447. In: Resnick D. (ed.), Diagnosis of Bone and Joint Disorders, Saunders, Edinburgh.

  • Ritis K., Tzoanopoulos D., Speletas M., Papadopoulos E., Arvanitidis K., Kartali S. & Sideras P. 2000. Amplification of IS6110 sequence for detection of Mycobacterium tuberculosis complex in HIV-negative patients with fever of unknown origin (FUO) and evidence of extrapulmonary disease. J. Intern. Med. 248: 415–424.

    Article  CAS  PubMed  Google Scholar 

  • Rohland N. & Hofreiter M. 2007. Ancient DNA extraction from bones and teeth. Nature Protoc. 2: 1756–1762.

    Article  CAS  Google Scholar 

  • Sankar S., Kuppanan S., Balakrishnan B. & Nandagopal B. 2011. Analysis of sequence diversity among IS6110 sequence of Mycobacterium tuberculosis: possible implications for PCR based detection. Bioinformation 6: 283–285.

    Article  PubMed  PubMed Central  Google Scholar 

  • Saubolle M.A., Kiehn T.E., White M.H., Rudinsky M.F. & Armstrong D. 1996. Mycobacterium haemophylum: microbiology and expanding clinical and geographic spectra of disease in humans. Clin. Microbiol. Rev. 9: 435–447.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Shimizu H., Mizuno Y., Nakamura I., Fukushima S., Endo K. & Matsumoto T. 2013. Vertebral osteomyelitis caused by nontuberculous mycobacteria: case reports and review. J. Infect. Chemother. 19: 972–977.

    Google Scholar 

  • Staššíková-Štukovská D., Thurzo M., Šefcáková A. & Lietava J. 2006. Archaeological and paleopathological interpretations of an atypical grave No. 22/86 excavated at the early medieval TBC detection in an 8th century skeleton from Slovakia 619 cemetery of Borovce (district Pieštany, Slovakia), pp. 208–221. In: In Service to Archaeology. Proceedings dedicated to Doc. PhDr. Karel Valoch, DrSc. Muzejní a vlastivědná spolecnost v Brně, Brno (In Slovak).

    Google Scholar 

  • Straus W.L., Ostroff S.M., Jernigan D.B., Kiehn T.E., Sordillo E.M., Armstrong D., Boone N., Schneider N., Kilburn J.O., Silcox V.A., LaBombardi V., Good R.C. 1994. Cinical and epidemiologic characteristics of Mycobacterium haemophilum, an emerging pathogen in immunocompromised patients. Ann. Intern. Med. 120: 118–125.

    Article  CAS  PubMed  Google Scholar 

  • Thacker T.C., Harris B., Palmer M.V. & Waters W.R. 2011. Improved specificity for detection of Mycobacterium bovis in fresh tissue using IS6110 real-time PCR. BMC Vet. Res. 7: 50.

    Article  PubMed  PubMed Central  Google Scholar 

  • Theodorou D.J., Theodorou S.J., Kikatsubata Y., Sartoris D.J. & Resnick D. 2001. Imaging characteristics and epidemiologic features of atypical mycobacterial infections involving the musculoskeletal system. AMR Am. J. Roentgenol. 176: 341–349.

    Article  CAS  Google Scholar 

  • Thierry D., Cave M.D., Eisenach K.D., Crawford J.T., Bates J.H., Gicquel B. & Guesdon J.L. 1990. IS6110, an IS-like element of Mycobacterium tuberculosis complex. Nucleic Acids Res. 18: 188.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Van Soolingen D., Hoogenboezem T., de Haas P.E.W., Hermans P.W.M., Koedam M.A., Teppema K.S., Brennan P.J., Besra G.S., Portaels F., Top J., Schouls L.M. & van Embden J.D.A. 1997. A novel pathogenic taxon of the Mycobacterium tuberculosis complex, Canetti: characterization of an exceptional isolate from Africa. Int. J. Syst. Bacteriol. 47: 1236–1245.

    Article  Google Scholar 

  • Van Soolingen D., van der Zanden A.G.M., de Haas P.E.W., Noordhoek G.T., Kiers A., Foudraine N.A., Portaels F., Kolk A.H.J., Kremer K. & van Embden J.D.A. 1998. Diagnosis of Mycobacterium microti infections among humans by using novel genetic markers. J. Clin. Microbil. 36: 1840–1845.

    Google Scholar 

  • White M.H., Papadopoulos E.B., Small T.N., Kiehn T.E. & Armstrong D. 1995. Mycobacterium haemophilum infections in bone marrow transplant recipients. Transplantation 60: 957–960.

    Article  CAS  PubMed  Google Scholar 

  • Wirth T., Hildebrand F., Allix-Béguec C., Wölbeling F., Kubica T., Kremer K., van Soolingen D., Rüsch-Gerdes S., Locht C., Brisse S., Meyer A., Supply P. & Niemann S. 2008. Origin, spread and demography of the Mycobacterium tuberculosis complex. PloS Pathog. 4: e1000160.

  • Wolinsky E. 1992. Mycobacterial diseases other than tuberculosis. Clin. Infect. Dis. 15: 1–12.

    Article  CAS  PubMed  Google Scholar 

  • Woods G. 1987. Mycobacteria other than Mycobacterium tuberculosis: review of microbiologic and clinical aspects. Rev. Infect. Dis. 9: 275–294.

    Article  CAS  PubMed  Google Scholar 

  • Zink A.R., Molnár E., Motamedi N., Pálfi G., Marcsik A. & Nerlich A.G. 2007. Molecular history of tuberculosis from ancient mummies and skeletons. Int. J. Osteoarchaeol. 17: 380–391.

    Article  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Department of Anthropology, Faculty of Natural Sciences, Comenius University in Bratislava, Mlynská dolina B2, SK-84215, Bratislava, Slovakia

    Klaudia Kyselicová & Radoslav Beňuš

  2. Department of Molecular Biology, Faculty of Natural Sciences, Comenius University in Bratislava, Mlynská dolina B2, SK-84215, Bratislava, Slovakia

    Lukáš Šebest, Csaba Bognár, Marián Baldovič & Ľudevít Kádaši

  3. Department of Biochemistry, Faculty of Natural Sciences, Comenius University in Bratislava, Mlynská dolina CH1, SK-84215, Bratislava, Slovakia

    Michal Šarkan

  4. Institute for Clinical and Translational Research, Biomedical Research Centre, Slovak Academy of Sciences, Dúbravská cesta 9, SK-84505, Bratislava, Slovakia

    Ľudevít Kádaši

Authors
  1. Klaudia Kyselicová
    View author publications

    Search author on:PubMed Google Scholar

  2. Lukáš Šebest
    View author publications

    Search author on:PubMed Google Scholar

  3. Csaba Bognár
    View author publications

    Search author on:PubMed Google Scholar

  4. Michal Šarkan
    View author publications

    Search author on:PubMed Google Scholar

  5. Marián Baldovič
    View author publications

    Search author on:PubMed Google Scholar

  6. Radoslav Beňuš
    View author publications

    Search author on:PubMed Google Scholar

  7. Ľudevít Kádaši
    View author publications

    Search author on:PubMed Google Scholar

Corresponding author

Correspondence to Klaudia Kyselicová.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Kyselicová, K., Šebest, L., Bognár, C. et al. Molecular detection of Mycobacterium tuberculosis complex in the 8th century skeletal remains from the territory of Slovakia. Biologia 71, 613–619 (2016). https://doi.org/10.1515/biolog-2016-0088

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1515/biolog-2016-0088

Key words