Vet Med - Czech, 2017, 62(3):153-168 | DOI: 10.17221/126/2016-VETMED

Detection of mycobacteria in the environment of the Moravian Karst (Bull Rock Cave and the relevant water catchment area): the impact of water sediment, earthworm castings and bat guanoOriginal Paper

H. Modra1, M. Bartos2, P. Hribova3, V. Ulmann4, D. Hubelova1, O. Konecny1, M. Gersl5, J. Kudelka5, D. Voros5, I. Pavlik*,1
1 Faculty of Regional Development and International Studies, Mendel University in Brno, Brno, Czech Republic
2 Faculty of Sciences, Masaryk University, Brno, Czech Republic
3 Faculty of Pharmacy, University of Veterinary and Pharmaceutical Sciences Brno, Brno, Czech Republic
4 Public Health Institute Ostrava, Ostrava, Czech Republic
5 Faculty of AgriSciences, Mendel University in Brno, Brno, Czech Republic

The presence of mycobacteria was studied in Bull Rock Cave ("Byci skala") and the water catchment area of Jedovnice Brook ("Jedovnicky potok") using direct microscopy after Ziehl-Neelsen (ZN) staining, culture examination and molecular techniques. Mycobacteria were detected in 47.1% of a total of 68 samples. The mycobacterial genes hsp65 and dnaA were detected and sequenced in 37 (74.0%) out of the 50 cave environmental samples and in 10 (55.6%) out of the 18 samples of water catchment sediments. Nine species of slowly growing mycobacteria (M. terrae, M. arupense, M. gordonae, M. lentiflavum, M. parascrofulaceum, M. parmense, M. saskatchewanense, M. simiae and M. xenopi) and two subsp. (M. avium subsp. avium and M. avium subsp. hominissuis) were detected. Fourteen species of rapidly growing mycobacteria (M. chelonae, M. chubuense, M. poriferae, M. flavescens, M. fortuitum, M. porcinum, M. rhodesiae, M. gilvum, M. goodii, M. peregrinum, M. mageritense, M. vanbaalenii, M. gadium and M. insubricum) were detected. The highest mycobacterial presence was documented by ZN staining and/or culture examinations in earthworm castings and bat guano (73.3% positivity out of the 15 samples) in the cave environment and in the water sediments collected under the outflow from the wastewater treatment plants (77.8% positivity out of nine samples). The highest total organic carbon (TOC) was detected in wooden material and earthworm castings with pH values between 5.0 and 7.7 in the cave environment and in water sediments collected under the outflow from the wastewater treatment plants with pH between 5.8 and 7.0. It could be concluded that the karst cave environment with its running surface water contaminated with different microorganisms or chemical substances creates favourable conditions not only for animals (especially earthworms) but also for mycobacteria. This fact is also demonstrated by the presence of these mycobacteria in the cave environment mainly in earthworm castings and bat guano.

Keywords: bat faeces; environmentally derived mycobacteria; potentially pathogenic mycobacteria; ecology; geomycobacteriology; biospeleology; cave fauna; epidemiology

Published: March 31, 2017  Show citation

ACS AIP APA ASA Harvard Chicago Chicago Notes IEEE ISO690 MLA NLM Turabian Vancouver
Modra H, Bartos M, Hribova P, Ulmann V, Hubelova D, Konecny O, et al.. Detection of mycobacteria in the environment of the Moravian Karst (Bull Rock Cave and the relevant water catchment area): the impact of water sediment, earthworm castings and bat guano. Vet Med - Czech. 2017;62(3):153-168. doi: 10.17221/126/2016-VETMED.
Share...
Download citation
  • BibTeX (.bib)
  • Bookends (.ris)
  • EasyBib (.ris)
  • EndNote (.enw)
  • EndNote 8 (.xml)
  • ISI WoS (.isi)
  • Medlars (.medlars)
  • Mendeley (.ris)
  • MODS (.xml)
  • Papers (.ris)
  • RefWorks (.txt)
  • RefManager (.ris)
  • RIS (.ris)
  • MS Word (.xml)
  • Zotero (.ris)
    • Open full article

    References

    1. Andreo B, Goldscheider N, Vadillo I, Vias JM, Neukum C, Sinreich M, Jimenez P, Brechenmacher J, Carrasco F, Hotzl H, Perles MJ, Zwahlen F (2006): Karst groundwater protection: First application of a Pan-European Approach to vulnerability, hazard and risk mapping in the Sierra de Libar (Southern Spain). Science of the Total Environment 357, 54-73. Go to original source... Go to PubMed...
    2. Blackwood KS, He C, Gunton J, Turenne CY, Wolfe J, Kabani AM (2000): Evaluation of recA sequences for identification of Mycobacterium species. Journal of Clinical Microbiology 38, 2846-2852. Go to original source... Go to PubMed...
    3. Breitbart M, Hoare A, Nitti A, Siefert J, Haynes M, Dinsdale E, Edwards R, Souza V, Rohwer F, Hollander D (2009): Metagenomic and stable isotopic analyses of modern freshwater microbialites in Cuatro Cienegas, Mexico. Environmental Microbiology 11, 16-34. Go to original source... Go to PubMed...
    4. Brooks RW, Parker BC, Gruft H, Falkinham III JO (1984): Epidemiology of infection by non-tuberculous mycobacteria. V. Numbers in eastern United States soils and correlation with soil characteristics. American Review of Respiratory Diseases 130, 630-633.
    5. Burkhardt R, Gregor VA, Novak K (1972): Exceptional flood in Bull Rock Cave (in Czech). Vlastivedne Zpravy z Adamova a Okoli 6, 4-8.
    6. Butscher C, Auckenthaler A, Scheidler S, Huggenberger P (2011): Validation of a numerical indicator of microbial contamination for karst springs. Ground Water 49, 66-76. Go to original source... Go to PubMed...
    7. Cernosvitov L (1935): Monographie des Tschechoslovakischen Lumbriciden. Archiv pro Prirodovedecky Vyzkum Cech 19, 86 pp.
    8. Coenye T, Vandamme P (2003): Intragenomic heterogeneity between multiple 16S ribosomal RNA operons in sequenced bacterial genomes. FEMS Microbiology Letters 228, 45-49. Go to original source... Go to PubMed...
    9. CSO - Czech Statistical Office (2016a): Spatial analytical materials. Available at https://www.czso.cz/csu/czso/csu_a_uzemne_analyticke_podklady (Accessed July 21, 2016).
    10. CSO - Czech Statistical Office (2016b): Public Database. Available at https://vdb.czso.cz/vdbvo2/faces/en/index. jsf (Accessed July 21, 2016).
    11. CSO - Czech Statistical Office (2016c): Population and Housing Census 2011. Available at https://www.czso.cz/csu/sldb/home (Accessed July 21, 2016).
    12. De Mandal S, Panda AK, Lalnunmawii E, Bisht SS, Kumar NS (2015a): Illumina-based analysis of bacterial community in Khuangcherapuk Cave of Mizoram, Northeast India. Genome Data 5, 13-14. Go to original source... Go to PubMed...
    13. De Mandal S, Sanga Z, Kumar NS (2015b): Metagenome sequencing reveals Rhodococcus dominance in Farpuk Cave, Mizoram, India, an Eastern Himalayan biodiversity hot spot region. Genome Announcements 3, doi: 10.1128/genomeA.00610-15. Go to original source... Go to PubMed...
    14. Einsiedl F, Radke M, Maloszewski P (2010): Occurrence and transport of pharmaceuticals in a karst groundwater system affected by domestic wastewater treatment plants. Journal of Contaminant Hydrology 117, 26-36. Go to original source... Go to PubMed...
    15. Engbaek HC, Vergmann B, Weis Bentzon B (1967): The sodium lauryl sulphate method in culturing sputum for mycobacteria. Scandinavian Journal of Respiratory Diseases 48, 268-284.
    16. Faimon J, Stelcl J, Sas D (2006): Anthropogenic CO2-flux into cave atmosphere and its environmental impact: A case study in the Cisarska Cave (Moravian Karst, Czech Republic). Science of the Total Environment 369, 231-245. Go to original source... Go to PubMed...
    17. Fischer OA, Matlova L, Bartl J, Dvorska L, Svastova P, Du Maine R, Melicharek I, Bartos M, Pavlik I (2003): Earthworms (Oligochaeta, Lumbricidae) and mycobacteria. Veterinary Microbiology 91, 325-338. Go to original source... Go to PubMed...
    18. Hertod-Todtenfeldt JF (1669): Tartaro-Mastix Moraviae, per quem rariora & admiranda a natura in faecundo huius regionis gremio effusa, comprimis tartarus, illiusque effectus morbosi curiose examinantur, & cura tam therapeutica quam prophylactica proponitur. Vienna Austriae 263, 258.
    19. Hill CA, Polyak VJ (2014): Karst piracy: A mechanism for integrating the Colorado River across the Kaibab uplift, Grand Canyon, Arizona, USA. Geosphere 10, 627-640. Go to original source...
    20. Hrabe S (1929): Zwei neue lumbriculiden-arten, sowie einige bemerkungen zur systematik einiger bereits bekannter. Zoologischer Anzeiger 84, 9-21.
    21. Hrabe S (1938): Trichodrilus moravicus und Claparedei, neue Lumbriculiden. Zoologischer Anzeiger 121, 73-85.
    22. Hromas J (2009): Caves (in Czech). In: Mackovcin P, Sedlacek M (eds): Protected Areas in the Czech Republic. Agentura Ochrany Prirody a Krajiny CR a EkoCentrum Brno, Praha. 608 pp.
    23. Hu M, Wang X, Wen X, Xia Y (2012): Microbial community structures in different wastewater treatment plants as revealed by 454-pyrosequencing analysis. Bioresource Technology 117, 72-79. Go to original source... Go to PubMed...
    24. Hubka V, Novakova A, Kolarik M, Jurjevic Z, Peterson SW (2015): Revision of Aspergillus section Flavipedes: seven new species and proposal of section Jani sect. nov. Mycologia 107, 169-208. Go to original source... Go to PubMed...
    25. Jurado V, Laiz L, Rodriguez-Nava V, Boiron P, Hermosin B, Sanchez-Moral S, Saiz-Jimenez C (2010): Pathogenic and opportunistic microorganisms in caves. International Journal of Speleology 39, 15-24. Go to original source...
    26. Jurende CJ (1835): Die Wunder der Unterwelt. Jurende's Vaterlandischer Pilger, Brunn 22, 61-101.
    27. Kapur V, Li LL, Hamrick MR, Plikaytis BB, Shinnick TM, Telenti A, Jacobs Jr WR, Banerjee A, Cole S, Yuen KY, Clarridge III JE, Kreiswirth BN, Musser JM (1995): Rapid Mycobacterium species assignment and unambiguous identification of mutations associated with antimicrobial resistance in Mycobacterium tuberculosis by automated DNA sequencing. Archives of Pathology and Laboratory Medicine 119, 131-138.
    28. Kazda J (ed.) (2000): The Ecology of Mycobacteria. Kluwer Academic Publishers. 72 pp. Go to original source...
    29. Kazda J, Pavlik I, Falkinham JO, Hruska K (eds) (2009): The Ecology of Mycobacteria: Impact on Animal's and Human's Health. 1st edn. Springer, New York. 520 pp. Go to original source...
    30. Korzeniewska E (2011): Emission of bacteria and fungi in the air from wastewater treatment plants - a review. Frontiers in Bioscience (Scholar Edition) 3, 393-407. Go to original source... Go to PubMed...
    31. Kriz M (1891): Die hohlen in den mahrischen Devonkalken und ihre vorzeit. I. Die slouperhohlen. Jahrbuch der Kaiserlich Koniglichen Geologischen Reichsanstalt, Wien 41, 443-570.
    32. Kriz M (1892): Die Hohlen in den mahrischen Devonkalken und ihre Vorzeit II-IV. Jahrbuch der Kaiserlich Koniglichen Geologischen Reichsanstalt, Wien 42, 463-626.
    33. LPSN - List of Prokaryotic Names with Standing in Nomenclature (2016): Available at http://www.bacterio.net/mycobacterium.html (Accessed July 9, 2016).
    34. Lukesova A, Novakova A (2009): Interactions between the soil micro-flora and invertebrates in Slovak and Moravian caves. In: 9th Central European Workshop on Soil Zoology, Ceske Budejovice, Czech Republic. April 17-20, 2007. 89-96.
    35. Mayer J (ed.) (1781): Versuch einer Beschreibung der Gegend um Sluppe in Mahren. Schriften der Berlinischen Gesellschaft Naturforschender Freunde, Bd. II, Berlin. 56-65. Go to original source...
    36. McNabb A, Eisler D, Adie K, Amos M, Rodrigues M, Stephens G, Black WA, Isaac-Renton J (2004): Assessment of partial sequencing of the 65-kilodalton heat shock protein gene (hsp65) for routine identification of Mycobacterium species isolated from clinical sources. Journal of Clinical Microbiology 42, 3000-3011. Go to original source... Go to PubMed...
    37. McNabb A, Adie K, Rodrigues M, Black WA, Isaac-Renton J (2006): Direct identification of mycobacteria in primary liquid detection media by partial sequencing of the 65-kilodalton heat shock protein gene. Journal of Clinical Microbiology 44, 60-66. Go to original source... Go to PubMed...
    38. Migon P (2011): Development of karst phenomena for geotourism in the Moravian Karst (Czech Republic). Geoturism, 3-24.
    39. Mohammadi Z, Shoja A (2014): Effect of annual rainfall amount on characteristics of karst spring hydrograph. Carbonates and Evaporites 29, 279-289. Go to original source...
    40. Moravkova M, Hlozek P, Beran V, Pavlik I, Preziuso S, Cuteri V, Bartos M (2008): Strategy for the detection and differentiation of Mycobacterium avium species in isolates and heavily infected tissues. Research in Veterinary Science 85, 257-264. Go to original source... Go to PubMed...
    41. Mukai T, Miyamoto Y, Yamazaki T, Makino M (2006): Identification of Mycobacterium species by comparative analysis of the dnaA gene. FEMS Microbiology Letters 254, 232-239. Go to original source... Go to PubMed...
    42. Nagel JA (1749): Beschreibung deren auf Allerhochsten Befehl Ihrer Rom. Kaas und Konigl. Maytt. Francisci I. untersuchten in dem Hertzogthum Crain befindlichen Seltenheiten der Natur. Manuscript in Vienna Court Library. 97 pp.
    43. Nossa CW, Oberdorf WE, Yang L, Aas JA, Paster BJ, Desantis TZ, Brodie EL, Malamud D, Poles MA, Pei Z (2010): Design of 16S rRNA gene primers for 454 pyrosequencing of the human foregut microbiome. World Journal of Gastroenterology 16, 4135-4144. Go to original source... Go to PubMed...
    44. Novakova A (2009): Microscopic fungi isolated from the Domica Cave system (Slovak Karst National Park, Slovakia). A review. International Journal of Speleology 38, 71-82. Go to original source...
    45. Parise M, Lollino P (2011): A preliminary analysis of failure mechanisms in karst and man-made underground caves in Southern Italy. Geomorphology 134, 132-143. Go to original source...
    46. Persing DH (ed.) (2011): Molecular Microbiology, Diagnostic Principles and Practice. 2nd edn. ASM Press, Washington, D.C. 936 pp.
    47. Pizl V (2008): Are earthworms (Oligochaeta, Lumbricidae) regular or accidental dwellers of cave systems in the Czech Republic and Slovakia (in Czech). Acta Carsologica Slovaca 46, 197-201.
    48. Prichystal A, Naplava M (1995): Mystery of Bull Rock Cave - the cave full of questions (in Czech). Amaprint, Trebic. 161 pp.
    49. Roblickova M, Kana V (2013): New palaeontological research in Barova (Sobolova) Cave, Moravian Karst: Preliminary report. Acta Musei Moraviae Scientiae Geologicae 98, 111-127.
    50. SAIF - State Agricultural Interventional Fund (2016): List of subsidy beneficiaries. Available at https://www.szif.cz/en/list_of_beneficiaries (Accessed July 20, 2016).
    51. Schuppler M, Mertens F, Schon G, Gobel UB (1995): Molecular characterization of nocardioform Actinomycetes in activated sludge by 16S rRNA analysis. Microbiology 141, 513-521. Go to original source... Go to PubMed...
    52. Seman M, Gaalova B, Cichova M, Proksova M, Haviarova D, Flakova R (2015): The occurrence of coliform bacteria in the cave waters of Slovak Karst, Slovakia. Folia Microbiologica 60, 269-278. Go to original source... Go to PubMed...
    53. Shabarova T, Villiger J, Morenkov O, Niggemann J, Dittmar T, Pernthaler J (2014): Bacterial community structure and dissolved organic matter in repeatedly flooded subsurface karst water pools. FEMS Microbiology Ecology 89, 111- 126. Go to original source... Go to PubMed...
    54. Slana I, Kaevska M, Kralik P, Horvathova A, Pavlik I (2010): Distribution of Mycobacterium avium subsp. avium and M. a. hominissuis in artificially infected pigs studied by culture and IS901 and IS1245 quantitative real time PCR. Veterinary Microbiology 144, 437-443. Go to original source... Go to PubMed...
    55. Stastna P, Bezdek J, Kovarik M (eds) (2003): The Animal's Species Described in Moravian Karst (in Czech). Obcanske Sdruzeni KORAX, Kulirov. 80 pp.
    56. Tang YW, Sussman M, Dongyou L, Poxton I, Schwartzman J (eds) (2014): Molecular Medical Microbiology, Academic Press. 2216 pp.
    57. Telenti A, Marchesi F, Balz M, Bally F, Bottger EC, Bodmer T (1993): Rapid identification of mycobacteria to the species level by polymerase chain reaction and restriction enzyme analysis. Journal of Clinical Microbiology 31, 175-178. Go to original source... Go to PubMed...
    58. Terzic J, Markovic T, Reberski JL (2014): Hydrogeological properties of a complex Dinaric Karst catchment: Miljacka Spring case study. Environmental Earth Sciences 72, 1129-1142. Go to original source...
    59. Thoen CO, Steele JH, Kaneene JB (eds) (2014): Zoonotic Tuberculosis: Mycobacterium bovis and Other Pathogenic Mycobacteria. 3rd edn. Wiley-Blackwell. 432 pp. Go to original source...
    60. Turenne CY, Semret M, Cousins DV, Collins DM, Behr MA (2006): Sequencing of hsp65 distinguishes among subsets of the Mycobacterium avium complex. Journal of Clinical Microbiology 44, 433-440. Go to original source... Go to PubMed...
    61. Ulmann V, Kracalikova A, Dziedzinska R (2015): Mycobacteria in water used for personal hygiene in heavy industry and collieries: A potential risk for employees. International Journal of Environmental Research and Public Health 12, 2870-2877. Go to original source... Go to PubMed...
    62. van der Wielen PW, Heijnen L, van der Kooij D (2013): Pyrosequence analysis of the hsp65 genes of nontuberculous mycobacterium communities in unchlorinated drinking water in the Netherlands. Applied and Environmental Microbiology 79, 6160-6166. Go to original source... Go to PubMed...
    63. Varaine F, Rich ML (eds) (2014): Tuberculosis: Practical Guide for Clinicians, Nurses, Laboratory Technicians and Medical Auxiliarie. Medecins Sans Frontieres Parnters in Health, Paris. 303 pp.
    64. Wankel H (1856): Ueber die Fauna der mahrischen Hohlen. Verhandlungen des Zoologisch-Botanischen Vereins in Wien, Wien 6, 467-470.
    65. Wankel H (ed.) (1882): Bilder aus der Mahrischen Schweiz und ihrer Vergangenheit, Druck Verlag von Adolf Holzhausen, Wien; translated in Czech by Grolich V, Svoboda O, Urban J (1988). 2nd edn. Muzejni a Vlastivedna Spolecnost v Blansku, Brno. 307 pp.
    66. Wayne LG, Kubica GP (1986): Genus Mycobacterium Lehmann and Neumann 1896, 363AL. In: Sneath PHA, Mair NS, Sharpe ME, Holt JG (eds.): Bergey's Manual of Systematic Bacteriology. The Williams and Wilkins Co., Baltimore. 1436-1457.
    67. Weisburg WG, Barns SM, Pelletier DA, Lane DJ (1991): 16S ribosomal DNA amplification for phylogenetic study. Journal of Bacteriology 173, 697-703. Go to original source... Go to PubMed...
    68. Wilhartitz IC, Kirschner AK, Brussaard CP, Fischer UR, Wieltschnig C, Stadler H, Farnleitner AH (2013): Dynamics of natural prokaryotes, viruses, and heterotrophic nanoflagellates in alpine karstic groundwater. Microbiology Open 2, 633-643. Go to original source... Go to PubMed...
    69. Zhang Y, Kelly WR, Panno SV, Liu WT (2014): Tracing fecal pollution sources in karst groundwater by Bacteroidales genetic biomarkers, bacterial indicators, and environmental variables. Science of the Total Environment 490, 1082-1090. Go to original source... Go to PubMed...

    This is an open access article distributed under the terms of the Creative Commons Attribution-NonCommercial 4.0 International (CC BY NC 4.0), which permits non-comercial use, distribution, and reproduction in any medium, provided the original publication is properly cited. No use, distribution or reproduction is permitted which does not comply with these terms.


    Return to the content