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Microbial Diversity in Paleolithic Caves: A Study Case on the Phototrophic Biofilms of the Cave of Bats (Zuheros, Spain)

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Abstract

The biological colonization of rocks in the Cave of Bats (Cueva de Los Murciélagos, Zuheros, Spain) was studied in order to reveal the diversity of microorganisms involved in the biofilm formation. The culturable, metabolically active fraction of biodeteriogens present on surfaces was investigated focusing on morphological, ultrastructural, and genetic features, and their presence related to the peculiar environmental conditions of the underground site. PCR-ITS analysis and 16S rDNA sequences were used to clusterize and characterize the isolated strains. The presence of bacterial taxa associated to the photosynthetic microflora and fungi within the biofilm contributed to clarify the relationships inside the microbial community and to explain the alteration observed at the different sites. These results will contribute to the application of more successful strategies for the preventive conservation of subterranean archaeological sites.

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References

  1. Albertano P, Bellezza S (2001) Cytochemistry of cyanobacterial exopolymers in biofilms from Roman hypogea. Nova Hedwig 123:501–518

    Google Scholar 

  2. Albertano P, Bruno L (2003) The importance of light in the conservation of hypogean monuments. In: Saiz-Jimenez C (ed) Molecular Biology and Cultural Heritage. Balkema, Lisse, pp 171–177

    Google Scholar 

  3. Albertano P, Bruno L, Bellezza S (2005) New strategy for the monitoring and control of cyanobacterial films on valuable lithic faces. Plant Biosyst 139:311–322

    Article  Google Scholar 

  4. Albertano P, Hernandez-Mariné M (2001) Algae in habitats with reduced and extreme radiation. Nova Hedwig 123:225–227

    Google Scholar 

  5. Albertano P, Moscone D, Palleschi G, Hermosín B, Saiz-Jimenez C, Sánchez-Moral S, Hernández-Mariné M, Urzi C, Groth I, Schroeckh V, Saarela M, Mattla Sandholm T, Gallon JR, Graziottin F, Bisconti F, Giuliani R (2003) Cyanobacteria attack rocks (CATS): control and preventive strategies to avoid damage caused by cyanobacteria and associated microorganisms in Roman hypogean monuments. In: Saiz-Jimenez C (ed) Molecular Biology and Cultural Heritage. Balkema, Lisse, pp 151–162

    Google Scholar 

  6. Albertano P, Urzì C (1999) Structural interactions among epilithic cyanobacteria and heterotrophic microorganisms in Roman hypogea. Microb Ecol 38:244–252

    Article  PubMed  Google Scholar 

  7. Anagnostidis K, Komàrek J (1985) Modern approach to classification system of cyanophytes. 1—Introduction. Arch Hydrobiol Algol Stud 38–39:291–302

    Google Scholar 

  8. Anagnostidis K, Komàrek J (1990) Modern approach to classification system of cyanophytes. 5—Stigonematales. Arch Hydrobiol Algol Stud 50–53:327–472

    Google Scholar 

  9. Barnett HL, Hunter BB (1972) Illustrated Genera of Imperfect Fungi. Burgess Publishing Company, Minneapolis

    Google Scholar 

  10. Bastian F, Alabouvette C (2009) Lights and shadows on the conservation of a rock art cave: the case of Lascaux Cave. Int J Speleol Spec Issue Cave Microbiol 38:55–60

    Google Scholar 

  11. Bastian F, Alabouvette C, Jurado V, Saiz-Jimenez C (2009) Impact of biocide treatments on the bacterial communities of the Lascaux Cave. Naturwissenschaften 96:863–868

    Article  CAS  PubMed  Google Scholar 

  12. Bellezza S, Albertano P (2003) A Chroococcalean species from Roman hypogean sites: characterisation of Gloeothece membranacea (Cyanobacteria, Synechoccaceae). Arch Hydrobiol Algol Stud 109:103–112

    Google Scholar 

  13. Borin S, Daffonchio D, Sorlini C (1997) Single strand conformation polymorphism analysis of PCR-tDNA fingerprinting to address the identification of Bacillus species. FEMS Microbiol Lett 157:87–93

    Article  CAS  PubMed  Google Scholar 

  14. Bruno L, Billi D, Urzì C, Albertano P (2006) Genetic characterization of epilithic cyanobacteria and their associated bacteria. Geomicrobiol J 23:293–297

    Article  CAS  Google Scholar 

  15. Cañaveras JC, Sanchez-Moral S, Soler V, Saiz-Jimenez C (2001) Microorganisms and microbially induced fabrics in cave walls. Geomicrobiol J 18:223–240

    Article  Google Scholar 

  16. Chelius MK, Beresford G, Horton H, Quirk M, Selby G, Simpson RT, Horrocks R, Moore JC (2009) Impacts of alterations of organic inputs on the bacterial community within the sediments of Wind Cave, South Dakota, USA. Int J Speleol 38:1–10

    Google Scholar 

  17. Normal C (1990) Raccomandazioni Normal: 9/88 Microflora autotrofa ed eterotrofa: tecniche di isolamento in coltura. C.N.R. - I.C.R, Roma

    Google Scholar 

  18. Cox EJ (1996) Identification of Freshwater Diatoms from Live Material. Chapman & Hall, London, p 158

    Google Scholar 

  19. Curtis TP, Sloan WT (2004) Prokaryotic diversity and its limits: microbial community structure in nature and implications for microbial ecology. Curr Opin Microbiol 7:221–226

    Article  PubMed  Google Scholar 

  20. Dupont J, Jacquet C, Dennetière B, Lacoste S, Bousta F, Orial G, Couloux A, Roquebert MF (2007) Invasion of the French Paleolithic painted cave of Lascaux by members of the Fusarium solani species complex. Mycologia 99:526–533

    Article  CAS  PubMed  Google Scholar 

  21. Ellis MB (1971) Dematiaceous Hyphomycetes. CAB International Mycological Institute, Kew

    Google Scholar 

  22. Ellis MB (1976) More Dematiaceous Hyphomycetes. CAB International Mycological Institute, Kew

    Google Scholar 

  23. Fassatiovà O (1986) Moulds and filamentous fungi in technical microbiology. In: Bushell ME (ed) Progress in Industrial Microbiology 22. Elsevier, Amsterdam

    Google Scholar 

  24. Gonzalez I, Laiz L, Hermosin B, Caballero B, Incerti C, Saiz-Jimenez C (1999) Bacteria isolated from rock art paintings: the case of Atlanterra shelter (South Spain). J Microbiol Meth 36:123–127

    Article  CAS  Google Scholar 

  25. Groth I, Saiz-Jimenez C (1999) Actinomycetes in hypogean environments. Geomicrobiol J 16:1–8

    Article  Google Scholar 

  26. Groth I, Schumann P, Laiz L, Sanchez-Moral S, Cañaveras JC, Saiz-Jimenez C (2001) Geomicrobiological study of the Grotta dei Cervi, Porto Badisco, Italy. Geomicrobiol J 18:241–258

    Article  CAS  Google Scholar 

  27. Groth I, Schumann P, Schuetze B, Gonzalez JM, Laiz L, Saiz-Jimenez C, Stackebrandt E (2005) Isoptericola hypogeus sp. nov., isolated from the Roman catacomb of Domitilla. Int J Syst Evol Microbiol 55:1715–1719

    Article  CAS  PubMed  Google Scholar 

  28. Groth I, Schumann P, Schutze B, Gonzalez JM, Laiz L, Suihko ML, Stackebrandt E (2006) Myceligenerans crystallogenes sp. nov., isolated from Roman catacombs. Int J Syst Evol Microbiol 56:283–287

    Article  CAS  PubMed  Google Scholar 

  29. Gurtler V, Stanisich VA (1996) New approaches to typing and identification of bacteria using the 16S–23S rDNA spacer region. Microbiology 142:3–16

    Article  PubMed  Google Scholar 

  30. Hernández-Mariné M, Asencio-Martínez A, Canals A, Ariño X, Aboal M, Hoffmann L (1999) Discovery of populations of the lime incrusting genus Loriella (Stigonematales) in Spanish caves. Arch Hydrobiol Algol Stud 94:121–138

    Google Scholar 

  31. Hernandez-Marine M, Clavero E, Roldan M (2003) Why there is such luxurious growth in the hypogean environments. Arch Hydrobiol Algol Stud 109:229–240

    Google Scholar 

  32. Hernanz A, Mas M, Gavilán B, Hernández B (2006) Raman microscopy and IR spectroscopy of prehistoric paintings from Los Murciélagos cave (Zuheros, Córdoba, Spain). J Raman Spectrosc 37:492–497

    Article  CAS  Google Scholar 

  33. Hoffmann L (2002) Caves and other low-light environments: Aerophytic photoautotrophic microorganisms. In: Bitton G (ed) Encyclopedia of Environmental Microbiology. John Wiley & Sons, New York, pp 835–843

    Google Scholar 

  34. Ikner LA, Toomey RS, Nolan G, Neilson JW, Pryor BM, Maier RM (2007) Culturable microbial diversity and the impact of tourism in Kartchner Caverns, Arizona. Microb Ecol 53:30–42

    Article  PubMed  Google Scholar 

  35. Jager K, Marialigeti K, Hauck M, And Barabas G (1983) Promicromonospora enterophila sp. nov., a New Species of Monospore Actinomycetes. Int J Syst Bacteriol 33:525–531

    Article  Google Scholar 

  36. John DM, Whitton BA, Brook AJ (2002) The freshwater algal flora of British Islands: an identification guide to freshwater and terrestrial algae. Cambridge University Press, Cambridge, UK, p 702

    Google Scholar 

  37. Jurado V, Gonzalez JM, Laiz L, Saiz-Jimenez C (2006) Aurantimonas altamirensis sp. nov., a novel member of the order Rhizobiales isolated from Altamira Cave. Int J Syst Evol Microbiol 56:2583–2585

    Article  CAS  PubMed  Google Scholar 

  38. Jurado V, Groth I, Gonzalez JM, Laiz L, Schuetze B, Saiz-Jimenez C (2005) Agromyces italicus sp. nov., Agromyces humatus sp. nov. and Agromyces lapidis sp. nov., isolated from Roman catacombs. Int J Syst Evol Microbiol 55:871–875

    Article  CAS  PubMed  Google Scholar 

  39. Jurado V, Laiz L, Gonzalez JM, Hernandez-Marine M, Valens M, Saiz-Jimenez C (2005) Phyllobacterium catacumbae sp. nov., a member of the order 'Rhizobiales' isolated from Roman catacombs. Int J Syst Evol Microbiol 55:1487–1490

    Article  CAS  PubMed  Google Scholar 

  40. Jurado V, Laiz L, Gonzalez JM, Saiz-Jimenez C (2007) Cave Research: understanding biodiversity through conservation studies. Coalition Newsletter 13:4–6

    Google Scholar 

  41. Jurado V, Laiz L, Rodriguez-Nava V, Boiron P, Hermosin B, Sanchez-Moral S, Saiz-Jimenez C (2010) Pathogenic microorganisms in caves. Int J Speleol 39:15–21

    Google Scholar 

  42. Komárek J, Anagnostidis K (1989) Modern approach to the classification system of cyanophytes. 4—Nostocales. Arch Hydrobiol Algol Stud 43:157–226

    Google Scholar 

  43. Komárek J, Anagnostidis K (1999) Cyanoprokaryota, 1. Teil. Chroococcales. In: Ettl H, Gärtner G, Heynig H, Mollenhauer D (eds) Süβwasserflora von Mitteleuropa Band 19/2. Gustav Fischer, Jena, pp 1–523

    Google Scholar 

  44. Komárek J, Anagnostidis K (2005) Cyanoprokaryota, 2. Teil. Oscillatoriales. In: Büdel B, Gärtner G, Krienitz L, Schagerl M (eds) Süβwasserflora von Mitteleuropa. Elsevier GmbH, Munchen, pp 1–720

    Google Scholar 

  45. Komárek J, Fott B (1983) Chlorococcales. In: Huber-Pestalozzi (ed) Das Phytoplankton des Süβwassers, Teil 7. Schweizerbart, Stuttgart, p 1043

    Google Scholar 

  46. Kuster E, Williams ST (1964) Selection of media for isolation of Streptomyces. Nature 22:928–929

    Article  Google Scholar 

  47. Monte M, Ferrari R (2000) Airborne microorganisms in a subterranean archaeological area of the basilica of San Lorenzo in Lucina (Rome). Aerobiologia 16:435–439

    Article  Google Scholar 

  48. Ohki K, Gantt E (1983) Fonctional phycobilisomes from Tolypothrix tenuis (cyanophyta) grown heterotrophically in the dark. J Phycol 19:359–364

    Article  Google Scholar 

  49. Pangallo D, Chovanová K, Drahovska H, De Leo F, Urzì C (2009) Application of fluorescence internal transcribed spacer-PCR (f-ITS) for the cluster analysis of bacteria isolated from air and deteriorated fresco surfaces. Int Biodeterior Biodegrad 63:868–872

    Article  CAS  Google Scholar 

  50. Rainey FA, Ward-Rainey N, Kroppenstedt RM, Stackebrandt E (1996) The genus Nocardiopsis represents a phylogenetically coherent taxon and a distinct actinomycete lineage: proposal of Nocardiopsaceae fam. nov. Int J Syst Bacteriol 46:1088–1092

    Article  CAS  PubMed  Google Scholar 

  51. Rindi F, Sherwood AR, Guiry MD (2005) Taxonomy and distribution of Trentepohlia and Printzina (Trentepohliales, Chlorophyta) in the Hawaiian Islands. Phycologia 44:270–284

    Article  Google Scholar 

  52. Rippka R, Deruelles J, Waterbury B, Herdman M, Stanier RY (1979) Generic assignments, strains histories and properties of pure cultures of cyanobacteria. J Gen Microbiol 111:1–61

    Google Scholar 

  53. Rippka R, Neilson A, Kunisawa R, Cohen-Bazire G (1971) Nitrogen fixation by unicellular blue-green algae. Arch Microbiol 76:341–348

    CAS  Google Scholar 

  54. Rogerio Candelera MA (2007) Conservation of Rock Art: a brief overview. Coalition Newsletter 13:2

    Google Scholar 

  55. Roldán M, Clavero E, Canals A, Gómez-Bolea A, Ariño X, Hernández-Mariné M (2004) Distribution of phototrophic biofilms in cavities (Garraf, Spain). Nova Hedwig 78:329–351

    Article  Google Scholar 

  56. Roldán M, Hernández-Mariné M (2009) Exploring the secrets of the three-dimensional architecture of phototrophic biofilms in caves. Int J Speleol 38:41–53

    Google Scholar 

  57. Saiz-Jimenez C (2005) The lessons of Lascaux. Coalition Newsletter 10:3

    Google Scholar 

  58. Saiz-Jimenez C (2009) Editorial Lascaux re-visited. Coalition Newsletter 18:2

    Google Scholar 

  59. Schabereiter-Gurtner C, Saiz-Jimenez C, Pinar G, Lubitz W, Rölleke S (2002) Culture-independent analyses of bacterial communities on paleolithic paintings and surrounding rock walls in karstic caves (Altamira, Tito Bustillo, La Garma and Llonin). Coalition Newsletter 5:7–10

    Google Scholar 

  60. Schabereiter-Gurtner C, Saiz-Jimenez C, Pinar G, Lubitz W, Rölleke S (2002) Altamira cave Paleolithic paintings harbor partly unknown bacterial communies. FEMS Microbiol Lett 211:7–11

    Article  CAS  PubMed  Google Scholar 

  61. Schabereiter-Gurtner C, Saiz-Jimenez C, Pinar G, Lubitz W, Rölleke S (2002) Phylogenetic 16S rRNA analysis reveals the presence of complex and partly unknown bacterial communities in Tito Bustillo cave, Spain, and on its Palaeolithic paintings. Environ Microbiol 4:392–400

    Article  CAS  PubMed  Google Scholar 

  62. Sugita T, Kikuchi K, Makimura K, Urata K, Someya T, Kamei K, Niimi M, Uehara Y (2005) Trichosporon species isolated from guano samples obtained from bat-inhabited caves in Japan. Appl Environ Microbiol 71:7626–7629

    Article  CAS  PubMed  Google Scholar 

  63. Urzì C, Brusetti L, Salamone P, Sorlini C, Stackebrandt E, Daffonchio D (2001) Biodiversity of Geodermatophilaceae isolated from altered stones and monuments in the Mediterranean basin. Environ Microbiol 3:471–479

    Article  PubMed  Google Scholar 

  64. Urzì C, De Leo F (2001) Sampling with adhesive tape strips: an easy and rapid method to monitor microbial colonization on monument surfaces. J Microbiol Meth 44:1–11

    Article  Google Scholar 

  65. Urzì C, De Leo F, Donato P, La Cono V (2003) Multiple approaches to study the structure and diversity of microbial communities colonizing artistic surfaces. In: Saiz-Jimenez C (ed) Molecular Biology and Cultural Heritage. Balkema, Lisse, pp 187–194

    Google Scholar 

  66. Urzì C, De Leo F, Schumann P (2008) Kribbella catacumbae sp. nov. and Kribbella sancticallisti sp. nov., isolated from whitish-grey patinas in the Catacombs of St Callistus in Rome, Italy. Int J Syst Evol Microbiol 58:2090–2097

    Article  PubMed  Google Scholar 

  67. Urzì C, Donato P, Lo Passo C, Albertano P (2002) Occurrence and biodiversity of Streptomyces strains isolated from Roman Hypogea. In: Galan E, Zezza F (eds) Protection and Conservation of the Cultural Heritage of the Mediterranean Cities. Balkema, Lisse, pp 269–272

    Google Scholar 

  68. Urzì C, Lisi S, Criseo G, Zagari M (1992) Comparazione di terreni per l’enumerazione e l’isolamento di funghi deteriogeni isolati da materiali naturali. Ann Microbiol Enzimol 42:185–193

    Google Scholar 

  69. Urzì C, Salamone P, Schumann P, Rohde M, Stackebrandt E (2004) Blastococcus saxobsidens sp. nov., isolated from marble and calcareous sandstone in quarries and archeological sites in the Mediterranean Basin, and emended description of the genus Blastococcus Ahrens and Moll and Blastococcus aggregatus Ahrens and Moll 1970. Int J Syst Evol Microbiol 4:253–259

    Article  Google Scholar 

  70. Wiley M, Sherwood M, Woolverton J (2009) Prescott Microbiologia, vol 2. MacGraw-Hill, Milan, p 112

    Google Scholar 

  71. Zhou JP, Gu YQ, Zou CS, Mo MH (2007) Phylogenetic diversity of bacteria in an earth-cave in Guizhou Province, Southwest of China. J Microbiol 45:105–112

    CAS  PubMed  Google Scholar 

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Acknowledgements

This work was supported by the EU Programs EESD, Project CATS contract EVK4-CT2000-00028, and the University of Messina through Progetti di Ricerca di Ateneo P.R.A. years 2004–2008. The Authors like to thank Prof. Cesareo Saiz Jimenez for his suggestions to improve the manuscript content and Paolo Donato and Violetta La Cono in the analysis of samples during the CATS project.

The support of staff of the Cueva de Zuheros is also acknowledged.

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Authors and Affiliations

  1. Department of Life Sciences “M. Malpighi”, University of Messina, Salita Sperone 31, Messina, Italy

    Clara Urzì & Filomena De Leo

  2. Laboratory of Biology of Algae, Department of Biology, University of Rome “Tor Vergata”, via della Ricerca scientifica, 00133, Rome, Italy

    Laura Bruno & Patrizia Albertano

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  1. Clara Urzì
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  2. Filomena De Leo
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Correspondence to Clara Urzì.

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Urzì, C., De Leo, F., Bruno, L. et al. Microbial Diversity in Paleolithic Caves: A Study Case on the Phototrophic Biofilms of the Cave of Bats (Zuheros, Spain). Microb Ecol 60, 116–129 (2010). https://doi.org/10.1007/s00248-010-9710-x

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