Environmental Science and Pollution Research

, Volume 23, Issue 12, pp 12216–12226 | Cite as

MiSeq HV4 16S rRNA gene analysis of bacterial community composition among the cave sediments of Indo-Burma biodiversity hotspot

  • Surajit De Mandal
  • Zothansanga
  • Amrita Kumari Panda
  • Satpal Singh Bisht
  • Nachimuthu Senthil Kumar
Research Article

Abstract

Caves in Mizoram, Northeast India, are potential hotspot diversity regions due to the historical significance of the formation of the Indo-Burman plateau and also because of their unexplored and unknown diversity. High-throughput paired end Illumina sequencing of the V4 region of 16S rRNA was performed to study the bacterial community of three caves situated in Champhai district of Mizoram, Northeast India. A total of 10,643 operational taxonomic units (OTUs) (based on 97 % cutoff) comprising of 21 major and 21 candidate phyla with a sequencing depth of 1,140,013 were found in this study. The overall taxonomic profile obtained by the RDP classifier and Greengenes OTU database revealed high diversity within the bacterial communities. Communities were dominated by Planctomycetes, Actinobacteria, Proteobacteria, Bacteroidetes, and Firmicutes, while members of Archaea were less varied and mostly comprising of Eukaryoarchea. Analysis revealed that Farpuk (CFP) cave sediment has low microbial diversity and is mainly dominated by Actinobacteria (80 % reads), whereas different bacterial communities were found in the caves of Murapuk (CMP) and Lamsialpuk (CLP). Analysis also revealed that a major portion of the identified OTUs was classified under rare biosphere. Importantly, all these caves recorded a high number of unclassified OTUs, which might represent new species. Further analysis with whole genome sequencing is needed to validate the unknown species as well as to determine their functional role.

Keywords

Cave Indo-Burma plateau Bacterial diversity Illumina sequencing 

Supplementary material

11356_2016_6423_MOESM1_ESM.docx (14 kb)
Table S1Top ten bacterial genera based on OTU number among the cave samples.
11356_2016_6423_MOESM2_ESM.docx (17 kb)
Table S2Top ten OTUs based on total read count number among the cave samples.
11356_2016_6423_MOESM3_ESM.docx (28 kb)
Table S3Pearson’s correlation coefficient between soil characteristics and bacterial phyla.
11356_2016_6423_MOESM4_ESM.docx (131 kb)
Figure S1Graphical representation of the relative abundance of bacterial diversity from phylum to species level of CFP using the Krona visualization tool.
11356_2016_6423_MOESM5_ESM.docx (183 kb)
Figure S2Graphical representation of the relative abundance of bacterial diversity from phylum to species level of CLP using Krona visualization tool.
11356_2016_6423_MOESM6_ESM.docx (187 kb)
Figure S3Graphical representation of the relative abundance of bacterial diversity from phylum to species level of CMC using the Krona visualization tool.

References

  1. Adetutu EM, Thorpe K, Bourne S, Cao X, Shahsavari E, Kirby G, Ball AS (2011) Phylogenetic diversity of fungal communities in areas accessible and not accessible to tourists in Naracoorte Caves. Mycologia 103:959–968CrossRefGoogle Scholar
  2. Adetutu EM, Thorpe K, Shahsavari E, Bourne S, Cao X, Fard RMN, Kirby G, Ball AS (2012) Bacterial community survey of sediments at Naracoorte Caves, Australia. Int J Speleol 41(2):137–147CrossRefGoogle Scholar
  3. Aravindraja C, Viszwapriya D, Karutha PS, Ultradeep (2013) 16S rRNA sequencing analysis of geographically similar but diverse unexplored marine samples reveal varied bacterial community composition. PLoS ONE 8(10):e76724. doi:10.1371/journal.pone.0076724 CrossRefGoogle Scholar
  4. Barton HA (2006) Introduction to cave microbiology: a review for the non-specialist. J Cave Karst Stud 68:43–54Google Scholar
  5. Barton HA (2014) Life in extreme environments: microbial life of cave systems. In: Wagner D (ed) Starving artists: bacterial oligotrophic heterotrophy in caves. De Gruyter, BerlinGoogle Scholar
  6. Barton AH, Northup ED (2007) Geomicrobiology in cave environments: past, current and future perspectives. J Cave Karst Stud 69:163–178Google Scholar
  7. Barton HA, Spear JR, Pace NR (2001) Microbial life in the underworld: biogenicity in secondary mineral formations. Geomicrobiol J 18:359–368CrossRefGoogle Scholar
  8. Brook BW, Sodhi NS, Ng PKL (2003) Catastrophic extinctions follow deforestation in Singapore. Nature 424:420–424CrossRefGoogle Scholar
  9. Caporaso JG, Bittinger K, Bushman FD, DeSantis TZ, Andersen GL, Knight R (2010a) PyNAST: a flexible tool for aligning sequences to a template alignment. Bioinformatics 26:266–267CrossRefGoogle Scholar
  10. Caporaso JG, Kuczynski J, Stombaugh J, Bittinger K, Bushman FD, Costello EK, Fierer N, Peña AG, Goodrich JK, Gordon JI, Huttley GA, Kelley ST, Knights D, Koenig JE, Ley RE, Lozupone CA, McDonald D, Muegge BD, Pirrung M, Reeder J, Sevinsky JR, Turnbaugh PJ, Walters WA, Widmann J, Yatsunenko T, Zaneveld J, Knight R (2010b) QIIME allows analysis of high-throughput community sequencing data. Nat Methods 7:335–336CrossRefGoogle Scholar
  11. Conrad R, Erkel C, Liesack W (2006) Rice Cluster I methanogens, an important group of Archaea producing greenhouse gas in soil. Curr Opin Biotechnol 17:262–267CrossRefGoogle Scholar
  12. Cuezva S, Fernandez-Cortes A, Porca E, Pašić L, Jurado V, Hernandez-Marine M, Serrano-Ortiz P, Hermosin B, Carlos Cañaveras J, Sanchez-Moral S, Saiz-Jimenez C (2012) The biogeochemical role of Actinobacteria in Altamira Cave, Spain. FEMS Microbiol Ecol 81:281–290CrossRefGoogle Scholar
  13. DeSantis TZ, Hugenholtz P, Larsen N, Rojas M, Brodie EL, Keller K, Huber T, Dalevi D, Hu P, Andersen GL (2006) Greengenes, a chimera-checked 16S rRNA gene database and workbench compatible with ARB. Appl Environ Microbiol 72:5069–5072CrossRefGoogle Scholar
  14. Edgar RC (2010) Search and clustering orders of magnitude faster than BLAST. Bioinformatics 26:2460–2461CrossRefGoogle Scholar
  15. Edgar RC, Haas BJ, Clemente JC, Quince C, Knight R (2011) UCHIME improves sensitivity and speed of chimera detection. Bioinformatics 27:2194–2200CrossRefGoogle Scholar
  16. Ehrich S, Behrens D, Lebedeva E, Ludwig W, Bock E (1995) A new obligately chemolithoautotrophic, nitriteoxidizing bacterium, Nitrospira moscoviensis sp. nov. and its phylogenetic relationship. Arch Microbiol 164(1):16–23CrossRefGoogle Scholar
  17. Engel AS, Meisinger DB, Porter ML, Payn R, Schmid M, Stern LA, Schleifer KH, Lee NM (2010) Linking phylogenetic and functional diversity to nutrient spiraling in microbial mats from Lower Kane Cave (USA). ISME J 4:98–110CrossRefGoogle Scholar
  18. Epure J, Meleg JN, Munteanu CM, Roban RD, Moldovan OT (2014) Bacterial and fungal diversity of quaternary cave sediment deposits. Geomicrobiol J 31:116–127CrossRefGoogle Scholar
  19. Fuerst JA, Sagulenko E (2011) Beyond the bacterium: Planctomycetes challenge our concepts of microbial structure and function. Nat Rev Microbiol 9:403–413CrossRefGoogle Scholar
  20. Galagan JE, Nusbaum C, Roy A, Endrizzi MG, MacDonald P, Fitzhugh W, Calvo S, Engels R, Smirnov S, Atnoor D, Brown A, Allen N, Naylor J, Stange-Thomann N, Dearellano K, Johnson R, Linton L, McEwan P, McKernan K, Talamas J, Tirrell A, Ye W, Zimmer A, Barber RD, Cann I, Graham DE, Grahame DA, Guss AM, Hedderich R, Ingram-Smith C (2002) The genome of M. Acetivorans reveals extensive metabolic and physiological diversity. Genome Res 12(4):532–542CrossRefGoogle Scholar
  21. Gebauer HD, Chhakchhuak B, Sootinck N (2001) Caves of Mizoram (speleological projects in NE-India) 5:61Google Scholar
  22. Gray CJ, Engel AS (2013) Microbial diversity and impact on carbonate geochemistry across a changing geochemical gradient in a karst aquifer. ISME J 7(2):325–337CrossRefGoogle Scholar
  23. Hammer O, Harper DAT, Ryan PD (2001) PAST: paleontological statistics software package for education and data analysis. Palaeontol Electron 4(1):9ppGoogle Scholar
  24. Hanada S, Hiraishi A, Shimada K, Matsuura K (1995) Chloroflexus aggregans sp. nov., a filamentous phototrophic bacterium which forms dense cell aggregates by active gliding movement. Int J Syst Evol Microbiol 45:676–681Google Scholar
  25. Hugenholtz P, Stackebrandt E (2004) Reclassification of Sphaerobacter thermophilus from the subclass Sphaerobacteridae in the phylum Actinobacteria to the class Thermomicrobia (emended description) in phylum Chloroflexi (emended description). Int J Syst Evol Microbiol 54:2049–2051CrossRefGoogle Scholar
  26. Jetten MS, Sliekers O, Kuypers M, Dalsgaard T, van Niftrik L, Cirpus I, van de Pas-Schoonen K, Lavik G, Thamdrup B, Le Paslier D, Op den Camp HJ, Hulth S, Nielsen LP, Abma W, Third K, Engström P, Kuenen JG, Jørgensen BB, Canfield DE, Sinninghe Damsté JS, Revsbech NP, Fuerst J, Weissenbach J, Wagner M, Schmidt I, Schmid M, Strous M (2003) Anaerobic ammonium oxidation by marine and freshwater planctomycete-like bacteria. Appl Microbiol Biotechnol 63:107–114CrossRefGoogle Scholar
  27. Jones RT, Robeson MS, Lauber CL, Hamady M, Knight R, Fierer N (2009) A comprehensive survey of soil acidobacterial diversity using pyrosequencing and clone library analyses. ISME J 3:442–453CrossRefGoogle Scholar
  28. Kersters K, De Vos P, Gillis M, Swings J, Vandamme P, Stackebrandt E (2006) In: Dwarkin M, Falkow S, Rosenberg E, Schleifer KH, Stackebrandt E (eds) Introduction to the Proteobacteria. The prokaryotes, Vol. 5. Springer, New York, pp 3–37Google Scholar
  29. Li J, Tan B, Mai K, Ai Q, Zhang W, Xu W, Liufu Z, Ma H (2005) Comparative study between probiotic bacterium Arthrobacter XE-7 and chloramphenicol on protection of Penaeus chinensis post-larvae from pathogenic vibrios. Aquaculture 253:140–147CrossRefGoogle Scholar
  30. Lozupone C, Knight R (2005) UniFrac: a new phylogenetic method for comparing microbial communities. Appl Environ Microbiol 71:8228–8235CrossRefGoogle Scholar
  31. Lü Z, Lu Y (2012) Complete genome sequence of a thermophilic methanogen, Methanocella conradii HZ254, isolated from Chinese rice field soil. J Bacteriol 194(9):2398–2399CrossRefGoogle Scholar
  32. MacLeod FA, Guiot SR, Costerton JW (1990) Layered structure of bacterial aggregates produced in an upflow anaerobic sludge bed and filter reactor. Appl Environ Microbiol 56:1598–1607Google Scholar
  33. Maymó-Gatell X, Chien YT, Gossett JM, Zinder SH (1997) Isolation of a bacterium that reductively dechlorinates tetrachloroethene to ethane. Science 276:1568–1571CrossRefGoogle Scholar
  34. Minyard ML, Bruns MA, Liermann LJ, Buss HL, Brantley SL (2012) Bacterial associations with weathering minerals at the Regolith-Bedrock interface, Luquillo Experimental Forest, Puerto Rico. Geomicrobiol J 29:792–803CrossRefGoogle Scholar
  35. Moss JA, Nocker A, Snyder RA (2011) Microbial characteristics of a submerged karst cave system in Northern Florida. Geomicrobiol J 28:719–931CrossRefGoogle Scholar
  36. Myers N, Mittermeier RA, Mittermeier CG, da Fonseca GAB, Kent J (2000) Biodiversity hotspots for conservation priorities. Nature 403:853–858CrossRefGoogle Scholar
  37. Northup DE, Melim LA, Spilde MN, Hathaway JM, Garcia MG, Moya M, Stone FD, Boston PJ, Dapkevicius MLNE, Riquelme C (2011) Lava cave microbial communities within mats and secondary mineral deposits: implications for life detection on other planets. Astrobiology 11:601–618CrossRefGoogle Scholar
  38. Ortiz M, Neilson JW, Nelson WM, Legatzki A, Byrne A, Yu Y, Wing RA, Soderlund CA, Pryor BM, Pierson LS, Maier RM (2013) Profiling bacterial diversity and taxonomic composition on speleothem surfaces in Kartchner Caverns, AZ. Microb Ecol 65:371–383CrossRefGoogle Scholar
  39. Ortiz M, Legatzki A, Neilson JW, Fryslie B, Nelson WM, Wing RA, Soderlund CA, Pryor BM, Maier RM (2014) Making a living while starving in the dark: metagenomic insights into the energy dynamics of a carbonate cave. ISME J 8:478–491CrossRefGoogle Scholar
  40. Pohlman JW, Iliffe TM, Cifuentes LA (1997) A stable isotope study of organic cycling and the ecology of an anchialine cave ecosystem. Mar Ecol Prog Ser 155:17–27CrossRefGoogle Scholar
  41. Reid A, Buckley M (2011) Washington, DC: American Academy of Microbiology. The rare biosphere: a report from the American Academy of MicrobiologyGoogle Scholar
  42. Saiz-Jimenez C (2012) Microbiological and environmental issues in show caves. World J Microbiol Biotechnol 28:2453–2464CrossRefGoogle Scholar
  43. Sakai S, Takaki Y, Shimamura S, Sekine M, Tajima T, Kosugi H, Ichikawa N, Tasumi E, Hiraki AT, Shimizu A, Kato Y, Nishiko R, Mori K, Fujita N, Imachi H, Takai K (2011) Genome sequence of a mesophilic hydrogenotrophic methanogen Methanocella paludicola, the first cultivated representative of the order Methanocellales. PLoS One 6(7):e22898CrossRefGoogle Scholar
  44. Schabereiter-Gurtner C, Saiz-Jimenez C, Piñar G, Lubitz W, Rölleke S (2002) Altamira cave Paleolithic paintings harbour partly unknown bacterial communities. FEMS Microbiol Lett 211:7–11CrossRefGoogle Scholar
  45. Schmid M, Twachtmann U, Klein M, Strous M, Juretschko S, Jetten M, Metzger JW, Schleifer KH, Wagner M (2000) Molecular evidence for genus level diversity of bacteria capable of catalyzing anaerobic ammonium oxidation. Syst Appl Microbiol 23:93–106CrossRefGoogle Scholar
  46. Sekiguchi Y, Yamada T, Hanada S, Ohashi A, Harada H, Kamagata Y (2003) Anaerolinea thermophila gen. nov., sp. nov. and Caldilinea aerophila gen. nov., sp. nov., novel filamentous thermophiles that represent a previously uncultured lineage of the domain Bacteria at the subphylum level. Int J Syst Evol Microbiol 53:1843–1851CrossRefGoogle Scholar
  47. Stomeo F, Makhalanyane TP, Valverde A, Pointing SB, Mark I, Steven MI, Cary CS, Tuffin MI, Cowan DA (2012) Abiotic factors influence microbial diversity in permanently cold soil horizons of a maritime-associated Antarctic Dry Valley. FEMS Microbiol Ecol 82:326–340CrossRefGoogle Scholar
  48. Strapoc D, Picardal FW, Turich C, Schaperdoth I, Macalady JL, Lipp JS, Lin YS, Ertefai TF, Schubotz F, Hinrichs KU, Mastalerz M, Schimmelmann (2008) A methane-producing microbial community in a coal bed of the Illinois basin. Appl Environ Microbiol 74(8):2424–2432CrossRefGoogle Scholar
  49. Tetu SG, Breakwell K, Elbourne LD, Holmes AJ, Gillings MR, Paulsen IT (2013) Life in the dark: metagenomic evidence that a microbial slime community is driven by inorganic nitrogen metabolism. ISME J 7:1227–1236CrossRefGoogle Scholar
  50. Thomsen TR, Finster K, Ramsing NB (2001) Biogeochemical and molecular signatures of anaerobic methane oxidation in a marine sediment. Appl Environ Microbiol 67:1646–1656CrossRefGoogle Scholar
  51. Waldron PJ, Petsch ST, Martini AM, Nüsslein K (2007) Salinity constraints on subsurface archaeal diversity and methanogenesis in sedimentary rock rich in organic matter. Appl Environ Microbiol 73(13):4171–4179CrossRefGoogle Scholar
  52. Ward NL, Challacombe JF, Janssen PH, Henrissat B, Coutinho PM, Wu M, Xie G, Haft DH, Sait M, Badger J, Barabote RD, Bradley B, Brettin TS, Brinkac LM, Bruce D, Creasy T, Daugherty SC, Davidsen TM, Deboy RT, Detter JC, Dodson RJ, Durkin AS, Ganapathy A, Gwinn-Giglio M, Han CS, Khouri H, Kiss H, Kothari SP, Madupu R, Nelson K, Nelson WC, Paulsen I, Penn K, Ren Q, Rosovitz MJ, Selengut JD, Shrivastava S, Sullivan SA, Tapia R, Thompson LS, Watkins KL, Yang Q, Yu C, Zafar N, Zhou L, Kuske CR (2009) Three genomes from the phylum Acidobacteria provide insight into their lifestyles in soils. Appl Environ Microbiol 75:2046–2056CrossRefGoogle Scholar
  53. Watson SW, Bock E, Valois FW, Waterbury JB, Schlosser U (1986) Nitrospira marina gen. nov. sp. nov.: a chemolithotrophic nitrite-oxidizing bacterium. Arch Microbiol 144(1):1–7CrossRefGoogle Scholar
  54. Wexler HM (2007) Bacteroides: the good, the bad, and the nitty-gritty. Clin Microbiol Rev 20(4):593–621CrossRefGoogle Scholar
  55. Wu Y, Tan L, Liu W, Wang B, Wang J, Cai Y, Lin X (2015) Profiling bacterial diversity in a limestone cave of the western Loess Plateau of China. Front Microbiol 6:244Google Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2016

Authors and Affiliations

  • Surajit De Mandal
    • 1
  • Zothansanga
    • 1
  • Amrita Kumari Panda
    • 2
  • Satpal Singh Bisht
    • 2
  • Nachimuthu Senthil Kumar
    • 1
  1. 1.Department of BiotechnologyMizoram UniversityAizawlIndia
  2. 2.Department of ZoologyKumaun UniversityNainitalIndia

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