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Microbial Ecology

, Volume 68, Issue 2, pp 416–425 | Cite as

Phylogenetic and Functional Analysis of Gut Microbiota of a Fungus-Growing Higher Termite: Bacteroidetes from Higher Termites Are a Rich Source of β-Glucosidase Genes

  • Meiling Zhang
  • Ning Liu
  • Changli Qian
  • Qianfu Wang
  • Qian Wang
  • Yanhua Long
  • Yongping Huang
  • Zhihua Zhou
  • Xing Yan
Host Microbe Interactions

Abstract

Fungus-growing termites, their symbiotic fungi, and microbiota inhibiting their intestinal tract comprise a highly efficient cellulose-hydrolyzing system; however, little is known about the role of gut microbiota in this system. Twelve fosmid clones with β-glucosidase activity were previously obtained by functionally screening a metagenomic library of a fungus-growing termite, Macrotermes annandalei. Ten contigs containing putative β-glucosidase genes (bgl110) were assembled by sequencing data of these fosmid clones. All these contigs were binned to Bacteroidetes, and all these β-glucosidase genes were phylogenetically closed to those from Bacteroides or Dysgonomonas. Six out of 10 β-glucosidase genes had predicted signal peptides, indicating a transmembrane capability of these enzymes to mediate cellulose hydrolysis within the gut of the termites. To confirm the activities of these β-glucosidase genes, three genes (bgl5, bgl7, and bgl9) were successfully expressed and purified. The optimal temperature and pH of these enzymes largely resembled the environment of the host’s gut. The gut microbiota composition of the fungus-growing termite was also determined by 454 pyrosequencing, showing that Bacteroidetes was the most dominant phylum. The diversity and the enzyme properties of β-glucosidases revealed in this study suggested that Bacteroidetes as the major member in fungus-growing termites contributed to cello-oligomer degradation in cellulose-hydrolyzing process and represented a rich source for β-glucosidase genes.

Keywords

Cellulase Cellobiose Bacteroidetes Cellulose Hydrolysis Symbiotic Fungus 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Notes

Acknowledgments

This work was supported by grants of the National Natural Science Foundation of China (31070098). We appreciate Professor Shubiao Wu from the University of New England for his critical reading and kind suggestions. We also give our thanks to Prashanth Singanallur and Lauren Christine Radlinski from the University of Illinois at Urbana-Champaign for proofreading. We appreciated Dr. Haokui Zhou from Department of Microbiology, The Chinese University of Hongkong and Lei zhang from Logic Informatics Co.,Ltd., for their kind help in data analysis.

Conflict of Interest

The authors have no conflict of interest to declare.

Supplementary material

248_2014_388_MOESM1_ESM.pptx (47.3 mb)
ESM 1 (PPTX 48413 kb)

References

  1. 1.
    Breznak JA (1994) Role of microorganisms in the digestion of lignocellulose by termites. Annu Rev Entomol 39:453–487CrossRefGoogle Scholar
  2. 2.
    Abo-Khatwa AN (1989) Termitomyces: a new source of potent cellulases. JKing Abulaziz Univ Science 1:51–59CrossRefGoogle Scholar
  3. 3.
    Breznak JA (1982) Intestinal microbiota of termites and other xylophagous insects. Annu Rev Microbiol 36:323–343CrossRefPubMedGoogle Scholar
  4. 4.
    Wood TG, Thomas RJ (1989) The mutualistic association between Macrotermitinae and Termitomyces. In: Wilding N, Collins NM, Hammond PM, Webber JF (eds) Insect-fungus interaction. Academic, London, pp 69–92CrossRefGoogle Scholar
  5. 5.
    Hongoh Y, Ekpornprasit L, Inoue T, Moriya S, Trakulnaleamsai S, Ohkuma M, Noparatnaraporn N, Kudo T (2006) Intracolony variation of bacterial gut microbiota among castes and ages in the fungus-growing termite Macrotermes gilvus. Mol Ecol 15(2):505–516CrossRefPubMedGoogle Scholar
  6. 6.
    Mackenzie LM, Muigai AT, Osir EO, Lwande W, Keller M, Yoledo G, Boga HI (2007) Bacterial diversity in the intestinal tract of the fungus-cultivating termite Macrotermes michaelseni (Sjostedt). African Journal of Biotechnology 6(6):658–667Google Scholar
  7. 7.
    Hongoh Y, Deevong P, Inoue T, Moriya S, Trakulnaleamsai S, Ohkuma M, Vongkaluang C, Noparatnaraporn N, Kudo T (2005) Intra- and interspecific comparisons of bacterial diversity and community structure support coevolution of gut microbiota and termite host. Appl Environ Microbiol 71(11):6590–6599PubMedCentralCrossRefPubMedGoogle Scholar
  8. 8.
    Shinzato N, Muramatsu M, Matsui T, Watanabe Y (2007) Phylogenetic analysis of the gut bacterial microflora of the fungus-growing termite Odontotermes formosanus. Biosci Biotechnol Biochem 71(4):906–915CrossRefPubMedGoogle Scholar
  9. 9.
    Long YH, Xie L, Liu N, Yan X, Li MH, Fan MZ, Wang QA (2010) Comparison of gut-associated and nest-associated microbial communities of a fungus-growing termite (Odontotermes yunnanensis). Insect Sci 17(3):265–276CrossRefGoogle Scholar
  10. 10.
    Rouland-Lefevre C, Inoue T, Johjima T (2006) Termitomyces/termite interactions. In: Konig H, Varma A (eds) Intestinal microorganisms of soil invertebrates. Springer, Berlin, pp 335–350CrossRefGoogle Scholar
  11. 11.
    Mathew GM, Ju YM, Lai CY, Mathew DC, Huang CC (2012) Microbial community analysis in the termite gut and fungus comb of Odontotermes formosanus: the implication of Bacillus as mutualists. FEMS Microbiol Ecol 79(2):504–517CrossRefPubMedGoogle Scholar
  12. 12.
    Tokuda G, Lo N, Watanabe H, Arakawa G, Matsumoto T, Noda H (2004) Major alteration of the expression site of endogenous cellulases in members of an apical termite lineage. Molecular Ecology 13(10):3219–3228CrossRefPubMedGoogle Scholar
  13. 13.
    Wu Y, Chi S, Yun C, Shen Y, Tokuda G, Ni J (2012) Molecular cloning and characterization of an endogenous digestive beta-glucosidase from the midgut of the fungus-growing termite Macrotermes barneyi. Insect Mol Biol 21(6):604–614CrossRefPubMedGoogle Scholar
  14. 14.
    Hyodo F, Inoue T, Azuma JI, Tayasu I, Tabe T (2000) Role of the mutualistic fungus in lignin degradation in the fungus-growing termite Macrotermes gilvus (Isoptera; Macrotermitinae). Soil Biol Biochem 32:653–658CrossRefGoogle Scholar
  15. 15.
    Hyodo F, Tayasu I, Inoue T, Azuma JI, Kudo T, Abe T (2003) Differential role of symbiotic fungi in lignin degradation and food provision for fungus-growing termites (Macrotermitinae: Isoptera). Functional Ecology 17:186–193CrossRefGoogle Scholar
  16. 16.
    Johjima T, Taprab Y, Noparatnaraporn N, Kudo T, Ohkuma M (2006) Large-scale identification of transcripts expressed in a symbiotic fungus (Termitomyces) during plant biomass degradation. Appl Microbiol Biotechnol 73(1):195–203CrossRefPubMedGoogle Scholar
  17. 17.
    Martin M, Martin J (1978) Cellulose digestion in the midgut of the fungus-growing termite Macrotermes natalensis: the role of acquired digestive enzyme. Science 199:1453–1455CrossRefPubMedGoogle Scholar
  18. 18.
    Matoub M, Rouland C (1995) Purification and properties of the xylanases from the termite Macrotermes bellicosus and its symbiotic fungus Termitomyces sp. Comp Biochem Physiol B Biochem Mol Biol 112(4):629–635CrossRefPubMedGoogle Scholar
  19. 19.
    Liu N, Yan X, Zhang M, Xie L, Wang Q, Huang Y, Zhou X, Wang S, Zhou Z (2011) Microbiome of fungus-growing termites: a new reservoir for lignocellulase genes. Appl Environ Microbiol 77(1):48–56PubMedCentralCrossRefPubMedGoogle Scholar
  20. 20.
    Liu N, Zhang L, Zhou H, Zhang M, Yan X, Wang Q, Long Y, Xie L, Wang S, Huang Y, Zhou Z (2013) Metagenomic insights into metabolic capacities of the gut microbiota in a fungus-cultivating termite (Odontotermes yunnanensis). PLoS One 8(7):e69184PubMedCentralCrossRefPubMedGoogle Scholar
  21. 21.
    Bastien G, Arnal G, Bozonnet A, Laguerre S, Ferreir F, GFaure R, Henrissat B, Lefevre G, Robe P, Bouchez O, Noirot C (2013) Mining for hemicellulases in the fungus-growing termite Pseudacanthotermes militaris using functional metagenomics. Biotechnol Biofuels 6:78. doi: 10.1186/1754-6834-1186-1178
  22. 22.
    DeSantis TZ, Jr., Hugenholtz P, Keller K, Brodie EL, Larsen N, Piceno YM, Phan R, Andersen GL (2006) NAST: a multiple sequence alignment server for comparative analysis of 16S rRNA genes. Nucleic Acids Res 34(Web Server issue):W394-399Google Scholar
  23. 23.
    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(7):5069–5072PubMedCentralCrossRefPubMedGoogle Scholar
  24. 24.
    Huber T, Faulkner G, Hugenholtz P (2004) Bellerophon: a program to detect chimeric sequences in multiple sequence alignments. Bioinformatics 20(14):2317–2319CrossRefPubMedGoogle Scholar
  25. 25.
    Schloss PD, Handelsman J (2005) Introducing DOTUR, a computer program for defining operational taxonomic units and estimating species richness. Appl Environ Microbiol 71(3):1501–1506PubMedCentralCrossRefPubMedGoogle Scholar
  26. 26.
    Wang Q, Garrity GM, Tiedje JM, Cole JR (2007) Naive Bayesian classifier for rapid assignment of rRNA sequences into the new bacterial taxonomy. Appl Environ Microbiol 73(16):5261–5267PubMedCentralCrossRefPubMedGoogle Scholar
  27. 27.
    Huson DH, Auch AF, Qi J, Schuster SC (2007) Megan analysis of metagenomic data. Genome Res 17:377–386PubMedCentralCrossRefPubMedGoogle Scholar
  28. 28.
    Mitra S, Klar B, Huson DH (2009) Visual and statistical comparison of metagenomes. Bioinformatics 25:1849CrossRefPubMedGoogle Scholar
  29. 29.
    Katoh K, Toh H (2008) Recent developments in the MAFFT multiple sequence alignment program. Brief Bioinformatics 9:286–298CrossRefPubMedGoogle Scholar
  30. 30.
    Mackenzie LM, Muigai AT, Osir EO, Lwande W, Keller M, Toledo G, Hi B (2007) Bacterial diversity in the intestinal tract of the fungus-cultivating termite Macrotermes michaelseni (Sjostedt). African Journal of Biotechnology 6:658–667Google Scholar
  31. 31.
    Kohler T, Dietrich C, Scheffrahn RH, Brune A (2012) High-resolution analysis of gut environment and bacterial microbiota reveals functional compartmentation of the gut in wood-feeding higher termites (Nasutitermes spp.). Appl Environ Microbiol 78(13):4691–4701PubMedCentralCrossRefPubMedGoogle Scholar
  32. 32.
    Warnecke F, Luginbuhl P, Ivanova N, Ghassemian M, Richardson TH, Stege JT, Cayouette M, McHardy AC, Djordjevic G, Aboushadi N, Sorek R, Tringe SG, Podar M, Martin HG, Kunin V, Dalevi D, Madejska J, Kirton E, Platt D, Szeto E, Salamov A, Barry K, Mikhailova N, Kyrpides NC, Matson EG, Ottesen EA, Zhang X, Hernandez M, Murillo C, Acosta LG, Rigoutsos I, Tamayo G, Green BD, Chang C, Rubin EM, Mathur EJ, Robertson DE, Hugenholtz P, Leadbetter JR (2007) Metagenomic and functional analysis of hindgut microbiota of a wood-feeding higher termite. Nature 450(7169):560–565CrossRefPubMedGoogle Scholar
  33. 33.
    Lawson PA, Falsen E, Inganas E, Weyant RS, Collins MD (2002) Dysgonomonas mossii sp. nov., from human sources. Syst Appl Microbiol 25(2):194–197CrossRefPubMedGoogle Scholar
  34. 34.
    Martens EC, Koropatkin NM, Smith TJ, Gordon JI (2009) Complex glycan catabolism by the human gut microbiota: the Bacteroidetes Sus-like paradigm. J Biol Chem 284(37):24673–24677PubMedCentralCrossRefPubMedGoogle Scholar
  35. 35.
    Dodd D, Mackie RI, Cann IK (2011) Xylan degradation, a metabolic property shared by rumen and human colonic Bacteroidetes. Mol Microbiol 79(2):292–304PubMedCentralCrossRefPubMedGoogle Scholar
  36. 36.
    Mackenzie AK, Pope PB, Pedersen HL, Gupta R, Morrison M, Willats WG, Eijsink VG (2012) Two SusD-like proteins encoded within a polysaccharide utilization locus of an uncultured ruminant Bacteroidetes phylotype bind strongly to cellulose. Appl Environ Microbiol 78(16):5935–5937PubMedCentralCrossRefPubMedGoogle Scholar
  37. 37.
    Carpenter KJ, Keeling PJ (2007) Morphology and phylogenetic position of Eucomonympha imla (Parabasalia: Hypermastigida). J Eukaryot Microbiol 54(4):325–332CrossRefPubMedGoogle Scholar
  38. 38.
    Hansen PS, Jensen TG, Gahrn-Hansen B (2005) Dysgonomonas capnocytophagoides bacteraemia in a neutropenic patient treated for acute myeloid leukaemia. APMIS 113(3):229–231CrossRefPubMedGoogle Scholar
  39. 39.
    Hironaga M, Yamane K, Inaba M, Haga Y, Arakawa Y (2008) Characterization and antimicrobial susceptibility of Dysgonomonas capnocytophagoides isolated from human blood sample. Jpn J Infect Dis 61(3):212–213PubMedGoogle Scholar
  40. 40.
    Hofstad T, Olsen I, Eribe ER, Falsen E, Collins MD, Lawson PA (2000) Dysgonomonas gen. nov. to accommodate Dysgonomonas gadei sp. nov., an organism isolated from a human gall bladder, and Dysgonomonas capnocytophagoides (formerly CDC group DF-3). Int J Syst Evol Microbiol 50(6):2189–2195CrossRefPubMedGoogle Scholar
  41. 41.
    Lawson PA, Carlson P, Wernersson S, Moore ER, Falsen E (2010) Dysgonomonas hofstadii sp. nov., isolated from a human clinical source. Anaerobe 16(2):161–164CrossRefPubMedGoogle Scholar
  42. 42.
    Matsumoto T, Kawakami Y, Oana K, Honda T, Yamauchi K, Okimura Y, Shiohara M, Kasuga E (2006) First isolation of Dysgonomonas mossii from intestinal juice of a patient with pancreatic cancer. Arch Med Res 37(7):914–916CrossRefPubMedGoogle Scholar
  43. 43.
    Collins MD, Lawson PA, Labrenz M, Tindall BJ, Weiss N, Hirsch P (2002) Nesterenkonia lacusekhoensis sp. nov., isolated from hypersaline Ekho Lake, East Antarctica, and emended description of the genus Nesterenkonia. Int J Syst Evol Microbiol 52(Pt 4):1145–1150PubMedGoogle Scholar
  44. 44.
    Martin MM, Matrtin JS (1979) The distribution and origins of the cellulolytic enzymes of the higher termite, Macrotermes natalensis. Physiological Zoology 52(1):11–21Google Scholar
  45. 45.
    Bhatia Y, Mishra S, Bisaria VS (2002) Microbial beta-glucosidases: cloning, properties, and applications. Crit Rev Biotechnol 22:375–407CrossRefPubMedGoogle Scholar
  46. 46.
    Suto M, Tomita F (2001) Induction and catabolite repression mechanisms of cellulase in fungi. J Biosci Bioeng 92(4):305–311CrossRefPubMedGoogle Scholar
  47. 47.
    Percival Zhang YH, Himmel ME, Mielenz JR (2006) Outlook for cellulase improvement: screening and selection strategies. Biotechnol Adv 24(5):452–481CrossRefPubMedGoogle Scholar
  48. 48.
    Feng Y, Duan CJ, Liu L, Tang JL, Feng JX (2009) Properties of a metagenome-derived beta-glucosidase from the contents of rabbit cecum. Biosci Biotechnol Biochem 73(7):1470–1473CrossRefPubMedGoogle Scholar
  49. 49.
    Jiang C, Ma G, Li S, Hu T, Che Z, Shen P, Yan B, Wu B (2009) Characterization of a novel beta-glucosidase-like activity from a soil metagenome. J Microbiol 47(5):542–548CrossRefPubMedGoogle Scholar
  50. 50.
    Kataeva IA, Seidel RD 3rd, Shah A, West LT, Li XL, Ljungdahl LG (2002) The fibronectin type 3-like repeat from the Clostridium thermocellum cellobiohydrolase CbhA promotes hydrolysis of cellulose by modifying its surface. Appl Environ Microbiol 68(9):4292–4300PubMedCentralCrossRefPubMedGoogle Scholar
  51. 51.
    Brulc JM, Antonopoulos DA, Miller ME, Wilson MK, Yannarell AC, Dinsdale EA, Edwards RE, Frank ED, Emerson JB, Wacklin P, Coutinho PM, Henrissat B, Nelson KE, White BA (2009) Gene-centric metagenomics of the fiber-adherent bovine rumen microbiome reveals forage specific glycoside hydrolases. Proc Natl Acad Sci U S A 106(6):1948–1953PubMedCentralCrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2014

Authors and Affiliations

  • Meiling Zhang
    • 1
  • Ning Liu
    • 3
    • 4
  • Changli Qian
    • 2
  • Qianfu Wang
    • 2
  • Qian Wang
    • 3
  • Yanhua Long
    • 5
  • Yongping Huang
    • 3
  • Zhihua Zhou
    • 2
  • Xing Yan
    • 2
    • 6
  1. 1.School of Life SciencesEast China Normal UniversityShanghaiChina
  2. 2.Key Laboratory of Synthetic Biology, Institute of Plant Physiology and EcologyShanghai Institutes for Biological Sciences, Chinese Academy of SciencesShanghaiChina
  3. 3.Key Laboratory of Insect Development and Evolutionary Biology, Institute of Plant Physiology and EcologyShanghai Institutes for Biological Sciences, Chinese Academy of SciencesShanghaiChina
  4. 4.Zhejiang Academy of Agricultural SciencesHangzhouChina
  5. 5.School of Life ScienceAnhui Agricultural UniversityHefeiChina
  6. 6.Key Laboratory of Synthetic Biology, Shanghai Institute of Plant Physiology and EcologyChinese Academy of SciencesShanghaiChina

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