Microbial Ecology

, Volume 71, Issue 1, pp 243–255 | Cite as

Phenotypic and Physiological Characterization of the Epibiotic Interaction Between TM7x and Its Basibiont Actinomyces

  • Batbileg Bor
  • Nicole Poweleit
  • Justin S. Bois
  • Lujia Cen
  • Joseph K. Bedree
  • Z. Hong Zhou
  • Robert P. Gunsalus
  • Renate Lux
  • Jeffrey S. McLean
  • Xuesong He
  • Wenyuan Shi
Human Microbiome


Despite many examples of obligate epibiotic symbiosis (one organism living on the surface of another) in nature, such an interaction has rarely been observed between two bacteria. Here, we further characterize a newly reported interaction between a human oral obligate parasitic bacterium TM7x (cultivated member of Candidatus Saccharimonas formerly Candidate Phylum TM7), and its basibiont Actinomyces odontolyticus species (XH001), providing a model system to study epiparasitic symbiosis in the domain Bacteria. Detailed microscopic studies indicate that both partners display extensive morphological changes during symbiotic growth. XH001 cells manifested as short rods in monoculture, but displayed elongated and hyphal morphology when physically associated with TM7x. Interestingly, these dramatic morphological changes in XH001 were also induced in oxygen-depleted conditions, even in the absence of TM7x. Targeted quantitative real-time PCR (qRT-PCR) analyses revealed that both the physical association with TM7x as well as oxygen depletion triggered up-regulation of key stress response genes in XH001, and in combination, these conditions act in an additive manner. TM7x and XH001 co-exist with relatively uniform cell morphologies under nutrient-replete conditions. However, upon nutrient depletion, TM7x-associated XH001 displayed a variety of cell morphologies, including swollen cell body, clubbed-ends, and even cell lysis, and a large portion of TM7x cells transformed from ultrasmall cocci into elongated cells. Our study demonstrates a highly dynamic interaction between epibiont TM7x and its basibiont XH001 in response to physical association or environmental cues such as oxygen level and nutritional status, as reflected by their morphological and physiological changes during symbiotic growth.


Obligate Epibiont Symbiosis Bacterial interaction TM7 Actinomyces 



We thank the members of the Shi and Lux laboratories for their feedback and invaluable discussion. We also thank Melissa Agnello for providing extensive editing of the manuscript. We thank the Chemistry and Biochemistry instrumentation facility at UCLA for providing access to the confocal microscope. This work was supported in part by grants from the National Institutes of Health (1R01DE023810-01) and Oral Health-Research Postdoctoral Training Program (B.B., UCLA School of Dentistry T90 award).

Compliance with Ethical Standards

Conflict of Interest

The authors declare that they have no conflict of interests.

Supplementary material

248_2015_711_MOESM1_ESM.docx (24 kb)
ESM 1 (DOCX 23 kb)
248_2015_711_MOESM2_ESM.pdf (4.6 mb)
Fig. S1 TM7x induces morphological changes in XH001. a XH001/TM7x co-culture grown under microaerophilic condition for 24 h showing clear micro-aggregation. Scale bar is 10 μm. b Monoculture of XH001. c, d Establishment of physical association between XH001 and TM7x via attachment assay (see supplementary methods). The re-attached XH001 cells were passaged two times (c) and four times (d) respectively in fresh medium. Scale bars are 5 μm (PDF 4683 kb)
248_2015_711_MOESM3_ESM.pdf (8.4 mb)
Fig. S2 Morphology of XH001 under different oxygen conditions. Phase contrast image of XH001 alone (a) under high oxygen condition (19.7 % O2, 5 % CO2). Phase contrast images of XH001 alone (b) and with TM7x (c) under normal atmospheric condition (20.9 % O2, 0.04 % CO2) after 24 h. df XH001 alone cells grown in microaerophilic condition (d) were shifted to anaerobic condition (e) and then back to the microaerophilic condition (f) before taking the phase contrast images. All scale bars indicate 5 μm (PDF 8569 kb)
248_2015_711_MOESM4_ESM.pdf (25.8 mb)
Fig. S3 FISH staining of XH001 alone and with TM7x. FISH probes specific to TM7x (white) and XH001 (red) were used to stain the fixed samples. Green represents syto9 staining of all bacteria. a XH001 monoculture grown under a microaerophilic condition for 24 h shows short rod morphology with XH001-specific probe (red) and universal DNA stain syto9 (green), but no staining with TM7x-specific probe, confirming our probe specificity. b, c XH001 alone (b) and with TM7x (c) grown under anaerobic condition for 24 h. Similar to a, we do not see any staining of TM7x probe in the XH001 alone cells, whereas in the co-culture, we saw elongated TM7x. Under anaerobic condition, XH001-specific probe stained the cells non-uniformly, suggesting that these cells were stressed and probably lost their cell content. All scale bars indicate 10 μm. (PDF 26435 kb)


  1. 1.
    Joseph S (2002) Symbiosis: mechanisms and model systems. Kluwer Academic Publisher, New YorkGoogle Scholar
  2. 2.
    Guerrero R, Pedros-Alio C, Esteve I et al (1986) Predatory prokaryotes: predation and primary consumption evolved in bacteria. Proc Natl Acad Sci 83:2138–2142. doi: 10.1073/pnas.83.7.2138 CrossRefPubMedCentralPubMedGoogle Scholar
  3. 3.
    Huber H, Hohn MJ, Rachel R et al (2002) A new phylum of Archaea represented by a nanosized hyperthermophilic symbiont. Nature 417:63–67. doi: 10.1038/417063a CrossRefPubMedGoogle Scholar
  4. 4.
    Lambina VA, Afinogenova AV, Romaĭ Penabad S et al (1982) Micavibrio admirandus gen. et sp. nov. Mikrobiologiia 51:114–117PubMedGoogle Scholar
  5. 5.
    Grice EA, Segre JA (2012) The human microbiome: our second genome. Annu Rev Genomics Hum Genet 13:151–170. doi: 10.1146/annurev-genom-090711-163814 CrossRefPubMedCentralPubMedGoogle Scholar
  6. 6.
    Human Microbiome Project Consortium (2012) Structure, function and diversity of the healthy human microbiome. Nature 486:207–214. doi: 10.1038/nature11234 CrossRefGoogle Scholar
  7. 7.
    Roesch LFW, Fulthorpe RR, Riva A et al (2007) Pyrosequencing enumerates and contrasts soil microbial diversity. ISME J 1:283–290. doi: 10.1038/ismej.2007.53 PubMedCentralPubMedGoogle Scholar
  8. 8.
    Schauer R, Bienhold C, Ramette A, Harder J (2010) Bacterial diversity and biogeography in deep-sea surface sediments of the South Atlantic Ocean. ISME J 4:159–170. doi: 10.1038/ismej.2009.106 CrossRefPubMedGoogle Scholar
  9. 9.
    Bryant MP, Wolin EA, Wolin MJ, Wolfe RS (1967) Methanobacillus omelianskii, a symbiotic association of two species of bacteria. Arch Für Mikrobiol 59:20–31CrossRefGoogle Scholar
  10. 10.
    Lancy P, Dirienzo JM, Appelbaum B et al (1983) Corncob formation between Fusobacterium nucleatum and Streptococcus sanguis. Infect Immun 40:303–309PubMedCentralPubMedGoogle Scholar
  11. 11.
    Overmann J (2010) The phototrophic consortium “Chlorochromatium aggregatum”—a model for bacterial heterologous multicellularity. Adv Exp Med Biol 675:15–29. doi: 10.1007/978-1-4419-1528-3_2 CrossRefPubMedGoogle Scholar
  12. 12.
    Davidov Y, Huchon D, Koval SF, Jurkevitch E (2006) A new Alpha-proteobacterial clade of Bdellovibrio-like predators: implications for the mitochondrial endosymbiotic theory. Environ Microbiol 8:2179–2188. doi: 10.1111/j.1462-2920.2006.01101.x CrossRefPubMedGoogle Scholar
  13. 13.
    Dashiff A, Junka RA, Libera M, Kadouri DE (2011) Predation of human pathogens by the predatory bacteria Micavibrio aeruginosavorus and Bdellovibrio bacteriovorus. J Appl Microbiol 110:431–444. doi: 10.1111/j.1365-2672.2010.04900.x CrossRefPubMedGoogle Scholar
  14. 14.
    He X, McLean JS, Edlund A et al (2015) Cultivation of a human-associated TM7 phylotype reveals a reduced genome and epibiotic parasitic lifestyle. Proc Natl Acad Sci U S A 112:244–249. doi: 10.1073/pnas.1419038112 CrossRefPubMedCentralPubMedGoogle Scholar
  15. 15.
    Lasken RS, McLean JS (2014) Recent advances in genomic DNA sequencing of microbial species from single cells. Nat Rev Genet 15:577–584. doi: 10.1038/nrg3785 CrossRefPubMedCentralPubMedGoogle Scholar
  16. 16.
    Albertsen M, Hugenholtz P, Skarshewski A et al (2013) Genome sequences of rare, uncultured bacteria obtained by differential coverage binning of multiple metagenomes. Nat Biotechnol 31:533–538. doi: 10.1038/nbt.2579 CrossRefPubMedGoogle Scholar
  17. 17.
    Marcy Y, Ouverney C, Bik EM et al (2007) Dissecting biological “dark matter” with single-cell genetic analysis of rare and uncultivated TM7 microbes from the human mouth. Proc Natl Acad Sci 104:11889–11894. doi: 10.1073/pnas.0704662104 CrossRefPubMedCentralPubMedGoogle Scholar
  18. 18.
    Podar M, Abulencia CB, Walcher M et al (2007) Targeted access to the genomes of low-abundance organisms in complex microbial communities. Appl Environ Microbiol 73:3205–3214. doi: 10.1128/AEM.02985-06 CrossRefPubMedCentralPubMedGoogle Scholar
  19. 19.
    Brown CT, Hug LA, Thomas BC et al (2015) Unusual biology across a group comprising more than 15% of domain Bacteria. Nature. doi: 10.1038/nature14486 Google Scholar
  20. 20.
    Luef B, Frischkorn KR, Wrighton KC et al (2015) Diverse uncultivated ultra-small bacterial cells in groundwater. Nat Commun 6:6372. doi: 10.1038/ncomms7372 CrossRefPubMedGoogle Scholar
  21. 21.
    Bik EM, Eckburg PB, Gill SR et al (2006) Molecular analysis of the bacterial microbiota in the human stomach. Proc Natl Acad Sci 103:732–737. doi: 10.1073/pnas.0506655103 CrossRefPubMedCentralPubMedGoogle Scholar
  22. 22.
    Dewhirst FE, Chen T, Izard J et al (2010) The human oral microbiome. J Bacteriol 192:5002–5017. doi: 10.1128/JB.00542-10 CrossRefPubMedCentralPubMedGoogle Scholar
  23. 23.
    Dinis JM, Barton DE, Ghadiri J et al (2011) In search of an uncultured human-associated TM7 bacterium in the environment. PLoS One 6:e21280. doi: 10.1371/journal.pone.0021280 CrossRefPubMedCentralPubMedGoogle Scholar
  24. 24.
    Eckburg PB, Bik EM, Bernstein CN et al (2005) Diversity of the human intestinal microbial flora. Science 308:1635–1638. doi: 10.1126/science.1110591 CrossRefPubMedCentralPubMedGoogle Scholar
  25. 25.
    Ferrari BC, Binnerup SJ, Gillings M (2005) Microcolony cultivation on a soil substrate membrane system selects for previously uncultured soil bacteria. Appl Environ Microbiol 71:8714–8720. doi: 10.1128/AEM.71.12.8714-8720.2005 CrossRefPubMedCentralPubMedGoogle Scholar
  26. 26.
    Fredricks DN, Fiedler TL, Marrazzo JM (2005) Molecular identification of bacteria associated with bacterial vaginosis. N Engl J Med 353:1899–1911. doi: 10.1056/NEJMoa043802 CrossRefPubMedGoogle Scholar
  27. 27.
    Gao Z, Tseng C, Pei Z, Blaser MJ (2007) Molecular analysis of human forearm superficial skin bacterial biota. Proc Natl Acad Sci 104:2927–2932. doi: 10.1073/pnas.0607077104 CrossRefPubMedCentralPubMedGoogle Scholar
  28. 28.
    Hanada A, Kurogi T, Giang NM et al (2014) Bacteria of the candidate phylum TM7 are prevalent in acidophilic nitrifying sequencing-batch reactors. Microbes Environ JSME 29:353–362. doi: 10.1264/jsme2.ME14052 CrossRefGoogle Scholar
  29. 29.
    Hugenholtz P, Tyson GW, Webb RI et al (2001) Investigation of candidate division TM7, a recently recognized major lineage of the domain Bacteria with no known pure-culture representatives. Appl Environ Microbiol 67:411–419. doi: 10.1128/AEM.67.1.411-419.2001 CrossRefPubMedCentralPubMedGoogle Scholar
  30. 30.
    Kianoush N, Adler CJ, Nguyen K-AT et al (2014) Bacterial profile of dentine caries and the impact of pH on bacterial population diversity. PLoS One 9:e92940. doi: 10.1371/journal.pone.0092940 CrossRefPubMedCentralPubMedGoogle Scholar
  31. 31.
    Kuehbacher T, Rehman A, Lepage P et al (2008) Intestinal TM7 bacterial phylogenies in active inflammatory bowel disease. J Med Microbiol 57:1569–1576. doi: 10.1099/jmm.0.47719-0 CrossRefPubMedGoogle Scholar
  32. 32.
    Paster BJ, Boches SK, Galvin JL et al (2001) Bacterial diversity in human subgingival plaque. J Bacteriol 183:3770–3783. doi: 10.1128/JB.183.12.3770-3783.2001 CrossRefPubMedCentralPubMedGoogle Scholar
  33. 33.
    Rheims H, Spröer C, Rainey FA, Stackebrandt E (1996) Molecular biological evidence for the occurrence of uncultured members of the Actinomycete line of descent in different environments and geographical locations. Microbiol Read Engl 142(Pt 10):2863–2870. doi: 10.1099/13500872-142-10-2863 CrossRefGoogle Scholar
  34. 34.
    Soro V, Dutton LC, Sprague SV et al (2014) Axenic culture of a Candidate division TM7 bacterium from the human oral cavity and biofilm interactions with other oral bacteria. Appl Environ Microbiol 80:6480–6489. doi: 10.1128/AEM.01827-14 CrossRefPubMedCentralPubMedGoogle Scholar
  35. 35.
    Brinig MM, Lepp PW, Ouverney CC et al (2003) Prevalence of bacteria of division TM7 in human subgingival plaque and their association with disease. Appl Environ Microbiol 69:1687–1694. doi: 10.1128/AEM.69.3.1687-1694.2003 CrossRefPubMedCentralPubMedGoogle Scholar
  36. 36.
    Kumar PS, Griffen AL, Barton JA et al (2003) New bacterial species associated with chronic periodontitis. J Dent Res 82:338–344CrossRefPubMedGoogle Scholar
  37. 37.
    Liu B, Faller LL, Klitgord N et al (2012) Deep sequencing of the oral microbiome reveals signatures of periodontal disease. PLoS One 7:e37919. doi: 10.1371/journal.pone.0037919 CrossRefPubMedCentralPubMedGoogle Scholar
  38. 38.
    Paster BJ, Russell MK, Alpagot T et al (2002) Bacterial diversity in necrotizing ulcerative periodontitis in HIV-positive subjects. Ann Periodontol Am Acad Periodontol 7:8–16. doi: 10.1902/annals.2002.7.1.8 CrossRefGoogle Scholar
  39. 39.
    Rylev M, Bek-Thomsen M, Reinholdt J et al (2011) Microbiological and immunological characteristics of young Moroccan patients with aggressive periodontitis with and without detectable Aggregatibacter actinomycetemcomitans JP2 infection. Mol Oral Microbiol 26:35–51. doi: 10.1111/j.2041-1014.2010.00593.x CrossRefPubMedGoogle Scholar
  40. 40.
    Becker MR, Paster BJ, Leys EJ et al (2002) Molecular analysis of bacterial species associated with childhood caries. J Clin Microbiol 40:1001–1009CrossRefPubMedCentralPubMedGoogle Scholar
  41. 41.
    Colombo AV, Silva CM, Haffajee A, Colombo APV (2006) Identification of oral bacteria associated with crevicular epithelial cells from chronic periodontitis lesions. J Med Microbiol 55:609–615. doi: 10.1099/jmm.0.46417-0 CrossRefPubMedGoogle Scholar
  42. 42.
    Kanasi E, Dewhirst FE, Chalmers NI et al (2010) Clonal analysis of the microbiota of severe early childhood caries. Caries Res 44:485–497. doi: 10.1159/000320158 CrossRefPubMedCentralPubMedGoogle Scholar
  43. 43.
    Ling Z, Kong J, Jia P et al (2010) Analysis of oral microbiota in children with dental caries by PCR-DGGE and barcoded pyrosequencing. Microb Ecol 60:677–690. doi: 10.1007/s00248-010-9712-8 CrossRefPubMedGoogle Scholar
  44. 44.
    Nagy KN, Sonkodi I, Szöke I et al (1998) The microflora associated with human oral carcinomas. Oral Oncol 34:304–308CrossRefPubMedGoogle Scholar
  45. 45.
    Sato T, Watanabe K, Kumada H et al (2012) Peptidoglycan of Actinomyces naeslundii induces inflammatory cytokine production and stimulates osteoclastogenesis in alveolar bone resorption. Arch Oral Biol 57:1522–1528. doi: 10.1016/j.archoralbio.2012.07.012 CrossRefPubMedGoogle Scholar
  46. 46.
    Aas JA, Paster BJ, Stokes LN et al (2005) Defining the normal bacterial flora of the oral cavity. J Clin Microbiol 43:5721–5732. doi: 10.1128/JCM.43.11.5721-5732.2005 CrossRefPubMedCentralPubMedGoogle Scholar
  47. 47.
    Paul D, Reddy D, Mukherjee D et al (2011) Actinomyces. In: Liu D (ed) Mol Detect Hum Bact. Pathog. CRC Press, p 23–30Google Scholar
  48. 48.
    Jones E, Oliphant E, Peterson P et al (2001) SciPy: open source scientific tools for python.
  49. 49.
    Walt S, Schonberger JL, Nunez-Iglesias J et al (2014) Scikit-image: image processing in pythong. PeerJ 2:e453. doi: 10.7717/peerj.453 CrossRefPubMedCentralPubMedGoogle Scholar
  50. 50.
    Pfaffl MW (2001) A new mathematical model for relative quantification in real-time RT-PCR. Nucleic Acids Res 29:e45CrossRefPubMedCentralPubMedGoogle Scholar
  51. 51.
    MJ Hill, PD Marsh (1990) Factors controlling the microflora of the huealthy mouth. In: Hum Microb Ecol. CRC Press. Inc., p 34–37Google Scholar
  52. 52.
    Loesche WJ, Gusberti F, Mettraux G et al (1983) Relationship between oxygen tension and subgingival bacterial flora in untreated human periodontal pockets. Infect Immun 42:659–667PubMedCentralPubMedGoogle Scholar
  53. 53.
    Schaal KP, Yassin AA (2012) Genus I. Actinomyces. In: Bergeys Man Syst Bacteriol. Springer, p 42–109Google Scholar
  54. 54.
    Jones TH, Vail KM, McMullen LM (2013) Filament formation by foodborne bacteria under sublethal stress. Int J Food Microbiol 165:97–110. doi: 10.1016/j.ijfoodmicro.2013.05.001 CrossRefPubMedGoogle Scholar
  55. 55.
    Herskowitz I (1988) Life cycle of the budding yeast Saccharomyces cerevisiae. Microbiol Rev 52:536–553PubMedCentralPubMedGoogle Scholar
  56. 56.
    Hirsch P (1974) Budding bacteria. Annu Rev Microbiol 28:391–440CrossRefPubMedGoogle Scholar
  57. 57.
    Greif D, Wesner D, Regtmeier J, Anselmetti D (2010) High resolution imaging of surface patterns of single bacterial cells. Ultramicroscopy 110:1290–1296. doi: 10.1016/j.ultramic.2010.06.004 CrossRefPubMedGoogle Scholar
  58. 58.
    Patil S, Valdramidis VP, Karatzas KAG et al (2011) Assessing the microbial oxidative stress mechanism of ozone treatment through the responses of Escherichia coli mutants. J Appl Microbiol 111:136–144. doi: 10.1111/j.1365-2672.2011.05021.x CrossRefPubMedGoogle Scholar
  59. 59.
    Glazebrook J (2005) Contrasting mechanisms of defense against biotrophic and necrotrophic pathogens. Annu Rev Phytopathol 43:205–227. doi: 10.1146/annurev.phyto.43.040204.135923 CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2015

Authors and Affiliations

  • Batbileg Bor
    • 1
  • Nicole Poweleit
    • 2
  • Justin S. Bois
    • 3
  • Lujia Cen
    • 1
  • Joseph K. Bedree
    • 1
  • Z. Hong Zhou
    • 2
    • 4
  • Robert P. Gunsalus
    • 2
  • Renate Lux
    • 1
  • Jeffrey S. McLean
    • 5
  • Xuesong He
    • 1
  • Wenyuan Shi
    • 1
  1. 1.Section of Oral Biology, School of DentistryUniversity of CaliforniaLos AngelesUSA
  2. 2.Department of Microbiology, Immunology, and Molecular GeneticsUniversity of CaliforniaLos AngelesUSA
  3. 3.Division of Biology and Biological EngineeringCalifornia Institute of TechnologyPasadenaUSA
  4. 4.California Nanosystems InstituteUniversity of CaliforniaLos AngelesUSA
  5. 5.Department of PeriodonticsUniversity of WashingtonSeattleUSA

Personalised recommendations