Skip to main content
Log in

Composition of the Cutaneous Bacterial Community in Japanese Amphibians: Effects of Captivity, Host Species, and Body Region

  • Host Microbe Interactions
  • Published:
Microbial Ecology Aims and scope Submit manuscript

A Correction to this article was published on 28 January 2021

This article has been updated

Abstract

The cutaneous microbiota plays a significant role in the biology of their vertebrate hosts, and its composition is known to be influenced both by host and environment, with captive conditions often altering alpha diversity. Here, we compare the cutaneous bacterial communities of 61 amphibians (both wild and captive) from Hiroshima, Japan, using high-throughput amplicon sequencing of a segment of the 16S rRNA gene. The majority of these samples came from a captive breeding facility at Hiroshima University where specimens from six species are maintained under highly standardized conditions for several generations. This allowed to identify host effects on the bacterial communities under near identical environmental conditions in captivity. We found the structure of the cutaneous bacterial community significantly differing between wild and captive individuals of newts, Cynops pyrrhogaster, with a higher alpha diversity found in the wild individuals. Community structure also showed distinct patterns when comparing different species of amphibians kept under highly similar conditions, revealing an intrinsic host effect. Bacterial communities of dorsal vs. ventral skin surfaces did not significantly differ in most species, but a trend of higher alpha diversity on the ventral surface was found in Oriental fire-bellied toads, Bombina orientalis. This study confirms the cutaneous microbiota of amphibians as a highly dynamic system influenced by a complex interplay of numerous factors.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4

Similar content being viewed by others

Change history

References

  1. Clarke BT (1997) The natural history of amphibian skin secretions, their normal function and potential medical applications. Biol Rev 72:365–379

    PubMed  CAS  Google Scholar 

  2. Rosenthal M, Goldberg D, Aiello A, Larson E, Foxman B (2011) Skin microbiota: microbial community structure and its potential association with health and disease. Infect Genet Evol 11:839–848

    PubMed  PubMed Central  Google Scholar 

  3. Grice EA, Segre JA (2011) The skin microbiome. Nat Rev Microbiol 9:244–253

    PubMed  PubMed Central  CAS  Google Scholar 

  4. Naik S, Bouladoux N, Wilhelm C, Molloy MJ, Salcedo R, Kastenmuller W et al (2012) Compartmentalized control of skin immunity by resident commensals. Science 337:1115–1119

    PubMed  PubMed Central  CAS  Google Scholar 

  5. Belden LK, Harris RN (2007) Infectious diseases in wildlife: the community ecology context. Front Ecol Environ 5:533–539

    Google Scholar 

  6. Reid G, Younes JA, Van Der Mei HC, Gloor GB, Knight R, Busscher HJ (2011) Microbiota restoration: natural and supplemented recovery of human microbial communities. Nat Rev Microbiol 9:27–38

    PubMed  CAS  Google Scholar 

  7. Fierer N, Ferrenberg S, Flores GE, Gonzalez A, Kueneman J, Legg T et al (2012) From animalcules to an ecosystem: application of ecological concepts of the human microbiome. Ann Rev Ecol Evol S 43:137–155

    Google Scholar 

  8. Woodhams DC, Brandt H, Baumgartner S, Kielgast J, Küpfer E, Tobler U et al (2014) Interacting symbionts and immunity in the amphibian skin mucosome predict disease risk and probiotic effectiveness. PLoS One 9, e96375

    PubMed  PubMed Central  Google Scholar 

  9. Daly JW, Myers CW, Whittaker N (1987) Further classification of skin alkaloids from Neotropical poison frogs (Dendrobatidae), with a general survey of toxic/noxious substances in the amphibia. Toxicon 25:1023–1095

    PubMed  CAS  Google Scholar 

  10. Woodhams DC, Ardipradja K, Alford RA, Marantelli G, Reinert LK, Rollins‐Smith LA (2007) Resistance to chytridiomycosis varies among amphibian species and is correlated with skin peptide defenses. Anim Conserv 10:409–417

    Google Scholar 

  11. Rollins-Smith LA (2009) The role of amphibian antimicrobial peptides in protection of amphibians from pathogens linked to global amphibian declines. BBA-Biomembranes 1788:1593–1599

    PubMed  CAS  Google Scholar 

  12. Becker MH, Harris RN (2010) Cutaneous bacteria of the redback salamander prevent morbidity associated with a lethal disease. PLoS One 5, e10957

    PubMed  PubMed Central  Google Scholar 

  13. Myers JM, Ramsey JP, Blackman AL, Nichols AE, Minbiole KP, Harris RN (2012) Synergistic inhibition of the lethal fungal pathogen Batrachochytrium dendrobatidis: the combined effect of symbiotic bacterial metabolites and antimicrobial peptides of the frog Rana muscosa. J Chem Ecol 38:958–965

    PubMed  Google Scholar 

  14. Bletz MC, Loudon AH, Becker MH, Bell SC, Woodhams DC, Minbiole KP, Harris RN (2013) Mitigating amphibian chytridiomycosis with bioaugmentation: characteristics of effective probiotics and strategies for their selection and use. Ecol Lett 16:807–820

    PubMed  Google Scholar 

  15. Walke JB, Becker MH, Loftus SC, House LL, Teotonio TL, Minbiole KP, Belden LK (2015) Community structure and function of amphibian skin microbes: an experiment with bullfrogs exposed to a chytrid fungus. PLoS One 10, e0139848

    PubMed  PubMed Central  Google Scholar 

  16. Harris RN, Lauer A, Simon MA, Banning JL, Alford RA (2008) Addition of antifungal skin bacteria to salamanders ameliorates the effects of chytridiomycosis. Dis Aquat Org 83:11

    Google Scholar 

  17. McKenzie VJ, Bowers RM, Fierer N, Knight R, Lauber CL (2012) Co-habiting amphibian species harbor unique skin bacterial communities in wild populations. ISME J 6:588–596

    PubMed  CAS  Google Scholar 

  18. Antwis RE, Haworth RL, Engelmoer DJ, Ogilvy V, Fidgett AL, Preziosi RF (2014) Ex situ diet influences the bacterial community associated with the skin of red-eyed tree frogs (Agalychnis callidryas). PLoS One 9, e85563

    PubMed  PubMed Central  Google Scholar 

  19. Becker MH, Richards-Zawacki CL, Gratwicke B, Belden LK (2014) The effect of captivity on the cutaneous bacterial community of the critically endangered Panamanian golden frog (Atelopus zeteki). Biol Conserv 176:199–206

    Google Scholar 

  20. Kueneman JG, Parfrey L, Woodhams DC, Archer HM, Knight R, McKenzie VJ (2014) The amphibian skin-associated microbiome across species, space and life history stages. Mol Ecol 23:1238–1250

    PubMed  Google Scholar 

  21. Loudon AH, Woodhams DC, Parfrey LW, Archer H, Knight R, McKenzie V, Harris RN (2014) Microbial community dynamics and effect of environmental microbial reservoirs on red-backed salamanders (Plethodon cinereus). ISME J 8:830–840

    PubMed  CAS  Google Scholar 

  22. Walke JB, Becker MH, Loftus SC, House LL, Cormier G, Jensen RV, Belden LK (2014) Amphibian skin may select for rare environmental microbes. ISME J 8:2207–2217

    PubMed  PubMed Central  CAS  Google Scholar 

  23. Toledo RD, Jared C (1995) Cutaneous granular glands and amphibian venoms. Comp Biochem Phys A 111:1–29

    Google Scholar 

  24. Bataille A, Lee-Cruz L, Tripathi B, Kim H, Waldman B (2016) Microbiome variation across amphibian skin regions: implications for chytridiomycosis mitigation efforts. Microb Ecol 71:221–232

    PubMed  Google Scholar 

  25. Brosius J, Dull TJ, Sleeter DD, Noller HF (1981) Gene organization and primary structure of a ribosomal RNA operon from Escherichia coli. J Mol Biol 148:107–127

    PubMed  CAS  Google Scholar 

  26. Kozich JJ, Westcott SL, Baxter NT, Highlander SK, Schloss PD (2013) Development of a dual-index sequencing strategy and curation pipeline for analyzing amplicon sequence data on the MiSeq Illumina sequencing platform. Appl Environ Microbiol 79:5112–5120

    PubMed  PubMed Central  CAS  Google Scholar 

  27. Caporaso JG, Kuczynski J, Stombaugh J, Bittinger K, Bushman FD, Costello EK et al (2010) QIIME allows analysis of high-throughput community sequencing data. Nat Methods 7:335–336

    PubMed  PubMed Central  CAS  Google Scholar 

  28. Aronesty E (2011) ea–utils: command–line tools for processing biological sequencing data. http://code.google.com/p/ea–utils. Accessed 1 Oct 2015

  29. Aronesty E (2013) TOBioiJ: comparison of sequencing utility programs. Open Bioinforma J 7:8

    Google Scholar 

  30. Edgar RC, Haas BJ, Clemente JC, Quince C, Knight R (2011) UCHIME improves sensitivity and speed of chimera detection. Bioinformatics 27:2194–2200

    PubMed  PubMed Central  CAS  Google Scholar 

  31. Rideout JR, He Y, Navas-Molina JA, Walters WA, Ursell LK, Gibbons SM (2014) Subsampled open-reference clustering creates consistent, comprehensive OTU definitions and scales to billions of sequences. Peer J 2, e545

    PubMed  PubMed Central  Google Scholar 

  32. Edgar RC (2010) Search and clustering orders of magnitude faster than BLAST. Bioinformatics 26:2460–2461

    CAS  PubMed  Google Scholar 

  33. Caporaso JG, Bittinger K, Bushman FD, DeSantis TZ, Andersen GL, Knight R (2010) PyNAST: a flexible tool for aligning sequences to a template alignment. Bioinformatics 26:266–267

    PubMed  CAS  Google Scholar 

  34. 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 73:5261–5267

    PubMed  PubMed Central  CAS  Google Scholar 

  35. Price MN, Dehal PS, Arkin AP (2010) FastTree 2—approximately maximum-likelihood trees for large alignments. PLoS One 5, e9490

    PubMed  PubMed Central  Google Scholar 

  36. Bokulich NA, Subramanian S, Faith JJ, Gevers D, Gordon JI, Knight R, Mills DA, Caporaso JG (2013) Quality–filtering vastly improves diversity estimates from Illumina amplicon sequencing. Nat Methods 10:57–59

    PubMed  CAS  Google Scholar 

  37. R Development Core Team (2011) R: a language and environment for statistical computing. Vienna: the R Foundation for Statistical Computing. ISBN: 3-900051-07-0. Available online at http://www.R-project.org/

  38. Lozupone C, Knight R (2005) UniFrac: a new phylogenetic method for comparing microbial communities. Appl Environ Microbiol 71:8228–8235

    PubMed  PubMed Central  CAS  Google Scholar 

  39. Belden LK, Hughey MC, Rebollar EA, Umile TP, Loftus SC, Burzynski EA, Minbiole KP, House LL, Jensen RV, Becker MH, Walke JB, Medina D, Ibáñez R, Harris RN (2015) Panamanian frog species host unique skin bacterial communities. Front Microbiol 6:1171

    PubMed  PubMed Central  Google Scholar 

  40. Rebollar EA, Hughey MC, Medina D, Harris RN, Ibáñez R, Belden LK (2016) Skin bacterial diversity of Panamanian frogs is associated with host susceptibility and presence of Batrachochytrium dendrobatidis. ISME J. doi:10.1038/ismej.2015.234

    Article  PubMed  PubMed Central  Google Scholar 

  41. Vences M, Dohrmann AB, Künzel S, Granzow S, Baines JF, Tebbe CC (2015) Composition and variation of the skin microbiota in sympatric species of European newts (Salamandridae). Amphibia-Reptilia 36:5–12

    Google Scholar 

  42. Kohl KD, Skopec MM, Dearing MD (2014) Captivity results in disparate loss of gut microbial diversity in closely related hosts. Conserv Physiol 2, cou009

    PubMed  PubMed Central  Google Scholar 

  43. Gibson BW, Tang DZ, Mandrell R, Kelly M, Spindel ER (1991) Bombinin-like peptides with antimicrobial activity from skin secretions of the Asian toad, Bombina orientalis. J Biol Chem 266:23103–23111

    PubMed  CAS  Google Scholar 

  44. Kohl KD, Yahn J (2016) Effects of environmental temperature on the gut microbial communities of tadpoles. Environ Microbiol, in press

  45. Shade A, Jones SE, Caporaso JG, Handelsman J, Knight R, Fierer N, Gilbert JA (2014) Conditionally rare taxa disproportionately contribute to temporal changes in microbial diversity. mBio 5, e01371-14

    PubMed  PubMed Central  Google Scholar 

  46. Sanford JA, Gallo RL (2013) Functions of the skin microbiota in health and disease. Semin Immunol 25:370–377

    PubMed  PubMed Central  CAS  Google Scholar 

Download references

Acknowledgments

We are grateful to Meike Kondermann for their help in the lab and to Christoph Tebbe for helpful advice. We express our appreciation to the Board of Education of Kagoshima prefecture for allowing us to use live crocodile newts and Amami Ishikawa’s frogs protected by law. We thank the strain maintenance team of the Institute for Amphibian Biology for providing captive Japanese fire-bellied newts. This work was supported by a grant of the Deutsche Forschungsgemeinschaft (VE247/9-1) and by a guest researcher fellowship of Hiroshima University to MV.

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Joana Sabino-Pinto.

Electronic Supplementary Material

Below is the link to the electronic supplementary material.

Supplementary Fig. S1

Full legend for the taxa bar plots. Percentages of each taxa represented for each subset of the data. (PDF 52.9 kb)

Supplementary Fig. S2

Results from LEfSe analysis showing taxa that significantly differ in abundance between wild and captive C. pyrrhogaster. Green taxa significantly characterize the wild community and blue taxa the captive community. (PDF 45.2 kb)

Supplementary Fig. S3

Results from LEfSe analysis showing taxa that significantly differ in abundance between the different analyzed species. Purple taxa significantly characterize the community of B. orientalis; pink taxon the community of B. japonicus; red taxon the community of C. pyrrhogaster; green taxon the community of O. splendida; and the blue taxa the community of R. japonica. (PDF 33.7 kb)

Supplementary Table S1

Core bacterial OTUs from the wild and captive individuals. Abundance reflects the abundance of the OTU on the dataset and is in percentage. (XLSX 11 kb)

Supplementary Table S2

Lower triangle reflects the pair-wise comparisons of the composition of the communities between species analyzed with PERMANOVA; P values are represented. Higher triangle reflects the distances between the communities: top value when determined with the Bray-Curtis distance matrix; bottom value when determined with the unweighted UniFrac distance matrix. (XLSX 9 kb)

Supplementary Table S3

Core bacterial OTUs from B. orientalis (Bom), B. japonicus (Buf), C. pyrrhogaster (Cyn), O. splendida (Odo), and R. japonica (Ran). Abundance reflects the abundance of the OTU on the dataset and is in percentage. (XLSX 13 kb)

Supplementary Table S4

Core bacterial OTUs from the dorsal and ventral sides of B. orientalis. Abundance reflects the abundance of the OTU on the dataset and is in percentage. (XLSX 12 kb)

Supplementary Table S5

Core bacterial OTUs from the dorsal and ventral sides of B. japonicus. Abundance reflects the abundance of the OTU on the dataset and is in percentage. (XLSX 12 kb)

Supplementary Table S6

Core bacterial OTUs from the dorsal and ventral sides of C. pyrrhogaster. Abundance reflects the abundance of the OTU on the dataset and is in percentage. (XLSX 11 kb)

Supplementary Table S7

Core bacterial OTUs from the dorsal and ventral sides of E. andersoni. Abundance reflects the abundance of the OTU on the dataset and is in percentage. (XLSX 11 kb)

Supplementary Table S8

Core bacterial OTUs from the dorsal and ventral sides of O. splendida. Abundance reflects the abundance of the OTU on the dataset and is in percentage. (XLSX 12 kb)

Supplementary Table S9

Core bacterial OTUs from the dorsal and ventral sides of R. japonica. Abundance reflects the abundance of the OTU on the dataset and is in percentage. (XLSX 11 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Sabino-Pinto, J., Bletz, M.C., Islam, M.M. et al. Composition of the Cutaneous Bacterial Community in Japanese Amphibians: Effects of Captivity, Host Species, and Body Region. Microb Ecol 72, 460–469 (2016). https://doi.org/10.1007/s00248-016-0797-6

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00248-016-0797-6

Keywords

Navigation