, Volume 39, Issue 4, pp 803–814 | Cite as

Habitat Characteristics in Created Vernal Pools Impact Spotted Salamander Water-Borne Corticosterone Levels

  • Alice R. MillikinEmail author
  • Sarah K. Woodley
  • Drew R. Davis
  • James T. Anderson
General Wetland Science


Spotted salamanders (Ambystoma maculatum) require vernal pools for breeding habitat. Limited protection and preservation of vernal pools makes suitable habitat creation important. Differences in corticosterone levels, a hormone associated with growth, development, and stress in amphibians, could indicate population health and habitat quality. Our objective was to determine if habitat characteristics in created vernal pools influence corticosterone levels of spotted salamander larvae. In May and June of 2015 and 2016, we sampled water-borne corticosterone levels of larval spotted salamanders in 34 created vernal pools constructed 1–5 years earlier. Using multiple regression, we determined the best model predicting corticosterone levels included larval total length, pool-water temperature, year sampled, and pool diameter. Pool-water pH, depth, and age; percent cover; and predator presence were not significant predictors. Annual variation in corticosterone levels and habitat characteristics, and positive associations with water temperature and salamander body size highlighted the importance of controlling for external influences. The negative association between pool diameter and corticosterone indicated that larvae in larger pools (up to 12.75-m maximum diameter) were less stressed and potentially healthier. These results indicate that pool diameter contributes to habitat quality and may be important when constructing vernal pools for spotted salamanders.


Ambystoma maculatum Caudata Habitat creation Hormones Stress 



This study was completed with approval from West Virginia University Institutional Animal Care and Use Committee (15-0409.3), the U.S. Forest Service, and the West Virginia Division of Natural Resources (Scientific Collecting Permit 2015.133, 2016.205). We thank J Rouda, J Strickland, M Mabry, A Magyan, A Bucher, and J Millikin for field and lab assistance, and D Brown for statistical advice. This research was funded by the U.S. Forest Service, Natural Resources Conservation Service, National Science Foundation (01A-1458952), West Virginia University Natural History Museum, National Institute of Food and Agriculture McStennis Project WVA00117, The Explorers Club Washington Group, Society of Wetland Scientists, Society of Wetland Scientists South Atlantic Chapter, West Virginia University Stitzel Graduate Enhancement Fund, and R and L Bowman. We also thank West Virginia Division of Natural Resources, Department of Biological Sciences at Duquesne University, and the Ruby Distinguished Doctoral Fellowship Program. This is scientific article number 3355 of the West Virginia Agricultural and Forestry Experiment Station, Morgantown.

Supplementary material

13157_2019_1130_MOESM1_ESM.docx (20 kb)
ESM 1 (DOCX 20.1 kb)
13157_2019_1130_MOESM2_ESM.docx (16 kb)
Fig. S1 Monthly average temperatures separated by sample year based on the 12 months before and including sampling: July 2014 – June 2015 and July 2015 – June 2016. Weather data are from the nearest weather station (Elkins, WV), which is 40.23 km away. Data from The Weather Underground [United States] Elkins-Randolph County Station, WV. (DOCX 15.5 kb)
13157_2019_1130_MOESM3_ESM.docx (16 kb)
Fig. S2 Monthly total precipitation separated by sample year based on the 12 months before and including sampling: July 2014 – June 2015 and July 2015 – June 2016. Weather data are from the nearest weather station (Elkins, WV), which is 40.23 km away. Data from The Weather Underground [United States] Elkins-Randolph County Station, WV. (DOCX 15.5 kb)


  1. Balcombe CK, Anderson JT, Fortney RH, Kordek WS (2005) Vegetation, invertebrate, and wildlife community rankings and habitat analysis of mitigation wetlands in West Virginia. Wetlands Ecology and Management 13:517–530CrossRefGoogle Scholar
  2. Barbour MG, Burk JH, Pitts WD, Gilliam FS, Schwartz MW (1999) Terrestrial plant ecology, Third Edition. Benjamin and Cummings, CaliforniaGoogle Scholar
  3. Baugh AT, Bastien B, Still M, Stowell N (2018) Validation of water-borne steroid hormones in a tropical frog (Physalaemus pustulosus). General and Comparative Endocrinology 261:67–80CrossRefGoogle Scholar
  4. Belden LK, Kiesecker JM (2005) Glucocorticosteroid hormone treatment of larval treefrogs increases infection by Alaria sp. trematode cercariae. The Journal of Parasitology 19:686–688CrossRefGoogle Scholar
  5. Bianchini K, Tattersall GJ, Sashaw J, Porteus CS, Wright PA (2012) Acid water interferes with salamander-green algae symbiosis during early embryonic development. Physiological and Biochemical Zoology 85:470–480CrossRefGoogle Scholar
  6. Bonier F, Martin PR, Moore IT, Wingfield JC (2009) Do baseline glucocorticoids predict fitness? Trends in Ecology & Evolution 24:634–642CrossRefGoogle Scholar
  7. Brodman R (1993) The effect of acidity on interactions of Ambystoma salamander larvae. Journal of Freshwater Ecology 8:209–214CrossRefGoogle Scholar
  8. Burnham KP, Anderson DR (2002) Model selection and multimodel inference: a practical information-theoretic approach. Springer Science & Business Media, LLC, New YorkGoogle Scholar
  9. Calhoun AJK, Arrigoni J, Brooks RP, Hunter ML, Richter SC (2014) Creating successful vernal pools: a literature review and advice for practitioners. Wetlands 34:1027–1038CrossRefGoogle Scholar
  10. Carr JA, Norris DO (1988) Interrenal activity during metamorphosis of the tiger salamander, Ambystoma tigrinum. General and Comparative Endocrinology 71:63–69CrossRefGoogle Scholar
  11. Chambers DL, Wojdak JM, Du P, Belden LK (2011) Corticosterone level changes throughout larval development in the amphibians Rana sylvatica and Ambystoma jeffersonianum reared under laboratory, mesocosm, or free-living conditions. Copeia 2011:530–538CrossRefGoogle Scholar
  12. Chambers DL, Wojdak JM, Du P, Belden LK (2013) Pond acidification may explain differences in corticosterone among salamander populations. Physiological and Biochemical Zoology 86:224–232CrossRefGoogle Scholar
  13. Charbonnier JF, Pearlmutter J, Vonesh JR, Gabor CR, Forsburg ZR, Grayson KL (2018) Cross-life stage effects of aquatic larval density and terrestrial moisture on growth and corticosterone in the spotted salamander. Diversity 10:68CrossRefGoogle Scholar
  14. Clark KL (1986) Responses of Ambystoma maculatum populations in Central Ontario to habitat acidity. The Canadian Field-Naturalist 100:463–469Google Scholar
  15. Cree A, Tyrrell CL, Preest MR, Thorburn D, Guillette LJ Jr (2003) Protecting embryos from stress: corticosterone effects and the corticosterone response to capture and confinement during pregnancy in a live-bearing lizard (Hoplodactylus maculatus). General and Comparative Endocrinology 134:316–329CrossRefGoogle Scholar
  16. Crespi EJ, Denver RJ (2005) Roles of stress hormones in food intake regulation in anuran amphibians throughout the life cycle. Comparative Biochemistry and Physiology, Part A 141:381–390CrossRefGoogle Scholar
  17. Dantzer B, Fletcher QE, Boonstra R, Sheriff MJ (2014) Measures of physiological stress: a transparent or opaque window into the status, management and conservation of species? Conservation Physiology 2:cou023CrossRefGoogle Scholar
  18. Davis DR, Gabor CR (2015) Behavioral and physiological antipredator responses of the San Marcos salamander, Eurycea nana. Physiology & Behavior 139:145–149CrossRefGoogle Scholar
  19. Denton RD, Richter SC (2013) Amphibian communities in natural and constructed ridge top wetlands with implications for wetland construction. The Journal of Wildlife Management 77:886–896CrossRefGoogle Scholar
  20. DiBello FJ, Calhoun AJK, Morgan DE, Shearin AF (2016) Efficiency and detection accuracy using print and digital stereo aerial photography for remotely mapping vernal pools in New England landscapes. Wetlands 36:505–514CrossRefGoogle Scholar
  21. Dickens MJ, Romero LM (2013) A consensus endocrine profile for chronically stressed wild animals does not exist. General and Comparative Endocrinology 191:177–189CrossRefGoogle Scholar
  22. Dupont W, Bourgeois P, Reinberg A, Vaillant R (1979) Circannual and circadian rhythms in the concentrations of corticosterone on the plasma of the edible frog (Rana esculenta L.). Journal of Endocrinology 80:117–125CrossRefGoogle Scholar
  23. Egan RS, Paton PWC (2004) Within-pond parameters affecting oviposition by wood frogs and spotted salamanders. Wetlands 24:1–13CrossRefGoogle Scholar
  24. Felix ZI, Wang Y, Schweitzer CJ (2010) Effects of experimental canopy manipulation on amphibian egg deposition. Journal of Wildlife Management 74:496–503CrossRefGoogle Scholar
  25. Fonner CW, Woodley SK (2015) Testing the predation stress hypothesis: behavioural and hormonal responses to predator cues in Allegheny Mountain dusky salamanders. Behaviour 152:797–819CrossRefGoogle Scholar
  26. Formanowicz DR, Bobka MS (1989) Predation risk and microhabitat preference: an experimental study of the behavioral responses of prey and predator. Am Midl Nat 121:379–386CrossRefGoogle Scholar
  27. Fox J, Weisberg S (2011) An {R} companion to applied regression, second edition. Sage, Thousand Oaks URL: Google Scholar
  28. Gabor CR, Bosch J, Fries JN, Davis DR (2013) A non-invasive water-borne hormone assay for amphibians. Amphibia-Reptilia 34:151–162CrossRefGoogle Scholar
  29. Gabor CR, Fisher MC, Bosch J (2015) Elevated corticosterone levels and changes in amphibian behavior are associated with Batrachochytrium dendrobatidis (Bd) infection and Bd lineage. PLoS One 10:e0122685CrossRefGoogle Scholar
  30. Gabor CR, Zabierek KC, Kim DS, da Barbiano LA, Mondelli MJ, Bendik NF, Davis DR (2016) A non-invasive water-borne assay of stress hormones in aquatic salamanders. Copeia 104:172–181CrossRefGoogle Scholar
  31. Glennemeier KA, Denver RJ (2002) Role for corticoids in mediating the response of Rana pipiens tadpoles in intraspecific competition. Journal of Experimental Zoology 292:32–40CrossRefGoogle Scholar
  32. Gormally BMG, Fuller R, McVey M, Romero LM (2018) DNA damage as an indicator of chronic stress: correlations with corticosterone and uric acid. Comparative Biochemistry and Physiology, Part A 227:116–122CrossRefGoogle Scholar
  33. Gosner KK, Black IH (1957) The effects of acidity on the development of New Jersey frogs. Ecology 38:256–262CrossRefGoogle Scholar
  34. Groff LA, Loftin CS, Calhoun AJK (2017) Predictors of breeding site occupancy by amphibians in montane landscapes. The Journal of Wildlife Management 81:269–278CrossRefGoogle Scholar
  35. Harris BN, Carr JA (2016) The role of the hypothalamus-pituitary-adrenal/interrenal axis in mediating predator-avoidance trade-offs. General and Comparative Endocrinology 230:110–142CrossRefGoogle Scholar
  36. Holmes AM, Emmans CJ, Jones N, Coleman R, Smith TE, Hosie CA (2016) Impact of tank background on the welfare of the African clawed frog, Xenopus laevis (Daudin). Applied Animal Behaviour Science 185:131–136CrossRefGoogle Scholar
  37. Homan RN, Regosin JV, Rodrigues DM, Reed JM, Windmiller BS, Romero LM (2003) Impacts of varying habitat quality on the physiological stress of spotted salamanders (Ambystoma maculatum). Animal Conservation 6:11–18CrossRefGoogle Scholar
  38. Homyack JA (2010) Evaluating habitat quality of vertebrates using conservation physiology tools. Wildlife Research 37:332–342CrossRefGoogle Scholar
  39. Hopkins WA, Mendonca MT, Congdon JD (1999) Responsiveness of the hypothalamo-pituitary-interrenal axis in an amphibian (Bufo terrestris) exposed to coal combustion wastes. Comparative Biochemistry and Physiology, Part C 122:191–196CrossRefGoogle Scholar
  40. Hossie TJ, MacFarlane S, Clement A, Murray DL (2017) Threat of predation alters aggressive interactions among spotted salamander (Ambystoma maculatum) larvae. Ecology and Evolution 8:3131–3138CrossRefGoogle Scholar
  41. Janin A, Lena JP, Deblois S, Joly P (2012) Use of stress-hormone levels and habitat selection to assess functional connectivity of a landscape for an amphibian. Conservation Biology 26:923–931CrossRefGoogle Scholar
  42. Kern MM, Nassar AA, Guzy JC, Dorcas ME (2013) Oviposition site selection by spotted salamanders (Ambystoma maculatum) in an isolated wetland. Journal of Herpetology 47:445–449CrossRefGoogle Scholar
  43. Kindt R, Coe R (2005) Tree diversity analysis. A manual and software for common statistical methods for ecological and biodiversity studies. World agroforestry Centre (ICRAF), Nairobi. In: ISBN 92–9059-179-XGoogle Scholar
  44. Mazerolle MJ (2017) AICcmodavg: model selection and multimodel inference based on (Q)AIC(c). R package version 2.1–1.
  45. McCormick SD, Romero M (2017) Conservation endocrinology. Bioscience 67:429–442CrossRefGoogle Scholar
  46. McEwin BS, Wingfield JC (2003) The concept of allostasis in biology and biomedicine. Hormones and Behavior 43:2–15CrossRefGoogle Scholar
  47. Moore IT, Jessop TS (2003) Stress, reproduction, and adrenocortical modulation in amphibians and reptiles. Hormones and Behavior 43:39–47CrossRefGoogle Scholar
  48. Novarro AJ, Gabor CR, Goff CB, Mezebish TD, Thompson LM, Grayson KL (2018) Physiological responses to elevated temperature across the geographic range of a terrestrial salamander. Journal of Experimental Biology 221:jeb178236CrossRefGoogle Scholar
  49. Petranka JW, Kennedy CA, Murray SS (2003a) Response of amphibians to restoration of a southern Appalachian wetland: a long-term analysis of community dynamics. Wetlands 23:1030–1042CrossRefGoogle Scholar
  50. Petranka JW, Kennedy CA, Murray SS (2003b) Response of amphibians to restoration of a southern Appalachian wetland: perturbations confound post-restoration assessment. Wetlands 23:278–290CrossRefGoogle Scholar
  51. Pough FH, Wilson RE (1977) Acid precipitation and reproductive success of Ambystoma salamanders. Water, Air, and Soil Pollution 7:307–316CrossRefGoogle Scholar
  52. R Core Team (2017) R: a language and environment for statistical computing. R Foundation for Statistical Computing, Vienna, Austria. URL
  53. Romero LM (2004) Physiological stress in ecology: lessons from biomedical research. Trends in Ecology and Evolution 19:249–255CrossRefGoogle Scholar
  54. Romero LM, Reed JM (2005) Collecting baseline corticosterone samples in the field: is under 3 min good enough? Comparative Biochemistry and Physiology, Part A 140:73–79CrossRefGoogle Scholar
  55. Romero L, Wikelski M (2001) Corticosterone levels predict survival probabilities of Galápagos marine iguanas during El Nino events. Proceedings of the National Academy of Sciences of the United States of America 98:7366–7370CrossRefGoogle Scholar
  56. Rowe CL, Dunson WA (1993) Relationships among biotic parameters and breeding effort by three amphibians in temporary wetlands of Central Pennsylvania. Wetlands 13:237–246CrossRefGoogle Scholar
  57. Sandeno CM (2011) Project status report – Barton Bench ecological restoration Greenbrier ranger district Monongahela National Forest. WV Department of Environmental Protection, Division of Mining and ReclamationGoogle Scholar
  58. Scheffers BR, Furman BL, Evans JP (2013) Salamanders continue to breed in ephemeral ponds following the removal of surrounding terrestrial habitat. Herpetological Conservation and Biology 8:715–723Google Scholar
  59. Scott AP, Ellis T (2007) Measurement of fish steroids in water–a review. General and Comparative Endocrinology 153:392–400CrossRefGoogle Scholar
  60. Skidds DE, Golet FC, Paton PW, Mitchell JC (2007) Habitat correlates of reproductive effort in wood frogs and spotted salamanders in an urbanizing watershed. Journal of Herpetology 41:439–450CrossRefGoogle Scholar
  61. The Weather Underground Elkins-Randolph County Station. Weather history for Elkins-Randolph county, WV. Weather Underground, The Weather Company. Accessed 15 Jan 2019
  62. Thomas JR, Magyan AJ, Freeman PE, Woodley SK (2017) Testing hypotheses about individual variation in plasma corticosterone in free-living salamanders. Journal of Experimental Biology 220:1210–1221CrossRefGoogle Scholar
  63. Troïanowski M, Mondy N, Dumet A, Arcanjo C, Lengagne T (2017) Effects of traffic noise on tree frog stress levels, immunity, and color signaling. Conservation Biology 31:1132–1140CrossRefGoogle Scholar
  64. United States Forest Service (2014) Mower Tract ecological restoration final reportGoogle Scholar
  65. Vasconcelos D, Calhoun AJK (2006) Monitoring created seasonal pools for functional success: a six-year case study of amphibian responses, Sears Island, Maine, USA. Wetlands 26:992–1003CrossRefGoogle Scholar
  66. Wack CL, DuRant SE, Hopkins WA, Lovern MB, Feldhoff RC, Woodley SK (2012) Elevated plasma corticosterone increases metabolic rate in a terrestrial salamander. Comparative Biochemistry and Physiology, Part A 161:153–158CrossRefGoogle Scholar
  67. Wickham H (2009) ggplot2: elegant graphics for data analysis. Springer-Verlag, New YorkCrossRefGoogle Scholar
  68. Wingfield JC, Maney DL, Breuner CW, Jacobs JD, Lynn S, Ramenofsky M, Richardson RD (1998) Ecological bases of hormone-behavior interactions: the “emergency life history stage.”. American Zoologist 38:191–206CrossRefGoogle Scholar
  69. Woodley SK, Freeman P, Ricciardella LF (2014) Environmental acidification is not associated with altered corticosterone levels in the stream-side salamander, Desmognathus ochrophaeus. General and Comparative Endocrinology 201:8–15CrossRefGoogle Scholar

Copyright information

© Society of Wetland Scientists 2019

Authors and Affiliations

  1. 1.School of Natural ResourcesWest Virginia UniversityMorgantownUSA
  2. 2.Biological SciencesDuquesne UniversityPittsburghUSA
  3. 3.School of Earth, Environmental, and Marine SciencesUniversity of Texas Rio Grande ValleySouth Padre IslandUSA

Personalised recommendations