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Chytridiomycosis-induced mortality in a threatened anuran

  • Andrea J. Adams ,

    Roles Conceptualization, Data curation, Formal analysis, Investigation, Methodology, Project administration, Validation, Visualization, Writing – original draft, Writing – review & editing

    Affiliations Yosemite National Park, El Portal, California, United States of America, Earth Research Institute, University of California Santa Barbara, Santa Barbara, California, United States of America

  • Allan Pessier,

    Roles Data curation, Formal analysis, Investigation, Methodology, Validation, Visualization, Writing – review & editing

    Affiliation Department of Veterinary Microbiology and Pathology, College of Veterinary Medicine, Washington State University, Pullman, Washington, United States of America

  • Peggy Cranston,

    Roles Data curation, Investigation, Methodology, Resources, Validation, Writing – review & editing

    Affiliation Mother Lode Field Office, U.S. Bureau of Land Management, Fair Oaks, California, United States of America

  • Robert L. Grasso

    Roles Conceptualization, Data curation, Funding acquisition, Investigation, Project administration, Resources, Supervision, Validation, Writing – review & editing

    Affiliation Yosemite National Park, El Portal, California, United States of America


Effectively planning conservation introductions involves assessing the suitability of both donor and recipient populations, including the landscape of disease risk. Chytridiomycosis, caused by the fungal pathogen Batrachochytrium dendrobatidis (Bd), has caused extensive amphibian declines globally and may hamper reintroduction attempts. To determine Bd dynamics in potential source populations for conservation translocations of the threatened California red-legged frog (Rana draytonii) to Yosemite National Park, we conducted Bd sampling in two populations in the foothills of the Sierra Nevada Mountains, California, U.S.A. At one of two sites, we observed lethally high Bd loads in early post-metamorphic life stages and confirmed one chytridiomycosis-induced mortality, the first such report for this species. These results informed source population site selection for subsequent R. draytonii conservation translocations. Conservation efforts aimed at establishing new populations of R. draytonii in a landscape where Bd is ubiquitous can benefit from an improved understanding of risk through disease monitoring and ex situ infection studies.


Successful conservation translocations hinge on adequate preparation and planning; along with habitat suitability and source population stability, disease susceptibility is a critical consideration [13]. The fungal pathogen Batrachochytrium dendrobatidis (hereafter Bd), the causative agent of the disease chytridiomycosis, is a primary cause of widespread amphibian declines globally [46], and can hamper amphibian reintroduction attempts [7].

The largest anuran native to the western United States (Wright and Wright 1949), the California red-legged frog (R. draytonii) has been extirpated from >70% of its former range, prompting calls for reintroduction feasibility studies [8]. Listed as threatened under the U.S. Endangered Species Act since 1996, R. draytonii was originally threatened by overharvest in the nineteenth and early twentieth centuries [9, 10]. Since the rapid urbanization of California, the species has declined due to habitat loss, pesticides, and introduced predators [1114]. The extent to which disease may have contributed to R. draytonii decline is unknown; however, higher Bd prevalence in the species has been associated with decreased survival [15].

No chytridiomycosis-induced mortality has been recorded for R. draytonii, though the closely-related congeners Rana muscosa, Rana sierrae, and Rana boylii have all experienced chytridiomycosis-induced die-offs in California [14, 16]. California red-legged frogs are generally presumed tolerant of Bd because they persist in areas where Bd is present [17, 18]. Bd prevalence in wild R. draytonii populations in California and Mexico ranges from 37% to 68% [14, 1820], but data regarding R. draytonii Bd susceptibility are sparse. In the only published laboratory study of Bd infection in R. draytonii to date, Bd-positive metamorphs of unknown Bd load with wild-caught Bd infection did not present with morbidity or mortality during 18 months of laboratory observation [21]. In ex situ R. draytonii infection trials, individuals did not experience chytridiomycosis-induced morbidity or mortality [22], indicating that adults may have some measure of innate and adaptive immunity to Bd.

Among species, host responses to Bd infection are highly variable, ranging from lethal susceptibility to tolerance, and, in some cases, complete resistance to infection [2325]. Within-species disease outcome is also variable, ranging from disease-induced localized extirpations to infection tolerance, population persistence, and recovery [26, 27]. Variation in host Bd resistance and tolerance can be influenced by host environment [28]; behavior [19]; genetically mediated immune factors [29, 30] or the lineage of Bd infecting the host [31, 32]. Because reconstruction of the immune system occurs during metamorphosis, immunosuppression can make recently metamorphosed individuals particularly vulnerable to disease [3335].

Donor population selection and the life stage of introduced individuals may be essential factors in determining reintroduction success when conducting conservation translocations in a landscape where Bd is essentially ubiquitous [2]. Here, we report on Bd sampling at two candidate R. draytonii source populations to evaluate their suitability for conservation translocations.

Materials and methods

Ethics statement

Rana draytonii capture and handling were conducted under permits issued by the U.S. Fish and Wildlife Service (TE-86906B-0), California Department of Fish and Wildlife (SC-5130), and Yosemite National Park (YOSE-2015-SCI-129 and YOSE-2016-SCI-101).

Study sites

The Sierra Nevada Mountains have become an epicenter of amphibian decline studies [13, 27, 3644]. Extant Sierra Nevada R. draytonii populations are scarce [45] and rarely accessible for proactive conservation efforts, primarily due to their presence on privately owned lands. We identified two candidate R. draytonii source populations for translocations: Bear Creek Pond (790 m elevation) and Spivey Pond (975 m elevation), in El Dorado County, California, USA, which are artificial creek impoundments fed by headwater springs (Fig 1). When the Spivey Pond R. draytonii population was discovered in 1997, it was the first report for the species in the Sierra Nevada in nearly 25 years, and is currently one of only six known populations of the species in the mountain range [46]. In 1998, the site was conserved as part of a 20-hectare parcel and became public land when it was sold to the U.S. Bureau of Land Management [46]. Prior to this study, Spivey Pond had only been sampled for Bd once: two adults were sampled in 2009, and although quantitative data are not available, a “strong Bd-positive signal” was detected on one of two adult frogs [47].

Fig 1. Map of the study area.

The map was generated in R [67] using the “ggmap” [78] and “ggplot2” [79] packages with map tiles by Stamen Design ( and data by OpenStreetMap, under ODbL, under CC BY 3.0 (

Effective population sizes at Bear Creek are the largest among five Sierra Nevada foothill populations sampled for mtDNA analysis (Ne = 19.67–41.23) [48]. Bear Creek and Spivey Pond R. draytonii share a common mtDNA haplotype, indicating connectivity between the two sites at some time in the past, though there does not appear to be any contemporary gene flow [48]. Both ponds have similar amphibian species assemblages, including R. draytonii, Anaxyrus boreas halophilus (California toad), and Pseudacris regilla (Pacific chorus frog); however, Spivey also has American bullfrogs (Rana catesbeiana; hereafter “bullfrogs”) and Sierra newts (Taricha sierrae), whereas Bear Creek does not.

Study species

In the Sierra Nevada, R. draytonii is limited to small, isolated populations in the northern portion of its range with restricted gene exchange [48]. The species is present in fewer than 30% of historical localities, most of which have very small population sizes [48]. In Yosemite National Park (Yosemite), R. draytonii had been extirpated prior to the translocation project and were not currently known to occur within 160 km as evidenced by visual encounter surveys and environmental DNA. However, the recent eradication of bullfrogs from Yosemite Valley has made the establishment of California red-legged frogs in Yosemite possible for the first time in over 60 years [49, 50].

American bullfrogs (Rana catesbeiana after Yuan, Zhou [51]; hereafter “bullfrogs”) were introduced to California from the eastern USA in the 19th and early 20th centuries. They are an invasive predator and competitor of many native aquatic species in the western USA and globally, and have been implicated in R. draytonii declines [13, 5261]. Bullfrogs are susceptible to chytridiomycosis infection but appear tolerant of most Bd strains, making them suitable vectors and reservoir hosts for the pathogen in the wild [19, 62, 63]. Bullfrogs were first observed at Spivey Pond in 2000, and in 2003, the pond was drained in order to reduce the population of the species, which has an obligate two-year tadpole stage [64]. Measures have also been taken to reduce adult bullfrogs at the site, including egg mass removal and direct lethal taking.

Field surveys and laboratory analyses

We nocturnally surveyed for all lifestages of R. draytonii at both populations, detecting adults via eye shine using 200 lumen LED flashlights, and detecting subadults (<50 mm snout-vent length (SVL)) opportunistically during both daytime and nighttime site visits. We surveyed Bear Creek and Spivey in June and October 2016, and conducted additional surveys at Bear Creek in October 2015 and June 2017. We captured individual adults and subadults with fresh pairs of nitrile gloves and sampled them for Bd following standardized protocols using a rayon-tipped swab (Hyatt et al. 2007). We used quantitative polymerase chain reaction (qPCR) to detect Bd DNA following Boyle et al. (2004). We measured Bd infection intensity (Bd load) in terms of zoospore equivalents (ZE), calculated by multiplying the genomic equivalents by 80 to account for the dilution factor in qPCR sample preparation necessitated by the use of standard DNA extraction methods for swabs collected from live animals [65]. We used a Welch t-test [66] on log-transformed Bd values (ZE) to compare mean Bd loads of adults and subadults on one sample date. Statistical analyses were conducted and all figures were created using R [67].

We collected one subadult (24 mm SVL) from Spivey Pond with lethargy and loss of righting reflex to determine the cause of morbidity and eventual mortality. We fixed the frog whole in ethanol, post-fixed in 10% neutral buffered formalin, and decalcified in hydrochloric acid. After decalcification, the body was serially sectioned and processed routinely for histologic examination in two paraffin blocks [68].

Results and discussion

We sampled 63 R. draytonii individuals (57 adults and 6 subadults) for Bd. Bd prevalence at Bear Creek was 85% (n = 41, 95% CI 71–94) and Spivey Pond was 86% (n = 22, 95% CI 65–97)—among the highest prevalence reported for this species [14, 18, 69]. At Spivey Pond in October 2016—the only date that more than one subadult was captured—subadult Bd loads were significantly higher than those of adults (Welch’s t(5), t = -7.6, p = 0.0006; Fig 2). The moribund subadult collected at Spivey had the highest Bd load (297,700 ZE) of all animals sampled.

Fig 2. Batrachochytrium dendrobatidis (Bd) loads in Bd-positive California red-legged frogs (Rana draytonii) at two translocation donor populations in El Dorado County, California, U.S.A.

Box widths are proportional to sample size, bold horizontal lines within each boxplot indicate the median, boxes show the interquartile (IQ) range, and whiskers show the range within 1.5 times the IQ range.

Histologic findings of the moribund subadult from Spivey were consistent with clinically significant (lethal) chytridiomycosis caused by Bd infection. Examination demonstrated diffuse epidermal hyperplasia and orthokeratotic hyperkeratosis with myriad intracorneal chytrid-type fungal thalli (Fig 3). Most chytrid thalli were empty from previous discharge of zoospores but forms included flask-shaped zoosporangia with prominent discharge tubes and internally septate colonial thalli consistent with the genus Batrachochytrium. The distribution of skin lesions and number of fungal thalli present was consistent with lethal chytridiomycosis in other anuran species [39, 7072]. There was no histologic evidence of another contributory disease process (e.g. ranavirus infection).

Fig 3. Histologic findings from a California red-legged frog (Rana draytonii) with chytridiomycosis.

Marked hyperkeratosis with numerous empty chytrid fungal thalli. Characteristic thallus forms include zoosporangia with prominent discharge tubes (asterisk) and internally septate colonial thalli (arrow). Bar = 30 microns.

In addition to the moribund frog collected for histological examination, two subadults with loads considered lethally high in other ranid species [>10,000 ZE; 73] did not exhibit symptoms of chytridiomycosis in the field and were therefore not collected. Outward symptoms of the disease are not typically observed until very late stage morbidity; therefore, observing lethally infected frogs when they are symptomatic is rare [74].

Twenty-two years of Spivey Pond R. draytonii monitoring indicate that life stages and egg mass counts were variable across years (Fig 4). Increased detection of frogs in 2014, 2015, and 2016 could have resulted from drought, which concentrates frogs into smaller aquatic habitats [19]. Chytridiomycosis mortalities in a California congener (R. boylii) have been attributed to lower flow rates during drought conditions, which may also increase pathogen transmission opportunities from crowding of aquatic habitats [19]. The presence of a bullfrog Bd vector and reservoir host can increase pathogen burden in native California ranids [19]. Bullfrogs have historically been present at Spivey, but not Bear Creek; however, bullfrogs have not been observed at Spivey since 2009 (Fig 4).

Fig 4. Twenty-two-year monitoring data for Spivey Pond collected and curated by the U.S. Bureau of Land Management.

Lifestages are for California red-legged frogs (Rana draytonii); bullfrogs are non-native American bullfrogs (Rana catesbeiana).

Subadults were encountered less frequently than adults, constituting only 17% of Spivey samples and 5% of Bear Creek samples collected (Fig 2). In addition to subadult Bd mortality, Sierra newt (Taricha sierrae) predation may keep subadult R. draytonii densities low at Spivey as compared to Bear Creek. A predator of R. draytonii, T. sierrae and can account for up to 90% of embryo mortality (Calef 1973; Licht 1974). At Spivey, T. sierrae have been observed on egg masses in high abundance (>100), presumably waiting for tadpoles to hatch; T. sierrae is not present at Bear Creek. At Bear Creek, the adult R. draytonii population is larger (>100 frogs), and in a typical year 25–35 egg masses are observed. Higher densities at Bear Creek may increase R. draytonii cannibalism and reduce the subadult population [75].


This is the first report of chytridiomycosis-induced mortality in R. draytonii. We observed adult Bd loads well below those considered lethal in other ranids [19, 73], but observed extremely high loads in subadults (Fig 2), including one moribund individual. Though our sample sizes are too small to definitively conclude that subadults are more likely to be infected compared to adults, our observation is consistent with high Bd loads and mortality in the subadult stage of other Bd-susceptible ranid species, including R. muscosa and R. boylii in California [16, 19], and R. onca in Nevada [35]. The subadult in close proximity to the moribund frog that exhibited Bd loads on the same order of magnitude (>270,000 ZE) but had no outward symptoms highlights the importance of Bd sampling and qPCR detection to determine degree of infection rather than behavioral observations alone.

Though we do not have ample evidence to conclude that chytridiomycosis is a major source of mortality in R. draytonii, our finding of a link between Bd infection and mortality has been a consideration in the ongoing conservation translocation project in Yosemite. More broadly, this report should be considered when reintroductions or other elements called for in the recovery plan for this threatened species—such as mitigation banking—are undertaken. Rana draytonii populations with higher Bd prevalence exhibit lower survivorship [15], and mathematical models largely suggest that the post-metamorphic juvenile life stage can be a disproportionately essential driver of amphibian population dynamics [76]. Future work should use ex situ Bd inoculations of early post-metamorphic R. draytonii to determine how commonly juvenile Bd mortality can occur.

National parks are often considered refugia for species that are unable to persist in the face of threats outside of protected areas [77], and anthropogenic stressors outside of Yosemite—such as invasive predators and competitors—currently limit California red-legged frog reintroduction efforts. Disease is a threat indifferent to geopolitical boundaries, and thus the need for reintroduction feasibility research both inside and outside of protected areas is imperative. In a landscape where pathogens—such as Bd—are ubiquitous, diligent monitoring can improve managers’ understanding of disease risk. We recommend that ex situ Bd exposure studies be conducted with R. draytonii to further examine the susceptibility of this species in the vulnerable early post-metamorphic life stage.


We thank Jeffrey Jones for assistance with data acquisition, and Sarah Kupferberg and Tom Smith for helpful comments on the manuscript. The findings and conclusions in this article are those of the authors and do not necessarily represent the views of the U.S. Government.


  1. 1. IUCN. Guidelines for reintroductions and other conservation translocations. Gland Switz Camb UK IUCNSSC Re-Introd Spec Group. 2013.
  2. 2. Brannelly LA, Hunter DA, Skerratt LF, Scheele BC, Lenger D, McFadden MS, et al. Chytrid infection and post-release fitness in the reintroduction of an endangered alpine tree frog. Anim Conserv. 2016;19(2):153–62.
  3. 3. Seddon PJ, Griffiths CJ, Soorae PS, Armstrong DP. Reversing defaunation: restoring species in a changing world. Science. 2014;345(6195):406–12. pmid:25061203
  4. 4. Skerratt LF, Berger L, Speare R, Cashins S, McDonald KR, Phillott AD, et al. Spread of chytridiomycosis has caused the rapid global decline and extinction of frogs. EcoHealth. 2007;4(2):125–34.
  5. 5. Collins JP, Crump ML, Lovejoy III TE. Extinction in our times: global amphibian decline. Oxford, England, UK: Oxford University Press; 2009. 273 p.
  6. 6. Stuart SN, Chanson JS, Cox NA, Young BE, Rodrigues AS, Fischman DL, et al. Status and trends of amphibian declines and extinctions worldwide. Science. 2004;306(5702):1783–6. Epub 2004/10/16. pmid:15486254.
  7. 7. McFadden M, Hunter D, Harlow P, Pietsch R, Scheele B. Captive management and experimental reintroduction of the booroolong frog on the South Western Slopes region, New South Wales, Australia. In: Soorae PS, editor. Global re-introduction perspectives: Additional case studies from around the globe. Abu Dhabi: IUCN/SSC Re-introduction Specialist Group; 2010. p. 77–80.
  8. 8. U.S. Fish and Wildlife Service. Recovery plan for the California red-legged frog (Rana aurora draytonii). Portland, Oregon: U.S. Fish and Wildlife Service, 2002.
  9. 9. Endangered and threatened wildlife and plants; determination of threatened status for the California red-legged frog, 50 CFR Part 17 (1996).
  10. 10. Jennings MR, Hayes MP. Pre-1900 overharvest of California red-legged frogs (Rana aurora draytonii): The inducement of bullfrog (Rana catesbeiana) introduction. Herpetologica. 1985;41(1):94–103. PubMed PMID: WOS:A1985AFM6200014.
  11. 11. Jennings MR, Hayes MP. Amphibian and reptile species of special concern in California. Rancho Cordova, California: California Department of Fish and Game, 1994.
  12. 12. Davidson C, Shaffer HB, Jennings MR. Spatial tests of the pesticide drift, habitat destruction, UV-B, and climate-change hypotheses for California amphibian declines. Conserv Biol. 2002;16(6):1588–601.
  13. 13. Moyle PB. Effects of introduced bullfrogs, Rana catesbeiana, on the native frogs of the San Joaquin Valley, California. Copeia. 1973;1973(1):18–22.
  14. 14. Adams AJ, Pessier AP, Briggs CJ. Rapid extirpation of a North American frog coincides with an increase in fungal pathogen prevalence: Historical analysis and implications for reintroduction. Ecology and Evolution. 2017;7(23):10216–32. pmid:29238549
  15. 15. Russell RE, Halstead BJ, Mosher BA, Muths E, Adams MJ, Grant EHC, et al. Effect of amphibian chytrid fungus (Batrachochytrium dendrobatidis) on apparent survival of frogs and toads in the western USA. Biol Conserv. 2019;236:296–304.
  16. 16. Rachowicz LJ, Knapp RA, Morgan JAT, Stice MJ, Vredenburg VT, Parker JM, et al. Emerging infectious disease as a proximate cause of amphibian mass mortality. Ecology. 2006;87(7):1671–83. pmid:16922318
  17. 17. Padgett-Flohr GE. Batrachochytrium dendrobatidis in central California amphibians [Dissertation]: Southern Illinois University Carbondale; 2009.
  18. 18. Fellers GM, Cole RA, Reinitz DM, Kleeman PM. Amphibian chytrid fungus (Batrachochytrium dendrobatidis) in coastal and montane California, USA anurans. Herpetological Conservation and Biology. 2011;6(3):383–94.
  19. 19. Adams AJ, Kupferberg SJ, Wilber MQ, Pessier AP, Grefsrud M, Bobzien S, et al. Extreme drought, host density, sex, and bullfrogs influence fungal pathogen infection in a declining lotic amphibian Ecosphere. 2017;8(3):e01740.
  20. 20. Peralta-García A, Adams AJ, Briggs CJ, Galina-Tessaro P, Valdez-Villavicencio JH, Hollingsworth BD, et al. Occurrence of Batrachochytrium dendrobatidis in anurans of the Mediterranean region of Baja California, México. Dis Aquat Org. 2018;127(3):193–200.
  21. 21. Padgett-Flohr GE. Pathogenicity of Batrachochytrium dendrobatidis in two threatened California amphibians: Rana draytonii and Ambystoma californiense. Herpetological Conservation and Biology. 2008;3(2):182–91.
  22. 22. Adams AJ, Bushell J, Grasso RL. To treat or not to treat? Experimental inoculation and translocation of a threatened frog. Submitted.
  23. 23. Fisher MC, Garner TWJ. The relationship between the emergence of Batrachochytrium dendrobatidis, the international trade in amphibians and introduced amphibian species. Fungal Biology Reviews. 2007;21(1):2–9.
  24. 24. Woodhams DC, Ardipradja K, Alford RA, Marantelli G, Reinert LK, Rollins-Smith LA. Resistance to chytridiomycosis varies among amphibian species and is correlated with skin peptide defenses. Anim Conserv. 2007;10(4):409–17.
  25. 25. Ribas L, Li M-S, Doddington BJ, Robert J, Seidel JA, Kroll JS, et al. Expression profiling the temperature-dependent amphibian response to infection by Batrachochytrium dendrobatidis. PLoS One. 2009;4(12):e8408. pmid:20027316
  26. 26. Briggs CJ, Knapp RA, Vredenburg VT. Enzootic and epizootic dynamics of the chytrid fungal pathogen of amphibians. Proc Natl Acad Sci USA. 2010;107(21):9695–700. pmid:20457916; PubMed Central PMCID: PMC2906864.
  27. 27. Knapp RA, Fellers GM, Kleeman PM, Miller DAW, Vredenburg VT, Rosenblum EB, et al. Large-scale recovery of an endangered amphibian despite ongoing exposure to multiple stressors. Proc Natl Acad Sci. 2016;113(42):11889–94. pmid:27698128
  28. 28. Raffel TR, Halstead NT, McMahon TA, Davis AK, Rohr JR. Temperature variability and moisture synergistically interact to exacerbate an epizootic disease. Proceedings of the Royal Society B: Biological Sciences. 2015;282(1801). pmid:25567647
  29. 29. Savage AE, Zamudio KR. Adaptive tolerance to a pathogenic fungus drives major histocompatibility complex evolution in natural amphibian populations. Proceedings of the Royal Society of London B: Biological Sciences. 2016;283(1827). pmid:27009220
  30. 30. Savage AE, Becker CG, Zamudio KR. Linking genetic and environmental factors in amphibian disease risk. Evolutionary Applications. 2015. pmid:26136822
  31. 31. Farrer RA, Weinert LA, Bielby J, Garner TW, Balloux F, Clare F, et al. Multiple emergences of genetically diverse amphibian-infecting chytrids include a globalized hypervirulent recombinant lineage. Proc Natl Acad Sci USA. 2011;108(46):18732–6. Epub 2011/11/09. pmid:22065772; PubMed Central PMCID: PMC3219125.
  32. 32. Byrne AQ, Vredenburg VT, Martel A, Pasmans F, Bell RC, Blackburn DC, et al. Cryptic diversity of a widespread global pathogen reveals expanded threats to amphibian conservation. Proc Natl Acad Sci. 2019;116(41):20382–7. pmid:31548391
  33. 33. Abu Bakar A, Bower DS, Stockwell MP, Clulow S, Clulow J, Mahony MJ. Susceptibility to disease varies with ontogeny and immunocompetence in a threatened amphibian. Oecologia. 2016;181(4):997–1009. pmid:27021312
  34. 34. Rollins-Smith LA, Ramsey JP, Pask JD, Reinert LK, Woodhams DC. Amphibian Immune Defenses against Chytridiomycosis: Impacts of Changing Environments. Integr Comp Biol. 2011;51(4):552–62. pmid:21816807
  35. 35. Waddle AW, Levy JE, Rivera R, van Breukelen F, Nash M, Jaeger JR. Population-level resistance to chytridiomycosis is life-stage dependent in an imperiled anuran. EcoHealth. 2019;16(4):701–11. pmid:31654279
  36. 36. Bradford DF. Mass mortality and extinction in a high-elevation population of Rana muscosa. J Herpetol. 1991;25(2):174–7.
  37. 37. Drost CA, Fellers GM. Collapse of a regional frog fauna in the Yosemite area of the California Sierra Nevada, USA. Conserv Biol. 1996;10(2):414–25.
  38. 38. Drost CA, Fellers GM. Decline of frog species in the Yosemite section of the Sierra Nevada. University of California-Davis Cooperative National Park Studies Unit, 1994.
  39. 39. Green DE, Kagarise Sherman C. Diagnostic histological findings in Yosemite toads (Bufo canorus) from a die-off in the 1970s. J Herpetol. 2001;35(1):92–103.
  40. 40. Briggs CJ, Vredenburg VT, Knapp RA, Rachowicz LJ. Investigating the population-level effects of chytridiomycosis: an emerging infectious disease of amphibians. Ecology. 2005;86(12):3149–59.
  41. 41. Jani AJ, Knapp RA, Briggs CJ. Epidemic and endemic pathogen dynamics correspond to distinct host population microbiomes at a landscape scale. Proceedings of the Royal Society B: Biological Sciences. 2017;284(1857):20170944. pmid:28637861
  42. 42. Joseph MB, Knapp RA. Disease and climate effects on individuals drive post-reintroduction population dynamics of an endangered amphibian. Ecosphere. 2018;9(11):e02499.
  43. 43. Knapp RA, Briggs CJ, Smith TC, Maurer JR. Nowhere to hide: impact of a temperature-sensitive amphibian pathogen along an elevation gradient in the temperate zone. Ecosphere. 2011;2(8):1–26.
  44. 44. Knapp RA, Matthews KR. Non-native fish introductions and the decline of the mountain yellow-legged frog from within protected areas. Conserv Biol. 2000;14(2):428–38.
  45. 45. Barry SJ, Fellers GM. History and status of the California red-legged frog (Rana draytonii) in the Sierra Nevada, California, USA. Herpetological Conservation and Biology. 2013;8(2):456–502.
  46. 46. American River Conservancy. Final Report on the Habitat Acquisition and Restoration of Spivey Pond, North Fork Weber Creek, El Dorado County. Columa, California: The American River Conservancy, 1999.
  47. 47. Tatarian P, Tatarian G. Chytrid infection of Rana draytonii in the Sierra Nevada, California, USA. Herpetological Review. 2010;41(3):325.
  48. 48. Richmond JQ, Backlin AR, Tatarian PJ, Solvesky BG, Fisher RN. Population declines lead to replicate patterns of internal range structure at the tips of the distribution of the California red-legged frog (Rana draytonii). Biol Conserv. 2014;172:128–37.
  49. 49. Karlstrom EL. The toad genus Bufo in the Sierra Nevada of California: ecological and systematic relationships: Berkeley: University of California Press; 1962.
  50. 50. Kamoroff C, Daniele N, Grasso RL, Rising R, Espinoza T, Goldberg CS. Effective removal of the American bullfrog (Lithobates catesbeianus) on a landscape level: long term monitoring and removal efforts in Yosemite Valley, Yosemite National Park. Biol Invasions. 2020;22(2):617–26.
  51. 51. Yuan Z-Y, Zhou W-W, Chen X, Poyarkov NA Jr., Chen H-M, Jang-Liaw N-H, et al. Spatiotemporal Diversification of the True Frogs (Genus Rana): A Historical Framework for a Widely Studied Group of Model Organisms. Syst Biol. 2016;65(5):824–42. pmid:27288482
  52. 52. Kupferberg SJ. Bullfrog (Rana catesbeiana) invasion of a California river: the role of larval competition. Ecology. 1997;78(6):1736–51.
  53. 53. Adams MJ, Pearl CA. Problems and opportunities managing invasive Bullfrogs: is there any hope? In: Gherardi F, editor. Biological invaders in inland waters: Profiles, distribution, and threats. Dordrecht: Springer Netherlands; 2007. p. 679–93.
  54. 54. Becerra López JL, Esparza Estrada CE, Romero Méndez U, Sigala Rodríguez JJ, Mayer Goyenechea IG, Castillo Cerón JM. Evidence of niche shift and invasion potential of Lithobates catesbeianus in the habitat of Mexican endemic frogs. PLOS ONE. 2017;12(9):e0185086. pmid:28953907
  55. 55. Flynn LM, Kreofsky TM, Sepulveda AJ. Introduced American Bullfrog Distribution and Diets in Grand Teton National Park. Northwest Sci. 2017;91(3):244–56, 13.
  56. 56. Fuller TE, Pope KL, Ashton DT, Welsh HH. Linking the distribution of an invasive amphibian (Rana catesbeiana) to habitat conditions in a managed river system in northern California. Restor Ecol. 2011;19(201):204–13.
  57. 57. Johnson PTJ, McKenzie VJ, Peterson AC, Kerby JL, Brown J, Blaustein AR, et al. Regional decline of an iconic amphibian associated with elevation, land-use change, and invasive species. Conserv Biol. 2011;25(3):556–66. pmid:21342266 PubMed PMID: WOS:000290491700018.
  58. 58. Kats LB, Farrer RP. Alien predators and amphibian declines: review of two decades of science and the transition to conservation. Divers Distrib. 2003;9(2):99–110. PubMed PMID: WOS:000181229500002.
  59. 59. Lawler SP, Dritz D, Strange T, Holyoak M. Effects of introduced mosquitofish and bullfrogs on the threatened California red-legged frog. Conserv Biol. 1999;13(3):613–22.
  60. 60. Schloegel LM, Ferreira CM, James TY, Hipolito M, Longcore JE, Hyatt AD, et al. The North American bullfrog as a reservoir for the spread of Batrachochytrium dendrobatidis in Brazil. Anim Conserv. 2010;13:53–61.
  61. 61. Sepulveda AJ, Layhee M, Stagliano D, Chaffin J, Begley A, Maxell B. Invasion of American bullfrogs along the Yellowstone River. Aquatic Invasions. 2015;10(1).
  62. 62. Garner TW, Perkins MW, Govindarajulu P, Seglie D, Walker S, Cunningham AA, et al. The emerging amphibian pathogen Batrachochytrium dendrobatidis globally infects introduced populations of the North American bullfrog, Rana catesbeiana. Biol Lett. 2006;2(3):455–9. Epub 2006/12/07. pmid:17148429; PubMed Central PMCID: PMC1686185.
  63. 63. Miaud C, Dejean T, Savard K, Millery-Vigues A, Valentini A, Curt Grand Gaudin N, et al. Invasive North American bullfrogs transmit lethal fungus Batrachochytrium dendrobatidis infections to native amphibian host species. Biol Invasions. 2016;18(8):2299–308.
  64. 64. Storer TI. A synopsis of the amphibia of California. University of California Publications in Zoology. 1925;27:1–342.
  65. 65. Adams AJ, LaBonte JP, Ball ML, Richards-Hrdlicka KL, Toothman MH, Briggs CJ. DNA extraction method affects the detection of a fungal pathogen in formalin-fixed specimens using qPCR. PLoS One. 2015;10(8):e0135389. pmid:26291624
  66. 66. Ruxton GD. The unequal variance t-test is an underused alternative to Student's t-test and the Mann–Whitney U test. Behav Ecol. 2006;17(4):688–90.
  67. 67. R Core Team. R: A language and environment for statistical computing. 2019. Available from:
  68. 68. Berger L, Speare R, Kent A. Diagnosis of chytridiomycosis in amphibians by histologic examination. Zoos Print J. 1999;15:184–90.
  69. 69. Padgett-Flohr GE, Hopkins RL 2nd. Batrachochytrium dendrobatidis, a novel pathogen approaching endemism in central California. Dis Aquat Org. 2009;83(1):1–9. pmid:19301630.
  70. 70. Berger L, Speare R, Daszak P, Green DE, Cunningham AA, Goggin CL, et al. Chytridiomycosis causes amphibian mortality associated with population declines in the rain forests of Australia and Central America. Proc Natl Acad Sci. 1998;95(15):9031–6. pmid:9671799
  71. 71. Pessier AP, Nichols DK, Longcore JE, Fuller MS. Cutaneous chytridiomycosis in poison dart frogs (Dendrobates spp.) and White's tree frogs (Litoria caerulea). J Vet Diagn Invest. 1999;11(2):194–9. pmid:10098698
  72. 72. Muths E, Corn PS, Pessier AP, Green DE. Evidence for disease-related amphibian decline in Colorado. Biol Conserv. 2003;110(3):357–65. PubMed PMID: WOS:000181189400005.
  73. 73. Kinney VC, Heemeyer JL, Pessier AP, Lannoo MJ. Seasonal pattern of Batrachochytrium dendrobatidis infection and mortality in Lithobates areolatus: Affirmation of Vredenburg's “10,000 zoospore rule”. PLoS One. 2011;6(3):e16708. pmid:21423745
  74. 74. Piovia-Scott J, Pope K, Joy Worth S, Rosenblum EB, Poorten T, Refsnider J, et al. Correlates of virulence in a frog-killing fungal pathogen: evidence from a California amphibian decline. The ISME Journal. 2015;9(7):1570–8. pmid:25514536
  75. 75. Alvarez JA. Rana draytonii (California red-legged frog). Cannibalism. Herpetological Review. 2013;44(1):126–7.
  76. 76. Petrovan SO, Schmidt BR. Neglected juveniles; a call for integrating all amphibian life stages in assessments of mitigation success (and how to do it). Biol Conserv. 2019;236:252–60.
  77. 77. Lawrence DJ, Larson ER, Liermann CAR, Mims MC, Pool TK, Olden JD. National parks as protected areas for U.S. freshwater fish diversity. Conservation Letters. 2011;4(5):364–71.
  78. 78. Kahle D, Wickham H. ggmap: A package for spatial visualization with Google Maps and OpenStreetMap. R package version 2.3. R Foundation for Statistical Computing, Vienna. 2013.
  79. 79. Wickham H. ggplot2: elegant graphics for data analysis: Springer; 2016.