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    ASIH Stoye Award, Herpetology III (Physiology); Amphibian Ecology and Genetics

    2021-07-22   13:45 - 15:30

    Moderator: Jaime Smith

    1.  13:45  VIRTUAL    When Island Lizards Colonize the Mainland: Thermal Matching, Potential, or Plasticity? Natalie Claunch*, University of Florida; Colin Goodman, University of Florida; Robert Guralnick, University of Florida; Robert Reed, U.S. Geological Survey; Christina Romagosa, University of Florida; Emily Taylor, California Polytechnic State University

    Understanding a species’ physiological tolerance is critical to predicting environments at risk of invasion by nonnative species. When physiological data are limited, climate-matching ecological niche models (ENMs) are used to predict areas at risk of invasion. These models may mis-calculate the spread of island-sourced invaders after initial establishment, because the restricted geography of islands may not reflect evolutionary constraints on thermal limits. Additionally, rapid physiological adaptation or plasticity may occur in species introduced to new environments. We investigated the possibility of thermal plasticity in the Northern curly-tailed lizard, which has established across Florida. We evaluated the Thermal Matching hypothesis (distribution reflects thermal conditions of the native range) by comparing the results of an ENM incorporating only native populations (native model) and one including both native and invasive populations (full model). We found confirmed established populations outside predicted suitable areas of the native-only ENM, and rejected the Thermal Matching hypothesis. We tested the Thermal Potential (lizards tolerate environments with thermal extremes not observed in the native range) and Thermal Plasticity (thermal tolerance reflects local thermal extremes) hypotheses by assessing critical thermal limits (CTmin, CTmax) of individuals from two established populations: Key Largo, matching native range latitudes, and Cocoa Beach, 160 km north of the native range. Although CTmax did not differ, lizards from the northern population had 1°C lower CTmin compared with the population within the latitude of the native range, meaning that lizards are either acclimatized or adapted to lower temperatures in Cocoa Beach, supporting the Thermal Plasticity hypothesis.

    2.  14:00  IN-PERSON    Leukocyte Profiles of Snakes Infected with Ophidiomycesophiodiicola. Cody Godwin*, Tennessee Technological University; Donald Walker, Middle Tennessee State University; Andrew Moore, Tennessee Technological University; Helen Plylar, Utah State University; Christopher Murray, Southeastern Louisiana University

    Snake fungal disease (SFD) has been increasing in prevalence across the eastern United States causing cutaneous lesions and swelling, infection of the respiratory passages, and development of heterophilic granulomas in and on snakes. Ophidiomyces ophiodiicolahas been identified as the pathogen responsible for the SFD. The aim of this study was to investigate the cellular immunological response of snakes to O. ophiodiicola. Wild snakes throughout Tennessee were swabbed for O. ophiodiicola, DNA was extracted and qPCR amplified to determine fungal load. Blood was extracted from sampled snakes and used for blood smears to assess leukocyte profiles via staining and manual cell count. Differences in leukocyte profiles between infected and non-infected snakes were analytically investigated using a Mann-Whitney U test incorporating all species combined and Nerodia sipedon independently. Linear models were used to determine if leukocyte profiles correlated with fungal load for all infected snakes and N. sipedon samples. Leukocyte profiles were not significantly different between infected and non-infected snakes, nor as fungal load varied for all snake species and N. sipedon alone. Azurophil, lymphocyte, and basophil ratios were not significantly different in infectedN. sipedonwhen O. ophiodiicola was quantified but heterophil and monocyte ratios exhibited trends with heterophils decreasing as O. ophiodiicola load increased and monocytes increasing as O. ophiodiicola load increased. These results suggest the innate immune response, and macrophagic leukocytes specifically, play an important role in the immune response to O. ophiodiicolainfection.

    3.  14:15  IN-PERSON    Endocrine Modulation of Retinal Sensitivity to Different Wavelengths in Hyla cinerea. Whitney Walkowski*, Louisiana State University Health Sciences Center; Ashley Santana, Louisiana State University Health Sciences Center; Twila Gaston, Louisiana State University Health Sciences Center; William Gordon, Louisiana State University Health Sciences Center; Nicolas Bazan, Louisiana State University Health Sciences Center; Hamilton Farris, Louisiana State University Health Sciences Center

    Many behavior patterns that are strongly modulated by hormones, such as mate choice, are mediated by visual processing. Yet, little is known about the effect of reproductive hormones on the retina, which is critical to understanding how visual signals are processed during these behaviors. This project examines the effects of reproductive status on retinal spectral sensitivity or the ability to detect different wavelengths associated with visual signals. Our hypothesis is that hormones modulate stimulus sensitivity in the retina during reproductively receptive phases, ultimately influencing mate choice behavior through modulation of color vision. We tested the effect that hormones have on color vision using the green tree frog (Hyla cinerea), which detect and respond to wavelengths spanning (and beyond) the human visible light spectrum. Experiments used electroretinograms (ERGs) to compare stimulus threshold and response amplitude in the retina of reproductive and non-reproductive female frogs. Additionally, we compared retinal activation before and after injections of human chorionic gonadotropin (hCG), which causes secretion of estrogen and progesterone in females and testosterone in males. Our findings indicated that for females reproductive state and hormone injections cause an increase in spectral sensitivity to particular ranges of wavelengths. These results strongly suggest that there may be functional consequences of endocrine modulation on visual communication during mate choice.

    4.  14:30  VIRTUAL    Invasive Bullfrogs Maintain High Levels of Immune Gene Diversity Despite Elevated Bd Infection Relative to Native Populations. Jacob LaFond*, University of Central Florida; Hollis Dahn, University of Toronto; Katherine Martin, University of Central Florida; Jonathan Ricmond, USGS; Bob Murphy, University of Toronto; Njal Rollinson, University of Toronto; Anna Savage, University of Central Florida

    To successfully become established, invasive populations must circumvent the negative effects of founder effect that often accompany an invasion. The maintenance of genetic diversity at key, polymorphic loci could help invasive populations persist in novel environments despite decreases in overall genomic diversity. The major histocompatibility complex (MHC) genes are a diverse family of genes that facilitate the adaptive immune response in vertebrates. Recent studies have demonstrated that maintenance of MHC diversity is associated with invasion success across multiple vertebrate taxa. The American bullfrog, Rana catesbeiana, is one of the most successful invasive amphibians. Ranacatesbeiana also has low susceptibility to the fungal zoonotic Batrachochytrium dendrobatidis (Bd) and has been implicated as a vector species responsible for the global spread of the pathogen. For our study, we sequenced the MHC class IIβ exon 2 locus of 286 individuals from 10 invasive and 10 native populations on the Illumina MiSeq 2 x 250 bp platform. We also sequenced the Cytochrome b (cytb) using Sanger sequencing, and tested for the presence of Bd. We recovered a total of 28 MHC alleles, and determined that invasive R. catesbeianapopulations saw nonsignificant decreases in MHC diversity, but significant decreases in cytb diversity compared to their native counterparts. We found that invasive populations also had a significantly higher Bd pathogen load than the native populations and that the 2 most common MHC alleles consistently correlated with a decrease in Bd infection, while 3 other alleles correlated with a significant increase in Bd infection.

    5.  14:45  VIRTUAL    Neonicotinoids Cross Blood-Brain Barrier and Delay Reaction Times in Northern Leopard Frogs, Rana pipiens. Kaitlyn Campbell*, University of South Dakota; Jacob Kerby, Ph.D., University of South Dakota

    Neonicotinoids are a new type of highly water-soluble insecticide used in agricultural practices to eliminate pests. These broad-spectrum toxicants target and bind almost irreversibly to postsynaptic nicotinic acetylcholine receptors in the central nervous system of invertebrates, resulting in overstimulation, paralysis, and death. Imidacloprid, the most commonly used neonicotinoid, is often transported to nearby wetlands through subsurface tile drains, agricultural runoff, or aerial wind drift, where it comes into contact with non-target organisms. Although it is widely accepted that neonicotinoids have selective toxicity for insects, our previous field study indicated amphibians collected from tile wetlands had higher imidacloprid brain levels compared to control sites, suggesting these insecticides can cross the blood-brain barrier and may have higher binding affinities than previously thought. To corroborate our field findings and determine the extent that imidacloprid crosses the blood-brain barrier, we exposed Rana pipiens to nominal concentrations (0, 0.1, 1, 5, or 10 µg/L) for 21-days and quantified imidacloprid, and its breakdown product, imidacloprid-olefin, in whole brain samples. Additionally, we conducted feeding trials to better understand how imidacloprid affects foraging behavior over time. Consistent with our field study, exposure groups had 8 to 190 times more imidacloprid in the brain relative to the control and breakdown products showed a dose-response relationship. Reaction times to a food stimulus were 1.8 to 3.8 times slower among treatment groups compared to the control. These results indicate imidacloprid can successfully cross the blood-brain barrier in amphibians and ecologically relevant concentrations can alter foraging behavior by decreasing reaction time.

    7.  15:00  VIRTUAL    Phylogenetics of Houston Toads (Bufo [=Anaxyrus] houstonensis): Genome-wide ddRADseq data reveal historic connectivity in an endangered relict. Shashwat Sirsi*, Texas State University; David Rodriguez, Texas State University; Michael Forstner, Texas State University

    Houston Toads (Bufo[=Anaxyrus] houstonensis) are endemic to Southeast-Central Texas, USA and currently face a high risk of extinction. Land-use changes have predominantly driven decline in B. houstonensis, with population supplementation predominant among recovery efforts for the species. However, there has been historic uncertainty regarding the taxonomic status and age of B. houstonensisrelative to B. americanus. Further, there are few genetic assessments to potentially inform population supplementation efforts. We used ~20,000 genome-wide nuclear DNA markers to examine phylogenetic relationships and divergence times between constituents of the B. americanusgroup and B. nebulifer,a sympatric Middle American species. Phylogenetic analyses support a Late Cretaceous divergence between Nearctic and Middle American representatives that is substantially older than previous estimates of their divergence. Admixture analysis indicated reproductive isolation between B. americanusand B. houstonensis while ABBA-BABA tests corroborated ancient gene flow, indicative of secondary contact, between the two taxa. Further, a minimum of three Management Units (i.e., genomic clusters) were identified within B. houstonensiswith a county-wide/regional spatial scale for population supplementation and protection/restoration of dispersal corridors. Additionally, observed genetic differentiation within B. houstonensis appeared consistent with the role of Pleistocene refugia in shaping contemporary genetic structure. Across analyses, we obtained a concordant signal on the influence of historic processes on population connectivity and ancient gene flow between and within populations of Nearctic toad taxa. Our study strengthens the need and ability forB. houstonensisconservation while re-affirming the evolutionary novelty of an endangered relict.

    8.  15:15  VIRTUAL    Immune priming prior to pathogen exposure sheds light on the relationship between host, microbiome, and pathogen in disease. Alyssa Kaganer*, Cornell University; Robert Ossiboff, Cornell University; Nicole Keith, Cornell University; Krysten Schuler, Cornell University; Pierre Comizzoli, Smithsonian Conservation Biology Institute; Matthew Hare, Cornell University; Robert Fleischer, Smithsonian Conservation Biology Institute; Brian Gratwicke, Smithsonian Conservation Biology Institute; Elizabeth Bunting, Cornell University

    Pathogens including Batrachochytrium dendrobatidis (Bd) threaten global biodiversity but efforts to protect at-risk amphibian species from Bd are hindered by limited understanding of amphibian, Bd, and microbiome interactions. We investigated these interactions during a vaccination evaluation of infection severity and outcome in eastern hellbender salamanders (Cryptobranchus alleganiensis alleganiensis) challenged with a high or low dose of Bd. We longitudinally monitored the commensal skin microbiome with 16S rRNA sequencing and separately characterized gene expression of host, pathogen, and microbiome during pre-vaccination, post-vaccination, early infection, and late infection using a metatranscriptome approach. Vaccination treatment did not affect hellbender survival or pathogen load; infected hellbenders in all vaccination groups maintained high load infections for 135 days. Despite no observed difference in disease severity, route of vaccination affected timing of host immune activation; orally primed animals upregulated innate and adaptive immune genes in response to vaccination while topically vaccinated animals did not. During Bd infection, sham vaccinated animals displayed robust immune gene activation in late infection, while topically vaccinated animals showed little immune activation, and orally vaccinated animals downregulated immune gene expression. Bd increased expression of pathogenicity-associated genes in late infection, and Bd load increased in all vaccination groups regardless of host immune gene expression. The commensal hellbender microbiome was altered in the presence of Bd, but there was no correlation between putative anti-Bd microbe abundance or richness and pathogen burden. Our observed ineffective immune response to Bd suggests hellbenders can survive clinical disease, which is not facilitated by interaction with antifungal skin microbiota.

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