1 Freeze Tolerance and Cryoprotection Of

Total Page:16

File Type:pdf, Size:1020Kb

1 Freeze Tolerance and Cryoprotection Of FREEZE TOLERANCE AND CRYOPROTECTION OF ERYTHROCYTES FROM DRYOPHYTES CHRYSOSCELIS Thesis Submitted to The College of Arts and Sciences of the UNIVERSITY OF DAYTON In Partial Fulfillment of the Requirements for The Degree of Master of Science in Biology By Loren Geiss Dayton, Ohio August 2018 1 FREEZE TOLERANCE AND CRYOPROTECTION OF ERYTHROCYTES FROM DRYOPHYTES CHRYSOSCELIS Name: Geiss, Loren Vanessa APPROVED BY: ________________________ Carissa M. Krane, Ph.D. Professor, Dept. of Biology, University of Dayton Faculty Advisor ________________________ Amit Singh, Ph.D. Associate Professor, Dept. of Biology, University of Dayton Committee Member ________________________ Yvonne Sun, Ph.D. Assistant Professor, Dept. of Biology, University of Dayton Committee Member ________________________ David L. Goldstein, Ph.D. Professor, Dept. of Biological Sciences, Wright State University Committee Member ii ABSTRACT FREEZE TOLERANCE AND CRYOPROTECTION OF ERYTHROCYTES FROM DRYOPHYTES CHRYSOSCELIS Name: Geiss, Loren Vanessa University of Dayton Advisor: Dr. Carissa M. Krane Dryophytes chrysoscelis (Cope’s gray treefrog) is a freeze-tolerant anuran that survives the freezing of its extracellular fluids. Treefrogs accumulate glycerol and urea in the plasma during cold acclimation and at the onset of freezing. It is hypothesized that glycerol and urea function as cryoprotectants by minimizing osmotically induced cell damage during freezing and thawing; and therefore, the post-freeze viability of red blood cells (RBCs) would improve when frozen in phosphate buffered saline (PBS) containing glycerol or urea. In the present study, erythrocytes were obtained from warm (22°C) and cold-acclimated (4°C) frogs and suspended in PBS. RBCs were frozen in PBS that ranged in osmolarity: PBS at 230 mM is hypoosmotic/hypotonic, 280 mM PBS is isosmotic/isotonic, and PBS was made hyperosmotic/hypertonic by the addition of 150 mM solutes. Post-freeze viability was determined with a hemolysis assay. Viability of erythrocytes from warm frogs was 31.6±4.9% in 230 mM PBS but was enhanced to 59.4±8.7% and 72.9±4.3%, respectively, when cells were frozen with glycerol (p<0.05) or urea (p<0.001). Post-freeze viability of cells from warm-acclimated frogs improved iii from 18.9±1.3% to 47.4±5.2% with the addition of urea to 280 mM PBS (p<0.01). RBCs from cold-acclimated frogs had 45.8±3.4% viability when frozen in 280 mM PBS which improved to 71.6±8.9% or 71.9±1.6%, respectively, when frozen with glycerol (p<0.01) or urea (p<0.001). The viability of RBCs from cold-acclimated frogs was not different when cells were frozen with glycerol, 71.6±8.9%, or left unfrozen (0°C), 88.4±3.5% (p>0.05). There was also no difference in viability between cells from cold-acclimated frogs that were frozen with urea, 71.9±1.6%, or left unfrozen, 86.7±3.8% (p>0.05). These data suggest that glycerol and urea are part of a complex cryoprotectant system in D. chrysoscelis. iv ACKNOWLEDGEMENTS My special thanks are in order to my past and present committee members Dr. Carissa Krane, Dr. Amit Singh, Dr. Yvonne Sun, Dr. David Goldstein, and Dr. Pothitos Pitychoutis for providing me with the time, knowledge, and guidance for this thesis. I would also like to thank Dr. Clara do Amaral who was a mentor and friend throughout my graduate career. I appreciate all the support from James Frisbie and members of the Krane Lab. Lastly, I would like to thank the fellow biology graduate students that have become my friends for their constant support and positivity. v TABLE OF CONTENTS ABSTRACT……………………………………………………………………………...iii ACKNOWLEDGEMENTS……………………………………………………………….v LIST OF FIGURES……………………………………………………………………..viii LIST OF TABLES………………………………………………………………………..ix LIST OF ABBREVIATIONS AND NOTATIONS……………………………………....x CHAPTER 1: LITERATURE REVIEW………………………………………………….1 Discovery of Freeze-Tolerant Anurans……………………….………………....1 Freezing and Thawing……………………………….…………………………..2 Geographic Differences of Freeze-Tolerant Anurans…………………….……..3 Cellular Osmotic Stress During Freezing and Thawing……………………..…..4 Cryoprotectants Mitigate Osmotic Stress………………………………..……....6 Aquaporins and Aquaglyceroporins………………..……………………………9 Aquaporins in Mammals………………………….……………………………12 Transport Mechanisms of AQP / Cell Signaling………………………..……...14 Aquaporins in D. chrysoscelis…………………………..……………………...15 Aquaporin Expression Based on Thermal State of D. chrysoscelis….………...16 Glycerol Induced HC-3 Expression……………………..……………………...17 Post-Freeze Viability of Erythrocytes…………………….……………………18 Hypothesis…………………………….………………………………………..19 vi References……………………………….……………………………………..21 CHAPTER 2: POST-FREEZE VIABILITY OF ERYTHROCYTES FROM DRYOPHYTES CHRYSOSCELIS………………………………………………………..26 Abstract ……………………………………………….……………..………...27 Introduction…………………………………………….………………….…...29 Materials and Methods……………………………….………………………...30 Results………………………………………………….………………………35 Discussion……………………………………………..………………………..38 Acknowledgements..…………………………………..………….……………44 References………………………………………………………….…………..45 CHAPTER 3: FUTURE DIRECTIONS AND BROADER IMPACTS………………....53 References………………………………………………………….…………..60 vii LIST OF FIGURES CHAPTER 1: LITERATURE REVIEW Figure 1. Glycerol regulates water flux during freezing and thawing……………………5 Figure 2. Structure of aquaporins………………………………………………………...11 CHAPTER 2: POST-FREEZE VIABILITY OF ERYTHROCYTES FROM DRYOPHYTES CHRYSOSCELIS Figure 1. Cryoprotection of D. chrysoscelis erythrocytes via 150 mM of solute…….….49 Figure 2. Cryoprotection of D. chrysoscelis erythrocytes with 150 or 1500 mM solutes……………………………………………………………………………………50 Figure 3. Cryoprotection of D. chrysoscelis erythrocytes with acute and long term exposure to solutes……………………………………………………………………….51 Figure 4. Freeze tolerance and cryoprotection of erythrocytes from warm and cold- acclimated D. chrysoscelis………………………………………………………….……52 CHAPTER 3: FUTURE DIRECTIONS AND BROADER IMPACTS Figure 1. Erythrocytes after incubation at 0C or -8C….…………..…………….…….56 viii LIST OF TABLES CHAPTER 1: LITERATURE REVIEW Table 1. Physiological variables of plasma from warm, cold, frozen, and thawed Dryophytes chrysoscelis…………………………………………………………………..8 Table 2. Mammalian aquaporins……………..…………………………………………..13 ix LIST OF ABBREVIATIONS AND NOTATIONS AQP Aquaporin CCCM Complete Cell Culture Media GLP Aquaglyceroporin GPCR G protein-coupled receptor NPA Asparagine-Proline-Alanine PBS Phosphate Buffered Saline RBC Red blood cell x CHAPTER 1 LITERATURE REVIEW Discovery of Freeze-Tolerant Anurans The goal of research is to answer scientific questions based on observations and human curiosity. One scientist, William Schmid, made the observation that some frogs can survive harsh winters by freezing and subsequently thawing when temperatures rise (Schmid 1982). This remarkable observation evoked other scientists to ask how freeze tolerance occurs and begin research that would answer their own questions regarding this phenomenon. Freeze tolerance is a unique strategy utilized by select organisms in environments that experience subfreezing temperatures. The ability of anurans to survive freezing is generally dependent on the proportion of body fluid that freezes and the osmolarity of body fluids (Costanzo et al. 1995). To date there are six known anuran species that are freeze-tolerant: Pseudacris crucifer (spring peeper, formerly Hyla crucifer), Rana sylvatica (wood frog), Pseudacris triseriata (western chorus frog), Dryophytes versicolor (gray treefrog, formerly Hyla versicolor) and its sister species Dryophytes chrysoscelis (Cope’s gray treefrog, formerly Hyla chrysoscelis), and lastly Dryophytes japonicus (Japanese treefrog, formerly Hyla japonica) (Schmid 1982; Storey and Storey 1986; Costanzo et al. 1992; Hirota et al. 2015). These freeze-tolerant anurans survive freezing of up to 65% of their extracellular 1 fluids over a period of days to months, and have a full recovery after thawing (Schmid 1982; Storey and Storey 1988; Storey and Storey 1992; Layne and Jones 2001). It is important to note that freeze tolerance is not limited to the six previously described anuran species. Other animals, such as specific species of turtles (Churchill and Storey 1991b; Costanzo and Claussen 1990; Storey and Storey 1992), snakes (Costanzo et al. 1988; Churchill and Storey 1991a; Storey and Storey 1992), and insects (Zachariassen and Hammel 1976; Storey and Storey 1996; Sinclair 1999, Sinclair et al. 2003), are also freeze-tolerant. A modern definition of freeze tolerance is a strategy of long-term freezing in which organisms have a maximum ice content they can survive when exposed to temperatures that are ecologically-relevant (Storey and Storey 1992). Freezing and Thawing As temperatures drop towards the subzero range, freeze-tolerant anurans burry themselves in damp leaf litter to be protected from wind and low air temperatures. Even in these microhabitats, temperatures can sink to -8°C (Schmid 1982; Storey and Storey 1992). Freeze-tolerant frogs can supercool when exposed to temperatures that are just below the freezing point, but internal ice inoculation occurs when the skin of a supercooled frog makes contact with environmental ice crystals (Layne et al. 1990; Layne 1991; Costanzo et al. 1995). In freeze-tolerant anurans ice crystals only form extracellularly, as intracellular ice formation would disrupt subcellular organization and be lethal (Mazur 1984; Storey and Storey 1992; Costanzo et al. 1995). Upon freezing, anurans become immobile, hypoxic, and ischemic (Storey and Storey 1988). Freeze- tolerant frogs in northern
Recommended publications
  • Cortisol Regulation of Aquaglyceroporin HC-3 Protein Expression in the Erythrocytes of the Freeze Tolerant Tree Frog Dryophytes Chrysoscelis
    University of Dayton eCommons Honors Theses University Honors Program 4-1-2019 Cortisol Regulation of Aquaglyceroporin HC-3 Protein Expression in the Erythrocytes of the Freeze Tolerant Tree Frog Dryophytes chrysoscelis Maria P. LaBello University of Dayton Follow this and additional works at: https://ecommons.udayton.edu/uhp_theses Part of the Biology Commons eCommons Citation LaBello, Maria P., "Cortisol Regulation of Aquaglyceroporin HC-3 Protein Expression in the Erythrocytes of the Freeze Tolerant Tree Frog Dryophytes chrysoscelis" (2019). Honors Theses. 218. https://ecommons.udayton.edu/uhp_theses/218 This Honors Thesis is brought to you for free and open access by the University Honors Program at eCommons. It has been accepted for inclusion in Honors Theses by an authorized administrator of eCommons. For more information, please contact [email protected], [email protected]. Cortisol Regulation of Aquaglyceroporin HC-3 Protein Expression in Erythrocytes from the Freeze Tolerant Tree Frog Dryophytes chrysoscelis Honors Thesis Maria P. LaBello Department: Biology Advisor: Carissa M. Krane, Ph.D. April 2019 Page | i Cortisol Regulation of Aquaglyceroporin HC-3 Protein Expression in the Erythrocytes of the Freeze Tolerant Tree Frog Dryophytes chrysoscelis Honors Thesis Maria P. LaBello Department: Biology Advisor: Carissa M. Krane, Ph.D. April 2019 Abstract Dryophytes chrysoscelis, commonly known as Cope’s gray treefrog, is a freeze tolerant anuran that freezes up to 65% of extracellular fluid during winter to survive. Glycerol is presumably used as a cryoprotectant during a period of cold-acclimation to protect cells from permanent damage due to hypoosmotic stress upon freezing and thawing. The passage of glycerol and water during cold-acclimation is mediated through aquaglyceroporin HC-3 in the nucleated erythrocytes (RBCs) of D.
    [Show full text]
  • Amphibiaweb's Illustrated Amphibians of the Earth
    AmphibiaWeb's Illustrated Amphibians of the Earth Created and Illustrated by the 2020-2021 AmphibiaWeb URAP Team: Alice Drozd, Arjun Mehta, Ash Reining, Kira Wiesinger, and Ann T. Chang This introduction to amphibians was written by University of California, Berkeley AmphibiaWeb Undergraduate Research Apprentices for people who love amphibians. Thank you to the many AmphibiaWeb apprentices over the last 21 years for their efforts. Edited by members of the AmphibiaWeb Steering Committee CC BY-NC-SA 2 Dedicated in loving memory of David B. Wake Founding Director of AmphibiaWeb (8 June 1936 - 29 April 2021) Dave Wake was a dedicated amphibian biologist who mentored and educated countless people. With the launch of AmphibiaWeb in 2000, Dave sought to bring the conservation science and basic fact-based biology of all amphibians to a single place where everyone could access the information freely. Until his last day, David remained a tirelessly dedicated scientist and ally of the amphibians of the world. 3 Table of Contents What are Amphibians? Their Characteristics ...................................................................................... 7 Orders of Amphibians.................................................................................... 7 Where are Amphibians? Where are Amphibians? ............................................................................... 9 What are Bioregions? ..................................................................................10 Conservation of Amphibians Why Save Amphibians? .............................................................................
    [Show full text]
  • Summary Conservation Action Plans for Mongolian Reptiles and Amphibians
    Summary Conservation Action Plans for Mongolian Reptiles and Amphibians Compiled by Terbish, Kh., Munkhbayar, Kh., Clark, E.L., Munkhbat, J. and Monks, E.M. Edited by Munkhbaatar, M., Baillie, J.E.M., Borkin, L., Batsaikhan, N., Samiya, R. and Semenov, D.V. ERSITY O IV F N E U D U E T C A A T T S I O E N H T M ONGOLIA THE WORLD BANK i ii This publication has been funded by the World Bank’s Netherlands-Mongolia Trust Fund for Environmental Reform. The fi ndings, interpretations, and conclusions expressed herein are those of the author(s) and do not necessarily refl ect the views of the Executive Directors of the International Bank for Reconstruction and Development / the World Bank or the governments they represent. The World Bank does not guarantee the accuracy of the data included in this work. The boundaries, colours, denominations, and other information shown on any map in this work do not imply any judgement on the part of the World Bank concerning the legal status of any territory or the endorsement or acceptance of such boundaries. The World Conservation Union (IUCN) have contributed to the production of the Summary Conservation Action Plans for Mongolian Reptiles and Amphibians, providing technical support, staff time, and data. IUCN supports the production of the Summary Conservation Action Plans for Mongolian Reptiles and Amphibians, but the information contained in this document does not necessarily represent the views of IUCN. Published by: Zoological Society of London, Regent’s Park, London, NW1 4RY Copyright: © Zoological Society of London and contributors 2006.
    [Show full text]
  • Phylogeography of the American Green Treefrog Species Group
    PHYLOGEOGRAPHY OF THE AMERICAN GREEN TREEFROG SPECIES GROUP by PAUL NATHANIEL PASICHNYK Presented to the Faculty of the Graduate School of The University of Texas at Arlington In Partial Fulfillment of the Requirements for the Degree of DOCTOR OF PHILOSOPHY THE UNIVERSITY OF TEXAS AT ARLINGTON November 2016 Copyright © by Paul N. Pasichnyk 2016 All Rights Reserved ii Acknowledgments A student’s graduate work is definitely not something that is done alone. I have received help from numerous people over the years and without their contributions, this would not have been possible. First, I would like to thank my parents for their tremendous support and motivation as I grew up and provided me with a foundation and love in education. Secondly, I would like to thank the person who has had to help me daily with tasks as simple as a pat on the back to as complex as reading a rough draft of a dissertation that might as well be in a foreign language, my wife Karen. My in-laws deserve special thanks as well, I do not believe that it is possible for others’ in-laws to be as supportive and helpful as mine. Then I would like to thank all the undergrads and friends who have helped see this to completion: Richard Hanna, Sari Mahon, Sarah Young, Nicole Lopez, Annamarie Slaven, Leslie Segovia, Pankaj BC, Matt Moseley, Christian Cox, Jeff Streicher, Claudia Marquez, Jesse Meik, Walter Schargel, Corey Roelke, Matt Watson, Rebbekah Watson, Thomas Eimermacher, Utpal Smart, David Sanchez, Jacobo Reyes-Velasco, Melissa Muenzler, Carl Franklin, Linda Taylor, Gloria Burlingham, Sherri Echols, and Paulette Williams.
    [Show full text]
  • Phylogeography Reveals an Ancient Cryptic Radiation in East-Asian Tree
    Dufresnes et al. BMC Evolutionary Biology (2016) 16:253 DOI 10.1186/s12862-016-0814-x RESEARCH ARTICLE Open Access Phylogeography reveals an ancient cryptic radiation in East-Asian tree frogs (Hyla japonica group) and complex relationships between continental and island lineages Christophe Dufresnes1, Spartak N. Litvinchuk2, Amaël Borzée3,4, Yikweon Jang4, Jia-Tang Li5, Ikuo Miura6, Nicolas Perrin1 and Matthias Stöck7,8* Abstract Background: In contrast to the Western Palearctic and Nearctic biogeographic regions, the phylogeography of Eastern-Palearctic terrestrial vertebrates has received relatively little attention. In East Asia, tectonic events, along with Pleistocene climatic conditions, likely affected species distribution and diversity, especially through their impact on sea levels and the consequent opening and closing of land-bridges between Eurasia and the Japanese Archipelago. To better understand these effects, we sequenced mitochondrial and nuclear markers to determine phylogeographic patterns in East-Asian tree frogs, with a particular focus on the widespread H. japonica. Results: We document several cryptic lineages within the currently recognized H. japonica populations, including two main clades of Late Miocene divergence (~5 Mya). One occurs on the northeastern Japanese Archipelago (Honshu and Hokkaido) and the Russian Far-East islands (Kunashir and Sakhalin), and the second one inhabits the remaining range, comprising southwestern Japan, the Korean Peninsula, Transiberian China, Russia and Mongolia. Each clade further features strong allopatric Plio-Pleistocene subdivisions (~2-3 Mya), especially among continental and southwestern Japanese tree frog populations. Combined with paleo-climate-based distribution models, the molecular data allowed the identification of Pleistocene glacial refugia and continental routes of postglacial recolonization. Phylogenetic reconstructions further supported genetic homogeneity between the Korean H.
    [Show full text]
  • 2021Harcourtajmres
    Bangor University MASTERS BY RESEARCH Interspecific Differences in Treefrog Response to Artificial Light at Night and Spectral Manipulation Harcourt, Alexander Award date: 2021 Link to publication General rights Copyright and moral rights for the publications made accessible in the public portal are retained by the authors and/or other copyright owners and it is a condition of accessing publications that users recognise and abide by the legal requirements associated with these rights. • Users may download and print one copy of any publication from the public portal for the purpose of private study or research. • You may not further distribute the material or use it for any profit-making activity or commercial gain • You may freely distribute the URL identifying the publication in the public portal ? Take down policy If you believe that this document breaches copyright please contact us providing details, and we will remove access to the work immediately and investigate your claim. Download date: 02. Oct. 2021 Interspecific Differences in Treefrog Response to Artificial Light at Night and Spectral Manipulation Understanding the effect of artificial light at night (ALAN) on biodiversity is a key research topic of the 21st Century. Evidence suggests that LED lighting may be particularly disruptive due to strong short-wavelength emissions. Spectral manipulation of LED lighting to reduce these emissions may mitigate some disturbance, although further research is required to assess its value in comparison with other techniques. The impact of LED lighting has been documented for many species, however, amphibians remain relatively under-studied. Amphibians may be particularly sensitive to the effects of ALAN due to specialised vision adapted for low-light environments and reduced mobility.
    [Show full text]
  • Phylogenetic Relationship Among Hylidae and Mitochondrial Protein-Coding Gene Expression in Response to Freezing and Ano
    See discussions, stats, and author profiles for this publication at: https://www.researchgate.net/publication/332083499 The complete mitochondrial genome of Dryophytes versicolor: Phylogenetic relationship among Hylidae and mitochondrial protein-coding gene expression in response to freezing and ano... Article in International Journal of Biological Macromolecules · March 2019 DOI: 10.1016/j.ijbiomac.2019.03.220 CITATIONS READS 5 209 6 authors, including: Jia-Yong Zhang Bryan E Luu Zhejiang Normal University McGill University 88 PUBLICATIONS 696 CITATIONS 29 PUBLICATIONS 193 CITATIONS SEE PROFILE SEE PROFILE Danna Yu Leping Zhang Zhejiang Normal University Westlake University 57 PUBLICATIONS 255 CITATIONS 18 PUBLICATIONS 104 CITATIONS SEE PROFILE SEE PROFILE Some of the authors of this publication are also working on these related projects: Hibernation Metabolomics View project New project: 1)The frog mitochondrial genome project: evolution of frog mitochondrial genomes and their gene expression View project All content following this page was uploaded by Jia-Yong Zhang on 08 April 2019. The user has requested enhancement of the downloaded file. International Journal of Biological Macromolecules 132 (2019) 461–469 Contents lists available at ScienceDirect International Journal of Biological Macromolecules journal homepage: http://www.elsevier.com/locate/ijbiomac The complete mitochondrial genome of Dryophytes versicolor: Phylogenetic relationship among Hylidae and mitochondrial protein-coding gene expression in response to freezing and
    [Show full text]
  • Social-Ecological Restoration in Paddy-Dominated Landscapes Ecological Research Monographs
    Ecological Research Monographs Nisikawa Usio · Tadashi Miyashita Editors Social-Ecological Restoration in Paddy-Dominated Landscapes Ecological Research Monographs Series Editor: Yoh Iwasa More information about this series at http://www.springer.com/series/8852 Nisikawa Usio • Tadashi Miyashita Editors Social-Ecological Restoration in Paddy- Dominated Landscapes Editors Nisikawa Usio Tadashi Miyashita Center for Toki and Ecological Graduate School of Agricultural and Life Restoration (CTER) Sciences Niigata University The University of Tokyo Sado, Japan Tokyo, Japan Institute of Nature and Environmental Technology (K-INET) Kanazawa University Kanazawa, Japan ISSN 2191-0707 ISSN 2191-0715 (electronic) Ecological Research Monographs ISBN 978-4-431-55329-8 ISBN 978-4-431-55330-4 (eBook) DOI 10.1007/978-4-431-55330-4 Library of Congress Control Number: 2014958758 Springer Tokyo Heidelberg New York Dordrecht London © Springer Japan 2014 This work is subject to copyright. All rights are reserved by the Publisher, whether the whole or part of the material is concerned, specifically the rights of translation, reprinting, reuse of illustrations, recitation, broadcasting, reproduction on microfilms or in any other physical way, and transmission or information storage and retrieval, electronic adaptation, computer software, or by similar or dissimilar methodology now known or hereafter developed. The use of general descriptive names, registered names, trademarks, service marks, etc. in this publication does not imply, even in the absence of a specific statement, that such names are exempt from the relevant protective laws and regulations and therefore free for general use. The publisher, the authors and the editors are safe to assume that the advice and information in this book are believed to be true and accurate at the date of publication.
    [Show full text]
  • Translocation of an Endangered Endemic Korean Treefrog Dryophytes Suweonensis
    A. Borzée, Y.-I. Kim, Y.-E. Kim & Y. Jang / Conservation Evidence (2018) 15, 6-11 Translocation of an endangered endemic Korean treefrog Dryophytes suweonensis Amaël Borzée1,2, Ye Inn Kim2, Ye Eun Kim2* & Yikweon Jang2,3*. 1 Laboratory of Behavioral Ecology and Evolution, School of Biological Sciences, Seoul National University, Seoul, 08826, Republic of Korea 2 Department of Life Sciences and Division of EcoScience, Ewha Womans University, Seoul, 03760, Republic of Korea 3 Interdisciplinary Program of EcoCreative, Ewha Womans University, Seoul, 03760, Republic of Korea SUMMARY Endangered species in heavily modified landscapes may be vulnerable to extinction if no conservation plan is implemented. The Suweon treefrog Dryophytes suweonensis is an endemic endangered species from the Korean Peninsula. In an attempt to conserve the species, a translocation plan was implemented in the city of Suwon. The receptor site was a specially modified island in a reservoir. Egg clutches were collected from four nearby sites, and were hatched and reared in a laboratory during 2015. One hundred and fifty froglets were released at the new site. In 2016, one year after the translocation, calling male D. suweonensis, and newly hatched tadpoles and juveniles were recorded. Juveniles were seen until the last week before hibernation in autumn 2016. However, only a single male was recorded calling in 2017. The population was consequently considered functionally extinct. Failure of the translocation most likely arose from mismanagement of the vegetation surrounding the wetlands, and the resulting inability of the site to fulfil the ecological requirements of the species. The project allowed the development of rearing protocols for the species, and defined its ecological requirements.
    [Show full text]
  • Phylogenetics, Classification, and Biogeography of the Treefrogs (Amphibia: Anura: Arboranae)
    Zootaxa 4104 (1): 001–109 ISSN 1175-5326 (print edition) http://www.mapress.com/j/zt/ Monograph ZOOTAXA Copyright © 2016 Magnolia Press ISSN 1175-5334 (online edition) http://doi.org/10.11646/zootaxa.4104.1.1 http://zoobank.org/urn:lsid:zoobank.org:pub:D598E724-C9E4-4BBA-B25D-511300A47B1D ZOOTAXA 4104 Phylogenetics, classification, and biogeography of the treefrogs (Amphibia: Anura: Arboranae) WILLIAM E. DUELLMAN1,3, ANGELA B. MARION2 & S. BLAIR HEDGES2 1Biodiversity Institute, University of Kansas, 1345 Jayhawk Blvd., Lawrence, Kansas 66045-7593, USA 2Center for Biodiversity, Temple University, 1925 N 12th Street, Philadelphia, Pennsylvania 19122-1601, USA 3Corresponding author. E-mail: [email protected] Magnolia Press Auckland, New Zealand Accepted by M. Vences: 27 Oct. 2015; published: 19 Apr. 2016 WILLIAM E. DUELLMAN, ANGELA B. MARION & S. BLAIR HEDGES Phylogenetics, Classification, and Biogeography of the Treefrogs (Amphibia: Anura: Arboranae) (Zootaxa 4104) 109 pp.; 30 cm. 19 April 2016 ISBN 978-1-77557-937-3 (paperback) ISBN 978-1-77557-938-0 (Online edition) FIRST PUBLISHED IN 2016 BY Magnolia Press P.O. Box 41-383 Auckland 1346 New Zealand e-mail: [email protected] http://www.mapress.com/j/zt © 2016 Magnolia Press All rights reserved. No part of this publication may be reproduced, stored, transmitted or disseminated, in any form, or by any means, without prior written permission from the publisher, to whom all requests to reproduce copyright material should be directed in writing. This authorization does not extend to any other kind of copying, by any means, in any form, and for any purpose other than private research use.
    [Show full text]
  • Distribution and Genetic Diversity of the Amphibian Chytrid in Japan
    Journal of Fungi Article Distribution and Genetic Diversity of the Amphibian Chytrid in Japan Koichi Goka 1,*, Jun Yokoyama 2 and Atsushi Tominaga 3 1 National Institute for Environmental Studies, 16-2 Onogawa, Tsukuba 305-8506, Japan 2 Department of Biology, Faculty of Sciences, Yamagata University, 1-4-12 Kojirakawa, Yamagata-shi, Yamagata 990-8560, Japan; [email protected] 3 Department of Natural Sciences, Faculty of Education, University of the Ryukyus, 1 Senbaru, Nishihara, Okinawa 901-0213, Japan; [email protected] * Correspondence: [email protected]; Tel.: +81-29-850-2480; Fax: +81-29-850-2582 Abstract: While research on frog chytrid fungus Batrachochytrium dendrobatidis (Bd), an infectious disease that threatens amphibian diversity, continues to advance worldwide, little progress has been made in Japan since around 2010. The reason for this is, which we pointed out in 2009, that the origin of frog chytrid fungus may be in the East Asian region, including Japan based on the Bd ITS-DNA variation, and as few cases of mass mortality caused by this fungus have been observed in wild amphibian populations in Japan, the interest of the Japanese government and the general public in Bd has waned. However, we believe that organizing the data obtained so far in Japan and distributing the status of frog chytrid fungus in Japan to the world will provide useful insight for future risk management of this pathogen. We collected more than 5500 swab samples from wild amphibians throughout Japan from 2009 to 2010. Then, we investigated the infection status using the Nested-PCR method.
    [Show full text]
  • Captive Wildlife Regulations, 2021, W-13.12 Reg 5
    1 CAPTIVE WILDLIFE, 2021 W-13.12 REG 5 The Captive Wildlife Regulations, 2021 being Chapter W-13.12 Reg 5 (effective June 1, 2021). NOTE: This consolidation is not official. Amendments have been incorporated for convenience of reference and the original statutes and regulations should be consulted for all purposes of interpretation and application of the law. In order to preserve the integrity of the original statutes and regulations, errors that may have appeared are reproduced in this consolidation. 2 W-13.12 REG 5 CAPTIVE WILDLIFE, 2021 Table of Contents PART 1 PART 5 Preliminary Matters Zoo Licences and Travelling Zoo Licences 1 Title 38 Definition for Part 2 Definitions and interpretation 39 CAZA standards 3 Application 40 Requirements – zoo licence or travelling zoo licence PART 2 41 Breeding and release Designations, Prohibitions and Licences PART 6 4 Captive wildlife – designations Wildlife Rehabilitation Licences 5 Prohibition – holding unlisted species in captivity 42 Definitions for Part 6 Prohibition – holding restricted species in captivity 43 Standards for wildlife rehabilitation 7 Captive wildlife licences 44 No property acquired in wildlife held for 8 Licence not required rehabilitation 9 Application for captive wildlife licence 45 Requirements – wildlife rehabilitation licence 10 Renewal 46 Restrictions – wildlife not to be rehabilitated 11 Issuance or renewal of licence on terms and conditions 47 Wildlife rehabilitation practices 12 Licence or renewal term PART 7 Scientific Research Licences 13 Amendment, suspension,
    [Show full text]