International Horseshoe Crab Conservation and Research Efforts: 2007- 2020 : Conservation of Horseshoe Crabs Species Globally

The first International Conference on Horseshoe Crab’s Conservation conducted at Dowling College, USA, (2007) and it’s proceedings published by Springer in 2009, prompted the continued research and conservation efforts presented at subsequent conferences and colloquium in Hong Kong, Taiwan, (2011); San Diego, CA, (2014), (CERF); Japan, Sasebo (2015) and an accepted inclusion for a special session on Horseshoe Crabs at the 2017 CERF Conference held in Providence, RI, USA. All these aforementioned conferences contributed manuscripts, posters, workshop “position papers”, and oral presentations the majority of which have not been published in total. In 2015, Carmichael et al. had published by Springer the majority of manuscripts from the 2011 Hong Kong / Taiwan conference. However, workshop results and all subsequent presentations and workshops were not. The Japan conference presented over 40 papers alone. A collection of all workshop summaries, poster presentations and new manuscript submittals (San Diego, CA; Sasebo, Japan; and Providence, RI) as well as products prepared for the IUCN World Congress in Hawaii, (2016), are included potential contributions for review in this compilation now available for global distribution in this Springer Nature publication. The “Proceedings of International Conferences on the Biology and Conservation of Horseshoe Crabs”, thus contains over 50 manuscripts and a diversified collection of documents, photos and memorabilia covering all four of the horseshoe crab species globally: their biology, ecology evolution, educational, and societal importance. This book exposes the impacts that humans have imposed on all four of these species, revealing through the coordinated effort of horseshoe crab scientists with the IUCN, of the worldwide need for a clear conservative effort to protect these paleo- survival organisms from a looming extinction event. Biologists, conservationists, educators, and health professionals will all welcome this book not only for exploration of its pharmacological interest, but also for the mystery of their longevity. This book also clarifies the future research needs and the conservation agenda for the species worldwide. Anyone working or studying estuaries on a global scale, will need to obtain this seminal work on horseshoe crabs. Preface Contents About the Editors An Homage to Carl Shuster’s Contributions to Horseshoe Crab Biology Dr. Carl N. Shuster Jr.: A Brief Biography Reflections on Dr. Carl N. Shuster Jr., by Mark L. Botton (Fordham University) and Bob Loveland (Rutgers University) Celebrating the Life of Dr. Carl N. Shuster Jr., by Glenn Gauvry (Ecological Research Development Group) and Gary Kreamer (Delaware Division of Fish and Wildlife, retired) Memories of Carl, by Mike Oates (Filmmaker and Documentarian) Personal Reflections on Dr. Carl N. Shuster Jr., by Gary Kreamer (Delaware Division of Fish and Wildlife, retired) Remembrances, by Dr. Dick Weber (University of Delaware, retired) An Appreciation, by Dr. David Smith (U.S. Geological Survey) Remembering Carl Shuster, by Dr. Ruth Carmichael (Dauphin Island Sea Lab) Personal Reflections, by Glenn Gauvry (Ecological Research and Development Group) Remembering Carl Shuster, by H. Jane Brockmann (University of Florida, retired) Dr. Carl N. Shuster Jr.: A Selected Bibliography Part I: Evolution Genetics & Adaptation Variation in the American Horseshoe Crab: A Spiny Variant from the Southeast 1 Introduction 2 Methods 3 Results 4 Discussion Literature Cited Genetic Structure of the Tri-Spine Horseshoe Crab in the Seto Inland Sea, Japan: Is the Current Population at Kasaoka in the Eastern Area Native or Re-established? 1 Introduction 2 Materials and Methods 3 Results 4 Discussion References Molecular Identification and Phylogenetic Analysis of Indian Mangrove Horseshoe Crab 1 Introduction 2 Materials and Methods 2.1 Collection of Horseshoe Crabs 2.2 Isolation of Horseshoe Crab DNA from the Gills 2.3 PCR Amplification of Isolated DNA 2.4 Purification of PCR Products 2.5 Molecular Identification and Phylogenetic Analysis 3 Result and Discussion 3.1 DNA Isolation and PCR Amplification 3.2 Molecular Characterization and Phylogenetic Analysis 4 Conclusion References Population Genetics and Movement Show Metapopulation Dynamics of Mid-Atlantic Region Horseshoe Crabs 1 Introduction 1.1 Population Genetics 1.2 Movement 1.3 Objectives 2 Methods 2.1 Population Genetics 2.2 Movement 3 Results 3.1 Population Genetics 3.2 Movement 4 Discussion 4.1 Population Genetics 4.2 Movement 4.3 Management Perspectives References Microbiome Associated with Laboratory Bred Horseshoe Crabs: A Case Study 1 Introduction 2 Methods 3 Results and Discussion References Population Genetic Structure of Juvenile Tri-Spine Horseshoe Crabs in Hong Kong 1 Introduction 2 Materials and Methods 2.1 Field Collection of Juvenile Horseshoe Crabs 2.2 DNA Extraction and Analysis 2.3 Data Treatment and Analysis 3 Results 4 Discussion Appendix 1. Genetic sequence similarities among different tri-spine horseshoe crab individuals in Hong Kong. Sampling locations: Ha Pak Nai (HPN), Pak Nai (PN), Shui Hau (SH), San Tau (ST), Sham Wat (SW), and Yi O (YO) References Applying Records of Extant and Extinct Horseshoe Crab Abnormalities to Xiphosurid Conservation 1 Introduction 1.1 Abnormalities: Causes and Forms 2 Methods 3 Results 3.1 Injury Patterns 3.2 Abnormal Fossil Specimens 4 Discussion 4.1 Patterns of Abnormalities on Extant Species 4.2 Fossil Xiphosurids 4.3 Conservation Implications References Part II: Population Dynamics & Ecology Activity Rhythm of the Tri-Spine Horseshoe Crab Tachypleus tridentatus in the Seto Inland Sea, Western Japan, Monitored with Acceleration Data-Loggers 1 Introduction 2 Materials and Methods 2.1 Experimental Conditions 2.2 Activity Recording 2.3 Data Analysis 3 Results 4 Discussion 4.1 Activity Rhythm of Tachypleus tridentatus 4.2 Ecological Functions of the Activity Rhythm in the Seto Inland Sea References Sediment-Molting Relationship of Juvenile Limulus polyphemus (Arthropoda, Merostomata) and Predation Against them 1 Introduction 2 Materials and Methods 2.1 Experimental Setup 2.2 Experimental Procedures Preference in Substrate Texture Introduction of Predators 3 Results 3.1 Preference in Substrate Texture Second and Third Instars Fourth to Sixth Instars 3.2 Introduction of Predators Juvenile Stone Crab Adult Female Blue Crabs Juvenile Female Blue Crab Hermit Crabs 4 Discussion 4.1 Frequency of Molting and Death 4.2 Chemosensory Reaction 4.3 Visual Acuity 4.4 Role of Temperature 5 Conclusion 6 Future Study 6.1 Sediment Size 6.2 Role of Temperature for Burrowing Activities in Juvenile Horseshoe Crabs References Conservation and Restoration of Estuaries and Coasts: Horseshoe Crabs as Flagship Species 1 Introduction 2 Sandy/Cobble Beaches 2.1 Ecosystem Services of Beaches 2.2 What Are the Causes of Loss/Decline of Beaches? Case Study: Shoreline Armoring, the Japanese Experience 2.3 Restoration of Beaches in General Beach Nourishment and Horseshoe Crabs: A Case History from New York City, USA 3 Saltmarshes, Seagrasses 3.1 Ecosystem Services of Marshes and Seagrasses 3.2 What Are the Causes of Loss/Decline in General? 3.3 Restoration Case Study: Nature-Based Restoration in Stratford, Connecticut, USA 4 Mangrove Forest Ecosystem 4.1 Relevance of Mangrove Restoration for Conservation of Horseshoe Crabs What Are the Causes of Mangrove Decline and Loss? Community-Based Ecological Mangrove Restoration: An Effective Approach to Mangrove Restoration 5 Intertidal Flats: Important for Juvenile Horseshoe Crab Growth and Development 5.1 Importance of Mudflats 5.2 Restoration and Conservation of Mudflats 6 Horseshoe Crabs Outreach: Connect People with Their Coastal Environments 6.1 Beaches/Marshes/Seagrasses 6.2 Mangroves Horseshoe Crabs, Mangroves, and Citizen Science in Singapore 6.3 Mudflats: Horseshoe Crabs and Marine Conservation in Hong Kong 7 Summary and Conclusions References Preliminary Characterization of Hemocyte and Immunity of Asian Horseshoe Crabs, Tachypleus tridentatus, and Carcinoscorpius rotundicauda in Captivity 1 Introduction 2 Materials and Methods 2.1 Collection and Culture of Horseshoe Crabs 2.2 Hemolymph Sampling and Preparation 2.3 Determination of Enzymatic Activity 2.4 Statistical Analysis 3 Results 3.1 Morphological Characteristics of Hemocytes 3.2 Enzymatic Activity Determination 4 Discussion 4.1 Classification of Horseshoe Crab Hemocytes 4.2 Characterization of Enzymatic Activity in Different Tissues References Visually Guided Behaviour of Juvenile Tri-spine Horseshoe Crab Tachypleus tridentatus: Implications for Designing Racks for Oyster Cultivation Compatible with Horseshoe Crab Conservation 1 Introduction 2 Materials and Methods 2.1 Effect of Obstacles on Foraging Behaviour in the Field 2.2 Preference of Degree of Shading 2.3 Statistical Analysis 3 Results 3.1 Effect of Obstacles on Foraging Behaviour in the Field 3.2 Preference of Degree of Shading 4 Discussion References Condition of Horseshoe Crabs as Bycatch from Gillnet Fisheries in Mayangan Waters of Subang Regency, West Java, Indonesia 1 Introduction 2 Materials and Methods 3 Results and Discussion 4 Conclusion References Occurrence and Morphometric Analysis of Horseshoe Crabs Along Bhadrak Coast of Odisha, India 1 Introduction 2 Materials and Methods 2.1 Study Site 2.2 Data Collection 2.3 Morphometric Measurements 3 Results 3.1 Total Samples of Horseshoe Crabs 3.2 Morphometric Analysis 3.3 Other Observations 4 Discussion References Preliminiary Investigation of Tachypleus tridentatus (Tri-spine HSC at Sone Tidal Flat, Fukuoka Japan) 1 Introduction 2 Spawning Survey 2.1 Summary of Survey Areas Yoshida Coast Ono River Nuki River Kusami River Matsuyama Coast Mashima Island 2.2 Survey Methods and Their Transition 2.3 Results 3 Juvenile Horseshoe Crab Survey 3.1 Changing Approach to Juvenile Horseshoe Crab Survey 3.2 Survey Method 3.3 Results 4 Conclusions References Distribution of Horseshoe Crabs, Carcinoscorpius rotundicauda, and Tachypleus gigas in Singapore, Including Coney Island 1 Introduction 2 Materials and Methods 2.1 Study Sites 2.2 Visual Search 3 Results and Discussion 3.1 Distribution of Both Species in Singapore 3.2 Changi Beach, Pasir Ris Park Beach, and Punggol Settlement 3.3 Coney Island 3.4 Comparison of T. gigas at Different Sites 3.5 Sungei Buloh Wetland Reserve 4 Conclusion References An Assessment of Horseshoe Crab Breeding Habitat and Trends on Long Island, New York, 2003–2017 1 Introduction 1.1 Horseshoe Crab General Background 1.2 Long Island Background (Fig. 2) 2 Methods 2.1 Study Area 2.2 Horseshoe Crab Inventory 2.3 Horseshoe Crab Analysis 2.4 Temperature Data Collection CERCOM? 2.5 Land-Use Data 2.6 Potential Breeding Area Collection and Analysis 3 Results 3.1 Horseshoe Crab Inventory and Classification 3.2 Sea Surface Water Temperature Data 3.3 Land-Use Analysis 3.4 Horseshoe Crab Potential Breeding Area Results Active Breeding Beach Zones (AB) Recovering Beach Zones (RB) Pending Beach Zones (PB) Nonbreeding Beaches (NBB) 3.5 Case Studies Manhasset Stony Brook Zone Northport Bay Zone Mastic Zone 4 Discussion 4.1 Land Use 4.2 Potential Breeding Area 4.3 Case Studies 4.4 Conclusions Appendix References Part III: Conservation Efforts & Managment Education Identifying the Major Threats to American Horseshoe Crab Populations, with Emphasis on Delaware Bay 1 Introduction 1.1 Background 1.2 Commercial Exploitation and Management 2 Loss and Degradation of Spawning Habitat 2.1 Habitat Loss Along the Cape May Peninsula 3 Horseshoe Crabs and Oyster Farms 3.1 Background Surveys Behavior Experiments Survey Results Behavioral Experiments: Results 3.2 Assessing the Impacts of Oyster Culture 4 Biomedical Use of Horseshoe Crabs 4.1 Discovery of a Reagent for Detection of Bacterial Endotoxin 4.2 FDA Elects to License and Regulate LAL as a Biological Product 4.3 The LAL Industry 4.4 Fishery Management Plan (FMP) and Biomedical Uses of the Horseshoe Crab 4.5 Estimated Mortality from LAL Processes 4.6 The Future of LAL Products 5 Summary and Conclusions References The Need to Establish Horseshoe Crab Global Biorepository 1 Background 2 Horseshoe Crab Biorepository 3 Nagoya Protocol 4 Data-Sharing Plan Benefits 5 Current Horseshoe Crab at Mid Repository References Economics of the Limulus/Tachypleus Amebocyte Lysate (LAL/TAL) Industry Relative to the Sustainability of Horseshoe Crabs Worldwide 1 Introduction 2 LAL Alternatives 3 Bait Alternatives 4 Horseshoe Crab Sustainability 5 Conclusions References LAL/TAL and Animal-Free rFC-Based Endotoxin Tests: Their Characteristics and Impact on the Horseshoe Crab Populations in the United States and Asia 1 Introduction and History of Bacterial Endotoxin Testing 2 Characteristics of LAL/TAL and rFC Tests 3 Regulation of Bacterial Endotoxin Tests 4 Effects of Horseshoe Crab Harvesting and Habitat Loss 5 Absence of Clear Ethical Directives 6 Conclusion References The Power of Citizen Science: 20 Years of Horseshoe Crab Community Research Merging Conservation, Education, and Management 1 Introduction 2 Methods 2.1 Mark-Recapture Study 2.2 Spawning Census Data 2.3 Genetic Characterization of LIS Horseshoe Crab Population 3 Results 3.1 Mark-Recapture Study 3.2 Spawning Census Results 3.3 Genetic Characterization of LIS Horseshoe Crab Population 4 Discussion References Relative Abundance of Horseshoe Crabs in the Delaware Bay Region: A Critical Factor for Adaptive Resource Management 1 Introduction 2 Methods 3 Results 3.1 Size 3.2 Spatial Distribution 3.3 Year Classes 3.4 Sex Ratios 3.5 Relative Abundance 4 Discussion References Detrimental Coastline Alteration at Horseshoe Crab Spawning Grounds in Pahang, South China Sea 1 Introduction 2 Changing of Sediment Properties in Light of Coastal Disturbances 3 Shoreline Topography Changes Assessment by Geospatial Data 4 Discussion 4.1 Natural Determinants of Morphodynamics in the Coastal Area 4.2 Anthropic Activities and Morphodynamic Changes to the Horseshoe Crab 4.3 Suggestions on Mitigation Efforts to Minimize the Coastal Impacts 5 Conclusion References Assessing the Use of Acoustic Sampling for Locating Horseshoe Crabs During Migrations Using Side Scan Sonar 1 Overview 2 Methods 3 Results 4 Discussion References Asian Horseshoe Crab Conservation: Knowledge, Attitudes, and Intentions of Local College Students in Northern Beibu Gulf, China 1 Introduction 2 Materials and Methods 2.1 Study Area 2.2 Survey Method 2.3 Statistical Analyses 3 Results 4 Discussion Supplementary Material Section A: Background Information of Respondents Section B: Concepts and Knowledge of Horseshoe Crabs Section C: Conservation Awareness of Horseshoe Crabs References Fishery Bycatch Jeopardizes Indian Horseshoe Crab Tachypleus gigas Along the Northeast Coast of Odisha, India 1 Introduction 2 Materials and Methods 2.1 Study Sites 2.2 Survey Observations 2.3 Data Analysis 3 Results 4 Discussion and Conservation Implications References Citizen Science Approach for Indonesian Horseshoe Crab Conservation 1 Introduction 2 Ecology and Value of Horseshoe Crabs in Indonesia 3 Citizen Science Approach for Conservation 4 Summary References Cumulative Modifications of the Endangered Horseshoe Crab Tachypleus tridentatus Habitat by Small-Scale Developments at Tsuyazaki Cove in Fukuoka, Japan 1 Introduction 2 Methods and Materials 2.1 Study Site 2.2 General Methods and Analysis 3 Results 3.1 Changes in the Potential Spawning Sites from 1948 to 2010 3.2 Deterioration of the Most Significant Spawning Site from 2005 to 2013 3.3 Decline of Breeding Pairs from 2005 to 2013 4 Discussion 4.1 Changes in the Potential Spawning Sites from 1948 to 2010 4.2 Deterioration of the Most Significant Spawning Site Resulting in the Decline of Breeding Pairs from 2005 to 2013 4.3 Importance of T. tridentatus Conservation 4.4 Recommendations for Restoring the Local Population 4.5 Conclusion References Exploitation Status of Two Species of Horseshoe Crabs from Subang, West Java, Indonesia 1 Introduction 2 Methods 2.1 Sampling Sites and Techniques 2.2 Physicochemical Parameters and Sediment Samples 2.3 Data Analysis 3 Results 3.1 Number of Horseshoe Crabs and Physicochemical Habitat Parameters 3.2 Length–Weight Relationship 3.3 Growth Models 3.4 Mortality and Exploitation Rates 4 Discussion 4.1 The Number of Horseshoe Crabs and Physicochemical Habitat Parameters 4.2 Growth Rate, Mortality, and Exploitation 4.3 Management Directions 4.4 Limitations of the Study References Captive Breeding Success and Pitfalls for Horseshoe Crab, Limulus polyphemus, at CERCOM, Molloy College 1 Salt Water Well Conditions and Infrastructure 2 Laboratory’s Bay Water Conditions and Infrastructure 3 Closed Aquaculture Systems 4 Captive Diet and Feeding Routines for Adult HSCs 5 Captive Spawning 6 Juvenile Rearing and Care 7 Conclusion References City-Wide Efforts in the Conservation of Horseshoe Crab (Tachypleus tridentatus) in Imari City, Saga, Japan 1 Conservation of Horseshoe Crabs in Imari 1.1 Regulations to Protect Horseshoe Crabs in Imari Bay 1.2 Regulations Related to Environment 1.3 Regarding Act on Protection of Cultural Properties 1.4 Protection of Cultural Properties as Preservation and Utilization 2 About the Preservation 2.1 The Necessity of Preservation 2.2 Awareness among Local Citizens Society for the Protection of Horseshoe Crabs, Imari City (Hereinafter, City Society) Saga Prefectural High School, Science and Biology Club (Hereinafter, Club) Makishima Society for Breeding Horseshoe Crabs and Fireflies (Hereinafter, Makishima Society) The City of Imari Saga Prefecture 2.3 Survey for Understanding the Actual Habitat Spawning Pair Number Survey Habitat Distribution Survey 2.4 For Breeding 2.5 Protection of Environment 2.6 Educational Activities 2.7 Coexistence of Development and Conservation 3 Utilization 3.1 Maintenance 3.2 A Resource for Local Town Planning Town Planning and Revitalization 3.3 Research Institutions and Exhibit Facility 3.4 Resource as Economic Activities 4 Becoming a Designation as a National Natural Monument 5 Records of the Survey Studies at Imari High School 6 Breeding 6.1 Method to Release Juveniles 7 Future Tasks Part IV: Biology & Physiology Temperature and Salinity Preferences of Adult American Horseshoe Crabs (Limulus polyphemus) in the Great Bay Estuary, New Hampshire U.S.A 1 Introduction 2 Materials and Methods 2.1 Horseshoe Crabs 2.2 Y-Maze Experimental Design 2.3 Cardiac Assay Experimental Design 2.4 Data Analyses 3 Results 3.1 Overall Movements in the Y-Maze 3.2 Y-Maze Experiments: Temperature Experiment 1: Warm Water Versus Ambient-Temperature Water Experiment 2: Warm Versus Cold Water 3.3 Y-Maze Experiments: Salinity 3.4 Cardiac Assay Results 4 Discussion References The Effects of Roundup® (a Glyphosate-Based Herbicide) on the Survival and Development of American Horseshoe Crab, Limulus polyphemus 1 Introduction 2 Methods 3 Results 4 Discussion References Feeding Ecology and Dietary Preferences of Tachypleus gigas from East Malay Peninsula 1 Introduction 2 Materials and Methods 2.1 Description of Study Area 2.2 Sample Collection and Preparation 2.3 Data Analysis 3 Results and Discussion 3.1 Macrobenthos Composition at the Nesting Grounds 3.2 Feeding Composition of T. Gigas 3.3 Natural Food Selectivity 4 Conclusion References Patterns of Flatworm (Bdelloura candida) Infestation in a Population of Horseshoe Crabs (Limulus polyphemus) from Jamaica Bay, New York 1 Introduction 2 Methods 2.1 Data Collection of Adult Horseshoe Crabs 2.2 Data Collection for Juvenile Horseshoe Crabs 2.3 Confirmation of Flatworm Species and Photography 3 Results 3.1 General Extent 3.2 Flatworm Species Identification 3.3 Correlation of Prosoma Width to Infestation 3.4 Sex and Carapace Condition and Flatworm Infestation 3.5 Prevalence of Flatworm Infestation on Juvenile Horseshoe Crabs 4 Discussion References Trophic Relationship of Sympatric Juvenile Asian Horseshoe Crabs in Beibu Gulf, Southwestern China 1 Introduction 2 Materials and Methods 2.1 Study Locations 2.2 Horseshoe Crab and Potential Prey Tissue Sample Collection 2.3 Organic Matter, Microalgal, and Plant Sample Collection 2.4 Sample Preparation and Stable Isotope Analyses 2.5 Data Analysis 3 Results 3.1 δ13C and δ15N Signatures in Juvenile Horseshoe Crabs 3.2 Potential Food Sources of Juvenile Horseshoe Crabs 4 Discussion 5 Conclusion References Tachypleus gigas and Carcinoscorpius rotundicauda Selection and Consumption of Tissue from Four Different Bivalve Species in Captivity 1 Introduction 2 Materials and Methods 2.1 Sample Collection 2.2 Feeding Experiment 2.3 Proximate Analysis of Bivalve Tissue 2.4 Data Analysis 3 Results 4 Discussion 5 Conclusion References Research Progress and Prospect of Tachypleus Amebocyte Lysate in China 1 Principles and Mechanisms of TAL 2 TAL Applications 3 Endotoxin Test Classification 4 TAL Disadvantages 5 Current Difficulties of China’s TAL Industry 5.1 Declining Horseshoe Crab Resources 5.2 Uneven Quality of TAL Products 6 Future Prospects of TAL in China 6.1 Substitutes Development for TAL 6.2 Horseshoe Crab Population Restoration by Artificial Breeding References Ecology of Juvenile American Horseshoe Crabs (Limulus polyphemus) at Plumb Beach, Jamaica Bay, New York 1 Introduction 2 Research Site 3 Population Dynamics, Growth, and Size Distribution of Juvenile Horseshoe Crabs 3.1 Methods 3.2 Results Live Juveniles Shed Carapaces 3.3 Discussion 4 Diet and Food Preferences of Juvenile Limulus polyphemus 4.1 Methods 4.2 Results Gut and Sediment Content Electivity 4.3 Discussion 5 Righting and Burrowing Behavior of Juvenile Limulus in Different Sand Substrates 5.1 Methods 5.2 Results 5.3 Discussion 6 General Discussion and Conclusions References