Hemolymph Proteomics and Gut Microbiota of Horseshoe Crabs Tachypleus tridentatus and Carcinoscorpius rotundicauda

Horseshoe crabs are a group of marine chelicerates that contain only four extant species, some of which are endangered. Their hemolymph has been widely used in medical applications for endotoxin detection. Nevertheless, there is limited information on the profiles of their hemolymph proteins and their gut microbial diversity. In this study, we performed the first detailed investigation of the hemolymph proteomics and gut microbiota of two Asian horseshoe crabs Tachypleus tridentatus and Carcinoscorpius rotundicauda. Among the identified proteins being cataloged in the juvenile and adult hemolymph, unexpectedly, sesquiterpenoid signaling pathway proteins including Heat shock protein 83 (HSP83), Chd64, and a juvenile hormone binding protein (JHBP) were revealed. This provides evidence for the presence of functional sesquiterpenoid hormonal systems in these marine chelicerates. consumption of certain horseshoe crab species often leads to tetrodotoxin poisoning and the horseshoe crab is thought to possess a tetrodotoxin resistance mechanism. As such, sodium channels were analyzed and found to have critical amino acid residues that are similar to the toxin resistant pufferfish sodium channels. The source of the toxin is unknown so we investigated the gut microbiota, and found that Clostridium and Vibrio were the most dominant bacteria in T. tridentatus and C. rotundicauda, respectively. Together, this study provides a framework for further understanding of sesquiterpenoids and gut microbiota of these marine chelicerates.

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Effects of Ocean Acidification on Molting, Oxidative Stress, and Gut Microbiota in Juvenile Horseshoe Crab Tachypleus tridentatus

Frontiers in Physiology ◽

10.3389/fphys.2021.813582 ◽

2022 ◽

Vol 12 ◽

Author(s):

Ximei Liu ◽

Jiani Liu ◽

Kai Xiong ◽

Caoqi Zhang ◽

James Kar-Hei Fang ◽

...

Keyword(s):

Oxidative Stress ◽

Gut Microbiota ◽

Ocean Acidification ◽

Horseshoe Crab ◽

Life Stage ◽

Growth Index ◽

Immune Defense ◽

Control Group ◽

Tachypleus Tridentatus ◽

Horseshoe Crabs

Anthropogenic elevation of atmospheric carbon dioxide (CO2) drives global-scale ocean acidification (OA), which has aroused widespread concern for marine ecosystem health. The tri-spine horseshoe crab (HSC) Tachypleus tridentatus has been facing the threat of population depletion for decades, and the effects of OA on the physiology and microbiology of its early life stage are unclear. In this study, the 1st instar HSC larvae were exposed to acidified seawater (pH 7.3, pH 8.1 as control) for 28 days to determine the effects of OA on their growth, molting, oxidative stress, and gut microbiota. Results showed that there were no significant differences in growth index and molting rate between OA group and control group, but the chitinase activity, β-NAGase activity, and ecdysone content in OA group were significantly lower than those of the control group. Compared to the control group, reactive oxygen species (ROS) and malondialdehyde (MDA) contents in OA group were significantly increased at the end of the experiment. Superoxide dismutase (SOD), catalase (CAT), and alkaline phosphatase (AKP) activities increased first and then decreased, glutathione peroxidase (GPX) decreased first and then increased, and GST activity changed little during the experiment. According to the result of 16S rRNA sequencing of gut microbiota, microbial-mediated functions predicted by PICRUSt showed that “Hematopoietic cell lineage,” “Endocytosis,” “Staphylococcus aureus infection,” and “Shigellosis” pathways significantly increased in OA group. The above results indicate that OA had no significant effect on growth index and molting rate but interfered with the activity of chitinolytic enzymes and ecdysone expression of juvenile horseshoe crabs, and caused oxidative stress. In addition, OA had adverse effects on the immune defense function and intestinal health. The present study reveals the potential threat of OA to T. tridentatus population and lays a foundation for the further study of the physiological adaptation mechanism of juvenile horseshoe crabs to environmental change.

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DNA Metabarcoding Revealed Interspecific Dietary Difference and Prey Selectivity in Juvenile Horseshoe Crabs Carcinoscorpius rotundicauda and Tachypleus tridentatus From Hong Kong

Frontiers in Marine Science ◽

10.3389/fmars.2021.752806 ◽

2021 ◽

Vol 8 ◽

Author(s):

Bo Yee Lee ◽

Kaori Wakabayashi ◽

Simon Yung Wa Sin ◽

Susumu Ohtsuka ◽

Ling Ming Tsang

Keyword(s):

Horseshoe Crab ◽

Trophic Ecology ◽

Food Sources ◽

Prey Selectivity ◽

Crab Species ◽

Invasive Approach ◽

Fecal Samples ◽

Tachypleus Tridentatus ◽

Dna Metabarcoding ◽

Horseshoe Crabs

Horseshoe crabs provide important ecological services including bioturbation and linking of food web in the shallow waters, but their populations are declining globally, leading to major concerns on conservation of these iconic animals. Baseline information of horseshoe crab ecology, such as their trophic role and food source, is pre-requisite for habitat protection plan and captive restocking program. Trophic ecology of Asian horseshoe crabs is relatively poorly understood and previous studies on their juveniles have suggested that they are selective feeders rather than opportunistic generalists. This study demonstrates a non-invasive approach, using DNA metabarcoding analyses of the nuclear 18S rRNA gene on fecal samples to assess the dietary compositions of Carcinoscorpius rotundicauda and Tachypleus tridentatus juveniles to (1) determine their dietary compositions and trophic roles in their ecosystem, (2) determine any prey selectivity, and (3) distinguish the interspecific dietary differences with potential implications on the habitat requirement and ecological partitioning between these two horseshoe crab species. The results based on relative read abundance (RRA) suggested that oligochaetes were the major prey items for both C. rotundicauda (41.6%) and T. tridentatus (32.4%). Bivalves and crustaceans were second major prey groups for C. rotundicauda (8.6 and 8.4%, respectively). Surprisingly, anthozoans contributed a considerable portion of T. tridentatus’s diet (22.8%), which has never been reported. Furthermore, the major prey groups identified in the fecal samples were not the dominant benthic organisms in the studied area as revealed by environmental DNA (eDNA) analyses on the sediment samples, implying that both species are selective feeders rather than dietary generalists. Significant differences observed in the dietary compositions of the two species might be partially due to the difference in habitat preference between the two species. This study provides new insights into the trophic ecology of the two Asian horseshoe crab species in the estuarine habitat and establishes a new framework for future detailed molecular dietary analyses on all developmental stages of horseshoe crabs around the world, which will allow us to evaluate the food sources needed for the survival of horseshoe crabs and facilitate future conservation strategies without killing the animals.

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The genome of horseshoe crab Tachypleus tridentatus reveals its evolutionary scenario and well-developed innate immunity

10.21203/rs.2.9427/v1 ◽

2019 ◽

Author(s):

Yan Zhou ◽

Yuan Liang ◽

Qing Yan ◽

Liang Zhang ◽

Dianbao Chen ◽

...

Keyword(s):

Innate Immunity ◽

Horseshoe Crab ◽

Divergence Time ◽

Coagulation Factor ◽

Gene Families ◽

Coagulation Cascade ◽

Last Common Ancestor ◽

Tachypleus Tridentatus ◽

Stat Signaling ◽

Horseshoe Crabs

Abstract Background Horseshoe crabs are ancient marine arthropods with a long evolutionary history. Results Here, we describe the 2.06 Gbp genome assembly of Tachypleus tridentatus with predicted 24,222 protein-coding genes. Comparative genomics show that T. tridentatus and the Atlantic horseshoe crab Limulus polyphemus had the most orthologs shared in only these two species, including genes involved in immune related Jak/STAT signaling pathway. Divergence time dating results show that the last common ancestor of Asian horseshoe crabs (including T. tridentatus) and L. polyphemus appeared at about 130 Mya (121-141) and the split of two Asian horseshoe crabs was dated to about 63 Mya (57-69). Hox gene analysis suggests two clusters in both horseshoe crab assemblies. Surprisingly, selective analysis of immune related gene families revealed high expansion of conservatively presented pattern recognition receptors. Genes involved in IMD and Jak/STAT signaling transduction pathways also exhibited a certain degree of expansion in both genomes. Intact coagulation cascade related genes were presented in T. tridentatus genome with a higher number of coagulation factor genes. Moreover, most antibacterial peptides reported have been identified in T. tridentatus with their potentially effective antimicrobial sites. Conclusions The well-developed innate immunity of T.tridentatus may affect the quality of the adaptive properties with regard to complicated marine environments.

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The draft genome of horseshoe crab Tachypleus tridentatus reveals its evolutionary scenario and well-developed innate immunity

10.21203/rs.2.9427/v3 ◽

2019 ◽

Author(s):

Yan Zhou ◽

Yuan Liang ◽

Qing Yan ◽

Liang Zhang ◽

Dianbao Chen ◽

...

Keyword(s):

Innate Immunity ◽

Horseshoe Crab ◽

Draft Genome ◽

Evolutionary Relationship ◽

Signalling Pathway ◽

Coagulation Cascade ◽

Last Common Ancestor ◽

Marine Environments ◽

Tachypleus Tridentatus ◽

Horseshoe Crabs

Abstract Background: Horseshoe crabs are ancient marine arthropods with a long evolutionary history extending back approximately 450 million years that rely entirely on their innate immune system and have developed multiple defence systems. However, the genetic mechanisms underlying their abilities of distinguishing and defending against invading microbes are still unclear. Results: Here, we describe the 2.06 Gbp genome assembly of Tachypleus tridentatus with 24,222 predicted protein-coding genes. Comparative genomics shows that T. tridentatus and the Atlantic horseshoe crab Limulus polyphemus have the most orthologues shared among two species, including genes involved in the immune-related JAK-STAT signalling pathway. Divergence time dating results show that the last common ancestor of Asian horseshoe crabs (including T. tridentatus) and L. polyphemus appeared approximately 130 Mya (121-141), and the split of the two Asian horseshoe crabs was dated to approximately 63 Mya (57-69). Hox gene analysis suggests two clusters in both horseshoe crab assemblies. Surprisingly, selective analysis of immune-related gene families revealed the high expansion of conserved pattern recognition receptors. Genes involved in the IMD and JAK-STAT signal transduction pathways also exhibited a certain degree of expansion in both genomes. Intact coagulation cascade-related genes were present in the T. tridentatus genome with a higher number of coagulation factor genes. Moreover, most reported antibacterial peptides have been identified in T. tridentatus with their potentially effective antimicrobial sites. Conclusions: The draft genome of T. tridentatus would provide an important source for eliminating the uncertainty in the evolutionary relationship of Chelicerata. The expansion of conserved immune signalling pathway genes, coagulation factors and intact antimicrobial peptides in T. tridentatus constitutes its robust and effective innate immunity for self-defence in marine environments with an enormous number of invading pathogens and may affect the quality of the adaptive properties with regard to complicated marine environments.

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The draft genome of horseshoe crab Tachypleus tridentatus reveals its evolutionary scenario and well-developed innate immunity

10.21203/rs.2.9427/v4 ◽

2019 ◽

Author(s):

Yan Zhou ◽

Yuan Liang ◽

Qing Yan ◽

Liang Zhang ◽

Dianbao Chen ◽

...

Keyword(s):

Innate Immunity ◽

Horseshoe Crab ◽

Draft Genome ◽

Evolutionary Relationship ◽

Signalling Pathway ◽

Coagulation Cascade ◽

Last Common Ancestor ◽

Marine Environments ◽

Tachypleus Tridentatus ◽

Horseshoe Crabs

Abstract Background: Horseshoe crabs are ancient marine arthropods with a long evolutionary history extending back approximately 450 million years, which may benefit from their innate immune systems. However, the genetic mechanisms underlying their abilities of distinguishing and defending against invading microbes are still unclear. Results: Here, we describe the 2.06 Gbp genome assembly of Tachypleus tridentatus with 24,222 predicted protein-coding genes. Comparative genomics shows that T. tridentatus and the Atlantic horseshoe crab Limulus polyphemus have the most orthologues shared among two species, including genes involved in the immune-related JAK-STAT signalling pathway. Divergence time dating results show that the last common ancestor of Asian horseshoe crabs (including T. tridentatus and C. rotundicauda) and L. polyphemus appeared approximately 130 Mya (121-141), and the split of the two Asian horseshoe crabs was dated to approximately 63 Mya (57-69). Hox gene analysis suggests two clusters in both horseshoe crab assemblies. Surprisingly, selective analysis of immune-related gene families revealed the high expansion of conserved pattern recognition receptors. Genes involved in the IMD and JAK-STAT signal transduction pathways also exhibited a certain degree of expansion in both genomes. Intact coagulation cascade-related genes were present in the T. tridentatus genome with a higher number of coagulation factor genes. Moreover, most reported antibacterial peptides have been identified in T. tridentatus with their potentially effective antimicrobial sites. Conclusions: The draft genome of T. tridentatus would would provide important evidence for further clarifying the taxonomy and evolutionary relationship of Chelicerata. The expansion of conserved immune signalling pathway genes, coagulation factors and intact antimicrobial peptides in T. tridentatus constitutes its robust and effective innate immunity for self-defence in marine environments with an enormous number of invading pathogens and may affect the quality of the adaptive properties with regard to complicated marine environments.

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The draft genome of horseshoe crab Tachypleus tridentatus reveals its evolutionary scenario and well-developed innate immunity

10.21203/rs.2.9427/v5 ◽

2020 ◽

Author(s):

Yan Zhou ◽

Yuan Liang ◽

Qing Yan ◽

Liang Zhang ◽

Dianbao Chen ◽

...

Keyword(s):

Innate Immunity ◽

Horseshoe Crab ◽

Draft Genome ◽

Evolutionary Relationship ◽

Signalling Pathway ◽

Coagulation Cascade ◽

Last Common Ancestor ◽

Marine Environments ◽

Tachypleus Tridentatus ◽

Horseshoe Crabs

Abstract Background: Horseshoe crabs are ancient marine arthropods with a long evolutionary history extending back approximately 450 million years, which may benefit from their innate immune systems. However, the genetic mechanisms underlying their abilities of distinguishing and defending against invading microbes are still unclear. Results: Here, we describe the 2.06 Gbp genome assembly of Tachypleus tridentatus with 24,222 predicted protein-coding genes. Comparative genomics shows that T. tridentatus and the Atlantic horseshoe crab Limulus polyphemus have the most orthologues shared among two species, including genes involved in the immune-related JAK-STAT signalling pathway. Divergence time dating results show that the last common ancestor of Asian horseshoe crabs (including T. tridentatus and C. rotundicauda ) and L. polyphemus appeared approximately 130 Mya (121-141), and the split of the two Asian horseshoe crabs was dated to approximately 63 Mya (57-69). Hox gene analysis suggests two clusters in both horseshoe crab assemblies. Surprisingly, selective analysis of immune-related gene families revealed the high expansion of conserved pattern recognition receptors. Genes involved in the IMD and JAK-STAT signal transduction pathways also exhibited a certain degree of expansion in both genomes. Intact coagulation cascade-related genes were present in the T. tridentatus genome with a higher number of coagulation factor genes. Moreover, most reported antibacterial peptides have been identified in T. tridentatus with their potentially effective antimicrobial sites. Conclusions: The draft genome of T. tridentatus would would provide important evidence for further clarifying the taxonomy and evolutionary relationship of Chelicerata. The expansion of conserved immune signalling pathway genes, coagulation factors and intact antimicrobial peptides in T. tridentatus constitutes its robust and effective innate immunity for self-defence in marine environments with an enormous number of invading pathogens and may affect the quality of the adaptive properties with regard to complicated marine environments.

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The draft genome of horseshoe crab Tachypleus tridentatus reveals its evolutionary scenario and well-developed innate immunity

10.21203/rs.2.9427/v6 ◽

2020 ◽

Author(s):

Yan Zhou ◽

Yuan Liang ◽

Qing Yan ◽

Liang Zhang ◽

Dianbao Chen ◽

...

Keyword(s):

Innate Immunity ◽

Horseshoe Crab ◽

Draft Genome ◽

Evolutionary Relationship ◽

Signalling Pathway ◽

Coagulation Cascade ◽

Last Common Ancestor ◽

Marine Environments ◽

Tachypleus Tridentatus ◽

Horseshoe Crabs

Abstract Background: Horseshoe crabs are ancient marine arthropods with a long evolutionary history extending back approximately 450 million years, which may benefit from their innate immune systems. However, the genetic mechanisms underlying their abilities of distinguishing and defending against invading microbes are still unclear. Results: Here, we describe the 2.06 Gbp genome assembly of Tachypleus tridentatus with 24,222 predicted protein-coding genes. Comparative genomics shows that T. tridentatus and the Atlantic horseshoe crab Limulus polyphemus have the most orthologues shared among two species, including genes involved in the immune-related JAK-STAT signalling pathway. Divergence time dating results show that the last common ancestor of Asian horseshoe crabs (including T. tridentatus and C. rotundicauda ) and L. polyphemus appeared approximately 130 Mya (121-141), and the split of the two Asian horseshoe crabs was dated to approximately 63 Mya (57-69). Hox gene analysis suggests two clusters in both horseshoe crab assemblies. Surprisingly, selective analysis of immune-related gene families revealed the high expansion of conserved pattern recognition receptors. Genes involved in the IMD and JAK-STAT signal transduction pathways also exhibited a certain degree of expansion in both genomes. Intact coagulation cascade-related genes were present in the T. tridentatus genome with a higher number of coagulation factor genes. Moreover, most reported antibacterial peptides have been identified in T. tridentatus with their potentially effective antimicrobial sites. Conclusions: The draft genome of T. tridentatus would would provide important evidence for further clarifying the taxonomy and evolutionary relationship of Chelicerata. The expansion of conserved immune signalling pathway genes, coagulation factors and intact antimicrobial peptides in T. tridentatus constitutes its robust and effective innate immunity for self-defence in marine environments with an enormous number of invading pathogens and may affect the quality of the adaptive properties with regard to complicated marine environments.

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