Drivers of species knowledge across the tree of life.

Knowledge of biodiversity is unevenly distributed across the Tree of Life. In the long run, such disparity in awareness unbalances our understanding of life on Earth, influencing policy decisions and the allocation of research and conservation funding. We investigated how humans accumulate knowledge of biodiversity by searching for consistent relationships between scientific (number of publications) and societal (number of views in Wikipedia) interest, and species-level morphological, ecological, and sociocultural factors. Across a random selection of 3019 species spanning 29 Phyla/Divisions, we show that sociocultural factors are the most important correlates of scientific and societal interest in biodiversity, including the fact that a species is useful or harmful to humans, has a common name, and is listed in the International Union for Conservation of Nature Red List. Furthermore, large-bodied, broadly distributed, and taxonomically unique species receive more scientific and societal attention, whereas colorfulness and phylogenetic proximity to humans correlate exclusively with societal attention. These results highlight a favoritism toward limited branches of the Tree of Life, and that scientific and societal priorities in biodiversity research broadly align. This suggests that we may be missing out on key species in our research and conservation agenda simply because they are not on our cultural radar.

Morphological and molecular study of Syllinae (Annelida, Syllidae) from Bermuda, with the description of five new species.

Although polychaetes from the Bermuda Archipelago have been studied since the beginning of the twentieth century, syllids have been particularly neglected in this area, which is surprising considering this family is usually a dominant group in marine benthic ecosystems. To fill this knowledge gap, we have carried out an extensive analysis of Bermudan Syllidae, combining morphological and molecular data including four nuclear and mitochondrial markers (cytochrome c oxidase subunit I, 18S rRNA, 16S rRNA and 28S rRNA). We have identified and established the phylogenetic position of five new species, Haplosyllis anitae n. sp., Haplosyllis guillei n. sp., Haplosyllis larsi n. sp., Haplosyllis vassiae n. sp. and Syllis laiae n. sp., together with Haplosyllis cf. cephalata. Overall, our results extend the knowledge on the diversity of Syllidae in Bermuda, increasing the number of species present in the area to 25. Our results also recover Opisthosyllis and Syllis as non-monophyletic genera, for which traditional diagnostic morphological features do not accurately reflect their evolutionary histories, and thus we propose that these groups should be reorganized based on molecular characters.

Distinct patterns of gene expression during regeneration and asexual reproduction in the annelid Pristina leidyi.

Regeneration, the ability to replace lost body parts, is a widespread phenomenon in the animal kingdom often connected to asexual reproduction or fission, since the only difference between the two appears to be the stimulus that triggers them. Both developmental processes have largely been characterized; however, the molecular toolkit and genetic mechanisms underlying these events remain poorly unexplored. Annelids, in particular the oligochaete Pristina leidyi, provide a good model system to investigate these processes as they show diverse ways to regenerate, and can reproduce asexually through fission under laboratory conditions. Here, we used a comparative transcriptomics approach based on RNA-sequencing and differential gene expression analyses to understand the molecular mechanisms involved in anterior regeneration and asexual reproduction. We found 291 genes upregulated during anterior regeneration, including several regeneration-related genes previously reported in other annelids such as frizzled, paics, and vdra. On the other hand, during asexual reproduction, 130 genes were found upregulated, and unexpectedly, many of them were related to germline development during sexual reproduction. We also found important differences between anterior regeneration and asexual reproduction, with the latter showing a gene expression profile more similar to that of control individuals. Nevertheless, we identified 35 genes that were upregulated in both conditions, many of them related to cell pluripotency, stem cells, and cell proliferation. Overall, our results shed light on the molecular mechanisms that control anterior regeneration and asexual reproduction in annelids and reveal similarities with other animals, suggesting that the genetic machinery controlling these processes is conserved across metazoans.

Modern venomics-Current insights, novel methods, and future perspectives in biological and applied animal venom research.

Venoms have evolved >100 times in all major animal groups, and their components, known as toxins, have been fine-tuned over millions of years into highly effective biochemical weapons. There are many outstanding questions on the evolution of toxin arsenals, such as how venom genes originate, how venom contributes to the fitness of venomous species, and which modifications at the genomic, transcriptomic, and protein level drive their evolution. These questions have received particularly little attention outside of snakes, cone snails, spiders, and scorpions. Venom compounds have further become a source of inspiration for translational research using their diverse bioactivities for various applications. We highlight here recent advances and new strategies in modern venomics and discuss how recent technological innovations and multi-omic methods dramatically improve research on venomous animals. The study of genomes and their modifications through CRISPR and knockdown technologies will increase our understanding of how toxins evolve and which functions they have in the different ontogenetic stages during the development of venomous animals. Mass spectrometry imaging combined with spatial transcriptomics, in situ hybridization techniques, and modern computer tomography gives us further insights into the spatial distribution of toxins in the venom system and the function of the venom apparatus. All these evolutionary and biological insights contribute to more efficiently identify venom compounds, which can then be synthesized or produced in adapted expression systems to test their bioactivity. Finally, we critically discuss recent agrochemical, pharmaceutical, therapeutic, and diagnostic (so-called translational) aspects of venoms from which humans benefit.

Evolution, Expression Patterns, and Distribution of Novel Ribbon Worm Predatory and Defensive Toxins.

Ribbon worms are active predators that use an eversible proboscis to inject venom into their prey and defend themselves with toxic epidermal secretions. Previous work on nemertean venom has largely focused on just a few species and has not investigated the different predatory and defensive secretions in detail. Consequently, our understanding of the composition and evolution of ribbon worm venoms is still very limited. Here, we present a comparative study of nemertean venom combining RNA-seq differential gene expression analyses of venom-producing tissues, tandem mass spectrometry-based proteomics of toxic secretions, and mass spectrometry imaging of proboscis sections, to shed light onto the composition and evolution of predatory and defensive toxic secretions in Antarctonemertes valida. Our analyses reveal a wide diversity of putative defensive and predatory toxins with tissue-specific gene expression patterns and restricted distributions to the mucus and proboscis proteomes respectively, suggesting that ribbon worms produce distinct toxin cocktails for predation and defense. Our results also highlight the presence of numerous lineage-specific toxins, indicating that venom evolution is highly divergent across nemerteans, producing toxin cocktails that might be finely tuned to subdue different prey. Our data also suggest that the hoplonemertean proboscis is a highly specialized predatory organ that seems to be involved in a variety of biological functions besides predation, including secretion and sensory perception. Overall, our results advance our knowledge into the diversity and evolution of nemertean venoms and highlight the importance of combining different types of data to characterize toxin composition in understudied venomous organisms.

How to thrive in unstable environments: Gene expression profile of a riparian earthworm under abiotic stress.

Nowadays, extreme weather events caused by climate change are becoming more frequent. This leads to the occurrence of extreme habitats to which species must adapt. This challenge becomes crucial for species living in unstable environments, such as the riparian earthworm Eiseniella tetraedra. Its cosmopolitan distribution exposes it to various environmental changes, such as freezing in subarctic regions or droughts in Mediterranean areas. Transcriptional changes under cold and desiccation conditions could therefore shed light on the adaptive mechanisms of this species. An experiment was performed for each condition. In the cold experiment, the temperature was lowered to -14 °C ± 2 °C (compared to 8 °C for control samples), and in the desiccation treatment, humidity was lowered from 60% to 15%. Comparisons of gene expression levels between earthworms under freezing conditions and control earthworms revealed a total of 84 differentially expressed genes and comparisons between the desiccation experiment and the control yielded 163 differentially expressed genes. However, no common responses were found between the two treatments. The results suggest that E. tetraedra can acclimate to low temperatures due to the upregulation of genes involved in glucose accumulation. However, downregulation of the respiratory chain suggests that this earthworm does not tolerate freezing conditions. Under desiccation conditions, genes involved in cell protection from apoptosis and DNA repair were upregulated. In contrast, lipid metabolism was downregulated, presumably to conserve resources by reducing the rate at which they are consumed.

The new COST Action European Venom Network (EUVEN)-synergy and future perspectives of modern venomics.

Venom research is a highly multidisciplinary field that involves multiple subfields of biology, informatics, pharmacology, medicine, and other areas. These different research facets are often technologically challenging and pursued by different teams lacking connection with each other. This lack of coordination hampers the full development of venom investigation and applications. The COST Action CA19144-European Venom Network was recently launched to promote synergistic interactions among different stakeholders and foster venom research at the European level.

Delegating Sex: Differential Gene Expression in Stolonizing Syllids Uncovers the Hormonal Control of Reproduction.

Stolonization in syllid annelids is a unique mode of reproduction among animals. During the breeding season, a structure resembling the adult but containing only gametes, called stolon, is formed generally at the posterior end of the animal. When stolons mature, they detach from the adult and gametes are released into the water column. The process is synchronized within each species, and it has been reported to be under environmental and endogenous control, probably via endocrine regulation. To further understand reproduction in syllids and to elucidate the molecular toolkit underlying stolonization, we generated Illumina RNA-seq data from different tissues of reproductive and nonreproductive individuals of Syllis magdalena and characterized gene expression during the stolonization process. Several genes involved in gametogenesis (ovochymase, vitellogenin, testis-specific serine/threonine-kinase), immune response (complement receptor 2), neuronal development (tyrosine-protein kinase Src42A), cell proliferation (alpha-1D adrenergic receptor), and steroid metabolism (hydroxysteroid dehydrogenase 2) were found differentially expressed in the different tissues and conditions analyzed. In addition, our findings suggest that several neurohormones, such as methyl farnesoate, dopamine, and serotonin, might trigger stolon formation, the correct maturation of gametes and the detachment of stolons when gametogenesis ends. The process seems to be under circadian control, as indicated by the expression patterns of r-opsins. Overall, our results shed light into the genes that orchestrate the onset of gamete formation and improve our understanding of how some hormones, previously reported to be involved in reproduction and metamorphosis processes in other invertebrates, seem to also regulate reproduction via stolonization.

Beach to Bench to Bedside: Marine Invertebrate Biochemical Adaptations and Their Applications in Biotechnology and Biomedicine.

The ocean covers more than 70% of the surface of the planet and harbors very diverse ecosystems ranging from tropical coral reefs to the deepest ocean trenches, with some of the most extreme conditions of pressure, temperature, and light. Organisms living in these environments have been subjected to strong selective pressures through millions of years of evolution, resulting in a plethora of remarkable adaptations that serve a variety of vital functions. Some of these adaptations, including venomous secretions and light-emitting compounds or ink, represent biochemical innovations in which marine invertebrates have developed novel and unique bioactive compounds with enormous potential for basic and applied research. Marine biotechnology, defined as the application of science and technology to marine organisms for the production of knowledge, goods, and services, can harness the enormous possibilities of these unique bioactive compounds acting as a bridge between biological knowledge and applications. This chapter highlights some of the most exceptional biochemical adaptions found specifically in marine invertebrates and describes the biotechnological and biomedical applications derived from them to improve the quality of human life.

Are Fireworms Venomous? Evidence for the Convergent Evolution of Toxin Homologs in Three Species of Fireworms (Annelida, Amphinomidae).

Amphinomids, more commonly known as fireworms, are a basal lineage of marine annelids characterized by the presence of defensive dorsal calcareous chaetae, which break off upon contact. It has long been hypothesized that amphinomids are venomous and use the chaetae to inject a toxic substance. However, studies investigating fireworm venom from a morphological or molecular perspective are scarce and no venom gland has been identified to date, nor any toxin characterized at the molecular level. To investigate this question, we analyzed the transcriptomes of three species of fireworms-Eurythoe complanata, Hermodice carunculata, and Paramphinome jeffreysii-following a venomics approach to identify putative venom compounds. Our venomics pipeline involved de novo transcriptome assembly, open reading frame, and signal sequence prediction, followed by three different homology search strategies: BLAST, HMMER sequence, and HMMER domain. Following this pipeline, we identified 34 clusters of orthologous genes, representing 13 known toxin classes that have been repeatedly recruited into animal venoms. Specifically, the three species share a similar toxin profile with C-type lectins, peptidases, metalloproteinases, spider toxins, and CAP proteins found among the most highly expressed toxin homologs. Despite their great diversity, the putative toxins identified are predominantly involved in three major biological processes: hemostasis, inflammatory response, and allergic reactions, all of which are commonly disrupted after fireworm stings. Although the putative fireworm toxins identified here need to be further validated, our results strongly suggest that fireworms are venomous animals that use a complex mixture of toxins for defense against predators.

Glowing Worms: Biological, Chemical, and Functional Diversity of Bioluminescent Annelids.

Bioluminescence, the ability to produce light by living organisms, has evolved independently in numerous lineages across the tree of life. Luminous forms are found in a wide range of taxonomic groups from bacteria to vertebrates, although the great majority of bioluminescent organisms are marine taxa. Within the phylum Annelida, bioluminescence is widespread, present in at least 98 terrestrial and marine species that represent 45 genera distributed in thirteen lineages of clitellates and polychaetes. The ecological diversity of luminous annelids is unparalleled, with species occupying a great variety of habitats including both terrestrial and marine ecosystems, from coastal waters to the deep-sea, in benthic and pelagic habitats from polar to tropical regions. This great taxonomic and ecological diversity is matched by the wide array of bioluminescent colors-including yellow light, which is very rare among marine taxa-different emission wavelengths even between species of the same genus, and varying patterns, chemical reactions and kinetics. This diversity of bioluminescence colors and patterns suggests that light production in annelids might be involved in a variety of different functions, including defensive mechanisms like sacrificial lures or aposematic signals, and intraspecific communication systems. In this review, we explore the world of luminous annelids, particularly focusing on the current knowledge regarding their taxonomic and ecological diversity and discussing the putative functions and chemistries of their bioluminescent systems.

From Mollusks to Medicine: A Venomics Approach for the Discovery and Characterization of Therapeutics from Terebridae Peptide Toxins.

Animal venoms comprise a diversity of peptide toxins that manipulate molecular targets such as ion channels and receptors, making venom peptides attractive candidates for the development of therapeutics to benefit human health. However, identifying bioactive venom peptides remains a significant challenge. In this review we describe our particular venomics strategy for the discovery, characterization, and optimization of Terebridae venom peptides, teretoxins. Our strategy reflects the scientific path from mollusks to medicine in an integrative sequential approach with the following steps: (1) delimitation of venomous Terebridae lineages through taxonomic and phylogenetic analyses; (2) identification and classification of putative teretoxins through omics methodologies, including genomics, transcriptomics, and proteomics; (3) chemical and recombinant synthesis of promising peptide toxins; (4) structural characterization through experimental and computational methods; (5) determination of teretoxin bioactivity and molecular function through biological assays and computational modeling; (6) optimization of peptide toxin affinity and selectivity to molecular target; and (7) development of strategies for effective delivery of venom peptide therapeutics. While our research focuses on terebrids, the venomics approach outlined here can be applied to the discovery and characterization of peptide toxins from any venomous taxa.

Nature's Palette: Characterization of Shared Pigments in Colorful Avian and Mollusk Shells.

Pigment-based coloration is a common trait found in a variety of organisms across the tree of life. For example, calcareous avian eggs are natural structures that vary greatly in color, yet just a handful of tetrapyrrole pigment compounds are responsible for generating this myriad of colors. To fully understand the diversity and constraints shaping nature's palette, it is imperative to characterize the similarities and differences in the types of compounds involved in color production across diverse lineages. Pigment composition was investigated in eggshells of eleven paleognath bird taxa, covering several extinct and extant lineages, and shells of four extant species of mollusks. Birds and mollusks are two distantly related, calcareous shell-building groups, thus characterization of pigments in their calcareous structures would provide insights to whether similar compounds are found in different phyla (Chordata and Mollusca). An ethylenediaminetetraacetic acid (EDTA) extraction protocol was used to analyze the presence and concentration of biliverdin and protoporphyrin, two known and ubiquitous tetrapyrrole avian eggshell pigments, in all avian and molluscan samples. Biliverdin was solely detected in birds, including the colorful eggshells of four tinamou species. In contrast, protoporphyrin was detected in both the eggshells of several avian species and in the shells of all mollusks. These findings support previous hypotheses about the ubiquitous deposition of tetrapyrroles in the eggshells of various bird lineages and provide evidence for its presence also across distantly related animal taxa.

Transcriptome sequencing and annotation of the polychaete Hermodice carunculata (Annelida, Amphinomidae).

BACKGROUND: The amphinomid polychaete Hermodice carunculata is a cosmopolitan and ecologically important omnivore in coral reef ecosystems, preying on a diverse suite of reef organisms and potentially acting as a vector for coral disease. While amphinomids are a key group for determining the root of the Annelida, their phylogenetic position has been difficult to resolve, and their publically available genomic data was scarce. RESULTS: We performed deep transcriptome sequencing (Illumina HiSeq) and profiling on Hermodice carunculata collected in the Western Atlantic Ocean. We focused this study on 58,454 predicted Open Reading Frames (ORFs) of genes longer than 200 amino acids for our homology search, and Gene Ontology (GO) terms and InterPro IDs were assigned to 32,500 of these ORFs. We used this de novo assembled transcriptome to recover major signaling pathways and housekeeping genes. We also identify a suite of H. carunculata genes related to reproduction and immune response. CONCLUSIONS: We provide a comprehensive catalogue of annotated genes for Hermodice carunculata and expand the knowledge of reproduction and immune response genes in annelids, in general. Overall, this study vastly expands the available genomic data for H. carunculata, of which previously consisted of only 279 nucleotide sequences in NCBI. This underscores the utility of Illumina sequencing for de novo transcriptome assembly in non-model organisms as a cost-effective and efficient tool for gene discovery and downstream applications, such as phylogenetic analysis and gene expression profiling.

Molecular Diversity and Gene Evolution of the Venom Arsenal of Terebridae Predatory Marine Snails.

Venom peptides from predatory organisms are a resource for investigating evolutionary processes such as adaptive radiation or diversification, and exemplify promising targets for biomedical drug development. Terebridae are an understudied lineage of conoidean snails, which also includes cone snails and turrids. Characterization of cone snail venom peptides, conotoxins, has revealed a cocktail of bioactive compounds used to investigate physiological cellular function, predator-prey interactions, and to develop novel therapeutics. However, venom diversity of other conoidean snails remains poorly understood. The present research applies a venomics approach to characterize novel terebrid venom peptides, teretoxins, from the venom gland transcriptomes of Triplostephanus anilis and Terebra subulata. Next-generation sequencing and de novo assembly identified 139 putative teretoxins that were analyzed for the presence of canonical peptide features as identified in conotoxins. To meet the challenges of de novo assembly, multiple approaches for cross validation of findings were performed to achieve reliable assemblies of venom duct transcriptomes and to obtain a robust portrait of Terebridae venom. Phylogenetic methodology was used to identify 14 teretoxin gene superfamilies for the first time, 13 of which are unique to the Terebridae. Additionally, basic local algorithm search tool homology-based searches to venom-related genes and posttranslational modification enzymes identified a convergence of certain venom proteins, such as actinoporin, commonly found in venoms. This research provides novel insights into venom evolution and recruitment in Conoidean predatory marine snails and identifies a plethora of terebrid venom peptides that can be used to investigate fundamental questions pertaining to gene evolution.

The cuticle modulates ultraviolet reflectance of avian eggshells.

Avian eggshells are variedly coloured, yet only two pigments, biliverdin and protoporphyrin IX, are known to contribute to the dramatic diversity of their colours. By contrast, the contributions of structural or other chemical components of the eggshell are poorly understood. For example, unpigmented eggshells, which appear white to the human eye, vary in their ultraviolet (UV) reflectance, which may be detectable by birds. We investigated the proximate mechanisms for the variation in UV-reflectance of unpigmented bird eggshells using spectrophotometry, electron microscopy, chemical analyses, and experimental manipulations. We specifically tested how UV-reflectance is affected by the eggshell cuticle, the outermost layer of most avian eggshells. The chemical dissolution of the outer eggshell layers, including the cuticle, increased UV-reflectance for only eggshells that contained a cuticle. Our findings demonstrate that the outer eggshell layers, including the cuticle, absorb UV-light, probably because they contain higher levels of organic components and other chemicals, such as calcium phosphates, compared to the predominantly calcite-based eggshell matrix. These data highlight the need to examine factors other than the known pigments in studies of avian eggshell colour.