Distinct gene expression dynamics in developing and regenerating crustacean limbs.

Regenerating animals have the ability to reproduce body parts that were originally made in the embryo and subsequently lost due to injury. Understanding whether regeneration mirrors development is an open question in most regenerative species. Here, we take a transcriptomics approach to examine whether leg regeneration shows similar temporal patterns of gene expression as leg development in the embryo, in the crustacean Parhyale hawaiensis. We find that leg development in the embryo shows stereotypic temporal patterns of gene expression. In contrast, the dynamics of gene expression during leg regeneration show a higher degree of variation related to the physiology of individual animals. A major driver of this variation is the molting cycle. We dissect the transcriptional signals of individual physiology and regeneration to obtain clearer temporal signals marking distinct phases of leg regeneration. Comparing the transcriptional dynamics of development and regeneration we find that, although the two processes use similar sets of genes, the temporal patterns in which these genes are deployed are different and cannot be systematically aligned.

The mitochondrial genome of Bottapotamon fukienense (Brachiura: Potamidae) is fragmented into two chromosomes.

BACKGROUND: China is the hotspot of global freshwater crab diversity, but their wild populations are facing severe pressures associated with anthropogenic factors, necessitating the need to map their taxonomic and genetic diversity and design conservation policies. RESULTS: Herein, we sequenced the mitochondrial genome of a Chinese freshwater crab species Bottapotamon fukienense, and found that it is fragmented into two chromosomes. We confirmed that fragmentation was not limited to a single specimen or population. Chromosome 1 comprised 15,111 base pairs (bp) and there were 26 genes and one pseudogene (pseudo-nad1) encoded on it. Chromosome 2 comprised 8,173 bp and there were 12 genes and two pseudogenes (pseudo-trnL2 and pseudo-rrnL) encoded on it. Combined, they comprise the largest mitogenome (23,284 bp) among the Potamidae. Bottapotamon was the only genus in the Potamidae dataset exhibiting rearrangements of protein-coding genes. Bottapotamon fukienense exhibited average rates of sequence evolution in the dataset and did not differ in selection pressures from the remaining Potamidae. CONCLUSIONS: This is the first experimentally confirmed fragmentation of a mitogenome in crustaceans. While the mitogenome of B. fukienense exhibited multiple signs of elevated mitogenomic architecture evolution rates, including the exceptionally large size, duplicated genes, pseudogenisation, rearrangements of protein-coding genes, and fragmentation, there is no evidence that this is matched by elevated sequence evolutionary rates or changes in selection pressures.

Reconstruction of Eriocheir sinensis Protein-Protein Interaction Network Based on DGO-SVM Method.

Eriocheir sinensis is an economically important aquatic animal. Its regulatory mechanisms underlying many biological processes are still vague due to the lack of systematic analysis tools. The protein-protein interaction network (PIN) is an important tool for the systematic analysis of regulatory mechanisms. In this work, a novel machine learning method, DGO-SVM, was applied to predict the protein-protein interaction (PPI) in E. sinensis, and its PIN was reconstructed. With the domain, biological process, molecular functions and subcellular locations of proteins as the features, DGO-SVM showed excellent performance in Bombyx mori, humans and five aquatic crustaceans, with 92-96% accuracy. With DGO-SVM, the PIN of E. sinensis was reconstructed, containing 14,703 proteins and 7,243,597 interactions, in which 35,604 interactions were associated with 566 novel proteins mainly involved in the response to exogenous stimuli, cellular macromolecular metabolism and regulation. The DGO-SVM demonstrated that the biological process, molecular functions and subcellular locations of proteins are significant factors for the precise prediction of PPIs. We reconstructed the largest PIN for E. sinensis, which provides a systematic tool for the regulatory mechanism analysis. Furthermore, the novel-protein-related PPIs in the PIN may provide important clues for the mechanism analysis of the underlying specific physiological processes in E. sinensis.

Molecular cloning, functional characterization and differential expression of two novel GABAAR-like subunits from red swamp crayfish Procambarus clarkii.

In this work, we cloned and functionally expressed two novel GABAA receptor subunits from Procambarus clarkii crayfish. These two new subunits, PcGABAA-α and PcGABAA-ß2, revealed significant sequence homology with the PcGABAA-ß subunit, previously identified in our laboratory. In addition, PcGABAA-α subunit also shared a significant degree of identity with the Drosophila melanogaster genes DmGRD (GABA and glycine-like receptor subunits of Drosophila) as well as PcGABAA-ß2 subunit with DmLCCH3 (ligand-gated chloride channel homolog 3). Electrophysiological recordings showed that the expression in HEK cells of the novel subunits, either alone or in combination, failed to form functional homo- or heteromeric receptors. However, the co-expression of PcGABAA-α with PcGABAA-ß evoked sodium- or chloride-dependent currents that accurately reproduced the time course of the GABA-evoked currents in the X-organ neurons from crayfish, suggesting that these GABA subunits combine to form two types of GABA receptors, one with cationic selectivity filter and the other preferentially permeates anions. On the other hand, PcGABAA-ß2 and PcGABAA-ß co-expression generated a chloride current that does not show desensitization. Muscimol reproduced the time course of GABA-evoked currents in all functional receptors, and picrotoxin blocked these currents; bicuculline did not block any of the recorded currents. Reverse transcription polymerae chain reaction (RT-PCR) amplifications and FISH revealed that PcGABAA-α and PcGABAA-ß2 are predominantly expressed in the crayfish nervous system. Altogether, these findings provide the first evidence of a neural GABA-gated cationic channel in the crayfish, increasing our understanding of the role of these new GABAA receptor subunits in native heteromeric receptors.

Characterization of a novel and testis-specific zinc finger protein during sexual development of Pacific white shrimp Litopenaeus vannamei.

Since females grow faster in penaeid shrimp, all-female aquaculture was proposed. Environmental conditions in the Pacific white shrimp did not found to affect genetic sex determination (ZZ/ZW system). The androgenic gland (AG)-secreting insulin-like androgenic gland hormone (IAG) is a key controlling factor in crustacean male differentiation. However, functional sex reversal (neo-male) in penaeid shrimp has not yet been achieved by manipulating the IAG-sexual switch. Therefore, understanding the molecular mechanisms of gonadal differentiation may help build appropriate tools to generate neo-male (ZW) for all-female breeding. This study describes the potential role of the novel penaeid-specific testicular zinc finger protein (pTZFP) in the gonads of Pacific white shrimp. First, pTZFP transcripts show a male-bias expression pattern in undifferentiated gonads, which is then exclusively expressed in the testis and absent or slightly expressed in the ovary and other tissues. Besides, the knockdown of pTZFP in undifferentiated males results in smaller testes but no sex reversal. Immunohistochemical (IHC) staining of proliferating cell nuclear antigen (PCNA) further confirmed that the smaller testes in pTZFP-deficient males are due to the lower proliferating activity of spermatogonia. These data reveal that pTZFP may be involved in testicular development but have fewer effects on gonadal differentiation. Moreover, testicular pTZFP transcription levels were not reduced with estradiol-17ß (E2) administration or AG excision. Therefore, our data suggest that pTZFP may regulate testicular development through downstream genes regulating spermatogonia proliferation. Moreover, our data provide an appropriate molecular marker for identifying the sex of undifferentiated gonads.

Anatomical and molecular insights into the antennal gland of the giant freshwater prawn Macrobrachium rosenbergii.

In this study, the complex organization of the AnG in the giant freshwater prawn Macrobrachium rosenbergii was revealed using various techniques, including conventional histology, histochemistry, scanning electron microscopy, and X-ray tomography. The results showed the diversity of cells in the AnG and the detailed organization of the labyrinth's tubule into four radiated areas from the central to peripheral zones. The study also demonstrated the expression of some vertebrate kidney-associated homolog genes, aquaporin (AQP), solute carrier family 22 (SLC-22), nephrin, and uromodulin, in the AnG by qPCR. The result of in situ hybridization further showed the localization of SLC-22 and AQP transcript in the bladder and labyrinth's epithelium, specifically in regions 2, 3, and 4. Additionally, the study revealed neuropeptide expressions in the AnG by qPCR and in situ hybridization, i.e., crustacean hyperglycemic hormone (CHH) and molt inhibiting hormone (MIH), implying that the AnG may have a role in hormone production. Moreover, male and female prawns exhibited different levels of AQP, SLC-22, nephrin, and CHH expressions during the premolt and intermolt stages, suggesting a crucial role relevant to the molting stages. In conclusion, this study clarified the complex structure of the AnG in M. rosenbergii and demonstrated for the first time the expression of vertebrate kidney-associated genes and the possible endocrine role of the AnG. Further investigation is needed to clarify the role of these genes, particularly during ecdysis. The implications of these findings could significantly advance our understanding of the AnG in decapod crustaceans.

Diving into broad-scale and high-resolution population genomics to decipher drivers of structure and climatic vulnerability in a marine invertebrate.

Species with widespread distributions play a crucial role in our understanding of climate change impacts on population structure. In marine species, population structure is often governed by both high connectivity potential and selection across strong environmental gradients. Despite the complexity of factors influencing marine populations, studying species with broad distribution can provide valuable insights into the relative importance of these factors and the consequences of climate-induced alterations across environmental gradients. We used the northern shrimp Pandalus borealis and its wide latitudinal distribution to identify current drivers of population structure and predict the species' vulnerability to climate change. A total of 1514 individuals sampled across 24° latitude were genotyped at high geographic (54 stations) and genetic (14,331 SNPs) resolutions to assess genetic variation and environmental correlations. Four populations were identified in addition to finer substructure associated with local adaptation. Geographic patterns of neutral population structure reflected predominant oceanographic currents, while a significant proportion of the genetic variation was associated with gradients in salinity and temperature. Adaptive landscapes generated using climate projections suggest a larger genomic offset in the southern extent of the P. borealis range, where shrimp had the largest adaptive standing genetic variation. Our genomic results combined with recent observations point to further deterioration in southern regions and an impending vulnerable status in the regions at higher latitudes for P. borealis. They also provide rare insights into the drivers of population structure and climatic vulnerability of a widespread meroplanktonic species, which is crucial to understanding future challenges associated with invertebrates essential to ecosystem functioning.

Freezing of movements and its correspondence with MLG1 neurons response to looming stimuli in the crab Neohelice.

Upon visually detecting a moving predator animals often freeze, i.e. stop moving, to minimize being uncovered and to gather detailed information of the object's movements and properties. In certain conditions the freezing behavior can be enough to avoid a predatory menace but, when the risk is high or increases to a higher level, animals switch strategy and engage in an escape response. The neural bases underlying escape responses to visual stimuli are extensively investigated both in vertebrates and arthropods. However, those involved in freezing behaviors are much less studied. Here, we investigated the freezing behavior displayed by the crab Neohelice granulata when confronted with a variety of looming stimuli simulating objects of distinct sizes approaching on a collision course at different speeds. The experiments were performed in a treadmill-like device. Animals engaged in exploratory walks respond to the looming stimulus with freezing followed by escaping. The analysis of the stimulus optical variables shows that regardless of the looming dynamic, the freezing decision is made when the angular size of the object increases by 1.4°. In vivo intracellular recording responses of Monostratified Lobula Giant Neurons (MLG1) to the same looming stimuli show that the freezing times correlate with the times predicted by a hypothetical spike counter of this neuron.

Considerations for cultivated crustacean meat: potential cell sources, potential differentiation and immortalization strategies, and lessons from crustacean and other animal models.

Cultivated crustacean meat (CCM) is a means to create highly valued shrimp, lobster, and crab products directly from stem cells, thus removing the need to farm or fish live animals. Conventional crustacean enterprises face increasing pressures in managing overfishing, pollution, and the warming climate, so CCM may provide a way to ensure sufficient supply as global demand for these products grows. To support the development of CCM, this review briefly details crustacean cell culture work to date, before addressing what is presently known about crustacean muscle development, particularly the molecular mechanisms involved, and how this might relate to recent work on cultivated meat production in vertebrate species. Recognizing the current lack of cell lines available to establish CCM cultures, we also consider primary stem cell sources that can be obtained non-lethally including tissues from limbs which are readily released and regrown, and putative stem cells in circulating hemolymph. Molecular approaches to inducing myogenic differentiation and immortalization of putative stem cells are also reviewed. Finally, we assess the current status of tools available to CCM researchers, particularly antibodies, and propose avenues to address existing shortfalls in order to see the field progress.

Comparison analysis of circulating hemocytes in decapod crustaceans.

Hemocytes are the primary immune cells of crustaceans. Few comparison studies have been done among different crustaceans and some key parameters of circulating hemocytes have not been investigated. Here, we compared the circulating hemocytes in six decapod crustaceans, Cherax quadrinatus, Procambarus clarkii, Penaeus vannamei, Penaeus monodon, Eriocheir sinensis, and Scylla paramamosain. Although the hemocytes of different species vary in size, they share common morphological characteristics. Based on their morphological features, circulating hemocytes can be basically classified into granular cells (GCs), semi-granular cells (SGCs), and hyaline cells (HCs). In the six decapods analyzed in this study, the proportion of GCs varied from 10 % to 30 %. P. vannamei, P. monodon, and P. clarkii had fewer GCs in circulation than the other three species. Correspondingly, proliferation was detected only in a small portion of cells in P. vannamei, P. monodon, and P. clarkii under physical conditions. The hemocyte renewal rates for P. clarkii, E. sinensis, and C. quadrinatus were 6.1 %, 5.1 %, and 1.5 % per day, while no steady new hemocyte production was found in S. paramamosain within six days. These data give a general picture of the similarities and differences of circulating hemocytes in decapods and provide a base for an in-depth study of their immune system.

Characterization of a pseudohemocyanin gene (PtPhc1) and its immunity function in response to Vibrio parahaemolyticus infection in the swimming crab Portunus trituberculatus.

Pseudohemocyanin is a member of the hemocyanin superfamily, but little research is available on its function in immunology. In this study, a Portunus trituberculatus pseudohemocyanin gene, named PtPhc1, was obtained by gene cloning. The PtPhc1 cDNA was 2312 bp in length, encoding 684 amino acids while exhibiting a characteristic hemocyanin structural domain. Tissue expression analysis revealed ubiquitous expression of PtPhc1 across all tissues, with the highest level of expression observed in the hepatopancreas. The expression pattern of PtPhc1 in response to Vibrio parahaemolyticus infection was clarified using RT-qPCR in swimming crabs. Notably, the expression peaked at 24 h, and increased 1435-fold compared to the control group in the hepatopancreas. While the expression level reached the maximum value at 72 h, which was 3.24 times higher than that of the control group in hemocytes. Remarkably, the reduction in PtPhc1 expression led to a noteworthy 30% increase in the mortality rate of P. trituberculatus when exposed to V. parahaemolyticus. In addition, in vitro bacterial inhibition assays exhibited a dose-dependent suppression of bacterial proliferation by recombinant PtPhc1 protein, with a notable inhibition rate of 48.33% against V. parahaemolyticus at a concentration of 0.03 mg/mL. To the best of our knowledge, the results establish the function of pseudohaemocyanin in immunity for the first time, contributing to a deeper comprehension of innate immune regulatory mechanisms in aquatic organisms and advancing strategies for disease-resistant breeding.

An i-type lysozyme from a crustacean, Pacifastacus leniusculus, functions as a clot-destabilising enzyme.

Lysozymes are hydrolytic enzymes, and they are ubiquitous among all living organisms. They are mostly associated with antibacterial properties through their muramidase activity, while other properties such as iso-peptidase activity are also common. Invertebrate-type (i-type) lysozymes include the enzyme Destabilase, which is present in the salivary secretions of the medicinal leach Hirundo medicinalis. Destabilase has the ability to hydrolyse the ε-(γ-glutamyl)-lysine iso-peptide bonds formed by transglutaminase in fibrin of vertebrate blood, thereby destabilising blood clots. We have identified an i-type lysozyme from the hemocytes of the freshwater crayfish Pacifastacus leniusculus, which was found to be upregulated at the protein level in response to an injection of the ß-1,3-glucan laminarin. Based on its sequence we predicted that this lysozyme would lack muramidase activity, and therefore we decided to determine its putative immune function. The P. leniusculus i-type lysozyme (Pl-ilys), is a protein with 159 amino acid residues, including a 29 residue signal peptide, with a predicted molecular weight of 16 kDa and a predicted pI of 5.6. It is expressed primarily in the hemocytes and to a lesser extent in the hematopoietic tissue. A recombinant mature Pl-ilys using an E. coli expression system was produced, and we could ascertain that this enzyme was deficient of muramidase activity. Moreover, no iso-peptidase activity could be detected against the substrate l-γ-glutamine-p-nitroanilide. Analysis of the conserved domains in Pl-ilys showed a putative destabilase domain, and thus we tested the clot dissolving activity of this enzyme. We could show that the purified P. leniusculus clotting protein which had been coagulated and clotted with transglutaminase was dissolved by the addition of Pl-ilys. Taken together our results indicate that Pl-ilys has a clot dissolving or destabilising activity in crustacean blood.

Insights into the DNA methylation of Portunus trituberculatus in response to Vibrio parahaemolyticus infection.

Vibrio parahaemolyticus is the main pathogen causing acute hepatopancreatic necrotic disease in crustaceans. To elucidate the epigenetic regulatory mechanism of crustacean resistance to V. parahaemolyticus infection, we conducted artificial infection studies on Portunus trituberculatus. The results showed that the mortality rate reached the highest at 12 h of artificial infection, which was 23.69 %. At 72 h after V parahaemolyticus infection, the expression level of DNA demethylase (ten-eleven-translocation protein) Tet was significantly decreased, the expression of DNA methyltransferase Dnmt3B fluctuated significantly. Based on the differential expression levels of Tet and Dnmt3B. We depict for DNA methylation profiles of the whole genome of P. trituberculatus at single-base resolution by using whole-genome bisulfite sequencing (WGBS) on hemolymph tissues. The overall DNA methylation level was low at 2.16% in P. trituberculatus hemolymph. A total of 2,590 differentially methylated regions (DMRs) were identified, of which 1,329 were hypermethylated and 1,261 were hypomethylated, and 1,389 genes were annotated in these DMRs. Differently methylated genes (DMGs) were significantly enriched in ribosomes (KO03010), protein kinases (KO01001), cell cycle (HSA04110), endocrine resistance (HSA01522) and FoxO signaling pathway (KO04068). Finally, we selected six differentially methylated genes for quantitative analysis. The results showed that DNA methylation not only has a negative regulatory effect on gene expression, but also has a positive regulatory effect. These results indicated that DNA methylation in the regulation of genes involved in immune responses contributes to the resistance of P. trituberculatus to V. parahaemolyticus, which is valuable for understanding how crustaceans regulate the innate immune system to defend against bacterial infections.

Metagenomic dissection of the intestinal microbiome in the giant river prawn Macrobrachium rosenbergii infected with Decapod iridescent virus 1.

Microbiome in the intestines of aquatic invertebrates plays pivotal roles in maintaining intestinal homeostasis, especially when the host is exposed to pathogen invasion. Decapod iridescent virus 1 (DIV1) is a devastating virus seriously affecting the productivity and success of crustacean aquaculture. In this study, a metagenomic analysis was conducted to investigate the genomic sequences, community structure and functional characteristics of the intestinal microbiome in the giant river prawn Macrobrachiumrosenbergii infected with DIV1. The results showed that DIV1 infection could significantly reduce the diversity and richness of intestinal microbiome. Proteobacteria represented the largest taxon at the phylum level, and at the species level, the abundance of Gonapodya prolifera and Solemya velum gill symbiont increased significantly following DIV1 infection. In the infected prawns, four metabolic pathways related to purine metabolism, pyrimidine metabolism, glycerophospholipid metabolism, and pentose phosphate pathway, and five pathways related to nucleotide excision repair, homologous recombination, mismatch repair, base excision repair, and DNA replication were significantly enriched. Moreover, several immune response related pathways, such as shigellosis, bacterial invasion of epithelial cells, Salmonella infection, and Vibrio cholerae infection were repressed, indicating that secondary infection in M. rosenbergii may be inhibited via the suppression of these immune related pathways. DIV1 infection led to the induction of microbial carbohydrate enzymes such as the glycoside hydrolases (GHs), and reduced the abundance and number of antibiotic-resistant ontologies (AROs). A variety of AROs were identified from the microbiota, and mdtF and lrfA appeared as the dominant genes in the detected AROs. In addition, antibiotic efflux, antibiotic inactivation, and antibiotic target alteration were the main antibiotic resistance mechanisms. Collectively, the data would enable a deeper understanding of the molecular response of intestinal microbiota to DIV1, and offer more insights into its roles in prawn resistance to DIVI infection.

Characterization of the molecular, cellular, and behavioral changes caused by exposure to a saline-alkali environment in the Chinese mitten crab, Eriocheir sinensis.

In the context of global warming, the accelerated evaporation of seawater will lead to a continuous expansion of saline-alkali land area. As an important economic freshwater crustacean, investigation on the mechanism of damage to Eriocheir sinensis (E. sinensis) under saline-alkali environment will provide a valuable precedent for understanding the detrimental effect of climate change on crustaceans. In this study, histopathological analysis and integrative omics analysis were employed to explore the injury mechanism on the cerebral nervous system of E. sinensis exposure to saline-alkali stress. Our findings revealed that under this stress E. sinensis exhibited behavioral disorders and damage to cerebral neurosecretory cells and key organelles. Additionally, several pathways related to detoxification metabolism, neurotransmitter synthesis, and antioxidant defense were significantly down-regulated. Collectively, these results show, for the first time, that saline-alkali stress can induce neurodegenerative disease-like symptoms in E. sinensis, and provide critical information for understanding the harmful effects of saline-alkali environments.

Crustacean endocrinology: Sexual differentiation and potential application for aquaculture.

Crustaceans, which represent a significant subset of arthropods, are classified into three major classes: Ostracoda, Malacostraca, and Branchiopoda. Among them, sex manipulation in decapod species from the Malacostraca class has been extensively researched for aquaculture purposes and to study reproductive physiology and sexual plasticity. Some decapods exhibit sexual dimorphism that influences their biological and economic value. Monosex culture, in which only one sex is cultivated, increases production yields while reducing the risk of invasiveness, as genetic leakage into natural waters is less likely to occur. Differences in yield are also observed when cultivating different sexes, with all-male cultures of Macrobrachium rosenbergii being more profitable than both mixed and all-female cultures. Research on decapod sexual differentiation has led to a better understanding of sex determination and sexual differentiation processes in arthropods. Similar to most mammals and other vertebrate classes, Malacostraca crustaceans, including decapods, exhibit a cell-non-autonomous mode of sexual development. Genetic factors (e.g., sex chromosomes) and endocrine factors (e.g., insulin-like androgenic gland factor and crustacean female sex hormone) play pivotal roles in the development of sexually dimorphic traits. This review synthesizes the existing understanding of sex determination mechanisms and the role of sex hormones in decapod species. Additionally, it provides an overview of the methyl farnesoate, which has been suggested to be involved in male sex differentiation in some crab species, as well as the phenomenon of male-to-female sex reversal in host decapods caused by parasitic crustaceans.

Paospora carinifang n. gen., n. sp. (Microsporidia: Spragueidae), a parasite of the ridgetail white prawn, Palaemon carinicauda.

A new microsporidian disease of the pond-reared ridgetail white prawn, Palaemon carinicauda, was found in China. Light microscopy, pathology, and scanning electron microscopy showed that the parasite infected the host's skeletal muscle tissue and formed spherical sporophorous vesicles (SPOVs). Electron microscopy revealed that its merogonic life stages developed in direct contact with the host cytoplasm. The sporogonic life stages underwent octosporoblastic sporogony with the formation of eight uninucleate spores in each SPOV. Fresh SPOVs were 5.4 ± 0.55 µm in diameter. The octospores were oval and measured 2.3 × 1.5 µm (fresh) and 1.96 × 1.17 µm (fixed). The isofilar polar filament was coiled with 9-10 turns and arranged in two rows. Phylogenetic analysis based on the SSU rRNA gene suggests that this microsporidium has close affinities with members of the genera Potaspora and Apotaspora, but represents an independent generic taxon. We therefore propose the establishment of a new genus and species (Paospora carinifang n. gen., n. sp.) within the family Spragueidae. We also propose a taxonomic revision to transfer Potaspora macrobrachium to this new genus and reclassify it as Paospora macrobrachium comb. nov.

Eusociality in snapping shrimps is associated with larger genomes and an accumulation of transposable elements.

Despite progress uncovering the genomic underpinnings of sociality, much less is known about how social living affects the genome. In different insect lineages, for example, eusocial species show both positive and negative associations between genome size and structure, highlighting the dynamic nature of the genome. Here, we explore the relationship between sociality and genome architecture in Synalpheus snapping shrimps that exhibit multiple origins of eusociality and extreme interspecific variation in genome size. Our goal is to determine whether eusociality leads to an accumulation of repetitive elements and an increase in genome size, presumably due to reduced effective population sizes resulting from a reproductive division of labor, or whether an initial accumulation of repetitive elements leads to larger genomes and independently promotes the evolution of eusociality through adaptive evolution. Using phylogenetically informed analyses, we find that eusocial species have larger genomes with more transposable elements (TEs) and microsatellite repeats than noneusocial species. Interestingly, different TE subclasses contribute to the accumulation in different species. Phylogenetic path analysis testing alternative causal relationships between sociality and genome architecture is most consistent with the hypothesis that TEs modulate the relationship between sociality and genome architecture. Although eusociality appears to influence TE accumulation, ancestral state reconstruction suggests moderate TE abundances in ancestral species could have fueled the initial transitions to eusociality. Ultimately, we highlight a complex and dynamic relationship between genome and social evolution, demonstrating that sociality can influence the evolution of the genome, likely through changes in demography related to patterns of reproductive skew.

Two faces of one coin: Beneficial and deleterious effects of reactive oxygen species during short-term acclimation to hypo-osmotic stress in a decapod crab.

Exposure to environmental changes often results in the production of reactive oxygen species (ROS), which, if uncontrolled, leads to loss of cellular homeostasis and oxidative distress. However, at physiological levels these same ROS are known to be key players in cellular signaling and the regulation of key biological activities (oxidative eustress). While ROS are known to mediate salinity tolerance in plants, little is known for the animal kingdom. In this study, we use the Mediterranean crab Carcinus aestuarii, highly tolerant to salinity changes in its environment, as a model to test the healthy or pathological role of ROS due to exposure to diluted seawater (dSW). Crabs were injected either with an antioxidant [N-acetylcysteine (NAC), 150 mg·kg-1] or phosphate buffered saline (PBS). One hour after the first injection, animals were either maintained in seawater (SW) or transferred to dSW and injections were carried out at 12-h intervals. After ≈48 h of salinity change, all animals were sacrificed and gills dissected for analysis. NAC injections successfully inhibited ROS formation occurring due to dSW transfer. However, this induced 55% crab mortality, as well as an inhibition of the enhanced catalase defenses and mitochondrial biogenesis that occur with decreased salinity. Crab osmoregulatory capacity under dSW condition was not affected by NAC, although it induced in anterior (non-osmoregulatory) gills a 146-fold increase in Na+/K+/2Cl- expression levels, reaching values typically observed in osmoregulatory tissues. We discuss how ROS influences the physiology of anterior and posterior gills, which have two different physiological functions and strategies during hyper-osmoregulation in dSW.

Effects of different salinity reduction intervals on osmoregulation, anti-oxidation and apoptosis of Eriocheir sinensis megalopa.

Eriocheir sinensis megalopa has a special life history of migrating from seawater to freshwater. In order to investigate how the megalopa adapt themselves to the freshwater environment, we designed an experiment to reduce the salinity of water from 30 ppt to 0 at rates of 30 ppt, 15 ppt, 10 ppt, and 5 ppt per 24 h to evaluate the effects of different degrees of hyposaline stress on the osmotic regulation ability and antioxidant system of the megalopa. Experimental results related to osmotic pressure regulation show that the gill tissue of megalopa in the treatment group of 30 ppt/24 h rapid reduction of salinity was damaged, while in the treatment group of 5 ppt/24 h it was intact. At the same time, the experiment also found that in each treatment group with different salinity reduction rates, compared with the control salinity, the NKA activity of megalopa increased significantly after the salinity was reduced to 20 ppt (p < 0.05). In addition, two genes involved in chloride ion transmembrane absorption have different expression patterns in the treatment groups with different salinity reduction rates. Among them, Clcn2 was significantly highly expressed only in the rapid salinity reduction intervals of 30 ppt/24 h and 15 ppt/24 h (p < 0.05). Slc26a6 was significantly highly expressed only in the slow salinity reduction intervals of 10 ppt/24 h and 5 ppt/24 h (p < 0.05). On the other hand, the results of antioxidant and apoptosis related experiments showed that in all treatment groups with different rates of salinity reduction, the activities of T-AOC, GSH-PX, and CAT basically increased significantly after salinity reduction compared to the control salinity. Moreover, the activities of T-AOC and CAT were significantly higher in the 10 ppt/24 h and 5 ppt/24 h treatment groups than in the 30 ppt/24 h and 15 ppt/24 h treatment groups. Finally, the experimental results related to apoptosis showed that the expression trends of Capase3 and Bax-2 were basically the same in the treatment groups with different salinity reduction rates, and their expressions were significantly higher in the 10 ppt/24 h and 5 ppt/24 h treatment groups than in the 30 ppt/24 h and 15 ppt/24 h treatment groups. In summary, the present study found that megalopa had strong hyposaline tolerance and were able to regulate osmolality at different rates of salinity reduction, but the antioxidant capacity differed significantly between treatment groups, with rapid salinity reduction leading to oxidative damage in the anterior gills and reduced antioxidant enzyme activity and apoptosis levels.