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To study the support design of deep thick coal seam roadway, we propose three support design schemes from the perspective of safety and support cost based on the 432 belt transportation roadway in Tangyang Coal Mine. Theoretical analysis, numerical simulation and engineering practice were used to study the values of the support parameters, displacement and deformation, plastic zone and the actual support effect of each scheme. The theoretical analysis results show that the anchor parameters and row spacing of the three support schemes are in line with the theoretical calculation values. Numerical simulation results show that: the relative displacement and deformation of the top and bottom plates and the two sides of support scheme I are 77.6 mm and 103.4 mm, respectively; the relative displacement and deformation of support scheme II are 79.7 mm and 105.5 mm, which are similar to those of support scheme I and have the least plastic failure elements; the maximum displacement and deformation of support scheme III are 83.8 mm and 111.1 mm, respectively. The support cost of support scheme II is only 64.7% of that of scheme I when the support effect is similar. The engineering practice results show that the actual support effect of support scheme II can realize the safety support of roadway.

Introduction: Whole-genome duplication (WGD) is one of the most sudden and dramatic events rarely reported in invertebrates, but its occurrence can lead to physiological, morphological, and behavioral diversification. WGD has also never been reported in barnacles, which is one of the most unique groups of crustaceans with extremely speciallized morphology (calcareous shells) and habits (intertidal sessile lifestyle). Objectives: To investigate whether WGD has occurred in barnacles and examine its potential role in driving the adaptive evolution and diversification of barnacles. Methods: Based on a newly sequenced and assembled chromosome-level barnacle genome, a novel WGD event has been identified in barnacles through a comprehensive analysis of interchromosomal synteny, the Hox gene cluster, and synonymous substitution distribution. Results: We provide ample evidences for WGD in the barnacle genomes. Comparative genomic analysis indicates that this WGD event predates the divergence of Thoracicalcarea, occurring more than 247 million years ago. The retained ohnologs from the WGD are primarily enriched in various pathways related to environmental information processing, shedding light on the adaptive evolution and diversification of intertidal sessile lifestyle. In addition, transcriptomic analyses show that most of these ohnologs were differentially expressed following the ebb of tide. And the cytochrome P450 ohnologs with differential expression patterns are subject to subfunctionalization and/or neofunctionalization for intertidal adaptation. Besides WGD, parallel evolution underlying intertidal adaptation has also occurred in barnacles. Conclusion: This study revealed an ancient WGD event in the barnacle genomes, which is potentially associated with the origin and diversification of thoracican barnacles, and may have contributed to the adaptive evolution of their intertidal sessile lifestyle.

Abstract Accurate and sensitive near‐infrared (NIR) luminescent lateral flow immunoassay (LFIA) has attracted considerable attention in the field of point‐of‐care testing (POCT). However, the detection accuracy and sensitivity are often compromised by the low fluorescence quantum efficiency ( 800 nm) completely eliminated background interference and allowed highly accurate and sensitive detection without any pre‐treatment steps. The limits of detection (LODs) for aflatoxin B1 (AFB1) in soy sauce, alpha hemolysin (Hla) of Staphylococcus aureus biomarker in joint fluid, and C‐reactive protein (CRP) in human haemolysed samples were 0.01 ng mL−1, 0.02 µg mL−1, and 0.156 mg L−1, respectively, commensurating with those of the corresponding gold standard assays and covering the detection range of interests. It is anticipated that the ultrabright NIR AIEgen nanoparticles will serve as a universally applicable signal probe for NIR‐LFIA diagnostics, promising to expand the range of applications for quantitative detection of complex samples.

Insulin-like peptide (ILP) is a member of the insulin superfamily with evolutionary conservation and is one of the most important factors affecting animal life activities. In this study, a full-length of ILP1 gene in Pacific white shrimp, Litopenaeus vannamei, was cloned, and the mRNA length consisted of 812 bp with an open reading frame (ORF) of 543 bp, encoding 180 amino acids. Sequence analysis showed that the predicted molecular weight of LvILP1 protein was 20.81 kDa, and the theoretical isoelectric point was 9.45 and the instability coefficient was 96.20. There was a signal peptide, no transmembrane structure. It was deduced that it was located outside the cell and is an alkaline unstable secreted protein. Structure prediction found that LvILP1 protein had the conserved IlGF domain of the insulin superfamily, which was composed of the N-terminal signal peptide, B-chain, C-peptide and A-chain, as well as six conserved cysteine sites and two cleavage sites. Phylogenetic analysis found that LvILP1 was most closely related to ILP7 in Penaeus monodon, and clustered with ILP1 of Crustaceans to form a branch, and then clustered with ILP7 of Invertebrate, Relaxin, Insulin and Insulin-like growth factor (IGF) of vertebrate; ILP7 of Invertebrate was evolutionarily closest to outgroup sea anemone ILP1, suggesting that it may be more similar to the ancestor gene of the insulin superfamily. Transcription factor prediction found that the possible transcription factors of LvILP1 are Forkhead box protein O3 (FoxO3), Glucocorticoid receptor (GR), CAAT region/enhancer binding protein (C/EBP) and Signal transduction and transcription activator protein (STAT); The protein interaction analysis found that LvILP1 interacted with Insulin receptor (IR) on the cell membrane, nerve signaling molecules (VGLUT1, SYT 1_3), Glycoprotein hormone beta 5 (GPHB5), Bursicon alpha, etc. By analyzing the biological function analysis of these transcription factors and interacting proteins, it is speculated that LvILP1 may play an important role in regulating shrimp growth and development, response to hormonal stimulation, nervous system homeostasis, carbohydrate homeostasis, postmolt tissue remodeling and reproductive development. The analysis showed that LvILP1 was expressed in the early developmental stage of shrimp, and expressed in all tissues of adults, but the expression level in the eye stalk was the highest. This study provides important information for in-depth understanding of the structure, evolution, function and expression of ILP in shrimp, as well as clues for molecular breeding and healthy farming of shrimp.

Vibriosis is one of the most serious diseases that commonly occurs in aquatic animals, thus, shaping a steady inherited resistance trait in organisms has received the highest priority in aquaculture. Whereas, the mechanisms underlying the development of such a resistance trait are mostly elusive. In this study, we constructed vibriosis-resistant and susceptible families of the Pacific white shrimp Litopenaeus vannamei after four generations of artificial selection. Microbiome sequencing indicated that shrimp can successfully develop a colonization resistance trait against Vibrio infections. This trait was characterized by a microbial community structure with specific enrichment of a single probiotic species (namely Shewanella algae), and notably, its formation was inheritable and might be memorized by host epigenetic remodeling. Regardless of the infection status, a group of genes was specifically activated in the resistant family through disruption of complete methylation. Specifically, hypo-methylation and hyper-expression of genes related to lactate dehydrogenase (LDH) and iron homeostasis might provide rich sources of specific carbon (lactate) and ions for the colonization of S. algae, which directly results in the reduction of Vibrio load in shrimp. Lactate feeding increased the survival of shrimp, while knockdown of LDH gene decreased the survival when shrimp was infected by Vibrio pathogens. In addition, treatment of shrimp with the methyltransferase inhibitor 5-azacytidine resulted in upregulations of LDH and some protein processing genes, significant enrichment of S. algae, and simultaneous reduction of Vibrio in shrimp. Our results suggest that the colonization resistance can be memorized as epigenetic information by the host, which has played a pivotal role in vibriosis resistance. The findings of this study will aid in disease control and the selection of superior lines of shrimp with high disease resistance.

Multiple projects are increasingly managed in a decentralized environment, resulting in an inapplicable centralized resource-constrained multi-project scheduling problem (CRCMPSP). A decentralized resource-constrained multi-project scheduling problem (DRCMPSP) is proposed to ensure the timely delivery of products and services within budget. DRCMPSP often faces difficulty in effectively addressing resource conflict. Moreover, activity durations are hard to determine in advance due to various unpredictable events. Therefore, this paper investigates the performance of priority rules (PRs) for global resource allocation of stochastic DRCMPSP. A two-stage heuristic based on PR is then developed to resolve global resource contention under a stochastic environment. 25 PRs from the literature were selected and incorporated into our approach to explore their efficiency on global resource allocation in different global objectives, that is, the expected average project delay (EAPD) and expected total makespan (ETMS). Computational experiments based on the Multi-Project Scheduling Problem LIBrary (MPSPLIB) dataset show that more efficient PRs always favor activities that try to obtain a low delay for each project for EAPD, whereas for ETMS, more efficient PRs tend to favor the longest projects. Furthermore, the hybrid PRs based on more efficient PRs are designed to allocate global resources, and experimental results show that the average percentage of 3 hybrid PRs better than that of their components is 1.14% for EAPD and 0.81% for ETMS, respectively.

In the culture of crustaceans, most species show sexual dimorphism. Monosex culture is an effective approach to achieve high yield and economic value, especially for decapods of high value. Previous studies have developed some sex control strategies such as manual segregation, manipulation of male androgenic gland and knockdown of the male sexual differentiation switch gene encoding insulin-like androgenic gland hormone (IAG) in decapods. However, these methods could not generate hereditable changes. Genetic manipulation to achieve sex reversal individuals is absent up to now. In the present study, the gene encoding IAG (EcIAG) was identified in the ridgetail white prawn Exopalaemon carinicauda. Sequence analysis showed that EcIAG encoded conserved amino acid structure like IAGs in other decapod species. CRISPR/Cas9-mediated genome editing technology was used to knock out EcIAG. Two sgRNAs targeting the second exon of EcIAG were designed and microinjected into the prawn zygotes or the embryos at the first cleavage with commercial Cas9 protein. EcIAG in three genetic males was knocked out in both chromosome sets, which successfully generated sex reversal and phenotypic female characters. The results suggest that CRISPR/Cas9-mediated genome editing technology is an effective way to develop sex manipulation technology and contribute to monosex aquaculture in crustaceans.

As one of the most important aquaculture species in the world, the improvement of growth traits of the Pacific white shrimp (Litopenaeus vannamei), has always been a primary focus. In this study, we conducted SNP-specific locus analysis and identified a growth-related gene, BAMBI, in L. vannamei. We analyzed the structure and function of LvBAMBI using genomic, transcriptomic, metabolomic, and RNA interference (RNAi) assays. The LvBAMBI possessed highly conserved structural domains and widely expressed in various tissues. Knockdown of LvBAMBI significantly inhibited the gain of body length and weight of the shrimp, underscoring its role as a growth-promoting factor. Specifically, knockdown of LvBAMBI resulted in a significant downregulation of genes involved in lipid metabolism, protein synthesis, catabolism and transport, and immunity. Conversely, genes related to glucose metabolism exhibited significant upregulations. Analysis of differential metabolites (DMs) in metabolomics further revealed that LvBAMBI knockdown may primarily affect shrimp growth by regulating biological processes related to lipid and glucose metabolism. These results suggested that LvBAMBI plays a crucial role in regulating lipid metabolism, glucose metabolism, and protein transport in shrimp. This study provides valuable insights for future research and utilization of BAMBI genes in shrimp and crustaceans.

Monocyte to macrophage differentiation factor 2 gene (Mmd2) encodes a member of the progestin and adipoQ receptor (PAQR) family, and plays a key role in growth and development. Our previous studies had found Mmd2 (Monocyte to macrophage differentiation factor 2) is a new candidate gene for growth traits in Pacific white shrimp (Litopenaeus vannamei). For the purpose of understanding the underlying mechanism of LvMmd2 affecting the growth of shrimp, we analyzed the gene structure, phylogeny, expression profiles and RNA interference of this gene in L. vannamei. We found the LvMmd2 gene sequence was highly conserved in transmembrane regions, it was widely expressed in different tissues, with the highest expression level in the eye stalk. Knockdown LvMmd2 could significantly promote body length and body weight gain, suggesting it is a growth suppressor. Through transcriptome analysis we identified 422 differentially expressed genes (DEGs) between the dsMmd2 group and control group, among which 337 genes were upregulated in the dsMmd2 group, including numerous muscle-related genes and protein synthesis genes. Further bioinformatics analysis showed that growth, metabolism, and immune-related signal pathway had changed significantly. The above results greatly increase our understanding on the conservative structure and function of LvMmd2 gene, and provide potential application prospects in genetics and breeding.

Abstract Background The deep-sea may be regarded as a hostile living environment, due to low temperature, high hydrostatic pressure, and limited food and light. Isopods, a species-rich group of crustaceans, are widely distributed across different environments including the deep sea and as such are a useful model for studying adaptation, migration, and speciation. Similar to other deep-sea organisms, giant isopods have larger body size than their shallow water relatives and have large stomachs and fat bodies presumably to store organic reserves. In order to shed light on the genetic basis of these large crustaceans adapting to the oligotrophic environment of deep-sea, the high-quality genome of a deep-sea giant isopod Bathynomus jamesi was sequenced and assembled. Results B. jamesi has a large genome of 5.89 Gb, representing the largest sequenced crustacean genome to date. Its large genome size is mainly attributable to the remarkable proliferation of transposable elements (84%), which may enable high genome plasticity for adaptive evolution. Unlike its relatives with small body size, B. jamesi has expanded gene families related to pathways of thyroid and insulin hormone signaling that potentially contribute to its large body size. Transcriptomic analysis showed that some expanded gene families related to glycolysis and vesicular transport were specifically expressed in its digestive organs. In addition, comparative genomics and gene expression analyses in six tissues suggested that B. jamesi has inefficient lipid degradation, low basal metabolic rate, and bulk food storage, suggesting giant isopods adopt a more efficient mechanism of nutrient absorption, storage, and utilization to provide sustained energy supply for their large body size. Conclusions Taken together, the giant isopod genome may provide a valuable resource for understanding body size evolution and adaptation mechanisms of macrobenthic organisms to deep-sea environments.

This paper studies the distribution characteristics of acoustic emission (AE) events in supporting roadways under different dynamic load conditions through numerical simulation and theoretical analysis. According to the principle of AE numerical simulation, the roadway model is established by FLAC3D software, and the supporting structure of the bolt cable is established by editing the Fish function. According to the model dynamic load application criterion, the relationship between the peak velocity of the particle, the distance from the source center to the dynamic impact damage point, and the dynamic load intensity is introduced. The velocity-distance-energy relationship is deduced from the relationship to determine the magnitude of the dynamic load energy. The simulation results show that the intensity of the dynamic load source and the distance of the dynamic load source directly impact the AE events. The larger the dynamic load intensity and the closer to the dynamic load source, the more concentrated the AE events. The roadway has a blocking effect on the transmission of the dynamic load stress waves. According to this characteristic, the roadway can be protected by a high-pressure relief roadway. Rock lithology greatly influences the transmission of dynamic load stress waves and the number of AE events. When the dynamic load stress wave passes through the rock strata of different lithologies, the attenuation of the dynamic load energy and the number of AE events are large. However, when the dynamic load stress wave passes through the rock strata of the same lithology, the attenuation of the dynamic load energy and AE events is small. The surrounding rock structure in the bottom corner area of the roadway is susceptible to disturbance from dynamic load sources above the roadway. The results are greatly significant for studying the AE characteristics of support roadways with disturbance-typeimpact failure.

The Ras superfamily of small guanosine triphosphatases (GTPases) are a large group of small GTP-binding proteins, which play crucial roles in basic cellular processes in all eukaryotes. In this study, by analyzing the gene structure, temporal and spatial expression patterns, a total of 108 Ras superfamily genes were identified in the genome of the Pacific white shrimp Litopenaeus vannamei. We found these genes included not only the classical Ras GTPase superfamily members, but also some unconventional and novel Ras GTPase proteins, which have unknown functions and unique expression patterns. All Ras superfamily genes of L. vannamei were highly conserved within the core G domain and closely related in phylogeny, but they might have two different evolutionary origins. In addition, different Ras GTPase genes exhibited distinct expression patterns in different tissues, development/molting stages and WSSV infection samples of L. vannamei, suggesting that they may have a high functional specialization, and play important roles in regulating the biological processes of cell differentiation, growth and development, immune response, etc. This study provides important clues for the structure, classification, evolution and function of Ras superfamily in shrimp.

In this paper, the dynamical behavior of the nonclassical diffusion equation is investigated. First, using the asymptotic regularity of the solution, we prove that the semigroup $ \{S(t)\}_{t\geq 0} $ corresponding to this equation satisfies the global exponentially $ \kappa- $dissipative. And then we estimate the upper bound of fractal dimension for the global attractors $ \mathscr{A} $ for this equation and $ \mathscr{A}\subset H^1_0(\Omega)\cap H^2(\Omega) $. Finally, we confirm the existence of exponential attractors $ \mathscr{M} $ by validated differentiability of the semigroup $ \{S(t)\}_{t\geq 0} $. It is worth mentioning that the nonlinearity $ f $ satisfies the polynomial growth of arbitrary order.

Insulin-like peptides (ILPs) play key roles in animal growth, metabolism and reproduction in vertebrates. In crustaceans, one type of ILPs, insulin-like androgenic gland hormone (IAG) had been reported to be related to the sex differentiations. However, the function of other types of ILPs is rarely reported. Here, we identified another type of ILPs in the ridgetail white prawn Exopalaemon carinicauda (EcILP), which is an ortholog of Drosophila melanogaster ILP7. Sequence characterization and expression analyses showed that EcILP is similar to vertebrate insulin/IGFs and insect ILPs in its heterodimeric structure and expression profile. Using CRISPR/Cas9 genome editing technology, we generated EcILP knockout (KO) prawns. EcILP-KO individuals have a significant higher growth-inhibitory trait and mortality than those in the normal group. In addition, knockdown of EcILP by RNA interference (RNAi) resulted in slower growth rate and higher mortality. These results indicated that EcILP was an important growth regulator in E. carinicauda.

Temperature serves as an important environmental factor in ecosystems. Understanding the cooperation of various tissues of animals in response to heat stress is the basis for clarifying the regulation mechanism of different species under heat stress. Herein, a comparative transcriptomic analysis was conducted on three tissues (hepatopancreas, gill and muscle) of the Pacific white shrimp Litopenaeus vannamei under heat stress. Three tissues displayed distinct gene expression patterns, suggesting a cooperation based on division of labor might have occurred among them. In hepatopancreas and gill, genes related to ATP generation and utilization were down-regulated, and energetically expensive protein turnover was almost shut down. While in muscle, genes related to ATP generation and utilization, and those involved in several energy-consuming processes were up-regulated. In consistent, significant accumulation of ATP and decrease of total protein concentration were detected in hepatopancreas and gill, while it was opposite in muscle. Therefore, we suggest that different tissues may cooperate with each other simultaneously via energy reallocation in response to heat stress. Less energy was channeled into protein turnover in gill and hepatopancreas, and more energy was required for muscle. This study not only provides a comprehensive understanding of the molecular mechanism of L. vannamei in response to high temperature, but also lays the foundation of mining thermotolerance genes and proposing effective strategies to cope with the high temperature environment.

Brachyurans, or crabs, are of commercial and ecological importance, but limited genomic resources exist. Here the authors present a chromosome-level genome and expression data for the Chinese mitten crab, to shed light on the biology of this group.

This study focuses on the role of simple sequence repeats (SSRs) in the evolution of the genomes of two penaeid shrimp species, Fenneropenaeus chinensis and Litopenaeus vannamei. They observed a significant expansion in SSRs in the genomes of these species (~23%) compared what is seen in other animal genomes (~1%) and suggest a possible mechanistic relationship between SSRs and transposable elements.

The Pacific white shrimp Litopenaeus vannamei is the most economically important crustacean in the world. The growth and development of shrimp muscle has always been the focus of attention. Myocyte Enhancer Factor 2 (MEF2), a member of MADS transcription factor, has an essential influence on various growth and development programs, including myogenesis. In this study, based on the genome and transcriptome data of L. vannamei, the gene structure and expression profiles of MEF2 were characterized. We found that the LvMEF2 was widely expressed in various tissues, mainly in the Oka organ, brain, intestine, heart, and muscle. Moreover, LvMEF2 has a large number of splice variants, and the main forms are the mutually exclusive exon and alternative 5′ splice site. The expression profiles of the LvMEF2 splice variants varied under different conditions. Interestingly, some splice variants have tissue or developmental expression specificity. After RNA interference into LvMEF2, the increment in the body length and weight decreased significantly and even caused death, suggesting that LvMEF2 can affect the growth and survival of L. vannamei. Transcriptome analysis showed that after LvMEF2 was knocked down, the protein synthesis and immune-related pathways were affected, and the associated muscle protein synthesis decreased, indicating that LvMEF2 affected muscle formation and the immune system. The results provide an important basis for future studies of the MEF2 gene and the mechanism of muscle growth and development in shrimp.

From the perspective of full-component utilization of woody fiber biomass resources, areca nut husk is an excellent woody fiber biomass feedstock because of its fast regeneration, significant regeneration ability, sustainability, low cost, and easy availability. In this study, fiber cell morphologies, chemical compositions, lignin structures, and carbohydrate contents of areca nut husks were analyzed and compared with those of rice straw, and the application potentials of these two materials as biomass resources were compared. We found that areca nut husk fibers were shorter and wider than those of rice straw; areca nut husk contained more lignin and less ash, as well as less holocellulose than rice straw; areca nut husk and rice straw lignin were obtained by ball milling and phase separation, and areca nut husk lignin was found to be a typical GHS-type lignin. Herein, the yield of lignocresol was higher than that of milled wood lignin for both raw materials, and the molecular size was more homogeneous. Tricin structural monomers were discovered in the lignin of areca nut husk, similar to those present in other types of herbaceous plants. Structures of areca nut husk MWL (AHMWL) and AHLC were comprehensively characterized by quantitative NMR techniques (that is, 1H NMR, 31P NMR, and 2D NMR). The molecular structure of AHLC was found to be closer to the linear structure with more functional groups exposed on the molecular surface, and the hydroxyl-rich p-cresol grafting structure was successfully introduced into the lignin structure. In addition, the carbohydrate content in the aqueous layer of the phase separation system was close to the carbohydrate content in the raw material, indicating that the phase separation method can precisely separate lignin from carbohydrates. These experimental results indicate that the phase separation method as a method for lignin utilization and structure study has outstanding advantages in lignin structure regulation and yield, and areca nut husk lignin is suitable for application in the same phase separation systems as short-period herbs, such as rice straw and wheat grass, and has the advantages of low ash content and high lignification degree, which will provide guidance for the high-value utilization of areca nut husk in the future.

Abstract Background Barnacles are specialized marine organisms that differ from other crustaceans in possession of a calcareous shell, which is attached to submerged surfaces. Barnacles have a wide distribution, mostly in the intertidal zone and shallow waters, but a few species inhabit the deep-sea floor. It is of interest to investigate how such sessile crustaceans became adapted to extreme deep-sea environments. We sequenced the transcriptomes of a deep-sea barnacle, Glyptelasma gigas collected at a depth of 731 m from the northern area of the Zhongjiannan Basin, and a shallow-water coordinal relative, Octolasmis warwicki. The purpose of this study was to provide genetic resources for investigating adaptation mechanisms of deep-sea barnacles. Results Totals of 62,470 and 51,585 unigenes were assembled for G. gigas and O. warwicki, respectively, and functional annotation of these unigenes was made using public databases. Comparison of the protein-coding genes between the deep- and shallow-water barnacles, and with those of four other shallow-water crustaceans, revealed 26 gene families that had experienced significant expansion in G. gigas. Functional annotation showed that these expanded genes were predominately related to DNA repair, signal transduction and carbohydrate metabolism. Base substitution analysis on the 11,611 single-copy orthologs between G. gigas and O. warwicki indicated that 25 of them were distinctly positive selected in the deep-sea barnacle, including genes related to transcription, DNA repair, ligand binding, ion channels and energy metabolism, potentially indicating their importance for survival of G. gigas in the deep-sea environment. Conclusions The barnacle G. gigas has adopted strategies of expansion of specific gene families and of positive selection of key genes to counteract the negative effects of high hydrostatic pressure, hypoxia, low temperature and food limitation on the deep-sea floor. These expanded gene families and genes under positive selection would tend to enhance the capacities of G. gigas for signal transduction, genetic information processing and energy metabolism, and facilitate networks for perceiving and responding physiologically to the environmental conditions in deep-sea habitats. In short, our results provide genomic evidence relating to deep-sea adaptation of G. gigas, which provide a basis for further biological studies of sessile crustaceans in the deep sea.

The airflow cylinder sieve directly affects the harvesting effect as one of the main components of the rice combine harvester. The process of grain separated from threshed rice mixture is a multi-physical random pattern. This paper constructed accurately the components particle models, and reproduced the screening process of mixture via CFD-DEM (Computational Fluid Dynamics and Discrete Element Method). Objectives of this work are to better understand the impact of airflow velocity, aperture and deflectors of cylinder sieve on separation effect of the mixture in an airflow cylinder sieve device. The optimal working parameters of sieve were developed by the OED (Orthogonal Experiment Design) and the weight of the index method. The simulations result show that airflow velocity and aperture have an impact on the loss ratio and efficiency; deflectors have implications on efficiency. It is beneficial for separation of grain from the mixture with the airflow velocity, the aperture and deflectors are of 11m/s, 11mm, and two turns, correspondingly. The experiments result carried out on the test bench with real threshed rice mixture and agree with simulations, which proved the feasibility of the results. This paper provided basic data for the design of screening unite of rice harvester’s equipment.

Considering the impact of informatization condition, vehicles on the road network are divided into connected automated vehicles (CAVs) and human-driven vehicles (HDVs), which follow the principle of system optimization and stochastic user equilibrium, respectively. Taking the road network reserve capacity maximization model under the condition of road capacity constraint as the upper-level programming and the traffic assignment model under heterogeneous flow environment as the lower level programming, then a bilevel programming model is constructed. Among them, the nonuniform demand growth multiplier is adopted for each OD pair to reflect the inconsistency of traffic demand structure growth, and the calculation of link capacity is related to the market penetration of CAVs. The incremental method, method of successive averages, and simulated annealing algorithm are used to solve the model, and the effects of different market penetration on road network capacity, travel time, and saturation are analyzed through a numerical example. The relevant data under different weights are normalized and the optimal deployment scheme of CAVs and HDVs in different periods is obtained by comprehensive evaluation. Meanwhile, the mixed equilibrium flow state is explored under the premise of given market penetration to verify the feasibility of the model and algorithm.

The underground rock was disturbed repeatedly during the excavation, resulting in its damage degree gradually accumulated. Therefore, the rock in practical engineering can be regarded as predamaged (PD) rock, and its mechanical behaviors are different from conventional intact rock. This paper proposed the calibrate method to evaluate the predamaged degree of rock based on acoustic emission (AE) ring counts, and the PD rock samples were prepared by uniaxial cyclic loading and unloading test. Then, the triaxial compression experiment of predamaged rock was carried out to study the influence of different predamaged degrees on mechanical behaviors and damage evolution characteristics. The results show that the peak stress and axial strain of PD rock decrease gradually with the increase of predamaged degree. The yield stage of PD rock was significantly shorter than that of intact rock, and it entered the dilation deformation earlier. Furthermore, the constitutive equation of the PD rock based on energy was constructed and the damage evolution characteristics of PD rock were analyzed. It was observed that the bigger the predamaged degree was, the earlier the rock entered the damage, and the failure mode was more complex and crushed. The research results can provide the theoretical basis for the stability control of engineering disturbed rock.

Genomic tRNA copy numbers determine cytoplasmic tRNA abundances, which in turn influence translation efficiency, but the underlying mechanism is not well understood. Using the sea cucumber Apostichopus japonicus as a model, we combined genomic sequence, transcriptome expression and ecological food resource data to study its codon usage adaptation. The results showed that, unlike intragenic non-coding RNAs, transfer RNAs (tRNAs) tended to be transcribed independently. This may be attributed to their specific Pol III promoters that lack transcriptional regulation, which may underlie the correlation between genomic copy number and cytoplasmic abundance of tRNAs. Moreover, codon usage optimization was mostly restrained by a gene's amino acid sequence, which might be a compromise between functionality and translation efficiency. Genes for stress responses were highly optimized for most echinoderms, while enzymes for saponin biosynthesis (LAS, CYPs and UGTs) were especially optimized in sea cucumbers, which might promote saponin synthesis as a defence strategy. The genomic tRNA content of A. japonicus was positively correlated with amino acid content in its natural food particles, which should promote its efficiency in protein synthesis. We propose that coevolution between genomic tRNA content and codon usage of sea cucumbers facilitates their saponin synthesis and survival using food resources with low nutrient content.

Somatic embryogenesis (SE) is a developmental process in which somatic cells undergo dedifferentiation to become plant stem cells, and redifferentiation to become a whole embryo. SE is a prerequisite for molecular breeding and is an excellent platform to study cell development in the majority of plant species. However, the molecular mechanism involved in M. sativa somatic embryonic induction, embryonic and maturation is unclear. This study was designed to examine the differentially expressed genes (DEGs) and miRNA roles during somatic embryonic induction, embryonic and maturation. The cut cotyledon (ICE), non-embryogenic callus (NEC), embryogenic callus (EC) and cotyledon embryo (CE) were selected for transcriptome and small RNA sequencing. The results showed that 17,251 DEGs, and 177 known and 110 novel miRNAs families were involved in embryonic induction (ICE to NEC), embryonic (NEC to EC), and maturation (EC to CE). Expression patterns and functional classification analysis showed several novel genes and miRNAs involved in SE. Moreover, embryonic induction is an active process of molecular regulation, and hormonal signal transduction related to pathways involved in the whole SE. Finally, a miRNA–target interaction network was proposed during M. sativa SE. This study provides novel perspectives to comprehend the molecular mechanisms in M. sativa SE.

Glycogen synthase kinase 3 (GSK3) is a vital multifunctional molecule that is widely distributed in invertebrates and vertebrates. GSK3 is a highly conserved serine/threonine (Ser/Thr) protein kinase, which plays an important role in insulin, Wnt, and various signaling pathways. In this study, a GSK3 gene were identified in the genome of the Pacific white shrimp, Litopenaeus vannamei, and analyzed its gene structure, phylogeny, and expression profiles. The deduced LvGSK3 protein contains a highly conserved Ser/Thr protein kinase catalytic (S_TKc) domain, the LvGSK3 gene exhibited high expression in different early developmental stages, most adult tissues, and premolting stages. RNA interference of LvGSK3 significantly retarded the increment of body weight and affected the expressions of molting-related genes compared with control groups. These results will improve our understanding of the conserved structure and functions of the LvGSK3 gene and show potential applications of shrimp growth.

Penaeid shrimp (family Penaeidae) represents one of the most economically and ecologically important groups of crustaceans. However, their genome sequencing and assembly have encountered extreme difficulties during the last 20 years. In this study, based on our previous genomic data, we investigated the genomic characteristics of four penaeid shrimp species and identified potential factors that result in their poor genome assembly, including heterozygosity, polyploidization, and repeats. Genome sequencing and comparison of somatic cells (diploid) of the four shrimp species and a single sperm cell (haploid) of Litopenaeus vannamei identified a common bimodal distribution of K-mer depths, suggesting either high heterozygosity or abundant homo-duplicated sequences present in their genomes. However, penaeids have not undergone whole-genome duplication as indicated by a series of approaches. Besides, the remarkable expansion of simple sequence repeats was another outstanding character of penaeid genomes, which also made the genome assembly highly fragmented. Due to this situation, we tried to assemble the genome of penaeid shrimp using various genome sequencing and assembly strategies and compared the quality. Therefore, this study provides new insights about the genomic characteristics of penaeid shrimps while improving their genome assemblies.

The Pacific white shrimp Litopenaeus vannamei is an important aquaculture species and a promising model for crustacean biology. Here, the authors provide a reference genome assembly, and show that gene expansion is involved in the regulation of frequent molting as well as benthic adaptation of the shrimp.

Abstract Background Red clover (Trifolium pratense L.) is an important cool-season legume plant, which is the most widely planted forage legume after alfalfa. Although a draft genome sequence was published already, the sequences and completed structure of mRNA transcripts remain unclear, which limit further explore on red clover. Results In this study, the red clover transcriptome was sequenced using single-molecule long-read sequencing to identify full-length splice isoforms, and 29,730 novel isoforms from known genes and 2194 novel isoforms from novel genes were identified. A total of 5492 alternative splicing events was identified and the majority of alter spliced events in red clover was corrected as intron retention. In addition, of the 15,229 genes detected by SMRT, 8719 including 186,517 transcripts have at least one poly(A) site. Furthermore, we identified 4333 long non-coding RNAs and 3762 fusion transcripts. Conclusions We analyzed full-length transcriptome of red clover with PacBio SMRT. Those new findings provided important information for improving red clover draft genome annotation and fully characterization of red clover transcriptome.

Insulin-like peptide (ILP) has been identified in various crustaceans, but whether it has a similar function in regulating hemolymph glucose as vertebrate insulin is unclear. We analyzed the components of hemolymph sugar in the Pacific white shrimp, Litopenaeus vannamei, and investigated the changes of hemolymph glucose concentration and the expressions of ILP and glucose metabolism genes under different treatments. We found glucose was a major component of hemolymph sugar in shrimp. Starvation caused hemolymph glucose to rise first and then decline, and the raised hemolymph glucose after exogenous glucose injection returned to basal levels within a short time, indicating that shrimp have a regulatory mechanism to maintain hemolymph glucose homeostasis. In addition, injections of bovine insulin and recombinant LvILP protein both resulted in a fast decline in hemolymph glucose. Notably, RNA interference of LvILP did not significantly affect hemolymph glucose levels, but it inhibited exogenous glucose clearance. Based on the detection of glucose metabolism genes, we found LvILP might maintain hemolymph glucose stability by regulating the expression of these genes. These results suggest that ILP has a conserved function in shrimp similar to insulin in vertebrates and plays an important role in maintaining hemolymph glucose homeostasis.

Leaf senescence is a complex organized developmental stage limiting the yield of crop plants, and alfalfa is an important forage crop worldwide. However, our understanding of the molecular mechanism of leaf senescence and its influence on biomass in alfalfa is still limited. In this study, RNA sequencing was utilized to identify differentially expressed genes (DEGs) in young, mature, and senescent leaves, and the functions of key genes related to leaf senescence. A total of 163,511 transcripts and 77,901 unigenes were identified from the transcriptome, and 5,133 unigenes were differentially expressed. KEGG enrichment analyses revealed that ribosome and phenylpropanoid biosynthesis pathways, and starch and sucrose metabolism pathways are involved in leaf development and senescence in alfalfa. GO enrichment analyses exhibited that six clusters of DEGs are involved in leaf morphogenesis, leaf development, leaf formation, regulation of leaf development, leaf senescence and negative regulation of the leaf senescence biological process. The WRKY and NAC families of genes mainly consist of transcription factors that are involved in the leaf senescence process. Our results offer a novel interpretation of the molecular mechanisms of leaf senescence in alfalfa.

Hydrate is a common issue in the natural gas production, which can be accelerated by the presence of H2S and CO2. This paper is to present the current experience in a gas project in Sichuan, including application of several surveillance technologies for the benefit of hydrate prevention and management, to support production optimization. This sour gas project has high H2S and CO2 content. With the high deliverability at the wells, the project has used a two-choke configuration in the surface system to manage the surface pressure to feed the gas into the production process. Given a design of the 2-choke system, the sour gas is choked, heated, and then choked again, and finally flows to the Tri-Ethylene Glycol (TEG) dehydration unit. Hydrate formation risk normally exists downstream of the first choke and the second choke if the heater is not efficient, and at the filtration process upstream of the dehydration. Hydrate prevention had been considered during the design phase of the surface production facilities. Methyl-Ethylene Glycol (MEG) is therefore selected and injected to the upstream of the first choke to mix with the gas flow stream. Real time data surveillance (i.e., pressure and temperature) with digital gauges are installed in the areas with high probability of hydrate formation. The most important next step is the real-time data (i.e., gas rate, water rate, pressure, temperature and MEG injection rate), which are updated and the hydrate formation curves are plotted to display on the central control computer. Based on the relative position of the pressure and temperature, the integrated digital control system can be used to optimize production by controlling gas rate, heater temperature and MEG injection. In brief, with this visualized monitoring system in place, hydrate prevention has been visually and effectively managed and MEG consumption has been optimized to minimize the operational cost.

Abstract Background Due to the great advantages in selection accuracy and efficiency, genomic selection (GS) has been widely studied in livestock, crop and aquatic animals. Our previous study based on one full-sib family of Litopenaeus vannamei (L. vannamei) showed that GS was feasible in penaeid shrimp. However, the applicability of GS might be influenced by many factors including heritability, marker density and population structure etc. Therefore it is necessary to evaluate the major factors affecting the prediction ability of GS in shrimp. The aim of this study was to evaluate the factors influencing the GS accuracy for growth traits in L. vannamei. Genotype and phenotype data of 200 individuals from 13 full-sib families were used for this analysis. Results In the present study, the heritability of growth traits in L. vannamei was estimated firstly based on the full set of markers (23 K). It was 0.321 for body weight and 0.452 for body length. The estimated heritability increased rapidly with the increase of the marker density from 0.05 K to 3.2 K, and then it tended to be stable for both traits. For genomic prediction on the growth traits in L. vannamei, three statistic models (RR-BLUP, BayesA and Bayesian LASSO) showed similar performance for the prediction accuracy of genomic estimated breeding value (GEBV). The prediction accuracy was improved with the increasing of marker density. However, the marker density would bring a weak effect on the prediction accuracy after the marker number reached 3.2 K. In addition, the genetic relationship between reference and validation population could influence the GS accuracy significantly. A distant genetic relationship between reference and validation population resulted in a poor performance of genomic prediction for growth traits in L. vannamei. Conclusions For the growth traits with moderate or high heritability, such as body weight and body length, the number of about 3.2 K SNPs distributed evenly along the genome was able to satisfy the need for accurate GS prediction in the investigated L.vannamei population. The genetic relationship between the reference population and the validation population showed significant effects on the accuracy for genomic prediction. Therefore it is very important to optimize the design of the reference population when applying GS to shrimp breeding.

Abstract Exoskeleton construction is an important issue in shrimp. To better understand the molecular mechanism of exoskeleton formation, development and reconstruction, the transcriptome of the entire developmental process in Litopenaeus vannamei, including nine early developmental stages and eight adult-moulting stages, was sequenced and analysed using Illumina RNA-seq technology. A total of 117,539 unigenes were obtained, with 41.2% unigenes predicting the full-length coding sequence. Gene Ontology, Clusters of Orthologous Group (COG), the Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis and functional annotation of all unigenes gave a better understanding of the exoskeleton developmental process in L. vannamei. As a result, more than six hundred unigenes related to exoskeleton development were identified both in the early developmental stages and adult-moulting. A cascade of sequential expression events of exoskeleton-related genes were summarized, including exoskeleton formation, regulation, synthesis, degradation, mineral absorption/reabsorption, calcification and hardening. This new insight on major transcriptional events provide a deep understanding for exoskeleton formation and reconstruction in L. vannamei. In conclusion, this is the first study that characterized the integrated transcriptomic profiles cover the entire exoskeleton development from zygote to adult-moulting in a crustacean, and these findings will serve as significant references for exoskeleton developmental biology and aquaculture research.

Alternative splicing (AS) can enhance transcript diversity dramatically and play an important role in stress adaptation. Limited researches of AS have been reported in the Pacific white shrimp (Litopenaeus vannamei), which is an important aquaculture species in the world. Here, we performed a genome-wide identification of AS events in L. vannamei based on eight transcriptomes. We identified 38,781 AS events in the shrimp genome, and some of them were validated by polymerase chain reaction experiments. These AS events correspond to 9,209 genes, accounting for 36% of protein-coding genes in the shrimp genome. The number of AS events increased after virus or bacteria infection and low salinity stress. Type 1 AS genes (AS was initially activated) were mainly enriched in substance and energy metabolism, such as carbon metabolism and amino metabolism. However, type 2 AS genes (AS events changed) displayed specific enrichment under different stress challenges. Specifically, type 2 AS genes under biotic stresses were mainly enriched in the pathogenic pathway and immune network, and the AS genes under low salinity stress were significantly enriched for betalain biosynthesis. In summary, our study indicates that AS events are complex in shrimp and may be related to stress adaptation. These results will provide valuable resource for functional genomic studies on crustaceans.

As the main structural protein of muscle fiber, myosin is essential for multiple cellular processes or functions, especially for muscle composition and development. Although the shrimp possess a well-developed muscular system, the knowledge about the myosin family in shrimp is far from understood. In this study, we performed comprehensive analysis on the myosin genes in the genome of the Pacific white shrimp, Litopenaeus vannamei. A total of 29 myosin genes were identified, which were classified into 14 subfamilies. Among them, Myo2 subfamily was significantly expanded in the penaeid shrimp genome. Most of the Myo2 subfamily genes were primarily expressed in abdominal muscle, which suggested that Myo2 subfamily genes might be responsible for the well-developed muscular system of the penaeid shrimp. In situ hybridization detection showed that the slow-type muscle myosin gene was mainly localized in pleopod muscle and superficial ventral muscle of the shrimp. This study provides valuable insights into the evolutionary and functional characterization of myosin genes in shrimps, which provides clues for us to understand the well-developed muscular system of shrimp.

Improvements of growth traits are always the focus in selective breeding programs for the Pacific white shrimp Litopenaeus vannamei (L. vannamei). Identification of growth-related genes or markers can contribute to the application of modern breeding technologies, and thus accelerate the genetic improvement of growth traits. The aim of this study was to identify the genes and molecular markers associated with the growth traits of L. vannamei. A population of 200 individuals was genotyped using 2b-RAD techniques for genome-wide linkage disequilibrium (LD) analysis and genome-wide association study (GWAS). The results showed that the LD decayed fast in the studied population, which suggest that it is feasible to fine map the growth-related genes with GWAS in L. vannamei. One gene designated as LvSRC, encoding the class C scavenger receptor (SRC), was identified as a growth-related candidate gene by GWAS. Further targeted sequencing of the candidate gene in another population of 322 shrimps revealed that several non-synonymous mutations within LvSRC were significantly associated with the body weight (P < 0.01), and the most significant marker (SRC_24) located in the candidate gene could explain 13% of phenotypic variance. The current results provide not only molecular markers for genetic improvement in L. vannamei, but also new insights for understanding the growth regulation mechanism in penaeid shrimp.

Many echinoderms are regenerative species that exhibit exceptional regenerative capacity, and sea cucumber is a representative organism that could regenerate the whole intestine after evisceration. There are many signaling pathways participate in the regeneration process, but it is not clear which is essential for the intestinal regeneration. In this study, we performed genome-wide comprehensive analyses on these regeneration-related signaling pathways, and found the Wnt signaling pathway was one of the most conservative pathways among regenerative species. Additionally, among these signaling pathways, we found that the Wnt signaling pathway was the only one under positive selection in regenerative echinoderms, and the only one enriched by differentially expressed genes during the intestinal regeneration. Thus, it suggests both coding sequence and gene expression of the Wnt signaling pathway have been shaped by natural selection to provide the genetic architecture for intestinal regeneration. Wnt7, Fz7, and Dvl are the three positively selected genes and also happen to be three upstream genes in the Wnt signaling pathway. They are all significantly upregulated at the early stages of regeneration, which may contribute significantly to the early activation of Wnt signaling and the initiation of intestinal regeneration. Expression knockdown of Wnt7 and Dvl by RNA interference significantly inhibit intestinal extension, implying that they are essential for intestinal regeneration. As an important regeneration-related gene, the downstream gene c-Myc is also conserved and highly expressed during the whole regeneration stages, which may make the Wnt/c-Myc signaling to be an important way to promote intestinal regeneration. Therefore, it is reasonable to conclude that the Wnt signaling pathway is the chosen one to play an important role in intestinal regeneration of sea cucumbers, or even in the regeneration of other echinoderms.

Chitin is among the most important components of the crustacean cuticular exoskeleton and intestinal peritrophic matrix. With the progress of genomics and sequencing technology, a large number of gene sequences related to chitin metabolism have been deposited in the GenBank database in recent years. Here, we summarized the genes and pathways associated with the biosynthesis and degradation of chitins in crustaceans based on genomic analyses. We found that chitin biosynthesis genes typically occur in single or two copies, whereas chitin degradation genes are all multiple copies. Moreover, the chitinase genes are significantly expanded in most crustacean genomes. The gene structure and expression pattern of these genes are similar to those of insects, albeit with some specific characteristics. Additionally, the potential applications of the chitin metabolism genes in molting regulation and immune defense, as well as industrial chitin degradation and production, are also summarized in this review.

The insulin signaling (IIS) pathway plays an important role in the metabolism, growth, development, reproduction, and longevity of an organism. As a key member of the IIS pathway, insulin-like growth factor binding proteins (IGFBPs) are widely distributed a family in invertebrates and vertebrates that are critical in various aspects of physiology. As an important mariculture species, the growth of Pacific white shrimp, Litopenaeus vannamei, is one of the most concerning characteristics in this area of study. In this study, we identified three IGFBP genes in the genome of L. vannamei and analyzed their gene structures, phylogenetics, and expression profiles. LvIGFBP1 was found to contain three domains (the insulin growth factor binding (IB) domain, the Kazal-type serine proteinase inhibitor (Kazal) domain, and the immunoglobulin C-2 (IGc2) domain), while LvIGFBP2 and LvIGFBP3 only contained a single IB domain. LvIGFBP1 exhibited high expression in most tissues and different developmental stages, while LvIGFBP2 and LvIGFBP3 were only slightly expressed in hemocytes. The RNA interference of LvIGFBP1 resulted in a significantly smaller increment of body weight than that of control groups. These results will improve our understanding of the conservative structure and function of IGFBPs and show potential applications for the growth of shrimp.

The sesquiterpenoid methyl farnesoate (MF), a juvenile hormone (JH) analog, plays important roles in many physiological processes of crustaceans, such as morphogenesis, molting and reproduction. Juvenile hormone esterase-like (JHE-like) carboxylesterase (CXE) is a key enzyme in MF degradation, playing a significant role in regulating MF titer. However, its function is barely known in shrimp. In this study, a total of 21 JHE-like CXEs (LvCXEs) were characterized in Pacific white shrimp Litopenaeus vannamei, based on the full genome and multi-transcriptomic data. LvCXE has a conserved triplet catalytic site (Ser-Glu-His) and a characteristic GxSxG motif. Most LvCXEs were highly expressed in the hepatopancreas, which was the main site for MF degradation. LvCXEs containing a GESAG motif showed a specific expansion in the L. vannamei genome. Those GESAG-containing LvCXEs presented differential expressions at different larvae stages and different molting stages of L. vannamei, which suggested their potential functions in development and molting. Additionally, when the transcription level of CXEs was inhibited, it could lead to failed molt and death of L. vannamei. When we further detected the expression levels of the key ecdysone responsive transcription factors including LvE75, LvBr-C, LvHr3 and LvFtz-f1 after the CXE inhibitor was injected into L. vannamei, they all showed apparent down-regulation. These results suggested that the expansion of LvCXEs in the L. vannamei genome should contribute to the regulation of metamorphosis at larvae stages and frequent molting during the growth of L. vannamei.

Shoot branching is one of the most variable determinants of crop yield, and the signaling pathways of plant branches have become a hot research topic. As an important transcription factor in the B3 family, NGATHA1 (NGA1), plays an important role in regulating plant lateral organ development and hormone synthesis and transport, but few studies of the role of this gene in the regulation of plant growth and stress tolerance have been reported. In this study, the NGA1 gene was isolated from Medicago truncatula (Mt) and its function was characterized. The cis-acting elements upstream of the 5′ end of MtNGA1 and the expression pattern of MtNGA1 were analyzed, and the results indicated that the gene may act as a regulator of stress resistance. A plant expression vector was constructed and transgenic Arabidopsis plants were obtained. Transgenic Arabidopsis showed delayed flowering time and reduced branching phenotypes. Genes involved in the regulation of branching and flowering were differentially expressed in transgenic plants compared with wild-type plants. Furthermore, transgenic plants demonstrated strong tolerances to salt- and mannitol-induced stresses, which may be due to the upregulated expression of NCED3 (NINE-CIS-EPOXYCAROTENOID DIOXYGENASE 3) by the MtNGA1 gene. These results provide useful information for the exploration and genetic modification use of MtNGA1 in the future.

Cadmium (Cd) toxicity seriously affects the growth and development of plants, so studies on uptake, translocation, and accumulation of Cd in plants are crucial for phytoremediation. However, the molecular mechanism of the plant response to Cd stress remains poorly understood. The main objective of this study was to reveal differentially expressed genes (DEGs) under lower (BT2_5) and higher (BT43) Cd concentration treatments in creeping bentgrass. A total of 463,184 unigenes were obtained from creeping bentgrass leaves using RNA sequencing technology. Observation of leaf tissue morphology showed that the higher Cd concentration damages leaf tissues. Four key transcription factor (TF) families, WRKY, bZIP, ERF, and MYB, are associated with Cd stress in creeping bentgrass. Our findings revealed that these four TFs play crucial roles during the creeping bentgrass response to Cd stress. This study is mainly focused on the molecular characteristics of DEGs under Cd stress using transcriptomic analysis in creeping bentgrass. These results provide novel insight into the regulatory mechanisms of respond to Cd stress and enrich information for phytoremediation.

Apart from sharing common ancestry with chordates, sea cucumbers exhibit a unique morphology and exceptional regenerative capacity. Here we present the complete genome sequence of an economically important sea cucumber, A. japonicus, generated using Illumina and PacBio platforms, to achieve an assembly of approximately 805 Mb (contig N50 of 190 Kb and scaffold N50 of 486 Kb), with 30,350 protein-coding genes and high continuity. We used this resource to explore key genetic mechanisms behind the unique biological characters of sea cucumbers. Phylogenetic and comparative genomic analyses revealed the presence of marker genes associated with notochord and gill slits, suggesting that these chordate features were present in ancestral echinoderms. The unique shape and weak mineralization of the sea cucumber adult body were also preliminarily explained by the contraction of biomineralization genes. Genome, transcriptome, and proteome analyses of organ regrowth after induced evisceration provided insight into the molecular underpinnings of visceral regeneration, including a specific tandem-duplicated prostatic secretory protein of 94 amino acids (PSP94)-like gene family and a significantly expanded fibrinogen-related protein (FREP) gene family. This high-quality genome resource will provide a useful framework for future research into biological processes and evolution in deuterostomes, including remarkable regenerative abilities that could have medical applications. Moreover, the multiomics data will be of prime value for commercial sea cucumber breeding programs.

Crustins form a large family of antimicrobial peptides (AMPs) in crustaceans composed of four sub-groups (Types I-IV). Type II crustins (Type IIa or “Crustins” and Type IIb or “Crustin-like”) possess a typical hydrophobic N-terminal region and are by far the most representative sub-group found in penaeid shrimp. To gain insight into the molecular diversity of Type II crustins in penaeids, we identified and characterized a Type IIb crustin in Litopenaeus vannamei (Crustin-like Lv) and compared Type II crustins at both molecular and transcriptional levels. Although L. vannamei Type II crustins (Crustin Lv and Crustin-like Lv) are encoded by separate genes, they showed a similar tissue distribution (hemocytes and gills) and transcriptional response to the shrimp pathogens Vibrio harveyi and White spot syndrome virus (WSSV). As Crustin Lv, Crustin-like Lv transcripts were found to be present early in development, suggesting a maternal contribution to shrimp progeny. Altogether, our in silico and transcriptional data allowed to conclude that (1) each sub-type displays a specific amino acid signature at the C-terminal end holding both the cysteine-rich region and the whey acidic protein (WAP) domain, and that (2) shrimp Type II crustins evolved from a common ancestral gene that conserved a similar pattern of transcriptional regulation.

Molting is one of the most important biological processes in shrimp growth and development. All shrimp undergo cyclic molting periodically to shed and replace their exoskeletons. This process is essential for growth, metamorphosis, and reproduction in shrimp. However, the molecular mechanisms underlying shrimp molting remain poorly understood. In this study, we investigated global expression changes in the transcriptomes of the Pacific white shrimp, Litopenaeus vannamei, the most commonly cultured shrimp species worldwide. The transcriptome of whole L. vannamei was investigated by RNA-sequencing (RNA-seq) throughout the molting cycle, including the inter-molt (C), pre-molt (D0, D1, D2, D3, D4), and post-molt (P1 and P2) stages, and 93,756 unigenes were identified. Among these genes, we identified 5,117 genes differentially expressed (log2ratio ≥1 and FDR ≤0.001) in adjacent molt stages. The results were compared against the National Center for Biotechnology Information (NCBI) non-redundant protein/nucleotide sequence database, Swiss-Prot, PFAM database, the Gene Ontology database, and the Kyoto Encyclopedia of Genes and Genomes database in order to annotate gene descriptions, associate them with gene ontology terms, and assign them to pathways. The expression patterns for genes involved in several molecular events critical for molting, such as hormone regulation, triggering events, implementation phases, skelemin, immune responses were characterized and considered as mechanisms underlying molting in L. vannamei. Comparisons with transcriptomic analyses in other arthropods were also performed. The characterization of major transcriptional changes in genes involved in the molting cycle provides candidates for future investigation of the molecular mechanisms. The data generated in this study will serve as an important transcriptomic resource for the shrimp research community to facilitate gene and genome annotation and to characterize key molecular processes underlying shrimp development.

Crustacea, particularly Decapoda, contains many economically important species, such as shrimps and crabs. Crustaceans exhibit enormous (nearly 500-fold) variability in genome size. However, limited genome resources are available for investigating these species. Exopalaemon carinicauda Holthuis, an economical caridean shrimp, is a potential ideal experimental animal for research on crustaceans. In this study, we performed low-coverage sequencing and de novo assembly of the E. carinicauda genome. The assembly covers more than 95% of coding regions. E. carinicauda possesses a large complex genome (5.73 Gb), with size twice higher than those of many decapod shrimps. As such, comparative genomic analyses were implied to investigate factors affecting genome size evolution of decapods. However, clues associated with genome duplication were not identified, and few horizontally transferred sequences were detected. Ultimately, the burst of transposable elements, especially retrotransposons, was determined as the major factor influencing genome expansion. A total of 2 Gb repeats were identified, and RTE-BovB, Jockey, Gypsy, and DIRS were the four major retrotransposons that significantly expanded. Both recent (Jockey and Gypsy) and ancestral (DIRS) originated retrotransposons responsible for the genome evolution. The E. carinicauda genome also exhibited potential for the genomic and experimental research of shrimps.