Pathological and ultrastructural changes of Bellamya bengalensis under chronic carboxylic acid exposure at environmentally relevant levels: Inferences from general unified threshold model for survival (GUTS) predictions and hepatopancreatic integrity assessment.

This study aimed to understand the effects of freshwater acidification, driven by industrial runoff, agricultural activities, and atmospheric deposition, on the freshwater mollusk Bellamya bengalensis. By systematically investigating the impact of two common carboxylic acids, acetic acid (AA) and benzoic acid (BA), this research employed diverse toxicological, pathological, and ecological assessments. We explored survival predictions through the generic unified threshold model of survival (GUTS-SD), examined oxidative stress responses, and investigated hepatopancreatic alterations. In the experimental design, Bellamya bengalensis were subjected to environmentally relevant sublethal concentrations (10%, 20% LC50) of AA (39.77 and 79.54 mg/l) and BA (31.41 and 62.82 mg/l) over 28 days. Acute toxicity tests revealed increased LC50 values, indicating heightened toxicity with prolonged exposure, particularly due to the greater potency of benzoic acid compared to acetic acid. The GUTS-SD model provided accurate predictions of time-specific effects on populations, presenting long-term exposure (100 days) LC50 values for AA (263.7 mg/l) and BA (330.9 mg/l). Sequentially, the integrated biomarker response (IBR) analysis across study intervals highlighted the 28-day interval as the most sensitive, with GST emerging as the most responsive enzyme to oxidative stress induced by AA and BA. Histopathological and ultrastructural assessments of the hepatopancreas showed severe alterations, including necrosis, vacuolation and disrupted micro-villi, which were especially pronounced in higher BA exposure concentrations. These findings highlight the health and survival impacts of carboxylic acid toxicity on Bellamya bengalensis, emphasizing the need for proactive measures to mitigate acidification in aquatic ecosystems. The broader ecological implications underscore the importance of effective management and conservation strategies to address ongoing environmental challenges.

Isolation, characterization and antimicrobial properties of hepatopancreas lectin of the freshwater crab Oziotelphusanaga.

Lectins are versatile proteins that specifically recognize and interact with sugar moieties expressed on the cell surface. The potential of lectin in drug targeting and delivery has instigated interest to identify natural lectins. Crabs have been identified as a rich source of lectin because the innate immune system is activated on encounter of pathogens and helps in the production of lectin. Although the presence of lectins in crab's hemolymph is well documented, little information about lectin in hepatopancreas, a vital organ for immunity and digestion in crustaceans, is currently available. A calcium dependent lectin (75 kDa) was purified from the hepatopancreas of the freshwater crab Oziotelphusa naga by bioadsorption and fetuin linked Sepharose 4B affinity chromatography technique. The isolated hepatopancreas lectin is calcium dependent and maximum agglutination was observed with rabbit erythrocytes. The hemagglutinating activity of the hepatopancreas lectin was effectively inhibited by sugars, such as α-lactose, GlcNAc, trehalose and NeuAc. Compared to sialylated N-glycosylated proteins including transferrin and apo transferrin, sialylated O-glycosylated proteins like fetuin exhibited stronger inhibitory effect. The ability of erythrocytes to bind hepatopancreas lectin has been diminished by desialylation of the potent inhibitor, indicating the significance of sialic acid in lectin-ligand interactions. The purified hepatopancreas lectin showed a broad spectrum of antimicrobial activity against bacteria Staphylococcus aureus, Klebsiella pneumoniae, Proteus mirabilis, Pseudomonas aeruginosa, E. coli and fungi Candida albicans and Aspergillus niger. The findings of this study demonstrate the significance of hepatopancreas lectin as a multifunctional defense protein that inhibits the growth of bacteria and fungi.

Las Bolitas Syndrome in Penaeus vannamei Hatcheries in Latin America.

In September 2023, several hatcheries in Latin America experienced significant mortality rates, up to 90%, in zoea stage 2 of Penaeus vannamei. Observations of fresh mounts revealed structures resembling lipid droplets, similar to those seen in a condition known as "las bolitas syndrome". Routine histopathological examinations identified detached cells and tissues in the digestive tracts of affected zoea, contrasting with the typical algal cell contents seen in healthy zoea. Polymerase chain reaction (PCR) testing for over 20 known shrimp pathogens indicated minimal differences between diseased and healthy batches. Both groups tested negative for acute hepatopancreatic necrosis disease (AHPND) but positive for Vibrio species and Rickettsia-like bacteria in the diseased samples. Histological analyses of the affected zoea revealed characteristic tissue degeneration in the hepatopancreas, forming spheres that eventually migrated into the upper gut, midgut, and midgut caeca, a pathology identified as bolitas syndrome (BS). Microbiological assessments revealed Vibrio species at concentrations of 106 CFU zoea/g in affected zoea, approximately two orders of magnitude higher than in healthy zoea. Bacterial isolation from both healthy and BS-affected zoea on thiosulphate-citrate-bile salts-sucrose (TCBS) agar and CHROMagar™ (Paris, France), followed by identification using API 20E, identified six strains of Vibrio alginolyticus. Despite similarities to "las bolitas syndrome" in fresh mounts, distinct histopathological differences were noted, particularly the presence of sloughed cells in the intestines and variations in hepatopancreatic lobes. This study highlights the critical need for further research to fully understand the etiology and pathology of bolitas syndrome in zoea stage 2 of P. vannamei to develop effective mitigation strategies for hatchery operations.

The role of the carapace in the accumulation of metals from seawater in the green crab (Carcinus maenas): Studies with radio-labeled calcium, zinc, and nickel.

The role of the carapace in the uptake and storage of newly accumulated metals was investigated in the green crab exposed to environmentally relevant concentrations of calcium ([Ca] = 389 mg L-1 or 9.7 mmol L-1), zinc ([Zn] = 82 µg L-1 or 1.25 µmol L-1), and nickel ([Ni] = 8.2 µg L-1 or 0.14 µmol L-1) in 12 °C seawater, using radio-tracers (45Ca, 65Zn, 63Ni). After 24-h exposure, carapace exhibited the highest concentration of newly accumulated Ca, whereas carapace and gills exhibited the highest concentrations of both newly accumulated Zn and Ni relative to other tissues. For all three metals, the carapace accounted for >85 % of the total body burden. Acute temperature changes (to 2 °C and 22 °C) revealed the highest overall temperature coefficient Q10 (2.15) for Ca uptake into the carapace, intermediate Q10 for Ni (1.87) and lowest Q10 (1.45) for Zn. New Ca uptake into the carapace continued linearly with time for 24 h, new Zn uptake gradually deviated from linearity, whereas Ni uptake reached a plateau by 6 h. Attachment of a rubber membrane to the dorsal carapace, thereby shielding about 20 % of the total crab surface area from the external water, eliminated both new Zn and Ni incorporation into the shielded carapace, whereas 36 % of new Ca incorporation persisted. When recently euthanized crabs were exposed, new Zn uptake into the carapace remained unchanged, whereas Ca and Ni uptake were reduced by 89 % and 71 %, respectively. We conclude that the carapace is a very important uptake and storage site for all three metals. All of the uptake of new Zn and new Ni, and most of the uptake of new Ca into this tissue comes directly from the external water. For Zn, the mechanism involves only physicochemical processes, whereas for Ca and Ni, life-dependent processes make the major contribution.

Transcriptome-based network analysis reveals hub immune genes and pathways of hepatopancreas against LPS in Amphioctopus fangsiao.

The hepatopancreas is the biggest digestive organ in Amphioctopus fangsiao (A. fangsiao), but also undertakes critical functions like detoxification and immune defense. Generally, pathogenic bacteria or endotoxin from the gut microbiota would be arrested and detoxified in the hepatopancreas, which could be accompanied by the inevitable immune responses. In recent years, studies related to cephalopods immune have been increasing, but the molecular mechanisms associated with the hepatopancreatic immunity are still unclear. In this study, lipopolysaccharide (LPS), a major component of the cell wall of Gram-negative bacteria, was used for imitating bacteria infection to stimulate the hepatopancreas of A. fangsiao. To investigate the immune process happened in A. fangsiao hepatopancreas, we performed transcriptome analysis of hepatopancreas tissue after LPS injection, and identified 2615 and 1943 differentially expressed genes (DEGs) at 6 and 24 h post-injection, respectively. GO and KEGG enrichment analysis showed that these DEGs were mainly involved in immune-related biological processes and signaling pathways, including ECM-receptor interaction signaling pathway, Phagosome signaling pathway, Lysosome signaling pathway, and JAK-STAT signaling pathways. The function relationships between these DEGs were further analyzed through protein-protein interaction (PPI) networks. It was found that Mtor, Mapk14 and Atm were the three top interacting DEGs under LPS stimulation. Finally, 15 hub genes involving multiple KEGG signaling pathways and PPI relationships were selected for qRT-PCR validation. In this study, for the first time we explored the molecular mechanisms associated with hepatopancreatic immunity in A. fangsiao using a PPI networks approach, and provided new insights for understanding hepatopancreatic immunity in A. fangsiao.

Exploring the impact of nonylphenol exposure on Litopenaeus vannamei at the histological and molecular levels.

Nonylphenol (NP) is one of the common pollutants in the environment that have toxic effects on aquatic animals. Nevertheless, little is known about the possible toxicity mechanism of NP on the hepatopancreas of Litopenaeus vannamei. In the present study, the detrimental effects of NP on the hepatopancreas of the L. vannamei were explored at the histological and transcriptomic levels. The findings indicated that after NP exposed for 3, 12, and 48 h, the hepatopancreas histology was changed significantly. Transcriptomic analysis showed that a total of 4302, 3651, and 4830 differentially expressed genes (DEGs) were identified at 3, 12, and 48 h following NP exposure. All these DEGs were classified into 12 clusters according to the expression patterns at different time points. GO and KEGG enrichment analyses of DEGs were also performed, immunological, metabolic, and inflammatory related pathways, including arachidonic acid metabolism (ko00590), the PPAR signaling pathway (ko03320), and the regulation of TRP channels by inflammatory mediators (ko04750) were significantly enriched. Six DEGs were selected for validation by quantitative real-time PCR (qRT-PCR) and the results confirmed the reliability of transcriptome data. All results indicated that NP is toxic to L. vannamei by damaging the histopathological structure and disrupting the biological function. The findings would provide a theoretical framework for lowering or limiting the detrimental impacts of NP on aquaculture and help us to further study the molecular toxicity of NP in crustaceans.

Changes of bacterial communities and bile acid metabolism reveal the potential "intestine-hepatopancreas axis" in shrimp.

The interaction between the gut and the liver plays a significant role in individual health and diseases. Mounting evidence supports that bile acids are important metabolites in the bidirectional communication between the gut and the liver. Most of the current studies on the "gut-liver axis" have focused on higher vertebrates, however, few was reported on lower invertebrates such as shrimp with an open circulatory system. Here, microbiomic and metabolomic analyses were conducted to investigate the bacterial composition and bile acid metabolism in intestine, hemolymph and hepatopancreas of Penaeus vannamei fed diets supplemented with octanoic acid and oleic acid. After six days of feeding, the bacterial composition in intestine, hemolymph and hepatopancreas changed at different stages, with significant increases in the relative abundance of several genera such as Pseudomonas and Rheinheimera in intestine and hepatopancreas. Notably, there was a more similar bacterial composition in intestine and hepatopancreas at the genus level, which indicated the close communication between shrimp intestine and hepatopancreas. Meanwhile, higher content of some bile acids such as lithocholic acid (LCA) and α-muricholic acid (α-MCA) in intestine and lower content of some bile acids such as taurohyocholic acids (THCA) and isolithocholic acid (IsoLCA) in hepatopancreas were detected. Furthermore, Spearman correlation analysis revealed a significant correlation between bacterial composition and bile acid metabolism in intestine and hepatopancreas. The microbial source tracking analysis showed that there was a high proportion of intestine and hepatopancreas bacterial community as the source of each other. Collectively, these results showed a strong crosstalk between shrimp intestine and hepatopancreas, which suggests a unique potential "intestine-hepatopancreas axis" in lower invertebrate shrimp with an open circulatory system. Our finding contributed to the understanding of the interplay between shrimp intestine and hepatopancreas in the view of microecology and provided new ideas for shrimp farming and disease control.

Identification of toll-like receptor family and the immune function of new Sptlr-6 gene of Scylla paramamosain.

As a crucial member of pattern-recognition receptors (PRRs), the Tolls/Toll-like receptors (TLRs) gene family has been proven to be involved in innate immunity in crustaceans. In this study, nine members of TLR gene family were identified from the mud crab (Scylla paramamosain) transcriptome, and the structure and phylogeny of different SpTLRs were analyzed. It was found that different SpTLRs possessed three conserved structures in the TIR domain. Meanwhile, the expression patterns of different Sptlr genes in examined tissues detected by qRT-PCR had wide differences. Compared with other Sptlr genes, Sptlr-6 gene was significantly highly expressed in the hepatopancreas and less expressed in other tissues. Therefore, the function of Sptlr-6 was further investigated. The expression of the Sptlr-6 gene was up-regulated by Poly I: C, PGN stimulation and Vibrio parahaemolyticus infection. In addition, the silencing of Sptlr-6 in hepatopancreas mediated by RNAi technology resulted in the significant decrease of several conserved genes involved in innate immunity in mud crab after V. parahaemolyticus infection, including relish, myd88, dorsal, anti-lipopolysaccharide factor (ALF), anti-lipopolysaccharide factor 2 (ALF-2) and glycine-rich antimicrobial peptide (glyamp). This study provided new knowledge for the role of the Sptlr-6 gene in defense against V. parahaemolyticus infection in S. paramamosain.

Complete genome of Vibrio harveyi isolate K2014767 from the hepatopancreas of captive Caribbean spiny lobster (Panulirus argus).

The complete genome sequence is reported for Vibrio harveyi isolate K2014767, isolated from a captive Caribbean spiny lobster (Panulirus argus) during a species-specific mortality event in a public display aquarium in the United States.

Comparison of Body Characteristics, Carotenoid Composition, and Nutritional Quality of Chinese Mitten Crab (Eriocheir sinensis) with Different Hepatopancreas Redness.

In this study, we investigated the body characteristics, carotenoid composition, and nutritional quality of Eriocheir sinensis with different hepatopancreas redness (a*). We distributed the crabs into two groups based on the hepatopancreas a* values and compared their body characteristics, chroma, carotenoid composition, and protein, lipid, total sugar, amino acid, and fatty acid content via paired t-test. The results revealed that the relationships between hepatopancreas a* values and crab quality are sex specific. In female crabs, the differences in nutritional characteristics were evident mainly in the hepatopancreases and ovaries. In the redder hepatopancreases, the content of zeaxanthin and ß-carotene increased, and the levels of C22:6n3 and C20:5n3 decreased (p < 0.05). In the ovaries, the content of astaxanthin, canthaxanthin, ß-carotene, umami, and sweet amino acids were lower in the redder hepatopancreas crabs (p < 0.05). In male crabs, there were positive relationships between hepatopancreas a* and amino acid and fatty acid content. The content of leucine, arginine, and total umami amino acids in muscles and of unsaturated fatty acids and n-6 polyunsaturated fatty acids in hepatopancreases and testicles increased with increasing hepatopancreas a* values (p < 0.05). Therefore, the redder the hepatopancreas, the higher the nutritional quality of male crabs.

Comparative Transcriptome Analysis of the Hepatopancreas from Macrobrachium rosenbergii Exposed to the Heavy Metal Copper.

The contamination of aquatic ecosystems by the heavy metal copper (Cu) is an important environmental issue and poses significant risks to the physiological functions of aquatic organisms. Macrobrachium rosenbergii is one of the most important freshwater-cultured prawns in the world. The hepatopancreas of crustaceans is a key organ for immune defense, heavy metal accumulation, and detoxification, playing a pivotal role in toxicological research. However, research on the molecular response of the hepatopancreas in M. rosenbergii to Cu exposure is still lacking. In this study, the transcriptomic response in the hepatopancreas of M. rosenbergii was studied after Cu exposure for 3 and 48 h. Compared with the control group, 11,164 (7288 up-regulated and 3876 down-regulated genes) and 10,937 (6630 up-regulated and 4307 down-regulated genes) differentially expressed genes (DEGs) were identified after 3 and 48 h exposure, respectively. Most of these DEGs were up-regulated, implying that gene expressions were largely induced by Cu. Functional enrichment analysis of these DEGs revealed that immunity, copper homeostasis, detoxification, DNA damage repair, and apoptosis were differentially regulated by Cu. Seven genes involved in immunity, detoxification, and metabolism were selected for validation by qRT-PCR, and the results confirmed the reliability of RNA-Seq. All these findings suggest that M. rosenbergii attempts to resist the toxicity of Cu by up-regulating the expression of genes related to immunity, metabolism, and detoxification. However, with the excessive accumulation of reactive oxygen species (ROS), the antioxidant enzyme system was destroyed. As a result, DNA damage repair and the cellular stress response were inhibited, thereby exacerbating cell damage. In order to maintain the normal function of the hepatopancreas, M. rosenbergii removes damaged cells by activating the apoptosis mechanism. Our study not only facilitates an understanding of the molecular response mechanisms of M. rosenbergii underlying Cu toxicity effects but also helps us to identify potential biomarkers associated with the stress response in other crustaceans.

Concentrations, sources and health risk of bisphenols in red swamp crayfish (Procambarus clarkii) from South-Eastern China.

Bisphenol analogs (BPs) are extensively employed in commercial and industrial products and they have been found in a variety of environmental matrices and human samples. The red swamp crayfish (Procambarus clarkii) has been a trendy food in China in recent decades. However, the levels of BPs in Chinese crayfish and the associated hazards of human exposure remain unknown. Thus, in this study, the levels of eight BPs in crayfish gathered from five major provinces engaged in crayfish within the Yangtze River Basin were analyzed. Additionally, the health risks for humans by ingesting crayfish were calculated. BPs were frequently detected in crayfish tissues, indicating the wide occurrence of these chemicals. In comparison to other substitutions, BPA remains the dominant bisphenol analog. Most of the BPs were observed to accumulate in the hepatopancreas compared to the muscle, so consuming the hepatopancreas of crayfish is not recommended. With the exception of BPS, the Estimated Daily Intakes (EDIs) of the remaining BPs exceeded the Tolerable Daily Intake (TDI) specified by the European Food Safety Authority (EFSA) by a factor of 1.75-69.0. The mean hazard index (HI) values exceeded 1 for both hepatopancreas and muscle in all provinces, and the mean HI values for hepatopancreas were significantly higher than those for muscle, indicating potential health risks for local consumers.

The multi-omics analysis in the hepatopancreas of Eriocheir sinensis provides novel insights into the response mechanism of heat stress.

Under global warming, heat stress can induce the excessive production of reactive oxygen species, causing irreversible damage to aquatic animals. It is essential to predict potentially harmful impacts on aquatic organisms under heat stress. Eriocheir sinensis, a typical crustacean crab, is widely distributed in China, American and Europe. Parent E. sinensis need migrate to the estuaries to reproduce in winter, and temperature is a key environmental factor. Herein, we performed a comprehensive transcriptomic and proteomic analysis in the hepatopancreas of E. sinensis under heat stress (20 °C and 30 °C), focusing on heat shock protein family, antioxidant system, energy metabolism and immune defense. The results revealed that parent E. sinensis generated adaptative responses to maintain physiological function under 20 °C stress via the transcriptional up-regulation of energy metabolism enzymes, mRNA synthesis and heat shock proteins. The transcriptional inhibition of key enzymes related to energy metabolism implied that 30 °C stress may lead to the dysfunction of energy metabolism in parent E. sinensis. Meanwhile, parent E. sinensis also enhanced the expression of ferritin and phospholipase D at translational level, and the glutathione s-transferase and heat shock protein 70 at both transcriptional and translational levels, speculating that parent E. sinensis can strengthen antioxidant and immune capacity to resist oxidative stress under 30 °C stress. This study elucidated the potential molecular mechanism in response to heat stress of parent E. sinensis hepatopancreas. The preliminary selection of heat tolerance genes or proteins in E. sinensis can provide a reference for the population prediction and the study of evolutionary mechanism under heat stress in crabs.

The Substitution of Fishmeal with Yeast Culture in the Yellow Catfish (Pelteobagrus fulvidraco) Diet: Growth, Serum Biochemical Indices, and Intestinal and Hepatopancreatic Histology.

Yeast culture is a complex fermentation product consisting of fermentation substrate, yeast cells and their metabolites. This study investigated the potential of yeast culture in replacing fishmeal in the diet of yellow catfish (Pelteobagrus fulvidraco). First, a basal diet was formulated to contain 160 g/kg fishmeal (CON), and then the dietary fishmeal was decreased to 120, 80, 40 and 0 g/kg via yeast culture inclusion, respectively, to form another four isonitrogenous and isolipidic diets (YC-12, YC-8, YC-4 and YC-0). Yellow catfish (3.00 ± 0.10 g) were fed with the above five diets with triplicates per treatment and 40 fish per replicate. After 8 weeks of feeding, the weight gain (WG), protein efficiency rate and protein retention in the YC-12 group and the feed conversion ratio (FCR) in the YC-12 and YC-8 groups showed no significant differences to the CON group (p > 0.05), but the WG in the YC-8, YC-4 and YC-0 groups was significantly lower, and the FCR in the YC-4 and YC-0 groups was significantly higher than in the CON group (p < 0.05). In terms of the whole-body composition, only the crude lipid content in the YC-0 group decreased significantly (p < 0.05). Compared with the CON group, the aspartate aminotransferase and alanine aminotransferase activities and D-lactic acid content in the YC-0 group were significantly increased, and the total cholesterol content was significantly reduced (p < 0.05). The activities of catalase, superoxide dismutase, and alkaline phosphatase, as well as the content of complement C3 and immunoglobulin M, were significantly increased, while the MDA content was significantly reduced in the YC-12 and YC-8 groups (p < 0.05). There were no significant differences in the intestinal amylase and lipase activity among all the groups (p > 0.05), while the trypsin activity in the YC-12 and YC-8 groups, as well as the diamine oxidase in the YC-4 and YC-0 groups, were significantly higher than those in the CON group (p < 0.05). In the intestine histology, there was a significant decrease in the intestinal villus height in the YC-4 and YC-0 groups as well as in the villus width in the YC-0 group (p < 0.05). In the hepatopancreas histology, lipid droplets appeared in the YC-4 and YC-0 groups, and severe cell vacuolation was observed in the YC-0 group. As a summary, in a practical diet containing 160 g/kg fishmeal, yeast culture can effectively replace 40 g/kg fishmeal without negatively affecting the growth performance, nutrient utilization, serum immune and antioxidant, intestinal and hepatopancreas histology of yellow catfish.

Morinda citrifolia fruit extract enhances the resistance of Penaeus vannamei to Vibrio parahaemolyticus infection.

Vibrio parahaemolyticus is a gram-negative facultative anaerobic bacterium implicated as the causative agent of several shrimp diseases. As part of the effort to provide biocontrol and cost-effective treatments, this research was designed to elucidate the effect of Morinda citrifolia fruit extract on the immunity of Penaeus vannamei postlarvae (PL) to V. parahaemolyticus. The methanol extract of M. citrifolia was vacuum evaporated, and the bioactive compounds were detected using gas chromatography‒mass spectrometry (GC‒MS). Thereafter, P. vannamei PL diets were supplemented with M. citrifolia at different concentrations (0, 10, 20, 30, 40, and 50 mg/g) and administered for 30 days before 24 h of exposure to the bacterium V. parahaemolyticus. A total of 45 bioactive compounds were detected in the methanol extract of M. citrifolia, with cyclononasiloxane and octadecamethyl being the most abundant. The survival of P. vannamei PLs fed the extract supplement was better than that of the control group (7.1-26.7% survival greater than that of the control group) following V. parahaemolyticus infection. Shrimp fed 50 mg/g M. citrifolia had the highest recorded survival. The activities of digestive and antioxidant enzymes as well as hepatopancreatic cells were significantly reduced, except for those of lipase and hepatopancreatic E-cells, which increased following challenge with V. parahaemolyticus. Histological assessment of the hepatopancreas cells revealed reduced cell degeneration following the administration of the plant extracts (expecially those fed 50 mg/g M. citrifolia) compared to that in the control group. Therefore, the enhanced immunity against V. parahaemolyticus infection in P. vannamei could be associated with the improved hepatopancreas health associated with M. citrifolia fruit extract supplementation.

Differential responses of hepatopancreas transcriptome between fast and slow growth in giant freshwater prawns (Macrobrachium rosenbergii) fed a plant-based diet.

Efficient utilisation of plant-based diets in the giant freshwater prawn, Marcrobrachium rosenbergii, varies according to individual, suggesting that it might be associated with differences in physiological and metabolic responses. Therefore, we aimed to investigate the individual differences in the growth response of shrimp fed to a soybean-based diet (SBM). Two hundred shrimp were fed SBM for 90 days, and specific growth rate (SGR) was determined individually. Fast- and slow-growing shrimp (F-shrimp vs. S-shrimp), with the highest and lowest 5% SGRs, respectively, were sampled to determine haemolymph chemistry and carcass composition. The hepatopancreas of these shrimps were used for transcriptome analysis through RNA sequencing (RNA-Seq). The results showed no significant differences in haemolymph chemistry parameters. In terms of carcass proximate composition, F-shrimp exhibited higher protein composition than did S-shrimp, suggesting that F-shrimp have higher protein anabolism. Using RNA-seq and real-time reverse transcription polymerase chain reaction (qRT-PCR), the expression levels of several genes encoding physiologic and metabolic enzymes were found to be upregulated in F-shrimp compared to in S-shrimp, suggesting that these enzymes/proteins mediated the efficient use of SBM-based diets for growth promotion in shrimp. Various DEGs associated with the immune system were observed, indicating a difference in immune processes between F- and S-shrimp. The expression of several housekeeping genes was found to be upregulated in S-shrimp. Collectively, the upregulated expression of several enzymes associated with physiological and/or metabolic processes and increased protein anabolism may be attributed to the efficient use of SBM for maximal growth in shrimp.

Flow cytometric analysis of hepatopancreatic cells from Armadillidium vulgare highlights terrestrial isopods as efficient environmental bioindicators in ex vivo settings.

Several studies have reported the high bioindication capacity of Isopoda (Crustacea, Oniscidea), which is related to their important ability to accumulate contaminants, usefulness in soil ecotoxicology and bioindication activities. Any change in the isopod population, diversity and life cycle can indicate relevant pollution levels. The analysis of target tissues, such as the hepatopancreas, is another emerging approach (from a cytologic/histological level) to detect contaminant accumulation from different sources. In this study, tissue disaggregation procedures were optimised in the hepatopancreas, and flow cytometry (FC) was applied to detect cell viability and several cell functions. After disaggregation, two hepatopancreatic cell types, small (S) and big (B), were still recognisable: they differed in morphology and behaviour. The analyses were conducted for the first time on isopods from sites under different conditions of ecological disturbance through cytometric re-interpretation of ecological-environmental parameters. Significant differences in cell functional parameters were found, highlighting that isopod hepatopancreatic cells can be efficiently analysed by FC and represent standardisable, early biological indicators, tracing environmental-induced stress through cytologic/histologic analyses.

Salvia miltiorrhiza polysaccharide promotes the health of crayfish (Procambarus clarkii) by promoting hemocyte phagocytosis, protecting hepatopancreas and enhancing intestinal barrier function.

Plant polysaccharides as immunomodulators are considered one of the effective measures to reduce antibiotic therapy in aquaculture. The immunomodulatory function of Salvia miltiorrhiza polysaccharides (SMP) has been demonstrated and begun to be applied in vertebrates, but its potential effect on crustaceans is unclear. In this study, crayfish (Procambarus clarkii) was fed with 0 %, 0.3 %, 0.7 %, 1.1 %, and 1.5 % SMP for 4 weeks to investigate the effects of SMP on hemocytes phagocytosis, hepatopancreatic function, and intestinal barrier function. The results revealed that hemocyte phagocytic activity was increased in all SMP groups. During the process of hemocytes phagocytic recognition and formation of phagosomes and phagolysosomes, the mRNA expression levels of mas, hem, rab3, ctsb, and lamp-1 were up-regulated mainly in the 0.3 % SMP group. During the clearance phase of phagocytosis, respiratory burst activity, ROS level, T-SOD, CAT, GST, and LZM activities were mainly increased in the 1.5 % SMP group. Hepatopancreas AKP and GOT activity were no significant change in all SMP groups. ACP activity was significantly enhanced in the 1.1 % SMP group. The GPT activity of 0.3-0.7 % SMP group was significantly decreased. The 0.7 % SMP group had the highest intestinal fold height. The highest index values of OTUs, Ace, Chao, and Shannon were in the 0.3 % SMP group. The dietary addition of 0.3 % SMP led to a tendency of increased relative abundance of Firmicutes and Bacteroidota at the phylum level, while the relative abundance of Proteobacteria at the phylum level decreased. In conclusion, dietary SMP could promote crayfish health by enhancing phagocytosis, protecting hepatopancreas and enhancing intestinal barrier function. This study contributes to the theoretical foundation for exploring the potential application of plant polysaccharides in crustaceans.

Effects of Alkalinity Exposure on Antioxidant Status, Metabolic Function, and Immune Response in the Hepatopancreas of Macrobrachium nipponense.

The oriental river prawn Macrobrachium nipponense is an important freshwater economic species in China, producing huge economic benefits. However, M. nipponense shows lower alkali tolerance than fish species, thus genetic selection is urgently needed in order to improve alkali tolerance in this species. In the present study, the effects of alkalinity exposure on the hepatopancreas of M. nipponense were measured under the alkali concentrations of 0 (control), 4, 8, and 12 mmol/L with the exposure time of 96 h through histological observations, measurement of antioxidant enzymes, metabolic profiling analysis, and transcriptome profiling analysis. The present study identified that the low concentration of alkali treatment (<4 mmol/L) did not result in morphological changes in the hepatopancreas and activity changes in antioxidant enzymes, while high-alkali treatment (>8 mmol/L) damaged the normal structures of the lumen and vacuoles and significantly stimulated the levels of superoxide dismutase, catalase, and total antioxidant capacity, indicating these antioxidant enzymes play essential roles in the protection of the body from the damage caused by the alkali treatment. Metabolic profiling analysis revealed that the main enriched metabolic pathways of differentially expressed metabolites in the present study were consistent with the metabolic pathways caused by environmental stress in plants and other aquatic animals. Transcriptome profiling analysis revealed that the alkali concentration of <8 mmol/L did not lead to significant changes in gene expression. The main enriched metabolic pathways were selected from the comparison between 0 mmol/L vs. 12 mmol/L, and some significantly up-regulated genes were selected from these metabolic pathways, predicting these selected metabolic pathways and genes are involved in the adaptation to alkali treatment in M. nipponense. The expressions of Ras-like GTP-binding protein, Doublesex and mab-3 related transcription factor 1a, and Hypothetical protein JAY84 are sensitive to changes in alkali concentrations, suggesting these three genes participated in the process of alkali adaptation in M. nipponense. The present study identified the effects of alkalinity exposure on the hepatopancreas of M. nipponense, including the changes in antioxidant status and the expressions of metabolites and genes, contributing to further studies of alkali tolerance in this species.

Genome-Wide Identification of Vitellogenin Gene Family and Comparative Analysis of Their Involvement in Ovarian Maturation in Exopalaemon carinicauda.

Vitellogenin (Vtg) is a precursor of yolk proteins in egg-laying vertebrates and invertebrates and plays an important role in vitellogenesis and embryonic development. However, the Vtg family remains poorly characterized in Exopalaemon carinicauda, a major commercial mariculture species found along the coasts of the Yellow and Bohai Seas. In this study, 10 Vtg genes from the genomes of E. carinicauda were identified and characterized. Phylogenetic analyses showed that the Vtg genes in crustaceans could be classified into four groups: Astacidea, Brachyra, Penaeidae, and Palaemonidae. EcVtg genes were unevenly distributed on the chromosomes of E. carinicauda, and a molecular evolutionary analysis showed that the EcVtg genes were primarily constrained by purifying selection during evolution. All putative EcVtg proteins were characterized by the presence of three conserved functional domains: a lipoprotein N-terminal domain (LPD_N), a domain of unknown function (DUF1943), and a von Willebrand factor type D domain (vWD). All EcVtg genes exhibited higher expression in the female hepatopancreas than in other tissues, and EcVtg gene expression during ovarian development suggested that the hepatopancreas is the main synthesis site in E. carinicauda. EcVtg1a, EcVtg2, and EcVtg3 play major roles in exogenous vitellogenesis, and EcVtg3 also plays a major role in endogenous vitellogenesis. Bilateral ablation of the eyestalk significantly upregulates EcVtg mRNA expression in the female hepatopancreas, indicating that the X-organ/sinus gland complex plays an important role in ovarian development, mostly by inducing Vtg synthesis. These results could improve our understanding of the function of multiple Vtg genes in crustaceans and aid future studies on the function of EcVtg genes during ovarian development in E. carinicauda.