Molecular simulation on the interaction between trehalose and asymmetric lipid bilayer mimicking the membrane of human red blood cells.

Trehalose is widely acknowledged for its ability to stabilize plasma membranes during dehydration. However, the exact mechanism by which trehalose interacts with lipid bilayers remains presently unclear. In this study, we conducted atomistic molecular dynamic simulations on asymmetric model bilayers that mimic the membrane of human red blood cells at various trehalose and water contents. We considered three different hydration levels mimicking the full hydration to desiccation scenarios. Results indicate that the asymmetric distribution of lipids did not significantly influence the computed structural characteristics at full and low hydration. At dehydration, however, the order parameter obtained from the symmetric bilayer is significantly higher compared to those obtained from asymmetric ones. Analysis of hydrogen bonds revealed that the protective ability of trehalose is well described by the water replacement hypothesis at full and low hydration, while at dehydration other interaction mechanisms associated with trehalose exclusion from the bilayer may involve. In addition, we found that trehalose exclusion is not attributed to sugar saturation but rather to the reduction in hydration levels. It can be concluded that the protective effect of trehalose is not only related to the hydration level of the bilayer, but also closely tied to the asymmetric distribution of lipids within each leaflet.

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

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

Elephantopus scaber L. Polysaccharides Alleviate Heat Stress-Induced Systemic Inflammation in Mice via Modulation of Characteristic Gut Microbiota and Metabolites.

Elephantopus scaber L. (ESL) is a Chinese herb that is used both as a food and medicine, often being added to soups in summer in south China to relieve heat stress (HS), but its exact mechanism of action is unknown. In this study, heat-stressed mice were gavaged with ESL polysaccharides (ESLP) at 0, 150, 300, and 450 mg/kg/d-1 (n = 5) for seven days. The gut microbiota composition, short-chain fatty acids (SCFAs), seven neurotransmitters in faeces, expression of intestinal epithelial tight junction (TJ) proteins (Claudin-1, Occludin), and serum inflammatory cytokines were measured. The low dose of ESLP (ESLL) improved the adverse physiological conditions; significantly reduced the cytokines (TNF-α, IL-1ß, IL-6) and lipopolysaccharide (LPS) levels (p < 0.05); upregulated the expression of Claudin-1; restored the gut microbiota composition including Achromobacter and Oscillospira, which were at similar levels to those in the normal control group; significantly increased beneficial SCFAs like butyric acid and 5-HT levels in the faeces of heat-stressed mice; and significantly decreased the valeric acid and glutamic acid level. The level of inflammatory markers significantly correlated with the above-mentioned indicators (p < 0.05). Thus, ESLL reduced the HS-induced systemic inflammation by optimizing gut microbiota (Achromobacter, Oscillospira) abundance, increasing gut beneficial SCFAs like butyric acid and 5-HT levels, and reducing gut valeric and glutamic acid levels.

The role of DNA methylation reprogramming during sex determination and sex reversal in the Pacific oyster Crassostrea gigas.

DNA methylation is instrumental in vertebrate sex reversal. However, the mechanism of DNA methylation regulation regarding sex reversal in invertebrates is unclear. In this study, we used whole genome bisulfite sequencing (WGBS) to map single-base resolution methylation profiles of the Pacific oyster, including female-to-male (FMa-to-FMb) and male-to-female (MFa-to-MFb) sex reversal, as well as sex non-reversed males and females (MMa-to-MMb and FFa-to-FFb). The results showed that global DNA methylation levels increase during female-to-male sex reversals, with a particular increase in the proportion of high methylation levels (mCGs >0.75) and a decrease in the proportion of intermediate methylation levels (0.25 < mCGs <0.75). This increase in DNA methylation was mainly associated with the elevated expression of DNA methylase genes. Genome-wide methylation patterns of females were accurately remodeled to those of males after sex reversal, while the opposite was true for the male-to-female reversal. Those findings directly indicate that alterations in DNA methylation play a significant role in sex reversal in Pacific oysters. Comparative analysis of the DNA methylomes of pre- and post- sex reversal gonadal tissues (FMb-vs-FMa or MFb-vs-MFa) revealed that differentially methylated genes were mainly involved in the biological processes of sex determination or gonadal development. However critical genes such as Dmrt1, Foxl2 and Sox-like, which are involved in the putative sex determination pathway in Pacific oysters, showed almost an absence of methylation modifications, varying greatly from vertebrates. Additionally, comparative analysis of the DNA methylomes of sexual reversal and sex non-reversal (FMa-vs-FFa or MFa-vs-MMa) revealed that heat shock protein genes, such as Hsp68-like and Hsp70B, were important for the occurrence of sex reversal. These findings shed light on the epigenetic mechanisms underlying the maintenance of gonadal plasticity and the reversal of organ architecture in oysters.

Comparative Transcriptome Analysis of the Response to Vibrio parahaemolyticus and Low-Salinity Stress in the Swimming Crab Portunus trituberculatus.

Vibrio parahaemolyticus is one of the main pathogenic bacteria of Portunus trituberculatus and causes mass mortality of P. trituberculatus in aquaculture. In addition, low-salinity stimulation makes P. trituberculatus more susceptible to V. parahaemolyticus infections. In order to elucidate the molecular mechanism of resistance to V. parahaemolyticus in P. trituberculatus, comparative transcriptomic analysis of blood cells stimulated by low salinity and V. parahaemolyticus was carried out in this study. Transcriptome sequencing of low-salinity stress and pathogen infection at different time points was completed using Illumina sequencing technology. A total of 5827, 6432, 5362 and 1784 differentially expressed genes (DEGs) involved in pathways related to ion transport and immunoregulation were found under low-salinity stress at 12, 24, 48 and 72 h compared with the control at 0 h. In contrast, 4854, 4814, 5535 and 6051 DEGs, which were significantly enriched in Toll and IMD signaling pathways, were found at 12, 24, 48 and 72 h compared with the control at 0 h under V. parahaemolyticus infection. Among them, 952 DEGs were shared in the two treatment groups, which were mainly involved in apoptosis and Hippo signaling pathway. Cluster analysis screened 103 genes that were differentially expressed in two factors that were negatively correlated, including immunoglobulin, leukocyte receptor cluster family, scavenger receptor, macroglobulin and other innate-immune-related genes. These results provide data support for the analysis of the mechanisms of immunity to V. parahaemolyticus under low-salinity stress in P. trituberculatus and help to elucidate the molecular mechanisms by which environmental factors affect immunity.

Starvation-induced changes in sex ratio involve alterations in sex-related gene expression and methylation in Pacific oyster Crassostrea gigas.

Aquatic animals are subject to varying degrees of starvation stress in their natural habitats due to food limitations. Consequently, starvation is a crucial environmental factor for sex determination in many species; however, limited research has been conducted on the effects of starvation on sex determination in shellfish. Here, four full sibling families of Pacific oysters were established and subjected to starvation stress. The results demonstrated that starvation caused the sex ratio (female to male) to change from 1:0.78 to 1:1.44 and resulted in a delay in gonadal development. Further studies revealed that the expression levels of DNA methylation-related genes Dnmt1 (DNA methyltransferase 1), Dnmt3a/b (DNA methyltransferase 3a/b) and Tet3 (tet methylcytosine dioxygenase 3) decreased under starvation stress. Conversely, the upregulation of Dmrt1 (doublesex and mab-3 related transcription factor 1), a gene typically associated with males, in females suggests that the increased proportion of males may be linked to starvation-induced high expression of this particular gene. In addition, the gene Dgkd (diacylglycerol kinase delta), which is involved in the regulation of second messenger protein kinase C, was differentially methylated between males and females, with the methylation level of this gene gradually increasing with male development, while the methylation level of this gene decreased under starvation stress. Correlation analysis of Dgkd methylation levels with expression levels showed a negative correlation between DNA methylation and gene expression. Finally dual fluorescence reporter experiments confirmed that DNA methylation suppressed Dgkd expression in vitro. In summary, the results suggest that starvation may regulate Dgkd gene expression through DNA methylation and thus affect Dmrt1 expression, thereby causing sex reversal in the Pacific oyster. The outcomes resolved how environmental factors are involved in sex reversal from an epigenetic perspective and provided a theoretical basis for sex control breeding in the Pacific oyster.

Water Extract of Chrysanthemum indicum L. Flower Inhibits Capsaicin-Induced Systemic Low-Grade Inflammation by Modulating Gut Microbiota and Short-Chain Fatty Acids.

Systemic low-grade inflammation induced by unhealthy diet has become a common health concern as it contributes to immune imbalance and induces chronic diseases, yet effective preventions and interventions are currently unavailable. The Chrysanthemum indicum L. flower (CIF) is a common herb with a strong anti-inflammatory effect in drug-induced models, based on the theory of "medicine and food homology". However, its effects and mechanisms in reducing food-induced systemic low-grade inflammation (FSLI) remain unclear. This study showed that CIF can reduce FSLI and represents a new strategy to intervene in chronic inflammatory diseases. In this study, we administered capsaicin to mice by gavage to establish a FSLI model. Then, three doses of CIF (7, 14, 28 g·kg-1·day-1) were tested as the intervention. Capsaicin was found to increase serum TNF-α levels, demonstrating a successful model induction. After a high dose of CIF intervention, serum levels of TNF-α and LPS were reduced by 62.8% and 77.44%. In addition, CIF increased the α diversity and number of OTUs in the gut microbiota, restored the abundance of Lactobacillus and increased the total content of SCFAs in the feces. In summary, CIF inhibits FSLI by modulating the gut microbiota, increasing SCFAs levels and inhibiting excessive LPS translocation into the blood. Our findings provided a theoretical support for using CIF in FSLI intervention.

Overview of omics applications in elucidating the underlying mechanisms of biochemical and biological factors associated with meat safety and nutrition.

Over the years, significant technological discoveries have facilitated the improvement of meat-related research. Recent studies of complex and interactive factors contributing to variations in meat safety are increasingly focused on data-driven omics approaches such as proteomics. This review highlighted omics advances in elucidating the biochemical and biological actions on meat safety. Also, the impacts of the nutritional characteristics of meat and meat products on human health are emphasized. Future perspectives should explore multi-omics and in situ investigations to elucidate the implications in microbiological studies, including nutritional and health-related assessments. Also, creating meat safety assessment and prediction models based on biomarkers of meat safety traits will help to mitigate application constraints, thereby evaluating meat quality more accurately. This could provide a scientific basis for increasing the meat industry's profitability and producing high-quality meat and meat products for consumers. SIGNIFICANCE OF THE REVIEW: This review highlighted omics advances in elucidating underlying mechanisms of biochemical and biological factors associated with meat safety. Also, the impacts of meat proteins on human health are emphasized. Future perspectives should explore multi-omics and in situ investigations to elucidate the implications in microbiological studies, including nutritional and health-related assessments. Also, creating meat safety assessment and prediction models based on biomarkers of meat safety traits will help to mitigate application constraints, thereby evaluating meat quality more accurately. This could provide a scientific basis for increasing the meat industry's profitability and producing high-quality meat and meat products for consumers.

Early gonadal differentiation is associated with the antagonistic action of Foxl2 and Dmrt1l in the Pacific oyster.

As the second largest phylum in the zoological kingdom next to arthropods, the mechanism of gonadal differentiation in mollusca is quite complex. Currently, although much has been carried out on gonadal differentiation in the Pacific oyster, there is still unknown information that needs to be further explored. Here, analysis of the Foxl2 and Dmrt1l expression in samples at different development periods of male and female gonads as well as in annual gonad samples revealed that Log10 (Foxl2/Dmrt1l) values were an effective method for sex identification in oysters. In differentiated gonadal tissue, Log10 (Foxl2/Dmrt1l) values greater than 2 were females and less than 1 for males. Subsequent sequential sampling of the same individuals verified that Log10 (Foxl2/Dmrt1l) values greater than 2 for resting gonads would develop as females and less than 1 would develop as males in the future. Relative expression analysis of Foxl2 and Dmrt1l in the annual samples revealed a negative correlation between Log10 (Foxl2) and Log10 (Dmrt1l). Double fluorescence reporter validation results showed that DMRT1L protein was able to bind the Foxl2 promoter and repress its activity with a weak dosage effect. Antagonism between Dmrt1l and Foxl2 is therefore not restricted to vertebrates, and the competing regulatory networks are of great significance in the maintenance of gonadal sex in oysters after sexual differentiation. This study provides novel ideas and insights into the study of early gonadal differentiation in the adult oyster.

DNA methylation differences between male and female gonads of the oyster reveal the role of epigenetics in sex determination.

DNA methylation involved in sex determination mechanism by regulating gene expression related to sex determination networks are common in vertebrates. However, the mechanism linking epigenetics in invertebrates and sex determination has remained elusive. Here, methylome of the male and female gonads in the oyster Crassostrea gigas were conducted to explore the role of epigenetics in invertebrate sex determination. Comparative analysis of gonadal DNA methylation of females and males revealed that male gonads displayed a higher level of DNA methylation and a greater number of hypermethylated genes. Luxury genes presented hypomethylation, while housekeeping genes got hypermethylation. Genes in the conserved signaling pathways, rather than the key master genes in the sex determination pathway, were the major targets of substantial DNA methylation modification. The negative correlation of expression and promoter methylation in the diacylglycerol kinase delta gene (Dgkd) - a ubiquitously expressed gene - indicated DNA methylation may fine turn the expression of Dgkd and be involved in the process of sex determination. Dgkd can be used as an epigenetic marker to distinguish male C. gigas based on the different methylation regions in the promoter region. The results suggest that DNA methylation mechanisms played potential functional impacts in the sex determination in oysters, which is helpful to deepen the understanding of sex determination in invertebrate.

Preliminary Report on Intestinal Flora Disorder, Faecal Short-Chain Fatty Acid Level Decline and Intestinal Mucosal Tissue Weakening Caused by Litchi Extract to Induce Systemic Inflammation in HFA Mice.

Certain foods are known as "heating" foods in Chinese medicine. Over-consumption of these foods can lead to symptoms known as "heating up". These symptoms have been shown to be symptoms of systemic low-grade inflammation. However, the mechanism by which these foods cause inflammation is not clear. In this preliminary study, we investigated dysbacteriosis of the gut microbiota as a possible cause of inflammation by litchi, a typical "heating" food. A human flora-associated (HFA) mouse model (donor: n = 1) was constructed. After gavaging the mice with litchi extract suspension at low, medium and high doses (400, 800, 1600 mg/kg·d-1, respectively) (n = 3) for 7 days, the serum levels of inflammatory cytokines, gut microbiota, the concentration of SCFAs and the integrity of the intestinal mucosal barrier were measured. The results revealed significant increases in the abundance of Prevotella and Bacteroides. A significant increase in the abundance of Bilophila and a decrease in Megasomonas was observed in the high-dose group. High-dose litchi intervention led to a decrease of most SCFA levels in the intestine. It also caused a more than two-fold increase in the serum TNF-α level and LPS level but a decrease in the IL-1ß and IL-6 levels. Medium- and high-dose litchi intervention caused widening of the intestinal epithelial cell junction complex and general weakening of the intestinal mucosal barrier as well as reduced energy conversion efficiency of the gut microbiota. These data suggest that litchi, when consumed excessively, can lead to a low degree of systematic inflammation and this is linked to its ability to cause dysbacteriosis of the gut microbiota, decrease SCFAs and weaken the intestinal mucosal tissues.

Genome-Wide Differential DNA Methylomes Provide Insights into the Infertility of Triploid Oysters.

Chromosomal incompatibility and gene expression changes would affect the development of polyploid gonad and gamete formation. The role of epigenetics like DNA methylation in reproductive development is fully demonstrated in diploid animals. The lack of polyploid species and the infertility of polyploid animals, especially the odd ploidy, limit the study of epigenetic regulation mechanism of polyploid reproduction. Fertile and infertile individuals exist in triploid Pacific oyster Crassostrea gigas, which provide an interesting model for studies on the effect of epigenetic regulation on gonadal development. The whole genome single base resolution DNA methylomes in gonads of triploid females α (F-3nα), triploid females ß (F-3nß), triploid males α (M-3nα), triploid hermaphrodite predominantly males (HPM-3n), diploid females (F-2n), and diploid males (M-2n) were generated by using bisulfite-sequencing. The overall DNA methylation profiles in gene regions and transposable regions of fertile and infertile triploid oysters were consistent with those of diploid oysters. The DNA methylation level of CG context decreased in infertile triploid oysters, with more hypomethylated than hypermethylated regions, and the opposite is true in fertile triploid oysters. Genes harbored with differentially methylated regions (DMRs) in infertile triploids were mainly related to the metabolism pathways and the signal pathways. Correlation analysis indicated that the expression of gene transcriptions was generally positively associated with DNA methylation in gene body regions, and DMRs in infertile triploid oysters played significant roles in gonadal development as a possible critical epigenetic regulator of gonadal development gene transcriptional activity. These findings indicate a potential relationship between DNA methylation variability and gene expression plasticity in newly formed polyploidy. As far as we know, this is the first study revealing the epigenetic regulation of gonadal development in invertebrates based on fertile and infertile models, meanwhile providing a new mentality to explore the regulatory mechanisms of infertility in triploids.

Characterization of a chitinase-1 gene (PtCht-1) from a marine crab Portunus trituberculatus and its response to immune stress.

Chitinases play an important role in many biological processes in crustaceans, including molting, digestion, and immunity. In order to further explore the immune defense mechanism of chitinase in Portunus trituberculatus, the PtCht-1 gene was cloned by RACE (rapid-amplification of cDNA ends). This cDNA with a full length of 1910 bp, and an ORF (open reading frame) 1749 bp, coded for 582 amino acid residues and was classified into P. trituberculatus chitinase GH18-group4. It had the typical structural characteristics of GH18 chitinase family. Real-time PCR was used to analyze the expression of PtCht-1 in different tissues, molting stages, after pathogen infection, and low salinity (11‰). PtCht-1 was expressed in all tissues, with the highest expression in the hepatopancreas. In the hepatopancreas of different molting stages, the expression level decreased successively during post-molt stages (A/B), pre-molt stage (D) and inter-molt stage (C). Under normal circumstances, after artificial infection with WSSV and Vibrio parahaemolyticus, the expression of PtCht-1 in hepatopancreas reached the maximum at 48 h, and in hemolymph at 72 h and 24 h, respectively. Overall PtCht-1 expression was up-regulated compared with the control group. Low salinity stress significantly inhibited the expression of PtCht-1, up to 42 folds. Under low salinity stress, the time when WSSV infection reached the peak was markedly delayed by at least 24 h. The results of this study indicate that PtCht-1, as an immune factor, is likely involved in pathogen defense of P. trituberculatus, the immune function of which may be inhibited to some extent after low salinity stress.

Vibrio parahaemolyticus Infection in Mice Reduces Protective Gut Microbiota, Augmenting Disease Pathways.

Vibrio parahaemolyticus (Vp), a major food-borne pathogen, is responsible for severe infections such as gastroenteritis and septicemia, which may be accompanied by life-threatening complications. While studies have evaluated factors that affect the virulence of the pathogen, none have investigated the interaction of Vp with gut microbiota. To address this knowledge gap, we compared the effect of Vp on gut bacterial community structure, immunity, liver and kidney function, in pseudo germ-free (PGF) mice and normal (control) mice. Significant damage to the ileum was observed in normal mice compared with the PGF mice. The inflammatory factors IL-1ß, IL-6, and TNF-α in normal mice were ∼2.5-fold higher than in the PGF mice, and liver (ALT, AST, ALP) and kidney (BUN) function indices were ∼1.6-fold higher. The Vp infection substantially reduced species composition and richness of the gut microbial communities. In particular, there was a shift in keystone taxa, from protective species of genera Bacteroides, Lactobacillus, Bifidobacterium, and Akkermansia in the gut of control mice to opportunistic pathogens Enterobacteriaceae, Proteus, Prevotella, and Sutterella in Vp-infected mice, thus affecting microbiota-related biological functions in the mice. Specifically, pathways involved in infectious diseases and ion channels were significantly augmented in infected mice, while the pathways involved in metabolism, digestion and cell growth declined. We propose that the normal mice are more prone to Vp infection because of the alteration in gut-microbe-mediated functions. All these effects reduce intestinal resistance, with marked damage to the gut lining and pathogen leakage into the blood culminating in liver and kidney damage. These findings greatly advance our understanding of the mechanisms underlying interactions between Vp, the gut microbiota and the infected host.

Complete Genome of Bacillus velezensis CMT-6 and Comparative Genome Analysis Reveals Lipopeptide Diversity.

The complete genome sequence of Bacillus velezensis type strain CMT-6 is presented for the first time. A comparative analysis between the genome sequences of CMT-6 with the genome of Bacillus amyloliquefaciens DSM7T, B. velezensis FZB42, and Bacillus subtilis 168 revealed major differences in the lipopeptide synthesis genes. Of the above, only the CMT-6 strain possessed an integrated synthetase gene for synthesizing surfactin, iturin, and fengycin. However, CMT-6 shared 14, 12, and 10 other lipopeptide-producing genes with FZB42, DSM7T, and 168 respectively. The largest numbers of non-synonymous mutations were detected in 205 gene sequences that produced these three lipopeptides in CMT-6 and 168. Comparing CMT-6 with DSM7T, 58 non-synonymous mutations were detected in gene sequences that contributed to produce lipopeptides. In addition, InDels were identified in yczE and glnR genes. CMT-6 and FZB42 had the lowest number of non-synonymous mutations with 8 lipopeptide-related gene sequences. And InDels were identified in only yczE. The numbers of core genes, InDels, and non-synonymous mutations in genes were the main reasons for the differences in yield and variety of lipopeptides. These results will enrich the genomic resources available for B. velezensis and provide fundamental information to construct strains that can produce specific lipopeptides.

Quantitative Trait Loci Mapping and Marker Identification for Low Salinity Tolerance Trait in the Swimming Crab (Portunus trituberculatus).

Low salinity is one of the most important abiotic factors that directly affect the abundance of the swimming crab, Portunus trituberculatus. Quantitative trait loci (QTL) mapping could be helpful in identifying the markers and genes involved in low salinity tolerance. In this study, two QTLs of low salt tolerance were mapped on linkage group 17 (LG17, 2.6-5.2 cM) based on a high-density linkage map. Ninety-five markers related to low salinity tolerance were identified via association analysis, and seventy-nine low salt-related candidate genes (including ammonium transport, aldehyde dehydrogenase, and glucosyltransferase) were screened from draft genome of the species via these markers. This represents the first report of QTL mapping for low salinity tolerance in the swimming crab, which may be useful to elucidate salinity adaptation mechanisms.

Effect of media and fermentation conditions on surfactin and iturin homologues produced by Bacillus natto NT-6: LC-MS analysis.

Lipopeptides possess excellent broad spectrum antimicrobial activity. Different lipopeptides have their own unique chemical structures, properties and biological activities. Quantitative analysis of the lipopeptides iturin and surfactin and their homologues produced by Bacillus natto NT-6 subjected to different culture media, shaking speed of rotary shaker, and liquid and solid fermentation methods was conducted using LC-MS. For iturins, liquid-state fermentation in Landy medium at a shaking speed of 160 r min-1 was the most suitable for maximal homologue production. Addition of 0.4% attapulgite powder increased production by 1.92-fold; activated carbon significantly reduced production. For surfactin homologues, solid-state fermentation in potato dextrose broth medium at shaking speed > 160 r min-1 was the best. Addition of 0.4% attapulgite powder increased production by 1.96-fold; activated carbon had no effect. Thus it is clear that fermentation conditions can be manipulated to maximize iturin and surfactin production.

Transcriptomic analysis provides insight into the mechanism of salinity adjustment in swimming crab Portunus trituberculatus.

BACKGROUND: Low salinity is one of the main factors limiting the distribution and survival of marine species. As a euryhaline species, the swimming crab (Portunus trituberculatus) is adaptive to relatively low salinity. However, the mechanisms underlying salinity stress responses in P. trituberculatus is not very clear. OBJECTIVES: The primary objective of this study was to describe the salinity adaptation mechanism in P. trituberculatus. METHODS: The crabs were exposed to low salinity stress, and gill tissue was sampled at 0, 12, 36, 48 and 72 h and subjected to high throughput sequencing. Subsequently, we tested the accuracy and quality of the sequencing results, and then carried out GO and KEGG bioinformatics on the differentially expressed genes (DEG). RESULTS: Each sample yielded more than 1.1 Gb of clean data and 23 million clean reads. The process was divided into early (0-12 h), middle (12-48 h), and late phase (48-72 h). A total of 1971 (1373 up-regulated, 598 down-regulated), 1212 (364 up-regulated, 848 down-regulated), and 555 (187 up-regulated, 368 down-regulated) DEGs with annotations were identified during the three stages, respectively. DEGs were mainly associated with lipid metabolism energy metabolism, and signal transduction from the three stages, respectively. CONCLUSION: A substantial number of genes were modified by salinity stress, along with a few important salinity acclimation pathways. This work provides valuable information on the salinity adaptation mechanism in P. trituberculatus. In addition, the comprehensive transcript sequences reported in this study provide a rich resource for identification of novel genes in this and other crab species.

Crustacean hyperglycemic hormone of Portunus trituberculatus: evidence of alternative splicing and potential roles in osmoregulation.

The crustacean hyperglycemic hormone (CHH) gene of Portunus trituberculatus (Pt-CHH) consists of four exons and three introns spanning 3849 bp in size and generating two mature mRNA, Pt-CHH1, and Pt-CHH2. The primary gene transcript produces a cDNA encoding for the putative Pt-CHH2 from exons 1, 2, 3, and 4 and an alternative transcript encodes for a putative Pt-CHH1 peptide from exons 1, 2, and 4. A promoter fragment of about 3 kb was obtained by genomic walking. The tissue-specific expression pattern is examined by reverse transcriptase chain reaction, and the results show that Pt-CHH1 is detected in the eyestalk, brain, muscle, and blood. However, Pt-CHH2 is detected in the ganglia thoracalis and gill. The results indicate that the expression of Pt-CHH2 in the gill might suggest a potential role in osmoregulation. The Pt-CHH transcript level in the gill increases when the crab is exposed to low salinity. The injection of dsRNA for Pt-CHH causes a significant reduction in Pt-CHH2 transcript level and the activity of Na+/K+-ATPase, and carbonic anhydrase (CA) show a serious decrease. In conclusion, this study provides molecular evidence to support the osmoregulatory function of Pt-CHH2.

QTL Mapping and Marker Identification for Sex-Determining: Indicating XY Sex Determination System in the Swimming Crab (Portunus trituberculatus).

Sex determination is an important area of research, which has always had an intriguing aspect in evolutionary and developmental biology. Quantitative trait locus (QTL) mapping for sex will be helpful in clarifying the sex determination system. In this study, the sex QTL mapping of the swimming crab (Portunus trituberculatus) was performed based on a high-density linkage map, and a highly significant QTL specifically mapped on a single linkage group (LG) was firstly identified (LG24, LOD > 14). Twenty markers in the QTL region showed significant associations with sex by association analysis, of which heterogametic genotypes in males supported the XY sex determination mechanism. Two sex-specific markers at the family level were identified via segregation distortion analysis, which were known to be the most closely linked to the sex of P. trituberculatus. Based on sex marker sequences (Marker3840, Marker20320, and Marker10494), three potential sex-related genes were identified, and the quantitative real-time PCR results suggested that these genes were important in spermatogenesis or sex characteristics in males. Our results will contribute to the fine-mapping of sex-determining genes and clarify the sex determination mechanism of P. trituberculatus.