Effects of Aeromonas hydrophila infection on the intestinal microbiota, transcriptome, and metabolomic of common carp (Cyprinus carpio).

Aeromonas hydrophila frequently has harmful effects on aquatic organisms. The intestine is an important defense against stress. In this study, we investigated the intestinal microbiota and transcriptomic and metabolomic responses of Cyprinus carpio subjected to A. hydrophila infection. The results showed that obvious variation in the intestinal microbiota was observed after infection, with increased levels of Firmicutes and Bacteroidetes and decreased levels of Proteobacteria. Several genera of putatively beneficial microbiota (Cetobacterium, Bacteroides, and Lactobacillus) were abundant, while Demequina, Roseomonas, Rhodobacter, Pseudoxanthomonas, and Cellvibrio were decreased; pathogenic bacteria of the genus Vibrio were increased after microbiota infection. The intestinal transcriptome revealed several immune-related differentially expressed genes associated with the cytokines and oxidative stress. The metabolomic analysis showed that microbiota infection disturbed the metabolic processes of the carp, particularly amino acid metabolism. This study provides insight into the underlying mechanisms associated with the intestinal microbiota, immunity, and metabolism of carp response to A. hydrophila infection; eleven stress-related metabolite markers were identified, including N-acetylglutamic acid, capsidiol, sedoheptulose 7-phosphate, prostaglandin B1, 8,9-DiHETrE, 12,13-DHOME, ADP, cellobiose, 1H-Indole-3-carboxaldehyde, sinapic acid and 5,7-dihydroxyflavone.

Comparative study on nutritional quality and serum biochemical indices of common carp (Cyprinus carpio) aged 11 to 13 months aged cultured in traditional ponds and land-based container aquaculture systems.

In the study, the physical characteristics, nutritional composition, mineral elements, volatile substances, and serum biochemistry of common carp (Cyprinus carpio) after rearing for two months were compared and analyzed to reveal the differences in muscle nutritional quality and serum biochemical indices between fish raised in traditional ponds (TP) and land-based container recirculating aquaculture systems (C-RAS). One hundred fish were selected from each aquaculture mode for the following experiments. Results show that: in terms of physical properties, C-RAS frozen seepage rate was significantly lower than that in TP (P < 0.05), the chewiness, gumminess, springiness, resilience, adhesiveness, cohesiveness, and shearing of C-RAS group were significantly higher than in TP (P < 0.01). Regarding muscle nutrients, the moisture, the crude lipid contents of C-RAS group were significantly lower than that of the TP group (P < 0.05), and the crude protein content of C-RAS group was extremely significantly higher than that in TP (P < 0.01). The saturated fatty acids (ΣSFA) and polyunsaturated fatty acids (ΣPUFA) in the muscle of C-RAS were significantly higher than that in TP (P < 0.01), and monounsaturated fatty acids (ΣMUFA) of C-RAS were significantly lower than that in TP (P < 0.01). Methionine content in C-RAS was significantly higher than that in TP (P < 0.05). In terms of mineral elements, the contents of K and Se in C-RAS were significantly higher than those in TP, and the content of Zn in C-RAS was significantly lower than that in TP (P < 0.05). In terms of volatile substances, nonanal, octanal, and benzaldehyde in C-RAS were significantly lower than those in TP(P < 0.01), 2,3-diethyl-5-methylpyrazine, ethyl 3-methylpentanoate, butyl formate were significantly higher than those in TP (P < 0.01). In terms of serum biochemistry, the glucose index in C-RAS was extremely significantly lower than that in TP group, total protein, albumin, alkaline phosphatase, and triglyceride were extremely significantly higher than those in TP (P < 0.01). Experiments show that the aquaculture mode has a large impact on the physical properties of the common carp, nutrients, mineral elements, volatile matter, and serum biochemical. In conclusion, the common carp cultured in C-RAS systems had better muscle nutritional quality, which provides a reference for future regulation of fish nutritional quality by C-RAS.

Investigation of hub ferroptosis-related genes and the immune landscape in recurrent pregnancy loss and unexplained infertility using bioinformatics analysis.

Background: Recurrent pregnancy loss (RPL) and unexplained infertility (UI) are common pregnancy disorders that affect women's physical and mental health and lack effective treatment. Endometrial factors are one factor that leads to RPL. The latest research indicates that ferroptosis and immunity are closely related to the normal physiological function of the endometrium and may play a role in the pathogenesis of RPL and UI. Therefore, the present study analyzed the relationship between ferroptosis genes and immune infiltration in RPL and UI. Methods: We downloaded the GSE165004 dataset and analyzed differences in ferroptosis-related genes (FRGs) between RPL and UI patients and healthy controls. Hub differentially expressed ferroptosis-related genes (DE-FRGs) were screened using the LASSO algorithm, the SVM-RFE algorithm and the protein-protein interaction (PPI) network. Differences in immune infiltration between healthy endometrium and RPL and UI endometrium was analyzed, and the relationship between hub DE-FRGs and immune cell infiltration was examined. Results: We extracted 409 FRGs and identified 36 up-regulated and 32 down-regulated DE-FRGs in RPL and UI. Twenty-one genes were screened using the LASSO regression algorithm, and 17 genes were screened using the SVM-RFE algorithm. We intersected the LASSO genes, SVM-RFE genes and PPI network proteins to obtain 5 hub DE-FRGs. Gene set enrichment analysis (GSEA) functional enrichment analysis results indicated that the cytokine-cytokine receptor interaction signaling pathway was the common pathway for hub DE-FRGs. T follicular helper cells were highly infiltrated in RPL and UI, and M1 and M2 macrophages were highly infiltrated. The expression levels of MAPK1 and RELA positively correlated with T follicular helper cells. Conclusions: Ferroptosis-related genes may disrupt endometrial functions and signaling pathways and lead to the occurrence of RPL and UI.

Immune-related gene ANGPT1 is an adverse biomarker for endometrial carcinoma.

BACKGROUND: Immunotherapy has made great strides in cancer treatment. Endometrial carcinoma (EC) has been 1 of the most common tumors among women. This study aimed to screen immune-related prognosis biomarkers for EC. METHODS: The transcriptome profiling and clinical data of EC were downloaded from The Carcinoma Genome Atlas (TCGA) public database, and differentially expressed genes (DEGs) were obtained through the limma package in R software. An immune-related genes (IRGs) list was collected from the ImmPort database. We constructed a free-scale gene co-expression network via weighted gene co-expression network analysis (WGCNA). Then, the intersection genes of the module genes which significantly related to EC, along with IRGs and DEGs were screened as the candidate genes for further analysis. We identified the hub gene via Venn analysis of the protein-protein interaction (PPI) network genes and the prognostic genes, and verified expression of the hub gene through Human Protein Atlas (HPA) and Gene Expression Omnibus (GEO) databases which provided the GSE17025 dataset. Furthermore, we used the CIBERSORT deconvolution algorithm to explore tumor immune cells infiltration in EC, and investigated correlations between the hub gene and immune cells. RESULTS: The differential expression analysis demonstrated that there were 900 up-regulated genes and 1,008 down-regulated genes in TCGA-UCEC (Uterine Corpus Endometrial Carcinoma) cohort. There were 74 candidate intersection genes in blue module genes, IRGs, and DEGs. Finally, angiopoietin 1 (ANGPT1) was identified as the hub gene in EC. Low expression of ANGPT1 was associated with better overall survival (OS) in EC patients. The expression of ANGPT1 was negatively correlated with regulatory T cells (Tregs), but positively correlated with resting memory cluster of differentiation 4 (CD4) T cells, activated dendritic cells (DCs), activated natural killer (NK) cells, and activated memory CD4 T cells (P<0.05, Spearman). A high-infiltrating regulatory T cell would improve the prognosis for EC patients. CONCLUSIONS: The gene ANGPT1 can increase the infiltration of T cells and improve the prognosis of EC patients.

Endogenous tassel-specific small RNAs-mediated RNA interference enables a novel glyphosate-inducible male sterility system for commercial production of hybrid seed in Zea mays L.

Hybrid crops produce higher yields than their inbred parents due to heterosis. For high purity of hybrid seeds, it is critical to eliminate self-pollination. Manual or mechanical removal of male parts (such as detasseling in maize) is labor-intensive, fuel and time-consuming, and can cause physical damage to female plants, resulting in significant seed yield reductions. Many male-sterility systems either require a maintainer for male-sterile line propagation or are often affected by environmental factors. Roundup® Hybridization System (RHS) utilizes glyphosate to induce male sterility, which effectively eliminates the need for maintainer lines and removal of male parts for commercial hybrid seed production. The first-generation RHS (RHS1) is based on low expression of a glyphosate-insensitive 5-enolpyruvylshikimate-3-phosphate synthase (CP4 EPSPS) in pollen. This report presents the second-generation RHS (RHS2) technology built on RNA interference (RNAi) combined with CP4 EPSPS. It utilizes maize endogenous male tissue-specific small interfering RNAs (mts-siRNAs) to trigger cleavage of the CP4 EPSPS mRNA specifically in tassels, resulting in glyphosate-sensitive male cells due to lack of the CP4 EPSPS protein. Male sterility is then induced by glyphosate application at the stages critical for pollen development, and the male-sterile plants are used as the female parent to produce hybrid seed. The endogenous mts-siRNAs are conserved across maize germplasms, and the inducible male sterility was replicated in representative germplasms through introgression of a CP4 EPSPS transgene containing the mts-siRNA target sequence. This technology combines the relative simplicity and convenience of a systemic herbicide spray methodology with targeted protein expression to create an inducible male sterility system for industrial production of row crop hybrid seeds in an environmentally-independent manner.

Improving hybrid seed production in corn with glyphosate-mediated male sterility.

BACKGROUND: Hybrid corn varieties exhibit benefits associated with heterosis and account for most of the corn acreage in the USA. Hybrid seed corn is produced by crossing a female parent which is male-sterile and therefore incapable of self-pollination with a male parent as the pollen donor. The majority of hybrid seed corn is produced by mechanical detasseling which involves physically removing the tassel, a process that is laborious and costly. RESULTS: Glyphosate-resistant corn was developed via expression of a glyphosate insensitive 5-enolpyruvyl-shikimate 3-phosphate synthase enzyme (CP4-EPSPS). Experimentation with molecular expression elements resulted in selective reduction of CP4-EPSPS expression in male reproductive tissues. The resulting plant demonstrated sterile tassel following glyphosate application with little to no injury to the rest of the plant. Using (14)C-glyphosate as a marker, we also examined the translocation of glyphosate to the tassel via spray application in a track sprayer to simulate field application. The results allowed optimization of spray parameters such as dose, spray timing and target to maximize tassel delivery of glyphosate for efficient sterilization. CONCLUSION: The Roundup hybridization system (RHS) is a novel process for hybrid seed production based on glyphosate-mediated male sterility. RHS replaces mechanical detasseling with glyphosate spray and greatly simplifies the process of hybrid seed corn production.

Metabolically engineered soybean seed with enhanced threonine levels: biochemical characterization and seed-specific expression of lysine-insensitive variants of aspartate kinases from the enteric bacterium Xenorhabdus bovienii.

Threonine (Thr) is one of a few limiting essential amino acids (EAAs) in the animal feed industry, and its level in feed rations can impact production of important meat sources, such as swine and poultry. Threonine as well as EAAs lysine (Lys) and methionine (Met) are all synthesized via the aspartate family pathway. Here, we report a successful strategy to produce high free threonine soybean seed via identification of a feedback-resistant aspartate kinase (AK) enzyme that can be over-expressed in developing soybean seed. Towards this goal, we have purified and biochemically characterized AK from the enteric bacterium Xenorhabdus bovienii (Xb). Site-directed mutagenesis of XbAK identified two key regulatory residues Glu-257 and Thr-359 involved in lysine inhibition. Three feedback-resistant alleles, XbAK_T359I, XbAK_E257K and XbAK_E257K/T359I, have been generated. This study is the first to kinetically characterize the XbAK enzyme and provide biochemical and transgenic evidence that Glu-257 near the catalytic site is a critical residue for the allosteric regulation of AK. Furthermore, seed-specific expression of the feedback-resistant XbAK_T359I or XbAK_E257K allele results in increases of free Thr levels of up to 100-fold in R(1) soybean seed when compared to wild-type. Expression of feedback-sensitive wild-type AK did not substantially impact seed Thr content. In addition to high Thr, transgenic seed also showed substantial increases in other major free amino acid (FAA) levels, resulting in an up to 3.5-fold increase in the total FAA content. The transgenic seed was normal in appearance and germinated well under greenhouse conditions.

RNA-binding protein-mediated translational repression of transgene expression in plants.

We have demonstrated that RNA-binding proteins from coliphages and yeast can function as translational repressors in plants. RNA sequences called translational operators were inserted at a cap-proximal position in the 5'-UTR of mRNAs of two reporter genes, gus or aroA:CP4. Translation of the reporter mRNAs was efficiently repressed when the RNA binding protein that specifically binds to its cognate operator was co-expressed. The efficiency of translational repression by RNA-binding protein positively correlated with the amount of binding protein in transformed plant cells. Detailed studies on coliphage MS2 coat protein-mediated translational repression also suggested that the efficiency of translational repression was position-dependent. A translational operator situated at the cap-proximal position was more efficient in conferring repression than one that was placed cap-distal. Translational repression can be an efficient means for regulation of transgene expression, thereby broadening current approaches for transgene regulation in plants.