Activation Mechanism of CcGSDMEb-1/2 and Regulation for Bacterial Clearance in Common Carp (Cyprinus carpio).

Gasdermin E (GSDME), to date, is considered the only direct executor of the pyroptosis process in teleost and plays an important role in innate immunity. In common carp (Cyprinus carpio), there contains two pairs of GSDME (GSDMEa/a-like and GSDMEb-1/2), and the pyroptotic function and regulation mechanism of GSDME still remain unclear. In this study, we identified two GSDMEb genes of common carp (CcGSDMEb-1/2), which contain a conserved N-terminal pore-forming domain, C-terminal autoinhibitory domain, and a flexible and pliable hinge region. We investigated the function and mechanism of CcGSDMEb-1/2 in association with inflammatory and apoptotic caspases in Epithelioma papulosum cyprinid cells and discovered that only CcCaspase-1b could cleave CcGSDMEb-1/2 through recognizing the sites 244FEVD247 and 244FEAD247 in the linker region, respectively. CcGSDMEb-1/2 exerted toxicity to human embryonic kidney 293T cells and bactericidal activity through its N-terminal domain. Interestingly, after i.p. infection by Aeromonas hydrophila, we found that CcGSDMEb-1/2 were upregulated in immune organs (head kidney and spleen) at the early stage of infection, but downregulated in mucosal immune tissues (gill and skin). After CcGSDMEb-1/2 were knocked down and overexpressed in vivo and in vitro, respectively, we found that CcGSDMEb-1/2 could govern the secretion of CcIL-1ß and regulate the bacterial clearance after A. hydrophila challenge. Taken together, in this study, it was demonstrated that the cleavage mode of CcGSDMEb-1/2 in common carp was obviously different from that in other species and played an important role in CcIL-1ß secretion and bacterial clearance.

Functional analysis of the CTNS gene exonic variants predicted to affect splicing.

Cystinosis is a severe, monogenic systemic disease caused by variants in CTNS gene. Currently, there is growing evidence that exonic variants in many diseases can affect pre-mRNA splicing. The impact of CTNS gene exonic variants on splicing regulation may be underestimated due to the lack of routine studies at the RNA level. Here, we analyzed 59 exonic variants in the CTNS gene using bioinformatics tools and identified candidate variants that may induce splicing alterations by minigene assays. We identified six exonic variants that induce splicing alterations by disrupting the ratio of exonic splicing enhancers/exonic splicing silencers (ESEs/ESSs) or by interfering with the recognition of classical splice sites, or both. Our results help in the correct molecular characterization of variants in cystinosis and inform emerging therapies. Furthermore, our work suggests that the combination of in silico and in vitro assays facilitates to assess the effects of DNA variants driving rare genetic diseases on splicing regulation and will enhance the clinical utility of variant functional annotation.

Identification of seven variants in the col4a1 gene that alter RNA splicing by minigene assay.

Type IV collagen is an integral component of basement membranes. Mutations in COL4A1, one of the key genes encoding Type IV collagen, can result in a variety of diseases. It is clear that a significant proportion of mutations that affect splicing can cause disease directly or contribute to the susceptibility or severity of disease. Here, we analyzed exonic mutations and intronic mutations described in the COL4A1 gene using bioinformatics programs and identified candidate mutations that may alter the normal splicing pattern through a minigene system. We identified seven variants that induce splicing alterations by disrupting normal splice sites, creating new ones, or altering splice regulatory elements. These mutations are predicted to impact protein function. Our results help in the correct molecular characterization of variants in COL4A1 and may help develop more personalized treatment options.

Performance of the Aspergillus galactomannan lateral flow assay with a digital reader for the diagnosis of invasive aspergillosis: a multicenter study.

PURPOSE: The objective of this multicenter study was to compare the diagnostic performance of lateral flow assay (LFA) and enzyme-linked immunosorbent assay (ELISA) to detect the Dynamiker Aspergillus Galactomannan levels in serum and bronchoalveolar lavage fluid (BALF) samples for I. METHODS: We registered 310 clinically suspected Aspergillus infection patients from December 2021 to February 2023 and classified them into subgroups as the "IA group" and "non-IA group" based on the latest EORTC/MSG guidelines. The immunoassays were analyzed by LFA and ELISA respectively. RESULTS: Galactomannan was examined using LFA, and serum and BALF samples demonstrated sensitivities of 82.57% and 89.47%, specificities of 90.76% and 92.00%, PPVs of 89.11% and 96.23%, and NPVs of 85.04% and 79.31%, respectively. Galactomannan was observed using two assays in serum and BALF samples and showed PPAs of 95.11% and 93.33%, NPAs of 89.19% and 96.30%, and TPAs of 92.47% and 94.25%, respectively. The ROC curve demonstrated that LFA had optimum diagnostic value when the index value (I value) = 0.5, the sensitivity was 84.94%, and the specificity was 90.97%. CONCLUSION: Compared to the ELISA method, the LFA has shown excellent performance for the diagnosis of IA in serum and BALF sample and can be used as an assay for the early diagnosis of patients with IA. The dynamic change in galactomannan levels may be useful for assessing treatment response.

H6N2 reassortant avian influenza virus isolate in wild birds in Jiangxi Province, China.

H6 avian influenza virus is widely prevalent in wild birds and poultry and has caused human infection in 2013 in Taiwan, China. During our active influenza surveillance program in wild waterfowl at Poyang Lake, Jiangxi Province, an H6N2 AIV was isolated and named A/bean goose/JiangXi/452-4/2013(H6N2). The isolate was characterized as a typical low pathogenic avian influenza virus (LPAIV) due to the presence of the amino acid sequence PQIETR↓GLFGAI at the cleavage site of the hemagglutinin (HA) protein. The genetic evolution analysis revealed that the NA gene of the isolate originated from North America and exhibited the highest nucleotide identity (99.29%) with a virus recovered from wild bird samples in North America, specifically A/bufflehead/California/4935/2012(H11N2). Additionally, while the HA and PB1 genes belonged to the Eurasian lineage, they displayed frequent genetic interactions with the North American lineage. The remaining genes showed close genetic relationships with Eurasian viruses. The H6N2 isolate possessed a complex genome, indicating it is a multi-gene recombinant virus with genetic material from both Eurasian and North American lineages.

EEG rhythm separation and time-frequency analysis of fast multivariate empirical mode decomposition for motor imagery BCI.

Motor imagery electroencephalogram (EEG) is widely employed in brain-computer interface (BCI) systems. As a time-frequency analysis method for nonlinear and non-stationary signals, multivariate empirical mode decomposition (MEMD) and its noise-assisted version (NA-MEMD) has been widely used in the preprocessing step of BCI systems for separating EEG rhythms corresponding to specific brain activities. However, when applied to multichannel EEG signals, MEMD or NA-MEMD often demonstrate low robustness to noise and high computational complexity. To address these issues, we have explored the advantages of our recently proposed fast multivariate empirical mode decomposition (FMEMD) and its noise-assisted version (NA-FMEMD) for analyzing motor imagery data. We emphasize that FMEMD enables a more accurate estimation of EEG frequency information and exhibits a more noise-robust decomposition performance with improved computational efficiency. Comparative analysis with MEMD on simulation data and real-world EEG validates the above assertions. The joint average frequency measure is employed to automatically select intrinsic mode functions that correspond to specific frequency bands. Thus, FMEMD-based classification architecture is proposed. Using FMEMD as a preprocessing algorithm instead of MEMD can improve the classification accuracy by 2.3% on the BCI Competition IV dataset. On the Physiobank Motor/Mental Imagery dataset and BCI Competition IV Dataset 2a, FMEMD-based architecture also attained a comparable performance to complex algorithms. The results indicate that FMEMD proficiently extracts feature information from small benchmark datasets while mitigating dimensionality constraints resulting from computational complexity. Hence, FMEMD or NA-FMEMD can be a powerful time-frequency preprocessing method for BCI.

Psychosocial factors affect the occurrence of nonsuicidal self-injury in adolescents with major depressive disorder through chain mediation.

In the adolescent group, about half of adolescents with major depressive disorder (MDD) have NSSI. Psychosocial factors are associated with the development of NSSI. Clarifying the relationship between psychosocial factors and NSSI in adolescents with MDD can help us achieve early prevent. Demographic data, Hamilton Depression Scale-24 (HAMA24), childhood trauma questionnaire, emotional intelligence scale and interpersonal reactivity index were collected from 187 adolescents with MDD. Use ANOVA, Chi-square test, Binary Logistic Regression, Pearson correlation analysis, Mediation effect analysis and the Structural Equation Model for data analysis. The results of ANOVA showed that there was significant difference between the two groups in HAMD24 total score, impulsiveness, emotional intelligence, and empathy (p < 0.05). In the regression analysis, women, depression degree, motor impulsiveness (MI), personal distress (PD) and appraisal of other's emotions empathy were the risk factors for MDD adolescents to produce NSSI behavior. Among the indicators that were significantly related to MDD and NSSI, MI and PD mediate the relationship between MDD and NSSI. The structural equation model showed that MDD, PD and MI had a direct impact on NSSI, but PD and MI had multiple intermediary effected in the relationship between MDD and NSSI. Emotional intelligence, emotional neglect and cognitive impulsiveness indirectly affected the occurrence of NSSI behavior. Impulsiveness, personal distress, emotional neglect, and emotional intelligence are important risk factors that affect NSSI behavior in adolescents with MDD, and they affect the occurrence of NSSI in adolescents with MDD through chain mediation.

Polarized M2 macrophages induced by glycosylated nano-hydroxyapatites activate bone regeneration in periodontitis therapy.

AIM: To investigate the association between periodontal macrophage polarization states and the alveolar bone levels, and to assess whether glycosylated nano-hydroxyapatites (GHANPs) could improve bone regeneration in periodontitis by inducing macrophage M2 polarization. MATERIALS AND METHODS: The change of macrophage polarization state in inflammatory periodontal tissues (with bone loss) was examined using clinical gingival samples. The relationship between macrophage phenotype and bone level in periodontal bone loss and repair was evaluated using a mouse periodontitis model. The effect of GHANPs on macrophage polarization was assessed by the in vitro model of lipopolysaccharide (LPS)-stimulated inflammation. The polarization-related markers were detected by immunofluorescence staining, real-time polymerase chain reaction and enzyme-linked immunosorbent assay analysis. The therapeutic effect of GHANPs on alveolar bone loss was explored in experimental periodontitis by histological staining and micro-CT analysis. RESULTS: A lower macrophage M2/M1 ratio was observed in periodontitis-affected human gingival tissues. The results of animal experiments demonstrated a positive correlation between a lower Arg-1/iNOS ratio and accelerated alveolar bone loss; also, the proportion of Arg-1-positive macrophages increased during bone repair and regeneration. The administration of GHANPs partially restored M2 macrophage polarization after LPS stimulation. GHANPs increased alveolar bone repair and regeneration in experimental periodontitis induced by ligation, potentially related to their macrophage M2 transition regulation. CONCLUSIONS: The findings of this study indicate that the induction of macrophage M2 polarization can be considered a viable approach for enhancing inflammatory bone repair. Additionally, GHANPs show potential in the clinical treatment of periodontitis.

A novel heterozygous variant of the SALL1 gene with atypical Townes-Brocks syndrome phenotypes in Chinese family.

Townes-Brocks syndrome (TBS) is an autosomal dominant disorder characterised by the triad of anorectal, thumb, and ear malformations. It may also be accompanied by defects in kidney, heart, eyes, hearing, and feet. TBS has been demonstrated to result from heterozygous variants in the SALL1 gene, which encodes zinc finger protein believed to function as a transcriptional repressor. The clinical characteristics of an atypical TBS phenotype patient from a Chinese family are described, with predominant manifestations including external ear dysplasia, unilateral renal hypoplasia with mild renal dysfunction, and hearing impairment. A novel heterozygous variant c.3060T>A (p.Tyr1020*) in exon 2 of the SALL1 gene was identified in this proband. Pyrosequencing of the complementary DNA of the proband revealed that the variant transcript accounted for 48% of the total transcripts in peripheral leukocytes, indicating that this variant transcript has not undergone nonsense-mediated mRNA decay. This variant c.3060T > A is located at the terminal end of exon 2, proximal to the 3' end of the SALL1 gene, and exerts a relatively minor impact on protein function. We suggest that the atypical TBS phenotype observed in the proband may be attributed to the truncated protein retaining partial SALL1 function.

Buying a Wealthy Dream: Determinants of Rural Residents' Lottery Purchase Behavior in China.

The expansion of illicit lottery activities has caused significant harm to both agricultural production and the livelihood of rural residents. An analysis of the factors that influence rural residents' participation in underground lotteries can provide crucial insight for regulating the lottery industry's development. This study examines the present state of rural residents' participation in underground lotteries, investigates the factors that impact their participation using the Double-Hurdle model, and further employs the ISM model to evaluate the correlations and hierarchical structure among the factors, using field survey data collected from 603 rural residents in S Province, China. The findings reveal that 53.07% of the respondents have participated in underground lotteries. Risk preference, information acquisition, social networks, age, education, family burden, percentage of agricultural labor, agricultural business scale, and household income significantly affect whether residents purchase lotteries (WPL). All factors, except the percentage of agricultural labor and agricultural business scale, also significantly impact underground lottery spending (LS). Among the significant influencing factors, rural residents' risk preference is the direct surface factor, whereas agricultural business scale, household income, information acquisition, and social networks are the middle indirect factors, and residents' age, education, family burden, and percentage of agricultural labor are the deep-rooted factors. It is recommended to regulate the lottery industry's development can be achieved by enhancing entertainment and cultural activities, expanding economic opportunities, enhancing rural education, increasing law awareness, and improving the public welfare lottery business model. The conclusions offer a valuable reference point for the standardized development of the lottery industry and the promotion of social stability in rural areas.

Regulation and resilience: Panarchy analysis in forest socio-ecosystem of Northeast National Forest Region, China.

This paper examines the socio-ecological resilience within China's Northeast National Forest Region (NNFR), focusing on the implications of climate change for forest management and carbon sequestration. It offers a critical assessment of the Natural Forest Protection Program (NFPP) and the associated logging ban policy, recognizing their pivotal contributions to forest conservation but also identifying the shortcomings of a one-size-fits-all approach. Integrating panarchy theory, the study proposes sustainable management practices that align ecological dynamics with societal needs, emphasizing nature-based solutions. The overarching aim is to bolster the long-term resilience and enhance the carbon sequestration potential of the NNFR's forests. It aims to inform global environmental strategy with lessons from the NNFR, advocating for integrated approaches that ensure both ecological sustainability and community prosperity. This approach seeks to provide a comprehensive and effective strategy for addressing environmental challenges, ensuring both ecological integrity and community well-being.

Calcium signal regulated carbohydrate metabolism in wheat seedlings under salinity stress.

This study aimed to explore the mechanism by which calcium (Ca) signal regulated carbohydrate metabolism and exogenous Ca alleviated salinity toxicity. Wheat seedlings were treated with sodium chloride (NaCl, 150 mM) alone or combined with 500 µM calcium chloride (CaCl2), lanthanum chloride (LaCl3) and/or ethylene glycol tetraacetic acid (EGTA) to primarily analyse carbohydrate starch and sucrose metabolism, as well as Ca signaling components. Treatment with NaCl, EGTA, or LaCl3 alone retarded wheat-seedling growth and decreased starch content accompanied by weakened ribulose-1,5-bisphosphate carboxylation/oxygenase (Rubisco) and Rubisco activase activities, as well as enhanced glyceraldehyde-3-phosphate dehydrogenase, phosphoglycerate kinase, alpha-amylase, and beta-amylase activities. However, it increased the sucrose level, up-regulated the sucrose phosphate synthase (SPS) and sucrose synthase (SuSy) activities and TaSPS and TaSuSy expression together, but down-regulated the acid invertase (SA-Inv) and alkaline/neutral invertase (A/N-Inv) activities and TaSA-Inv and TaA/N-Inv expression. Except for unchanged A/N-Inv activities and TaA/N-Inv expression, adding CaCl2 effectively blocked the sodium salt-induced changes of these parameters, which was partially eliminated by EGTA or LaCl3 presence. Furthermore, NaCl treatment also significantly inhibited Ca-dependent protein kinases and Ca2+-ATPase activities and their gene expression in wheat leaves, which was effectively relieved by adding CaCl2. Taken together, CaCl2 application effectively alleviated the sodium salt-induced retardation of wheat-seedling growth by enhancing starch anabolism and sucrose catabolism, and intracellular Ca signal regulated the enzyme activities and gene expression of starch and sucrose metabolism in the leaves of sodium salt-stressed wheat seedlings.

Identified eleven exon variants in PKD1 and PKD2 genes that altered RNA splicing by minigene assay.

BACKGROUND: Autosomal dominant polycystic kidney disease (ADPKD) is a common monogenic multisystem disease caused primarily by mutations in the PKD1 gene or PKD2 gene. There is increasing evidence that some of these variants, which are described as missense, synonymous or nonsense mutations in the literature or databases, may be deleterious by affecting the pre-mRNA splicing process. RESULTS: This study aimed to determine the effect of these PKD1 and PKD2 variants on exon splicing combined with predictive bioinformatics tools and minigene assay. As a result, among the 19 candidate single nucleotide alterations, 11 variants distributed in PKD1 (c.7866C > A, c.7960A > G, c.7979A > T, c.7987C > T, c.11248C > G, c.11251C > T, c.11257C > G, c.11257C > T, c.11346C > T, and c.11393C > G) and PKD2 (c.1480G > T) were identified to result in exon skipping. CONCLUSIONS: We confirmed that 11 variants in the gene of PKD1 and PKD2 affect normal splicing by interfering the recognition of classical splicing sites or by disrupting exon splicing enhancers and generating exon splicing silencers. This is the most comprehensive study to date on pre-mRNA splicing of exonic variants in ADPKD-associated disease-causing genes in consideration of the increasing number of identified variants in PKD1 and PKD2 gene in recent years. These results emphasize the significance of assessing the effect of exon single nucleotide variants in ADPKD at the mRNA level.

Dynamic network characteristics of adolescents with major depressive disorder: Attention network mediates the association between anhedonia and attentional deficit.

Attention deficit is a critical symptom that impairs social functioning in adolescents with major depressive disorder (MDD). In this study, we aimed to explore the dynamic neural network activity associated with attention deficits and its relationship with clinical outcomes in adolescents with MDD. We included 188 adolescents with MDD and 94 healthy controls. By combining psychophysics, resting-state electroencephalography (EEG), and functional magnetic resonance imaging (fMRI) techniques, we aimed to identify dynamic network features through the investigation of EEG microstate characteristics and related temporal network features in adolescents with MDD. At baseline, microstate analysis revealed that the occurrence of Microstate C in the patient group was lower than that in healthy controls, whereas the duration and coverage of Microstate D increased in the MDD group. Mediation analysis revealed that the probability of transition from Microstate C to D mediated anhedonia and attention deficits in the MDD group. fMRI results showed that the temporal variability of the dorsal attention network (DAN) was significantly weaker in patients with MDD than in healthy controls. Importantly, the temporal variability of DAN mediated the relationship between anhedonia and attention deficits in the patient group. After acute-stage treatment, the response prediction group (RP) showed improvement in Microstates C and D compared to the nonresponse prediction group (NRP). For resting-state fMRI data, the temporal variability of DAN was significantly higher in the RP group than in the NRP group. Overall, this study enriches our understanding of the neural mechanisms underlying attention deficits in patients with MDD and provides novel clinical biomarkers.

Characterization and pathogenicity of the porcine epidemic diarrhea virus isolated in China.

Piglet diarrhea caused by the porcine epidemic diarrhea virus (PEDV) is a common problem on pig farms in China associated with high morbidity and mortality rates. In this study, three PEDV isolates were successfully detected after the fourth blind passage in Vero cells. The samples were obtained from infected piglet farms in Jilin (Changchun), and Shandong (Qingdao) Provinces of China and were designated as CH/CC-1/2018, CH/CC-2/2018, and CH/QD/2018. According to the analysis of the complete S protein gene sequence, the CH/CC-1/2018 and CH/CC-2/2018 were allocated to the G2b branch, while CH/QD/2018 was located in the G1a interval and was closer to the vaccine strain CV777. Successful detection and identification of the isolated strains were carried out using electron microscopy and indirect immunofluorescence. Meanwhile, animal challenge experiments and viral RNA copies determination were used to compare the pathogenicity. The results showed that CH/CC-1/2018 in Changchun was more pathogenic than CH/QD/2018 in Qingdao. In conclusion, the discovery of these new strains is conducive to the development of vaccines to prevent the pandemic of PEDV, especially that the CH/CC-1/2018, and CH/CC-2/2018 were not related to the classical vaccine strain CV777.

The role of CcPTGS2a in immune response against Aeromonas hydrophila infection in common carp (Cyprinus carpio).

Prostaglandin-endoperoxide synthase 2 (PTGS2), a crucial enzyme in prostaglandin synthesis, catalyzes the conversion of arachidonic acid to prostaglandins and plays a significant role in the inflammatory response. This investigation aimed to determine the regulatory role of PTGS2a in the innate immune response to bacterial infection in fish. To achieve this objective, the CcPTGS2a gene was identified and characterized in common carp (Cyprinus carpio), and its function in immune defense was investigated. According to the sequence and structural analysis results, CcPTGS2a had an open reading frame of 1806 bp that encoded 602 amino acids. It was estimated that the protein's theoretical molecular weight was 69.0 kDa, and its isoelectric point was 8.10. The structure of CcPTGS2a was observed to be conserved, with an epidermal growth factor domain and a peroxidase domain present. Moreover, the amino acid sequence of CcPTGS2a exhibited significant homology with the amino acid sequences of several fish species. CcPTGS2a mRNA was detected in the healthy tissues of common carp, with higher expression in the head kidney, spleen, gills, and liver. Following the challenges with Aeromonas hydrophila and lipopolysaccharide, CcPTGS2a mRNA showed unique geographic and temporal expression patterns, with significant increases detected in the head kidney, gills, spleen, and liver. Additionally, the recombinant CcPTGS2a protein exhibited detectable bacterial binding to various bacteria. As determined by subcellular localization analysis, CcPTGS2a was predominantly localized in the nucleus and cytoplasm. Furthermore, it was discovered that the overexpression of CcPTGS2a stimulated the up-regulation of ferroptosis-related genes and inflammatory cytokine mRNA expression in fish and EPC (Epithelioma papulosum cyprinid) cells while concurrently reducing the bacterial load of A. hydrophila. In contrast, the interference of CcPTGS2a decreased the mRNA expression of ferroptosis-related genes and inflammatory cytokines in fish and EPC cells and increased the bacterial load of A. hydrophila. Notably, A. hydrophila stimulation resulted in the up-regulation of CcPTGS2a protein expression in EPC cells. These results suggested that CcPTGS2a was involved in the immune response to bacterial infections in common carp.

Two CcCCL19bs orchestrate an antibacterial immune response in Yellow River carp (Cyprinus carpio haematopterus).

Chemokines are a group of chemotactic cytokines with an essential role in homeostasis as well as immunity via specific G protein-coupled receptors and atypical receptors. In our study, two Yellow River carp (Cyprinus carpio haematopterus) CCL19b genes (CcCCL19bs), tentatively named CcCCL19b_a and CcCCL19b_b, were cloned. The open reading frames (ORFs) of CcCCL19b_a and CcCCL19b_b were both 333 bp that encoded a 12 kDa protein with 110 amino acid residues. CcCCL19bs contained a signal peptide and a SCY domain with four typical conserved cysteine residues. The two CcCCL19b proteins shared high similarities with each other in both secondary and three-dimensional structure. Phylogenetic analysis showed that CcCCL19bs and other CCL19bs from tetraploid cyprinid fish were clustered into one clade. CcCCL19bs were highly expressed in gill and intestine in healthy fish, and a significant up-regulation of gene expression after Aeromonas hydrophila infection and poly(I:C) stimulation was observed in gill, liver, and head kidney. Furthermore, chemotaxis and antibacterial activity of CcCCL19bs were studied. The results indicated that recombinant CcCCL19b_a and CcCCL19b_b protein (rCcCCL19b_a and rCcCCL19b_b) exhibited significant attraction to primary head kidney leukocytes (HKLs). Meanwhile, both of rCcCCL19bs could promote the proliferation of HKLs, and significantly up-regulate the expressions of IL-1ß, CCR7, and IL-6, and down-regulate the expression of IL-10 in primary HKLs. In vitro, rCcCCL19bs could bind and aggregate A. hydrophila and Staphylococcus aureus. The rCcCCL19bs exhibited significant antibacterial activity against A. hydrophila, but not S. aureus. Moreover, they inhibited the growth of A. hydrophila and S. aureus. In vivo, overexpression of CcCCL19bs contributed to the bacterial clearance. These studies suggested that CcCCL19bs orchestrate an antibacterial immune response.

Functional roles of CcGSDMEa-like in common carp (Cyprinus carpio) after Aeromonas hydrophila infection.

GSDMs could punch holes in cell membrane and participate in the immune response to bacterial infections. In current study, the molecular and structural characteristics of CcGSDMEa-like were analyzed, and the role of CcGSDMEa-like in the inflammatory response against Aeromonas hydrophila was studied. The results showed that the CcGSDMEa-like shared the conserved structural characteristics with GSDMEs of other teleosts. The CcGSDMEa-like mRNA and protein expression levels were significantly affected by A. hydrophila challenge. When the CcGSDMEa-like was overexpressed, the expression of CcIL-1ß were significantly increased in fish and EPC cells, and bacterial contents were significantly decreased in fish tissues. While, when the CcGSDMEa-like was knocked down, the expression and secretion of CcIL-1ß were significantly decreased in vivo and in vitro, and the bacterial contents were increased in vivo after A. hydrophila infection 12 h and 24 h. In brief, CcGSDMEa-like could regulate the content of bacteria in fish through mediating the expression and secretion of CcIL-1ß. Bactericidal assay and cytotoxicity assay showed that CcGSDMEa-like had no bactericidal activity to Escherichia coli, and did not disrupt cytomembrane integrity of HEK293T cells. This study suggested that CcGSDMEa-like could play roles in the antibacterial and inflammatory processes in fish.

Characterization of hepcidin gene and protection of recombinant hepcidin supplemented in feed against Aeromonas hydrophila infection in Yellow River carp (Cyprinus carpio haematopterus).

Hepcidin is a small peptide of defensins with antibacterial activity, and plays an important role in innate immunity against pathogenic microorganisms, which can also participate in the regulation of iron metabolism. The hepcidin gene in Yellow River carp (Cyprinus carpio haematopterus) (CcHep) was cloned and identified. The total length of CcHep cDNA was 480 bp, containing an open reading frame (ORF) that encoded 91 amino acids (aa), which contained a 24-aa signal peptide, a 42-aa propeptide, and a 25-aa mature peptide. The mature peptide had a typical RX (K/R) R motif and eight conserved cysteine residues forming four pairs of disulfide bonds. Homology and phylogenetic tree analysis showed that CcHep had the closest relationship with that of crucian carp. The expression levels of hepcidin mRNA in healthy and Aeromonas hydrophila stimulated fish were measured by real-time fluorescence quantitative PCR. The results showed that CcHep mRNA was expressed in different tissues of healthy fish with the highest relative expression level in liver, followed by kidney and intestine, and the lowest expression level was observed in heart. The hepcidin gene was extremely significantly up-regulated in head kidney, intestine, liver, skin, spleen, and gill at 6 h and 12 h after A. hydrophila infection. Furthermore, the immunoregulation effect of dietary recombinant protein was evaluated. The recombinant hepcidin protein (rCcHep) was successfully expressed by Pichia pastoris X-33 and showed strong antibacterial activity against A. hydrophila, Escherichia coli, Vibrio anguillarum and Bacillus subtilis in vitro. In order to evaluate the preventive effect of rCcHep, fish were fed with basal diet or diet supplemented with different doses of rCcHep, and then challenged with A. hydrophila. The results showed that immune genes were up-regulated to varying degrees, and feed additive groups exhibited a significantly improved up-regulation expressions of Lysozyme, Toll-like receptor 5 (TLR 5), Major histocompatibility complex classⅡ (MHCⅡ), while inhibited up-regulation expressions of Interleukin 1ß (IL-1ß), Interleukin 8 (IL-8), and Tumor necrosis factor α (TNF-α) in liver and spleen compared to the control. Meanwhile, the relative immune protection rate in 120 mg/kg feed additive group was 28%, and the bacterial clearance rate in tissues of this group was higher than that of the control. Collectively, these results indicated that rCcHep had antibacterial activity and showed an immune protection effect against A. hydrophila, and could be considered as a dietary supplement to apply in aquaculture.

Accumulation of Ag(0) Single Atoms at Water/Mineral Interfaces during Sunlight-Induced Reduction of Ionic Ag by Phenolic DOM.

Silver (Ag) undergoes a complex and dynamic Ag+/Ag0 cycle under environmental conditions. The Ag+ → Ag nanoparticles (AgNPs) transformation due to the combined actions of sunlight, O2, and dissolved organic matter has been a well-known environmental phenomenon. In this study, we indicate that this process may be accompanied by a pronounced accumulation of Ag(0) single atoms (Ag-SAs) on the minerals' surfaces. According to spherical aberration-corrected scanning transmission electron microscopy and high-energy-resolution X-ray adsorption fine structure analyses, humic acid (HA) and phenol (PhOH) can induce Ag-SAs accumulation, whereas oxalic acid causes only AgNPs deposition. Ag-SAs account for more than 20 wt % of total Ag(0) on the γ-Al2O3 surfaces during HA- and PhOH-mediated photolysis processes. HA also causes Ag-SAs to accumulate on two other prevalent soil minerals, SiO2 and Fe2O3, and the fractions of Ag-SAs are about 15 wt %. Our mechanism studies suggest that a phenolic molecule acts as a reducing agent of Ag+ and a stabilizer of Ag-SAs, protecting Ag-SAs against autocatalytic nucleation.