The glycosylation sites in RBD of spike protein attenuate the immunogenicity of PEDV AH2012/12.

Porcine epidemic diarrhea (PED) is a highly contagious swine intestinal disease caused by PED virus (PEDV). Vaccination is a promising strategy to prevent and control PED. Previous studies have confirmed that glycosylation could regulate the immunogenicity of viral antigens. In this study, we constructed three recombinant PEDVs which removed the glycosylation sites in RBD. Viral infection assays revealed that similar replication characteristics between the recombinant viruses and parental PEDV. Although animal challenging study demonstrated that the glycosylation sites in RBD do not affect the pathogenicity of PEDV, we found that removing the glycosylation sites on the RBD regions could promote the IgG and neutralization titer in vivo, suggesting deglycosylation in RBD could enhance the immunogenicity of PEDV. These findings demonstrated that removal of the glycosylation sites in RBD is a promising method to develop PEDV vaccines.

Resourcefulness Among Initial Ischemic Stroke Patients: A Longitudinal Study of 12 Months.

Objective: To explore distinct longitudinal trajectories of resourcefulness among initial ischemic stroke patients from diagnosis to 12 months, and to identify whether sociodemographic factors, disease-related factors, self-efficacy, family function, and social support can predict patterns in the trajectories of resourcefulness. Methods: A prospective longitudinal study was conducted. Initial ischemic stroke patients who met inclusion and exclusion criteria were followed up when still in hospital (Preparing for discharge, Baseline, T1), at 1 month (T2), at 3 months (T3), at 6 months (T4), at 9 months (T5) and 12 months (T6) (±1 week) after discharge. General information, National Institute of Health Stroke Scale (NIHSS), Modified Rankin Scale (mRS), General Self-Efficacy Scale (GSES), General Family Functioning Subscale (FAD-GF), and Social Support Rate Scale (SSRS) were used in T1. The Resourcefulness Scale© was evaluated at 6 time points. Growth mixture modeling was used to identify trajectory patterns of resourcefulness. Logistic regression was used to identify predictors of resourcefulness trajectories. Results: Three longitudinal trajectories of resourcefulness were identified and named as the high-stable class (38.9%, n=71), fluctuation class (41.2%, n=75), and low-stable class (19.9%, n=36), respectively. Dwelling areas (x2=6.805, P=0.009), education (x2=44.865, P=0.000), monthly income (x2=13.063, P=0.001), NIHSS scores (x2=44.730, P=0.000), mRS scores (x2=51.788, P=0.000), Hcy (x2=9.345, P=0.002), GSES (x2=56.933, P=0.000), FAD-GF (x2=41.305, P=0.000) and SSRS (x2=52.373, P=0.000) were found to be statistically significant for distinguishing between different resourcefulness trajectory patterns. Lower education (OR=0.404), higher NIHSS(OR=6.672) scores, and higher mRS(OR=21.418) scores were found to be risk factors for lower resourcefulness, whereas higher education(OR=0.404), GSES(OR=0.276), FAD-GF(OR=0.344), and SSRS(OR=0.358) scores were identified as protective factors enhancing resourcefulness. Conclusion: This study obtained three patterns of trajectories and identified their predictive factors in initial ischemic stroke. The findings will assist health care professionals in identifying subgroups of patients and when they may be at risk of low resourcefulness and provide timely targeted intervention to promote resourcefulness.

Sequential catalytic lignin valorization and bioethanol production: an integrated biorefinery strategy.

BACKGROUND: The effective valorization of lignin and carbohydrates in lignocellulose matrix under the concept of biorefinery is a primary strategy to produce sustainable chemicals and fuels. Based on the reductive catalytic fractionation (RCF), lignin in lignocelluloses can be depolymerized into viscous oils, while the highly delignified pulps with high polysaccharides retention can be transformed into various chemicals. RESULTS: A biorefinery paradigm for sequentially valorization of the main components in poplar sawdust was constructed. In this process, the well-defined low-molecular-weight phenols and bioethanol were co-generated by tandem chemo-catalysis in the RCF stage and bio-catalysis in fermentation stage. In the RCF stage, hydrogen transfer reactions were conducted in one-pot process using Raney Ni as catalyst, while the isopropanol (2-PrOH) in the initial liquor was served as a hydrogen donor and the solvent for lignin dissolution. Results indicated the proportion of the 2-PrOH in the initial liquor of RCF influenced the chemical constitution and yield of the lignin oil, which also affected the characteristics of the pulps and the following bioethanol production. A 67.48 ± 0.44% delignification with 20.65 ± 0.31% of monolignols yield were realized when the 2-PrOH:H2O ratio in initial liquor was 7:3 (6.67 wt% of the catalyst loading, 200 °C for 3 h). The RCF pulp had higher carbohydrates retention (57.96 ± 2.78 wt%), which was converted to 21.61 ± 0.62 g/L of bioethanol with a yield of 0.429 ± 0.010 g/g in fermentation using an engineered S. cerevisiae strain. Based on the mass balance analysis, 104.4 g of ethanol and 206.5 g of lignin oil can be produced from 1000 g of the raw poplar sawdust. CONCLUSIONS: The main chemical components in poplar sawdust can be effectively transformed into lignin oil and bioethanol. The attractive results from the biorefinery process exhibit great promise for the production of valuable biofuels and chemicals from abundant lignocellulosic materials.

Analysis of spike and accessory 3c genes mutations of less virulent and FIP-associated feline coronaviruses in Beijing, China.

Mutations in S and 3c genes of feline coronavirus (FCoV) have been associated with the development of feline infectious peritonitis (FIP). In the present study, FCoV S and 3c genes mutations were analyzed in healthy and FIP cats. M1058L mutation was found in 13.64% (3/22) feces from FIP cats, but not in feces from healthy cats (0/39). The intact 3c gene was found in feces from both healthy cats (19/19) and FIP cats (12/12). All parenteral samples from FIP cats carried one or more of the M1058L mutation, S1060A mutation and mutated 3c gene. FCoV reverse-transcriptase polymerase chain reaction (RT-PCR) of parenteral samples (including ascites, pleural effusions and tissue) is recommended as the gold standard for clinical diagnosis of FIP rather than detection of the M1058L mutation, but when cats have severe gastrointestinal symptoms and lesions, detection of the M1058L mutation in feces may be helpful in diagnosing FIP.

K-CapsNet: K-Nearest Neighbor Based Convolution Capsule Network for Cerenkov Luminescence Tomography Reconstruction.

Cerenkov luminescence tomography (CLT) has received significant attention as a promising imaging modality that can display the three-dimensional (3D) distribution of radioactive probes. However, the reconstruction of CLT suffers from severe ill-posed problem. It is difficult for traditional model-based method to obtain satisfactory result. Recently, deep learning-based method have shown great potential for accurate and efficient CLT reconstruction. In this study, a KNN-based convolution capsule network, named K-CapsNet, is proposed for cerenkov luminescence tomography. In K-CapsNet, the surface photon intensity is encoded in capsule form. The KNN-based convolution and K-means clustering are proposed for efficient encoding. Numerical simulation experiments have been carried out to verify the performance of K-CapsNet, and the results show that it performs superior in source localization and morphological restoration compared with existing methods.

FISTA-NET: Deep Algorithm Unrolling for Cerenkov luminescence tomography.

Cerenkov luminescence tomography (CLT) is a highly sensitive and promising imaging technique that can be used to reconstruct the three-dimensional distribution of radioactive probes in living animals. However, the accuracy of CLT reconstruction is limited by the simplified radiative transfer equation and ill-conditioned inverse problem. To address this issue, we propose a model-based deep learning network that combines the neural network with a model-based approach to enhance the performance of CLT reconstruction. The Fast Iterative Shrinkage Thresholding Algorithm (FISTA), a traditional model-based approach, is expanded into a deep network (known as FISTA-NET). Each layer in the network represents an iteration of the algorithm steps, and connecting these layers can form a deep neural network. In addition, different from the traditional FISTA, the key parameters in FISTA, such as gradient step size and threshold value, can be learned through training data without manual production. To evaluate the performance of FISTA-NET, numerical simulation experiments were conducted, which demonstrate its excellent positioning and shape recovery abilities.Clinical Relevance-This indicates that FISTA-NET strategy can significantly improve the quality of CLT reconstruction, which is further beneficial to the assessment of disease activity and treatment effect based on CLT.

Effectiveness of health management among individuals at high risk of stroke: An intervention study based on the health ecology model and self-determination theory (HEM-SDT).

Background: Stroke is the second leading cause of death in adults worldwide. However, up to 80% of strokes can be prevented by modifying risk factors. Objective: The study aims to assess the effectiveness of the Health Ecology Model and Self-Determination Theory (HEM-SDT) based health management intervention among individuals at high risk of stroke. Methods: A randomized controlled trial was conducted in Zhengzhou from May 1st, 2020, to December 31st, 2020. A total of 229 participants were recruited for the study, with 116 individuals at high risk of stroke being randomly assigned to the HEM-SDT health management group, while 113 participants were enrolled in the control group, following their current routine practices. The Generalized Estimating Equation model (GEE) was used to analyze the differences in health knowledge, belief and, behavior between the two groups at the beginning of the intervention, and at 6-month intervals after the intervention. The chi-square test was utilized to assess the control rate of risk factors. Results: After 6 months of intervention, there were significant improvements in health knowledge, behavior, and belief among the participants. The study found significant differences in the interaction effects between time and group for health knowledge (Mean, SD = 25.62 ± 3.88, 95%CI: 7.944-9.604, P＜0.001), health belief (Mean, SD = 87.18 ± 14.21, 95%CI: 23.999-29.887, P＜0.001), and health behavior (Mean, SD = 173.28 ± 24.22, 95%CI: 22.332-36.904, P＜0.001). Additionally, the rates of hypertension, hyperglycemia, dyslipidemia, high or medium risk condition of stroke, obesity, hyperhomocysteinemia, smoking, alcohol consumption, and lack of exercise also showed statistical significance (P＜0.05) after the intervention. Conclusion: The HEM-SDT health management model improves the health knowledge, behavior, and beliefs in people at high risk of stroke and remarkably it shows improvement in modifiable risk factors. It can be recommended for systematic health management in people at high-risk of stroke.

Regularized reconstruction based on joint smoothly clipped absolute deviation regularization and graph manifold learning for fluorescence molecular tomography.

Objective.Fluorescence molecular tomography (FMT) is an optical imaging modality that provides high sensitivity and low cost, which can offer the three-dimensional distribution of biomarkers by detecting the fluorescently labeled probe noninvasively. In the field of preclinical cancer diagnosis and treatment, FMT has gained significant traction. Nonetheless, the current FMT reconstruction results suffer from unsatisfactory morphology and location accuracy of the fluorescence distribution, primarily due to the light scattering effect and the ill-posed nature of the inverse problem.Approach.To address these challenges, a regularized reconstruction method based on joint smoothly clipped absolute deviation regularization and graph manifold learning (SCAD-GML) for FMT is presented in this paper. The SCAD-GML approach combines the sparsity of the fluorescent sources with the latent manifold structure of fluorescent source distribution to achieve more accurate and sparse reconstruction results. To obtain the reconstruction results efficiently, the non-convex gradient descent iterative method is employed to solve the established objective function. To assess the performance of the proposed SCAD-GML method, a comprehensive evaluation is conducted through numerical simulation experiments as well asin vivoexperiments.Main results.The results demonstrate that the SCAD-GML method outperforms other methods in terms of both location and shape recovery of fluorescence biomarkers distribution.Siginificance.These findings indicate that the SCAD-GML method has the potential to advance the application of FMT inin vivobiological research.

Dual-Targeting Biomimetic Nanomaterials for Photo-/Chemo-/Antiangiogenic Synergistic Therapy.

Avoiding the low specificity of phototheranostic reagents at the tumor site is a major challenge in cancer phototherapy. Meanwhile, angiogenesis in the tumor is not only the premise of tumor occurrence but also the basis of tumor growth, invasion, and metastasis, making it an ideal strategy for tumor therapy. Herein, biomimetic cancer cell membrane-coated nanodrugs (mBPP NPs) have been prepared by integrating (i) homotypic cancer cell membranes for evading immune cell phagocytosis to increase drug accumulation, (ii) protocatechuic acid for tumor vascular targeting along with chemotherapy effect, and (iii) near-infrared phototherapeutic agent diketopyrrolopyrrole derivative for photodynamic/photothermal synergetic therapy. The mBPP NPs exhibit high biocompatibility, superb phototoxicity, excellent antiangiogenic ability, and double-trigging cancer cell apoptosis in vitro. More significantly, mBPP NPs could specifically bind to tumor cells and vasculature after intravenous injection, inducing fluorescence and photothermal imaging-guided tumor ablation without recurrence and side effects in vivo. The biomimetic mBPP NPs could cause drug accumulation at the tumor site, inhibit tumor neovascularization, and improve phototherapy efficiency, providing a novel avenue for cancer treatment.

Application of the Electromagnetic Method to the Spatial Distribution of Subsurface Saline and Fresh Water in the Coastal Mudflat Area of Jiangsu Province.

Interfacial zones straddling terrestrial and marine realms, colloquially known as mudflats, epitomize a dynamic nexus between these environments and are fundamental to the coastal ecosystem. The investigation of these regions is paramount for facilitating infrastructural developments including ports, wharfs, cross-sea bridges, and the strategic utilization of freshwater resources sequestered from mainland islands amid ongoing economic progress. Terrestrial realms conventionally employ electromagnetic techniques as efficacious modalities to delineate subterranean geological information, encompassing structural details and water-bearing strata. However, the peculiar topographic and geological nuances of mudflat regions pose substantial challenges for the efficacious application of electromagnetic methodologies. The present paper endeavors to address these challenges by suggesting innovative modifications to the existing instrumentation and evolving novel data acquisition techniques specifically tailored for electromagnetic exploration within mudflat environments. This paper delves into the electrical characteristics of water-bearing layers within mudflats, and ascertains details pertaining to the subterranean structure and the spatial distribution of fresh and saline water resources, through the holistic interpretation of a multitude of profiles.

Sparse reconstruction based on dictionary learning and group structure strategy for cone-beam X-ray luminescence computed tomography.

As a dual-modal imaging technology that has emerged in recent years, cone-beam X-ray luminescence computed tomography (CB-XLCT) has exhibited promise as a tool for the early three-dimensional detection of tumors in small animals. However, due to the challenges imposed by the low absorption and high scattering of light in tissues, the CB-XLCT reconstruction problem is a severely ill-conditioned inverse problem, rendering it difficult to obtain satisfactory reconstruction results. In this study, a strategy that utilizes dictionary learning and group structure (DLGS) is proposed to achieve satisfactory CB-XLCT reconstruction performance. The group structure is employed to account for the clustering of nanophosphors in specific regions within the organism, which can enhance the interrelation of elements in the same group. Furthermore, the dictionary learning strategy is implemented to effectively capture sparse features. The performance of the proposed method was evaluated through numerical simulations and in vivo experiments. The experimental results demonstrate that the proposed method achieves superior reconstruction performance in terms of location accuracy, target shape, robustness, dual-source resolution, and in vivo practicability.

Generalized conditional gradient method with adaptive regularization parameters for fluorescence molecular tomography.

Fluorescence molecular tomography (FMT) is an optical imaging technology with the ability of visualizing the three-dimensional distribution of fluorescently labelled probes in vivo. However, due to the light scattering effect and ill-posed inverse problems, obtaining satisfactory FMT reconstruction is still a challenging problem. In this work, to improve the performance of FMT reconstruction, we proposed a generalized conditional gradient method with adaptive regularization parameters (GCGM-ARP). In order to make a tradeoff between the sparsity and shape preservation of the reconstruction source, and to maintain its robustness, elastic-net (EN) regularization is introduced. EN regularization combines the advantages of L1-norm and L2-norm, and overcomes the shortcomings of traditional Lp-norm regularization, such as over-sparsity, over-smoothness, and non-robustness. Thus, the equivalent optimization formulation of the original problem can be obtained. To further improve the performance of the reconstruction, the L-curve is adopted to adaptively adjust the regularization parameters. Then, the generalized conditional gradient method (GCGM) is used to split the minimization problem based on EN regularization into two simpler sub-problems, which are determining the direction of the gradient and the step size. These sub-problems are addressed efficiently to obtain more sparse solutions. To assess the performance of our proposed method, a series of numerical simulation experiments and in vivo experiments were implemented. The experimental results show that, compared with other mathematical reconstruction methods, GCGM-ARP method has the minimum location error (LE) and relative intensity error (RIE), and the maximum dice coefficient (Dice) in the case of different sources number or shape, or Gaussian noise of 5%-25%. This indicates that GCGM-ARP has superior reconstruction performance in source localization, dual-source resolution, morphology recovery, and robustness. In conclusion, the proposed GCGM-ARP is an effective and robust strategy for FMT reconstruction in biomedical application.

Boosting the epoxidation of squalene to produce triterpenoids in Saccharomyces cerevisiae.

BACKGROUND: Polycyclic triterpenoids (PTs) are common in plants, and have attracted considerable interest due to their remarkable biological activities. Currently, engineering the ergosterol synthesis pathway in Saccharomyces cerevisiae is a safe and cost-competitive way to produce triterpenoids. However, the strict regulation of ERG1 involved in the epoxidation of squalene limits the triterpenoid production. RESULTS: In this study, we found that the decrease in ERG7 protein level could dramatically boost the epoxidation of squalene by improving the protein stability of ERG1. We next explored the potential factors that affected the degradation process of ERG1 and confirmed that ERG7 was involved in the degradation process of ERG1. Subsequently, expression of four different triterpene cyclases utilizing either 2,3-oxidosqualene or 2,3:22,23-dioxidosqualene as the substrate in ERG7-degraded strains showed that the degradation of ERG7 to prompt the epoxidation of squalene could significantly increase triterpenoid production. To better display the potential of the strategy, we increased the supply of 2,3-oxidosqualene, optimized flux distribution between ergosterol synthesis pathway and ß-amyrin synthesis pathway, and modified the GAL-regulation system to separate the growth stage from the production stage. The best-performing strain ultimately produced 4216.6 ± 68.4 mg/L of ß-amyrin in a two-stage fed-fermentation (a 47-fold improvement over the initial strain). CONCLUSIONS: This study showed that deregulation of the native restriction in ergosterol pathway was an effective strategy to increase triterpenoid production in yeast, which provided a new insight into triterpenoids biosynthesis.

Molecular identification of a novel antimicrobial peptide in giant Triton snail Charonia tritonis: mRNA profiles for tissues and its potential antibacterial activity.

Antimicrobial peptides (AMPs) play an important role in innate immunity against microorganisms. AMPs is an effective antibacterial agent, and the chances of causing pathogens to develop is very low. However, there is little information about AMPs in the giant Triton snail Charonia tritonis. In this research, an antimicrobial peptide gene (termed Ct-20534) was identified in C. tritonis. The open reading frame of Ct-20534 is 381 bp in size and it encodes a basic peptide precursor containing 126 amino acids. Ct-20534 gene was found to be expressed in all five tissues examined by real-time fluorescence quantitative PCR (qPCR), but the highest expression was found in the proboscis. This is the first report that antibacterial peptides have been found in C. tritonis, and it has been proved that Ct-20534 has antibacterial activity against Gram-positive bacteria and Gram-negative bacteria, among which the activity of Staphylococcus aureus is most significantly inhibited, this suggests that the newly discovered antimicrobial peptides in C. tritonis may play an important role in the immune system and bacterial resistance of C. tritonis. This study presents the discovery of a newly identified antibacterial peptide from C. tritonis, with its structural properties fully characterized and potent antibacterial activity confirmed. The results provide essential fundamental data for the development of preventive and therapeutic measures against aquatic animal diseases, which in turn can promote the sustainable and stable growth of the aquaculture industry and create economic benefits. Additionally, this research lays the foundation for future development of novel anti-infective drugs.

Low-level tragus stimulation improves autoantibody-induced hyperadrenergic postural tachycardia syndrome in rabbits.

Background: Recent studies have demonstrated that antiadrenergic autoantibodies are involved in the pathophysiology of postural orthostatic tachycardia syndrome (POTS). Objective: The purpose of this study was to test the hypothesis that transcutaneous low-level tragus stimulation (LLTS) ameliorates autoantibody-induced autonomic dysfunction and inflammation in a rabbit model of autoimmune POTS. Methods: Six New Zealand white rabbits were co-immunized with peptides from the α1-adrenergic and ß1-adrenergic receptors to produce sympathomimetic antibodies. The tilt test was performed on conscious rabbits before immunization, 6 weeks after immunization, and 10 weeks after immunization with 4-week daily LLTS treatment. Each rabbit served as its own control. Results: An enhanced postural heart rate increase in the absence of significant change in blood pressure was observed in immunized rabbits, confirming our previous report. Power spectral analysis of heart rate variability during the tilt test showed a predominance of sympathetic over parasympathetic activity in immunized rabbits as reflected by markedly increased low-frequency power, decreased high-frequency power, and increased low-to-high-frequency ratio. Serum inflammatory cytokines were also significantly increased in immunized rabbits. LLTS suppressed the postural tachycardia, improved the sympathovagal balance with increased acetylcholine secretion, and attenuated the inflammatory cytokine expression. Antibody production and activity were confirmed with in vitro assays, and no antibody suppression by LLTS was found in this short-term study. Conclusion: LLTS improves cardiac autonomic imbalance and inflammation in a rabbit model of autoantibody-induced hyperadrenergic POTS, suggesting that LLTS may be used as a novel neuromodulation therapy for POTS.

Transcutaneous vagus nerve stimulation attenuates autoantibody-mediated cardiovagal dysfunction and inflammation in a rabbit model of postural tachycardia syndrome.

PURPOSE: Previous studies demonstrated M2 muscarinic acetylcholine receptor-activating autoantibodies (M2R-AAb) were present in some patients with postural tachycardia syndrome (POTS). This study examines how these autoantibodies might contribute to the pathophysiology of POTS, and whether low-level tragus stimulation (LLTS) can ameliorate autoantibody-mediated autonomic dysregulation in the rabbit. METHODS: Five New Zealand white rabbits were immunized with a M2R second extracellular loop peptide to produce cholinomimetic M2R-AAb. Tilt test and infusion studies were performed on conscious rabbits before immunization, 6 weeks after immunization, and 8 weeks after immunization with 2-week daily LLTS treatment. Each rabbit served as its own control. RESULTS: Compared to preimmune state, an enhanced heart rate increase and decreased parasympathetic activity upon tilting were observed in immunized rabbits. Furthermore, these rabbits demonstrated an attenuated heart rate-slowing response to infusion of the M2R orthosteric agonist arecaidine propargyl ester (APE), suggesting an inhibitory allosteric effect of M2R-AAb. There was also a significant increase in serum inflammatory cytokines in immunized rabbits. LLTS treatment suppressed the postural tachycardia, improved the sympathovagal balance with increased acetylcholine secretion, reduced the levels of inflammatory cytokines, and reversed the attenuated heart rate response to APE in immunized rabbits. No suppression of M2R-AAb expression by LLTS was found during this short-term study period. Receptor-modulating activity of M2R-AAb produced in immunized rabbits was confirmed with in vitro bioassay. CONCLUSIONS: Autoantibody inhibition of cholinergic ligand activity may be involved in the development of cardiovagal dysfunction and inflammation associated with POTS, both of which can be improved by vagal stimulation.

A reliability and validity study of the electronic health literacy scale among stroke patients in China.

BACKGROUND: Patients with stroke usually use smartphones to obtain online information to maintain their health. But their ability to identify, evaluate and apply this information is still unknown. AIM: This study was designed to examine the reliability and validity of the electronic Health Literacy Scale among patients with stroke in China. DESIGN: This is a cross-sectional survey. METHODS: A demographic questionnaire, the electronic Health Literacy Scale (e-HLS) and the eHealth Literacy Scale (eHEALS) were administered to a sample of 648 patients with ischemic stroke recruited from December 2020 to March 2021 in a tertiary hospital. RESULTS: The Cronbach'α coefficient on the e-HLS-CHI was 0.907. Kappa consistency coefficient of test-retest reliability was 0.691 (p < .05). Three factors were extracted by Exploratory Factor Analysis (EFA), accounting for 90.84% of the total variance. Confirmatory Factory Analysis (CFA) revealed that three factors of e-HLS-CHI fit well (NFI = 0.979, RFI = 0.955, IFI = 0.987, TLI = 0.972, CFI = 0.987, RMSEA = 0.070, CMIN/DF = 2.586). Good simultaneous validity was suggested by the positive correlation of 0.94 (p < .001) between the e-HLS-CHI and eHEALS. When using eHEALS as the standard, the area under the ROC curve of e-HLS-CHI was 0.896 (95% CI: 0.831-0.960, p < .001). The sensitivity and specificity were 97.8% and 70.4% respectively. CONCLUSIONS: The e-HLS can be used to evaluate electronic health literacy of patients with stroke in China after translation and cultural adaption.

Giant Triton Snail Charonia tritonis Macrophage-Expressed Gene 1 Protein Ct-Mpeg1: Molecular Identification, Expression Analysis, and Antimicrobial Activity.

Macrophage-expressed gene 1 proteins (Mpeg1/Perforin-2 (PRF2)) are a family of pore-forming proteins (PFPs) which can form pores and destroy the cell membrane of invading pathogens. However, little information is available regarding the function of Mpeg1 in the giant triton snail Charonia tritonis. In this study, a homolog of Mpeg1 (Ct-Mpeg1) was identified in C. tritonis. The predicted protein of Ct-Mpeg1 contains several structural features known in Mpegs, including a membrane attack complex/perforin (MACPF) domain and single transmembrane region. The Ct-Mpeg1 gene was constitutively expressed in almost all tissues examined except in the proboscis, with the highest expression level observed in the mantle. As a typical pore-forming protein, Ct-Mpeg1 has antibacterial activities against Vibrio (including Vibrio alginolyticus and Vibrio parahaemolyticus). In addition, rCt-Mpeg1 challenge to V. alginolyticus represses the expression of most outer membrane protein synthesis-related genes and genes involved in the TCA cycle pathway, which will lead to reduced outer membrane protein synthesis and less energy capacity. This is the first report to characterize the macrophage-expressed gene 1 protein in C. tritonis, and these results suggest that macrophage-expressed gene 1 protein Ct-Mpeg1 is an important immune molecule of C. tritonis that is involved in the bacterial infection resistance of Vibrio, and this study may provide crucial basic data for the understanding of the innate immunity system of C. tritonis.

Molecular and Functional Characterization of a Novel Kunitz-Type Toxin-like Peptide in the Giant Triton Snail Charonia tritonis.

It has been reported that the giant triton snail (Charonia tritonis) inserts its large proboscis and then injects venom or acid saliva from its salivary gland into its prey, the crown-of-thorns starfish Acanthaster planci (COTS), paralyzing it. A full-length cDNA sequence of the C. tritonis Ct-kunitzin gene was obtained by RACE PCR based on a transcriptomic database constructed by our laboratory (data not published), which contains an open reading frame (ORF) sequence with a length of 384 bp including a 1-32aa Kunitz domain. The Ct-kunitzin peptide was synthesized by solid-phase polypeptide methods according to its conserved amino acid sequence, with a molecular weight of 3746.0 as well as two disulfide bonds. Renatured Ct-kunitzin was injected into mice ventricles to evaluate its potential function. Compared with the normal control group (physiological saline), the spontaneous locomotor activity of the Ct-kunitzin group decreased significantly. There was a significant effect on Ct-kunitzin on mice grip strength in the grip strength test. In addition, Ct-kunitzin exhibited remarkable biological activity in suppressing pain in the pain thresholds test. There were no significant differences between the Ct-kunitzin group and the normal control group in terms of various hematological indexes and histopathological observations. When tested in COTS, the most significant histological change was the destruction, disorganization, and significant reduction in the amount of COTS tube feet tissues. Altogether, the potential paralyzing effect on mice suggests that Ct-kunitzin is a possible agent for novel drug development.

Molecular and Functional Characterization of a Short-Type Peptidoglycan Recognition Protein, Ct-PGRP-S1 in the Giant Triton Snail Charonia tritonis.

Peptidoglycan recognition proteins (PGRPs) are a family of pattern recognition receptors (PRRs) involved in host antibacterial responses, and their functions have been characterized in most invertebrate and vertebrate animals. However, little information is available regarding the potential function of PGRPs in the giant triton snail Charonia tritonis. In this study, a short-type PGRP gene (termed Ct-PGRP-S1) was identified in C. tritonis. Ct-PGRP-S1 was predicted to contain several structural features known in PGRPs, including a typical PGRP domain (Amidase_2) and Src homology-3 (SH3) domain. The Ct-PGRP-S1 gene was constitutively expressed in all tissues examined except in proboscis, with the highest expression level observed in the liver. As a typical PRR, Ct-PGRP-S1 has an ability to degrade peptidoglycan (PGN) and was proven to have non-Zn2+-dependent amidase activity and antibacterial activity against Vibrioalginolyticus and Staphylococcus aureus. It is the first report to reveal the peptidoglycan recognition protein in C. tritonis, and these results suggest that peptidoglycan recognition protein Ct-PGRP-S1 is an important effector of C. tritonis that modulates bacterial infection resistance of V. alginolyticus and S. aureus, and this study may provide crucial basic data for the understanding of an innate immunity system of C. tritonis.