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The physiological response of the clam<i> Ruditapes</i><i> philippinarum</i> and scallop<i> Chlamys</i><i> farreri</i> to varied concentrations of microplastics exposure
No full textMicroplastics (MPs) pollution's impact on the marine ecosystem is widely recognized. This study compared the effects of polyethylene (PE) and polyethylene terephthalate (PET) on two bivalve species, Ruditapes philippinarum (clam) and Chlamys farreri (scallop), at two particle concentrations (10 and 1000 mu g/L). MPs were found in the digestive glands and gills of both species. Although clearance rates showed no significant changes, exposure to different MPs caused oxidative stress, energy disruption, and lipid metabolism disorders in both clam and scallop. Histopathological damage was observed in gills and digestive glands. IBR values indicated increasing toxicity with concentration, with PET being more toxic than PE. WOE model suggested increasing hazard with concentration, highlighting higher PET toxicity on clam digestive glands. In contrast, PE hazard increased in gills, showing different species responses. R. philippinarum exhibited higher sensitivity to MPs than C. farreri, providing insights for assessing ecological risk under realistic conditions and stress conditions
MAPK/ERK 信号通路对菲律宾蛤仔血淋巴细胞胞外陷阱的形成及调控机制研究
No full text菲律宾蛤仔（Ruditapes philippinarum）作为近海生态系统的基础物种之一，同时也是重要的经济养殖物种。近年来，由于病原体侵袭和环境恶化，菲律宾蛤仔大规模死亡现象频繁发生，严重影响了近海生态系统的稳定性和养殖产业的可持续发展。鉴于贝类独特的开放循环系统，海水环境中的有害致病菌或污染物可直接与血淋巴细胞接触，从而对机体的细胞免疫产生负面影响。胞外陷阱（Extracellular traps，ETs）作为一种新兴的固有免疫防御手段，在抵御病原微生物入侵方面发挥了重要作用，已成为细胞免疫反应中不可或缺的一部分。然而，ETs的过度释放亦能触发免疫毒性，并造成负面影响。目前，关于海洋贝类ETs的研究多聚焦于现象表征与免疫功能评估，而ETs发生的潜在调控机制尚有待于深入探索。本研究旨在探究革兰氏阴性细菌细胞壁成分脂多糖（LPS）以及内分泌干扰物壬基酚（NP）对菲律宾蛤仔血淋巴细胞ETs发生的影响，并探讨其潜在的分子调控机制，以期能够为理解海洋贝类在应对环境污染和病原体侵袭时的免疫响应机制提供理论依据。取得的主要研究结果如下：
1. 组蛋白H2A参与细菌诱导的ETs抗菌免疫应答
从菲律宾蛤仔中克隆并表征了一种组蛋白H2A（命名为RpH2A），RpH2A开放阅读框（ORF）包含387 bp，为编码128个氨基酸的多肽，其预测的蛋白质结构域高度保守，仅包含一个H2A结构域；RpH2A的氨基酸序列与其他物种的H2A序列具有较高的同源性（57.1%-96.1%）。对菲律宾蛤仔进行鳗弧菌攻毒实验，发现RpH2A的mRNA表达量在刺激6小时和12小时后显著上调，分别达到对照组的8.08倍（p < 0.01）和8.50倍（p < 0.01）。体外原核表达了RpH2A重组蛋白（rRpH2A），并探究了其免疫活性，结果发现：rRpH2A蛋白对脂多糖（LPS）和肽聚糖（PGN）表现出较强的亲和力，且其结合活性具有剂量依赖性。此外，rRpH2A对模式细菌大肠杆菌具有杀菌作用，其杀菌能力随着蛋白浓度的升高而增强。此外，大肠杆菌（1 MOI）能够诱导血淋巴细胞释放ETs，并伴有活性氧（ROS）爆发。共定位分析发现，具有抗菌活性的RpH2A分布于ETs结构周围。这些ETs能够显著抑制大肠杆菌的生长，其抑菌效应可能由位于ETs中的RpH2A介导。本研究表明，组蛋白RpH2A可作为抗菌肽，在菲律宾蛤仔血淋巴细胞ETs的抗菌免疫反应中发挥重要作用。
2. LPS通过TLR-Raf-MEK-ERK-ROS信号通路调控ETs的形成
选取革兰氏阴性细菌细胞壁主要成分脂多糖（LPS）作为体外刺激物，以探讨菲律宾蛤仔血淋巴细胞中细菌诱导胞外陷阱（ETs）形成的分子机制。结果发现：LPS（1 μg/L）能够以时间依赖的方式诱导血淋巴ETs的形成，其中DNA构成了基本骨架，而抗菌肽RpH2A则是重要的蛋白组分。LPS刺激后，血淋巴细胞中髓过氧化物酶（MPO）和ROS的生成显著增强，并以依赖于MPO和ROS的方式诱导血淋巴细胞ETs的形成。进一步分析发现，LPS刺激导致血淋巴细胞中磷酸化细胞外调节蛋白激酶（ERK）的丰度升高，Raf抑制剂和MEK抑制剂能够抑制LPS诱导的ERK磷酸化上调，表明LPS通过经典的三级激酶级联，即Raf-MEK-MAPK通路激活ERK信号。ERK抑制剂能够显著抑制LPS诱导的ROS爆发，并限制ETs的形成；阻断ROS的产生也能减缓LPS诱导的ERK磷酸化及ETs的形成，这表明ERK-ROS正反馈环参与了LPS诱导的ETs发生。此外，血淋巴细胞中Toll样受体（TLR）的mRNA表达水平在LPS刺激后显著上调，通过RNA干扰（RNAi）敲减TLR的表达能够抑制LPS诱导的ERK磷酸化和ETs形成，表明TLR可能参与了LPS的识别，并通过ERK调控ETs的发生。向菲律宾蛤仔体内注射LPS后，观察到血淋巴细胞中ROS生成增多、ERK蛋白磷酸化增强，以及鳃组织中出现纤维状的ETs样结构。LPS诱导产生的ETs能够捕获灿烂弧菌、哈维氏弧菌和副溶血弧菌，并表现出广谱的抑菌作用。综上，本研究初步解析了细菌诱导ETs形成的分子机制：血淋巴细胞通过膜受体TLR识别细菌LPS，启动胞内MAPK级联反应调控ROS产生，进而促进ETs的释放以抵御细菌的入侵。
3. NP通过MAPK/ERK信号促进ROS爆发和糖酵解调控ETs的形成
选取海水中常见的内分泌干扰物壬基酚（NP）作为刺激物，探究了其对血淋巴细胞ETs形成的影响，以解析NP对菲律宾蛤仔的免疫毒性作用机制。结果发现：NP（10 μg/L、50 μg/L和100 μg/L）以剂量依赖的方式显著降低血淋巴细胞的活力。正常情况下，血淋巴细胞呈圆形，胞内颗粒和伪足清晰可见，而NP（100 μg/L）诱导ETs激活的血淋巴细胞破裂，伪足不明显，并伴有纤维样结构的形成。NP诱导的ETs发生依赖于MPO和ROS。转录组学分析发现，NP暴露后血淋巴细胞中MAPK、PI3K-AKT以及丙酮酸代谢和糖酵解/葡萄糖生成等信号通路呈上调趋势，而mTOR信号通路则呈下调趋势。在NP诱导的ETs形成过程中，MAPK信号通路被激活，表现为ERK蛋白的磷酸化水平上调。抑制ERK可导致NP诱导的NADPH氧化酶（NOX）表达降低，这进一步抑制ROS的产生，最终限制了ETs的释放，这表明ERK-NOX-ROS信号轴在NP诱导的ETs形成中发挥了作用。此外，NP暴露增加了血淋巴细胞对葡萄糖的摄取，同时也增强了糖酵解限速酶（己糖激酶和丙酮酸激酶）的活性及葡萄糖转运蛋白（GLUT）的表达。使用糖酵解抑制剂能够显著阻碍NP诱导的ETs形成，表明糖酵解在ETs形成中具有关键作用。NP暴露能够上调PI3K和AKT蛋白的磷酸化水平，而下调mTOR蛋白的磷酸化水平。抑制ERK能够逆转NP暴露后PI3K和mTOR的磷酸化状态。同时，NP暴露还上调了转录因子c-Myc和HIF-1α的mRNA表达，这表明MAPK/ERK可能通过PI3K/AKT通路和mTOR信号，调控这些转录因子介导糖酵解反应，为ETs的形成提供能量。本研究初步证实了NP通过ERK-NOX-ROS轴和糖酵解调控血淋巴细胞ETs的形成，从而对菲律宾蛤仔血淋巴细胞产生免疫毒性。
综上所述，本研究探索了不同类型刺激物作用下菲律宾蛤仔血淋巴细胞ETs的发生，初步揭示了ETs形成的分子调控机制，相关结果不仅有助于深化对海洋贝类免疫系统功能的认识，也为海洋贝类疾病的防治策略提供了新的理论基础
Surface-Enhanced Raman Scattering-Based Lateral Flow Assay Strips Using Highly Symmetric Gold Nanostars
No full textThe applications for lateral flow assay (LFA) strips in point-of-care testing have been significantly constrained by their insufficient sensitivity and reproducibility. To address these inherent issues, we developed surface-enhanced Raman scattering (SERS)-based LFA strips, in which highly symmetric Au nanostars (Sym-AuNS) were employed as the sensing element. Due to the uniform tip-sharp nanostructure and a certain number of branches on the surface of Sym-AuNS, it generates a uniform hotspot distribution, thus producing a strong and stable SERS signal. As a proof of concept, human IgG was chosen as the target to evaluate the performance of the proposed SERS-LFA strips. In human serum spiked samples, the limit of detection for human IgG detection was achieved as low as 38 ng/mL, which exhibited a 2-fold, 3-fold, and 13-fold sensitivity improvement compared with the SERS-LFA strips using conventional gold nanostars (AuNS), enzyme-linked immunosorbent assays (ELISA), and the conventional LFA strips, respectively. Furthermore, the SERS-LFA strips demonstrated high assay reproducibility, with a relative standard deviation of 7.75% for five repeated tests, much lower than those of SERS-LFA strips using AuNS (24.6%), ELISA (12.42%), and conventional LFA strips (31.32%). These results demonstrate that the construction of sensitive and reproducible SERS-LFA strips was obtained, and this platform using Sym-AuNS as SERS nanoparticles paves the way for a promising approach in immunoassay technology
Improving the Stability and Anti-Infective Activity of Sea Turtle AMPs Using Multiple Structural Modification Strategies
No full textAntimicrobial peptides (AMPs) are regarded as promising candidates for combating antimicrobial resistance. Previously we identified an AMP named Cm-CATH2 from the green sea turtle, which exhibited potent antibacterial activity and attractive potential in application. However, natural AMPs including Cm-CATH2 frequently suffer from structural instability and sensitivity to physiological conditions, limiting their effectiveness. Herein, we explored various strategies to enhance the efficacy and stability of Cm-CATH2, including peptide truncation, non-natural amino acid substitutions, disulfide bond-based cyclization, and stapled peptide techniques. The results demonstrated that the truncated NCM4 significantly improved the antimicrobial capability of Cm-CATH2 while also enhancing its anti-inflammatory and antibiofilm activities with minimal cytotoxicity. Further ornithine-substituted peptide oNCM markedly enhanced the stability of NCM4 without compromising its antimicrobial efficacy. This study successfully designed a lead peptide oNCM with significant development potential, while providing valuable insights into the advantages and limitations associated with diverse strategies for enhancing the stability of AMPs
A method for breeding fast-growing strains of unicinctus unicinctus based on intestinal flora
No full text本发明提供一种基于肠道菌群的单环刺螠速生品系选育方法，属水产新品种创制领域。通过对比同批次生长速率差异个体中肠道菌群的异同，筛选出与速生相关的特殊菌群。以此作为指标选育速生群体。本发明的选育方法与常规迭代选育相比，选育速度大幅加快，可实现无创筛选，为单环刺螠新品系的选育和新品种开发提供了新途径
A fluorescent probe for detecting biological thiols and its preparation method and application
No full text本发明公开了一种用于检测生物硫醇的荧光探针及其制备方法和应用，属于化学技术领域。该荧光探针的结构如式I所示。制备方法是：将2‑对羟基苯乙烯基‑异喹啉溶解于乙腈中，加入4‑溴甲基苯硼酸和碳酸钾，氩气保护下回流，析出黄色固体，抽滤，用去离子水洗涤固体，即得。本发明的有益之处在于：该荧光探针能够高选择性、高灵敏性响应生物硫醇，并具有良好的细胞渗透性和生物兼容性，可用于溶液和离体活细胞中生物硫醇检测以及活细胞内人参皂苷等具有抗氧化活性药物筛选
Crawler grass chopper
No full text本实用新型公开了履带碎草机，涉及碎草机技术领域，该设备包括履带机，所述履带机的一端设有力挖铲，固定机构的内部设有将互花米草打碎的碎草机构。本实用新型通过设置力挖铲和浮筒，可以使履带机在易陷滩涂中行驶时，操作力挖铲将其插入泥中，力挖铲往后发力，可辅助履带机向前移动，同时，履带机两侧装有浮筒，使履带机不会在易陷滩涂中陷车，进一步提高履带机在泥滩中的通行能力，达到了履带机在易陷滩涂中的通行能力更好的效果，从而解决现有互花米草治理设备容易陷车的难题，本实用新型中设置碎草机构, 启动驱动电机带动连接杆进行转动时使圆盘刀进行转动，通过定刀与转动的圆盘刀相配合，从而打碎防护罩下的互花米草，对其进行碎草治理
A digestion device for measuring COD of seawater
No full text本实用新型公开了一种测定海水COD的消解装置，包括外箱体，外箱体的内部设置有水浴锅，水浴锅顶部的右侧通过弧形塑料卡扣固定有多个进液软管，多个进液软管的进液端均贯穿外箱体并延伸至外箱体的右侧，水浴锅顶部的左侧也通过弧形塑料卡扣固定有多个出液软管，且多个出液软管的出液端均贯穿外箱体并延伸至外箱体的左侧，本实用新型涉及海水测定技术领域。该测定海水COD的消解装置，光催化与温控同时增加反应效率，提高消解效率，采用半导体光催化氧化技术，代替了国标法的高温高压消解，可以在低温或常温下对有机物进行氧化，并且氧化速率快、效率高，能使多数不能或难以降解的有机物完全矿化、且操作条件易于控制
Theoretical and numerical study on the fluctuation of the potential energy landscape
No full textThe diffusion of a Brownian particle exploring a meta-stable potential energy landscape with fluctuations is studied theoretically and numerically. The inverse harmonic part of the potential is replaced by a Gaussian approximated curve in the neighborhood of the saddle point and a new type of Monte-Carlo simulation algorithm is presented to simulate the random fluctuations of the potential energy. The probability of successful barrier surmounting and terminal arrival was calculated and compared with previous results. It is shown that, the replacement we have made is reasonable for the study of diffusion, which is primarily affected by the fluctuation of the barrier height, while other variations have little influence on the diffusion process. The Gaussian permutation and random Gaussian algorithms proposed here are expected to bring new ideas to the research of more complex problems