群体感应在水产养殖中的应用前景

群体感应,也被称为"细胞间的交流",是细菌通过化学信号的传递进行彼此交流的一种方式。本文介绍了群体感应的机理和类型以及信号分子的检测方法,并结合实际情况,对其在水产养殖领域的应用前景进行了探讨,以期为群体感应在水产养殖中的应用提供理论依据。     

细菌生物被膜的研究进展及与群体感应的关系

细菌生物被膜是细菌耐药性形成的重要机制之一,是许多感染性疾病难以控制的主要原因,也是食品加工中存在的重大污染源。在生物被膜形成过程中,细菌的群体感应系统起重要作用。文章在现有的理论和研究基础上,就细菌生物被膜的特性以及群体感应系统在细菌生物被膜形成过程中的作用进行综述,旨在为通过群体感应抑制剂抑制生物被膜的形成提供全新的研究思路。     

opaR基因缺失对AHPND致病菌生物学特性及毒力质粒接合转移的影响

急性肝胰腺坏死病(acute hepatopancreatic necrosis disease, AHPND)严重影响了我国乃至全球对虾养殖业的发展。近期研究发现,致病菌毒力质粒携带trb型Ⅳ型分泌系统(type Ⅳ secretion system, T4SS)可在高菌体密度下介导毒力质粒发生接合转移,从而导致AHPND致病菌多样性。为探究群体感应与T4SS表达以及毒力质粒接合转移的关系,本研究以副溶血弧菌(Vibrio parahaemolyticus)群体感应系统的高密度调控子OpaR为研究对象,在致AHPND副溶血弧菌20130629002S01::cat(Vp2S01::cat)菌株基础上,利用同源重组技术构建opaR基因缺失株Vp2S01::catΔopaR。比较了出发菌株和缺失株在生长性能、运动性和生物被膜形成能力等方面的差异,分析了opaR基因对T4SS基因表达量以及质粒接合转移效率的影响。结果显示,opaR基因缺失不影响细菌的生长特性和群集运动,但泳动能力显著增加,生物被膜形成能力显著下降;接合转移实验显示,opaR基因缺失显著提高T4SS表达水平和接合转移效率。本研究为解析群体感应系统调控AHPND致病菌T4SS表达以及毒力质粒接合转移机制提供了基础数据,可为控制AHPND致病菌毒力质粒传播提供技术支撑。     

嗜水气单胞菌QseC在响应去甲肾上腺素中的作用

QseBC是一种普遍存在于细菌的双组分调控系统(two-component regulatory system,TCS),能感应细胞外环境因子信号,在调控细菌毒力中发挥着重要作用。为了研究嗜水气单胞菌(Aeromonas hydrophila)QseC在识别和响应宿主应激激素去甲肾上腺素(norepinephrine,NE)及其对毒力因子表达的调控作用,本实验首先构建了嗜水气单胞菌NJ-35缺失突变株ΔqseC与互补株qseC+,分析了在NE诱导下野生株和突变株的体外毒力因子表达和鱼体死亡率的变化。结果表明,qseC的缺失抑制了NE对嗜水气单胞菌的促生长作用,降低了NE对生物膜形成能力和溶血活性的增强作用及其对罗非鱼的致死率,而对运动性、脂肪酶与蛋白酶活性没有产生显著影响。本实验明确了嗜水气单胞菌QseC能够响应NE,从而调节菌株的毒力,对全面认识嗜水气单胞菌的致病机理及其与宿主的互作机制有着重要意义,并为探索疾病防控新技术奠定相关理论基础。     

硬骨鱼类虹彩细胞的研究进展

虹彩细胞是硬骨鱼中一类常见的色素细胞,含有大量的虹彩小体,其内含有由鸟嘌呤晶体构成的高折射率的反射小板,反射小板与低折射率的细胞质交替排列形成交替层的结构,光被交替层反射后通过多层薄膜干涉,使细胞呈现出彩虹般的颜色。虹彩细胞能在激素、交感神经系统或外界环境刺激的作用下通过改变反射小板的间距来实现颜色的变化。鸟嘌呤是虹彩细胞呈色和颜色变化的基础,其合成途径已被详细解析。虹彩细胞由多能的神经嵴细胞分化而来,从神经嵴细胞分化为成熟的虹彩细胞的谱系特化过程受到多个基因的调控,目前已在模式生物斑马鱼中发现了一些关键基因,并通过对这些基因间相互作用的研究初步解析了控制虹彩细胞特化的核心基因调控网络,同时也发现了其他影响虹彩细胞特化、分化及存活的基因和信号通路。本文综述了硬骨鱼类虹彩细胞呈色及颜色变化的机制、鸟嘌呤合成途径及调控基因以及影响虹彩细胞发育和存活的基因和信号通路的研究进展,并对未来的研究方向进行了展望,期望能对虹彩细胞有更深入的认识,并为研究虹彩细胞发育过程的基因调控提供参考。     

绵羊TIMP1基因克隆及真核表达载体的构建

为探究TIMP1基因在绵羊卵巢组织中的生物学功能及在繁殖调控中的作用机制。研究以绵羊卵巢组织为材料,提取RNA后采用RT-PCR技术检测TIMP1基因在产多胎和产单胎绵羊卵巢组织中的差异表达情况,另外利用PCR技术扩增得到绵羊TIMP1基因编码序列,共编码207个氨基酸。构建TIMP1基因的真核表达载体,利用限制性内切酶双酶切后插入pEGFP-C1载体中。提取质粒利用脂质体转染至HEK 293F细胞中,验证真核表达载体的表达情况,为进一步探索TIMP1基因的生物学功能及繁殖调控机制提供良好的基础材料。     

鱼类体色变异的遗传基础研究进展简述

虹彩细胞是硬骨鱼中一类常见的色素细胞,含有大量的虹彩小体,其内含有由鸟嘌呤晶体构成的高折射率的反射小板,反射小板与低折射率的细胞质交替排列形成交替层的结构,光被交替层反射后通过多层薄膜干涉,使细胞呈现出彩虹般的颜色。虹彩细胞能在激素、交感神经系统或外界环境刺激的作用下通过改变反射小板的间距来实现颜色的变化。鸟嘌呤是虹彩细胞呈色和颜色变化的基础,其合成途径已被详细解析。虹彩细胞由多能的神经嵴细胞分化而来,从神经嵴细胞分化为成熟的虹彩细胞的谱系特化过程受到多个基因的调控,目前已在模式生物斑马鱼中发现了一些关键基因,并通过对这些基因间相互作用的研究初步解析了控制虹彩细胞特化的核心基因调控网络,同时也发现了其他影响虹彩细胞特化、分化及存活的基因和信号通路。本文综述了硬骨鱼类虹彩细胞呈色及颜色变化的机制、鸟嘌呤合成途径及调控基因以及影响虹彩细胞发育和存活的基因和信号通路的研究进展,并对未来的研究方向进行了展望,期望能对虹彩细胞有更深入的认识,并为研究虹彩细胞发育过程的基因调控提供参考。

     

绵羊FHL2蛋白原核表达、蛋白纯化及多克隆抗体的制备

通过表达和纯化重组的绵羊FHL2抗原蛋白和制备其多克隆抗体,用于研究该蛋白的生物学功能及绵羊繁殖调控的作用机制。研究采用PCR技术扩增绵羊FHL2蛋白的编码序列,利用限制性内切酶双酶切后插入pET28a-sumo中,加入0.5 M IPTG诱导蛋白表达,利用Ni-NTA纯化获得48 ku的重组FHL2蛋白。以重组FHL2蛋白为抗原,混合佐剂皮下注射免疫BALB/c小鼠,获得抗FHL2的多克隆抗体,间接ELISA检测结果显示抗体效价达到1:6 400。通过转染pEGFP-C1-FHL2质粒至HEK 293F细胞,提取细胞总蛋白,利用制备的抗FHL2多克隆抗体检测蛋白的结合力,具有良好的免疫原性,为研究FHL2蛋白的生物学功能及繁殖调控机制提供良好的基础材料。     

rpoS对鳗弧菌MHK3株Hcp表达和杀菌能力的影响

溶血素共调节蛋白(Hcp)的合成和分泌是六型分泌系统(T6SS)行使功能的重要特征。RNA聚合酶的σ亚基RpoS参与调节细菌的生长和应激反应。为了探究RpoS对鳗弧菌(Vibrio anguillarum) T6SS的调控作用,本研究构建了MHK3?rpoS突变株,检测了部分表型特征的变化;利用lacZ半乳糖苷酶报告基因检测了突变株hcp1和hcp2在转录水平的变化;进行Western blot并定量分析突变株Hcp在翻译水平的变化;并通过细菌拮抗实验检测了突变株杀菌能力的变化。结果显示,MHK3?rpoS突变株的生长情况、泳动性、明胶酶活性及酪蛋白酶活性与MHK3野生株相比无显著差异(P>0.05),而平台早期的菌膜形成能力有显著上升(P<0.05);在各生长时期,MHK3?rpoS的hcp1和hcp2在转录水平的表达量均较MHK3有显著上升(P<0.01),最高时分别为MHK3的1.79倍和1.94倍;在翻译水平上,MHK3?rpoS在胞内和胞外Hcp的分泌均有显著升高(P<0.05),最高时分别为MHK3的1.59倍和1.31倍;同时,MHK3?rpoS对大肠杆菌(Escherichia coli) E5的杀菌能力约为MHK3的1%。研究表明,rpoS对鳗弧菌MHK3株的生长情况、泳动性、明胶酶活性及酪蛋白酶活性均无显著调控作用,但对平台早期的菌膜形成能力具有一定的负调控作用,在转录和翻译水平上也负调控Hcp的表达。而在杀菌能力上,rpoS发挥一定的正调控作用。说明菌株的杀菌能力强弱并非直接与Hcp的表达和分泌量正相关。本研究为进一步阐明T6SS的调控机制及其介导的杀菌作用机制提供了新思路,并丰富了其理论基础。     

杂色鲍的血细胞

通过光镜、电镜技术,比较了正常状态和受到细菌刺激后杂色鲍血细胞的变化.结果显示,正常状态下的杂色鲍血细胞分为三种类型:颗粒细胞、透明细胞和小细胞;杂色鲍血细胞具有很强的免疫功能,对外界刺激敏感;受到细菌刺激后杂色鲍的血细胞在结构和形态上均发生显著的变化.并推测,颗粒细胞的颗粒中含有大量的水解酶类和抗菌肽等免疫因子,发挥主要的免疫作用.     

Mechanisms of quorum sensing and strategies for quorum sensing disruption in aquaculture pathogens

In many countries, infectious diseases are a considerable threat to aquaculture. The pathogenicity of micro‐organisms that infect aquaculture systems is closely related to the release of virulence factors and the formation of biofilms, both of which are regulated by quorum sensing (QS). Thus, QS disruption is a potential strategy for preventing disease in aquaculture systems. QS inhibitors (QSIs) not only inhibit the expression of virulence‐associated genes but also attenuate the virulence of aquaculture pathogens. In this review, we discuss QS systems in important aquaculture pathogens and focus on the relationship between QS mechanisms and bacterial virulence in aquaculture. We further elucidate QS disruption strategies for targeting aquaculture pathogens. Four main types of QSIs that target aquaculture pathogens are discussed based on their mechanisms of action.     

Regulation of Vibrio alginolyticus virulence by the LuxS quorum-sensing system

Quorum sensing (QS) is a bacterial intercommunication system that controls the expression of multiple genes in response to population density. The LuxS QS system regulates the expression of several virulence factors in a wide variety of pathogenic bacteria. LuxS has been characterized to be responsible for producing a type of autoinducer, AI-2, which stimulates the expression of the luciferase operon in Vibrio harveyi. Vibrio alginolyticus is established as an opportunistic pathogen of several marine animals, and its LuxS QS system remains undefined. To investigate the pathogenic role of luxS in V. alginolyticus, the luxS mutants of both the standard strain ATCC 33787 and a fish-clinical isolate MVP01, named MYJS and MYJM, respectively, were constructed. The mutation resulted in reduced lethality to Pagrus major. Intraperitoneal LD(50) of MYJS and MYJM increased by 15- and 93-fold, respectively. The two luxS mutants exhibited a lower growth rate and defective flagellar biosynthesis. They also showed a significant decrease in protease production and an increase in both extracellular polysaccharide production and biofilm development. The results suggest that the LuxS QS system plays an important role in regulating the expression of virulence factors in V. alginolyticus.     

Effects of Quillaja saponins on growth,feed efficiency,digestive enzyme activities and metabolism of common carp (Cyprinus carpio L)

Common carp were fed diets containing various levels of Quillaja saponins (QS) (0, 150, 300 and 450 mg kg^?1 dry diet) in a completely computerized respirometric system for 4 weeks. Fish fed diets containing QS exhibited significantly higher ABW and specific growth rate than did those fed the control diet; those fed diets containing QS 150 grew fastest but were not significantly different from those fed diets with QS 300 and QS 00450. All the utilization efficiency indices, namely food conversion efficiency (FCE), protein productive value and PG were increased by QS supplementation. There were no significant differences in the average routine metabolic rate between treatments, indicating that dietary QS at the levels tested were not toxic to the carp. Increases in amylase and trypsin specific activities were observed at QS 300 and QS 450. Enzymes of carbohydrate metabolism such as G6PDH, 6‐phosphogluconate dehydrogenase and pyruvate kinase were not significantly affected by dietary QS. Activities of the aerobic enzyme Cox and to a limited extent that of the anaerobic enzyme lactate dehydrogenase were significantly increased by dietary QS but the net effect was a shift towards aerobic metabolism, indicating absence of stress and favouring the anabolic processes. Thus, Quillaja saponin was beneficial as a feed supplement in the common carp.     

芽孢杆菌R1共培养对嗜水气单胞菌NJ-1毒力基因表达的影响

为探讨芽孢杆菌潜在的淬灭靶点,本研究通过芽孢杆菌R1与嗜水气单胞菌NJ-1共培养,并对NJ-1生物量、信号分子产生量、毒力因子基因和群体感应(QS)关键调控基因表达量进行检测。结果显示:1NJ-1单独培养时,信号分子BHL和HHL的产量与NJ-1生长趋势一致,在27 h时达到峰值并启动Lux R表达,进而调控毒力因子金属蛋白酶、丝氨酸蛋白酶、肠毒素、气溶素和血溶素等表达,其在27~30 h时呈现峰值表达,通路Qse B在27~30 h启动表达,48 h时达到峰值,而种间调控Lux S基因表达一直处于低水平状态;2与芽孢杆菌R1共培养时,仅痕量AHLs信号分子被检出,毒力因子相关基因表达量均显著下调;Lux R表达受抑制,Lux S在36 h时表达量显著上调,未影响Qse B正常表达;3AHLs与R1共培养可被显著降解。研究表明,芽孢杆菌R1抑制NJ-1毒力因子相关基因表达可能与其对NJ-1多QS系统进行调控相关联。     

Effects of cyclic and regular feeding of a Quillaja saponin supplemented diet on growth and metabolism of common carp (Cyprinus carpio L.)

Fifteen common carp weighing 19.2±2.9 g were reared individually in chambers of a respirometer system which allowed feeding, and continuous measurement of oxygen consumption. The fish were divided into three groups of 5 each, and were given control feed (C group), control feed supplemented with 150 mg kg⁻¹ of Quillaja saponins (QS) (S150 group), and control and the saponin supplemented diet during alternate weeks (S150b group), for 56 days. Supplementation with QS at a level of 150 mg kg⁻¹ caused a significant (P<0.05) increase in metabolic growth rate, food conversion efficiency (FCE), protein productive value (PPV) and apparent lipid conversion (ALC) of carp, even as the metabolic rate was lower than the C group. The efficiency of energy utilisation indicated by parameters such as energy expenditure (EE), energy retention (ER) and amount of oxygen consumed per unit body mass gain (OPM) were also significantly (P<0.05) better in the S150 group. The performance of the S150b group was intermediate between the C and S150 group in this experiment. Possible reasons for the effects of dietary QS are discussed. It was concluded that there has to be continuous dietary supply of QS at the level of 150 mg kg⁻¹, for maximum positive effects of these substances on growth of common carp. This revised version was published online in August 2006 with corrections to the Cover Date.     

具有群体感应抑制作用的地衣芽孢杆菌T-1的动物安全及生态评价

摘要：该研究以具有群体感应抑制作用的地衣芽孢杆菌（Bacillus licheniformis）T-1为试验对象,参照国标的急性毒性实验方法检测其对动物的安全性及对水体生态环境的影响。安全评价试验采用异育银鲫、斑马鱼、小鼠为测试动物,结果显示：异育银鲫腹腔注射浓度为11200mg/L（2.6×1011 CFU/Ml）菌株T–1菌液后,连续观察96h,异育银鲫均健康生长,未出现不良症状或死亡;将斑马鱼浸泡在浓度为2240mg/L（5.2×1010 CFU/mL）的菌株 T–1菌液中,连续观察96h,斑马鱼均健康生长,未出现不良症状或死亡;SPF级小鼠(ICR品系)灌胃16800 mg/kg（3.9×1011 CFU/mL）体重浓度的菌株T-1,观察9d小白鼠健康生长无不良症状。水体生态评价采用小球藻、大型蚤及萼花臂尾轮虫为测试对象,不同浓度的菌株T-1在96h内,对小球藻生长无毒且促进其生长,对大型蚤生长无不良影响,对萼花臂尾轮虫生长无害,且在高浓度下对萼花臂尾轮虫的生长繁殖有促进作用。研究表明,具有群体感应抑制作用的地衣芽孢杆菌T-1对异育银鲫、斑马鱼及小鼠安全无毒害;对水体生态环境中的浮游生物也无不良影响,具有开发成微生态制剂的潜力。     

大鲵细菌性疾病研究进展

为深入了解和治疗大鲵(Andrias davidianus)细菌性疾病,针对已知的大鲵细菌性疾病的致病病原菌、发病的临床特征及诊断防治措施进行了简单概述,并提出了相应研究展望。     

白藜芦醇生物学活性及其在水产动物中的应用

白藜芦醇(resveratrol,RES)是一种多酚类防御素,广泛存在于葡萄、虎杖及蓝莓等多种植物之中.RES因具有抗氧化、抗炎、抗细菌、免疫调节等生物学活性而被作为新型绿色饲料添加剂应用于水产行业中.本文综述了RES的理化性质、生物利用率、生物学活性及其调节机制,并总结了RES在水产养殖中的应用.结果 表明,适宜浓度...     

Rate models for a marine biological filter

An empirically based model is proposed for the kinetics of oxidation of ammonia and nitrite in a marine biological filter at low concentration. Assuming first order kinetics, the rate constants for Nitrosomonas and Nitrobacter respectively are correlated as functions of pH, temperature and dissolved oxygen. A series of experiments using a recirculating batch biological filter was conducted to validate the model over a limited range of conditions. Predictions of the model agree satisfactorily with literature values of the rate constants and show favourable comparison with the observed experimental data.     

夏季美济礁潟湖养殖区海水细菌多样性及其与环境因子的相关性

美济礁地貌独特、生物资源丰富, 其海域微生物与渔业资源正被开发利用。为了解夏季美济礁潟湖海水养殖区细菌多样性及其与环境因子的相关性, 试验通过高通量测序技术分析了该海区海水细菌的群落结构组成, 探索了主要环境影响因子, 并采用 PICRUSt 对菌群功能进行了预测分析。美济礁潟湖海水细菌群落组成丰富, 分属于 29 个门、57 个纲、152 个目、256 个科、439 个属, 优势菌门为变形菌门(Proteobacteria)、蓝细菌门(Cyanobacteria)、 拟杆菌门(Bacteroidetes)、放线菌门(Actinobacteria)。各个站点的 3 个水层之间的 alpha 多样性指数指数均存在差异, 但均未达到显著水平。通过冗余分析发现, 主要环境影响因子为盐度、COD/NO₃-N、活性硅酸盐、COD。通过 PICRUSt 功能预测, 微生物功能以新陈代谢(氨基酸代谢、碳水化合物代谢、能量产生与转换)、遗传信息处理(蛋白质翻译与修饰、DNA 复制重组和修复)及信息处理(信号转导机制、辅酶运输、无机离子运输)为主。该研究分析夏季美济礁潟湖养殖区海水细菌群落结构及其与环境因子的关系, 为认识南海微生物多样性与合理开发利用、海域渔业的开发、生态系统的保护修复提供基础数据。