Purification and characterization of a 630 kDa bacterial killing metalloprotease (KilC) isolated from plaice Pleuronectes platessa (L.), epidermal mucus     

Tvete T  Haugan K 《Journal of fish diseases》2008,31(5):343-352

Antibacterial chemicals in the mucus of fish such as lysozyme, lectins, peptides and proteases provide an efficient first line of defence against pathogens. This study shows that there are at least three antibacterial proteins in plaice skin mucus in addition to lysozyme. One of these proteins is responsible for approximately 74% of the antibacterial activity and is a 630 kDa protease complex designated KilC (bacterial killing metalloprotease C). Purified KilC kills the bacteria Staphylococcus aureus, Escherichia coli, Bacillus subtilis and Pseudomonas aeruginosa efficiently. The protease activity of KilC is dependent upon the divalent cation Mg(2+) and shows pH dual optima of 5.0 and 8.0. The enzyme has a temperature optimum of 25 degrees C and is made up of at least five different sized peptides. Studies with protease inhibitors show that the catalytic site of KilC may be cysteine- or serine protease-like. KilC may kill bacterial cells by acting directly upon the bacteria or by producing low molecular weight bioactive compounds such as peptides.  相似文献   

The influence of salmon surface mucus on the growth of Flavobacterium columnare     

Staroscik AM  Nelson DR 《Journal of fish diseases》2008,31(1):59-69

Flavobacterium columnare is the causative agent of columnaris disease. The presence of lesions on the gills, skin and fins of diseased fish suggests that F. columnare is able to utilize fish skin mucus as a substrate for growth and that exposure to this material would alter the expression of genes involved in the colonization of the outer surfaces of the fish. Growth, biofilm formation, extracellular protease production and changes in protein expression of F. columnare strain C#2 cultured in media supplemented with juvenile Atlantic salmon skin mucus were compared with the same media without mucus. C#2 was able to grow by using mucus as the sole nutrient source. Growth in mucus-containing media induced cells to grow as a biofilm and extracellular protease activity increased in mucus-containing cultures. SDS-PAGE protein profiles showed that expression of six extracellular proteins increased in mucus-containing media. These results demonstrate that salmon surface mucus promotes the growth of F. columnare and that exposure to mucus alters the growth characteristics of this bacterium with regard to protease production and biofilm formation. Further characterization of mucus-induced physiological changes will increase our understanding of the basis of virulence of this economically important fish pathogen.  相似文献   

鳜浸泡嗜水气单胞菌全菌疫苗后皮肤黏液抗体的变化   总被引：5，自引：0，他引：5       下载免费PDF全文

罗霞  潘厚军  巩华  刘瑞明  石存斌  吴淑勤 《中国水产科学》2007,14(5):823-828

应用间接ELISA方法,研究经嗜水气单胞菌(Aeromonas hydrophila)疫苗浸泡免疫后鳜(Siniperca chuatsi)皮肤黏液中抗体消长规律,以揭示鳜局部黏膜免疫在免疫保护中的作用及浸泡免疫的保护效果。结果表明,鳜皮肤黏液中抗体滴度在免疫后第7天达到峰值211,抗体从开始形成到消失持续21 d;添加佐剂(IMS1312、葡聚糖、莨菪碱、食盐)可以提高鳜皮肤黏液及血清中抗体滴度和相对免疫保护率,其中添加IMS1312组免疫保护率最高,达77.8%,免疫保护与抗体滴度成正相关(R2=0.79,P<0.05);以溶菌酶为代表的非特异性免疫在免疫后1周内即抗体未形成时,对鱼体起主要保护作用。此外,通过比较鳜血清与皮肤黏液中的抗体消长规律,发现血清中抗体滴度峰值出现时间较迟(第14天),抗体持续时间较长(42 d),初步推断鳜皮肤黏膜中可能存在相对独立于全身免疫系统的局部黏膜免疫系统。  相似文献   

黄鳝血清、肠黏液免疫球蛋白的分离纯化及部分特性分析   总被引：4，自引：0，他引：4  

张静  安利国  张福淼  谢萍  杨桂文 《水产科学》2007,26(3):130-133

采用硫酸铵盐析、SephadexG-200凝胶层析等方法从黄鳝血清和肠道黏液中分离纯化出黄鳝血清和黄鳝肠黏液免疫球蛋白。SDS—PAGE分析表明，二者的理化性质相同；经β-巯基乙醇处理后解离形成重链和轻链两个亚单位，分子量分别为74kD和29kD。试验结果表明，黄鳝血清和肠黏液中存在免疫球蛋白，二者具有相同的层析和SDS-PAGE电泳特性。  相似文献   

Epidermal proteases of the Japanese eel     

F. Aranishi  M. Nakane 《Fish physiology and biochemistry》1997,16(6):471-478

A fluorescent-sensitive assay was used to demonstrate the protease activity in the dorsal skin of Japanese eel (Anguilla japonica). Two distinct extracts were separately prepared from skin mucus and epidermal cell layers, with no mutual contamination. The epidermal extract was sensitive to various substrates, whereas there was no, or only marginal, susceptibility to the same substrates for the mucous extract. Optimum hydrolysis pHs of the epidermal extract was variable and below pH 7.0, and the optimum hydrolysis temperatures were between 40 and 50 °C. In addition, Tos-Phe-Ch₂Cl, chymostatin, CdCl₂, CuCl₂, HgCl₂ and ZnCl₂ inhibited protease activities to different extents. Several other reagents specifically affected the protease activities, and their induced effects were useful for the identification of epidermal proteases. The findings indicate that a proteolytic factor, exhibiting various enzymological specificities, is retained within epidermal cell layers of Japanese eel. This factor is composed of 4 distinct proteases, such as cathepsins L and B-like proteases, a serine protease and an aminopeptidase.  相似文献   

不同环境条件对溶藻弧菌粘附大黄鱼肠粘液的影响   总被引：9，自引：3，他引：9  

鄢庆枇  陈强  邹文政  马甡  庄峙厦  王小如 《水产学报》2006,30(2):254-259

研究了不同环境条件下溶藻弧菌对大黄鱼肠粘液的粘附作用。采用细菌计数法测定溶藻弧菌的粘附作用。粘附的细菌经SYBR GreenⅠ染色后在荧光显微镜下观察，用数码相机拍照后在电脑上计数。试验结果表明，溶藻弧菌对大黄鱼肠粘液的粘附量随菌浓度的升高而升高，并在1—1．5h内趋于饱和；粘附作用在15—30℃、pH偏酸时较强；盐度在5—35范围内对前肠粘液的粘附作用影响不明显，后肠粘液的粘附作用在此范围内随盐度增大而加强，在盐度为0时，溶藻弧菌对前、后肠粘液都无粘附作用；56℃热处理5min及60℃处理1h均能大幅减弱溶藻弧菌对两种肠粘液的粘附作用，表明溶藻弧菌表面的某些热敏结构在粘附作用中起着重要作用。根据以上结果，可以认为溶藻弧菌能够很好地粘附于大黄鱼肠粘液层，其粘附作用受温度、盐度、pH值等环境因子影响很大，溶藻弧菌表面的某些热敏结构可能在粘附过程中起着重要作用。研究结果表明，溶藻弧菌对大黄鱼肠粘液的粘附作用是可控制的，这对于鱼类养殖疾病的控制有一定的参考价值。  相似文献   

Effects of steroid hormones on immunoglobulin M (IgM) in rainbow trout, Oncorhynchus mykiss   总被引：3，自引：0，他引：3  

Y.Y. Hou  Y. Suzuki  K. Aida 《Fish physiology and biochemistry》1999,20(2):155-162

The immunosuppressive effects of steroid hormones were evaluated as the response against implanted steroid hormones, cortisol (F), testosterone (T), estradiol-17 (E2), and 11- ketotestosterone (11-KT), in juvenile rainbow trout. In long term experiments (5 weeks), fish were given a single intraperitoneal implant of F or T. A clear suppressive effect of plasma IgM levels with F and T was not necessarily obtained, although mucus IgM levels were reduced corresponding to the elevated plasma steroid hormone levels. In short term experiments (1 week), intraperitoneal implantation of T, 11-KT and E2 suppressed plasma and mucus IgM levels, although the effects were not dose-dependent. When administered through diet, F and T caused a suppression of plasma IgM levels; F administration at both high and low dosages caused a significant decrease in plasma IgM levels, while only a high dose of T caused the suppression. These results suggest that sex steroid hormones, as well as F, have immunosuppressive functions in rainbow trout.  相似文献   

母牛发情周期和妊娠早期宫颈-阴道粘液电导率变化规律的研究   总被引：3，自引：0，他引：3  

殷国荣  王建军 《中国农业科学》1992,25(4):77-81

 本文应用自行设计安装的MFR-1型母牛发情鉴定及早期妊娠诊断仪，将特制探测电极插入母牛阴道穹窿部，测定了94头母牛宫颈-阴道粘液的电导率（CVMEC）。结果表明，母牛在发情周期和妊娠早期CVMEC呈规律性变化。发情周期正常的母牛，CVMEC在间情期处于较低水平（5.64mS/cm），前情期开始上升（8.93mS/cm），发情期达峰值（13.40mS/cm），后情期开始下降（9.84mS/cm），最后降至间情期水平。输精后，未妊与妊娠母牛CVMEC的变化规律不同。在发情周期的各时期，未妊母牛高于妊娠母牛；未妊母牛在输精后21天左右返情，CVMEC再度出现峰值；妊娠早期母牛则保持与间情期相近的较低水平（5.08mS/cm）。笔者认为，母牛CVMEC的规律性变化的生理学基础是体内激素水平和宫颈-阴道粘液的周期性变化，据此进行发情鉴定和早期妊娠诊断是可行的。与目前使用的方法相比，CVMEC法更具有实用价值。  相似文献   

鸭跖草营养器官的解剖学研究     

马淑英  王萍 《吉林农业大学学报》1996,18(1):34-37

本文报道了药用植物鸭跖草的根、茎和叶的解剖结构。鸭跖草的木质部为六原型，韧皮部与木质部排列，根皮层中分布有粘液细胞和晶体细胞。其内皮层具凯氏带。茎中维管束散生，许多近表皮薄壁组织细胞中含有分散的或成束的多个晶体，茎中央薄壁组织中含有粘液细胞和单宁细胞。叶为两面叶，叶肉组织分栅栏组织和海绵组织，栅栏组织细胞一层，叶中含有粘液细胞和晶体细胞。  相似文献   

黄鳝体表粘液中基因组DNA的提取与分析     

胡彬  曹哲明  丁炜东  邴旭文 《安徽农业大学学报》2016,43(2):305-308

黄鳝基因组DNA一直采用抽血或剪取肌肉来提取,但是采血或剪取肌肉后黄鳝成活率很低。为了保留亲本,尝试从黄鳝体表粘液中提取DNA,并采用紫外分光光度计对提取到的DNA的纯度和浓度进行了检测。结果表明,用蛋白酶K消化,酚仿抽提后从粘液中得到的DNA的纯度与从肌肉中提取到的相差不大,但是提取到的量较少。为了验证提取的DNA是不是黄鳝基因组DNA,用黄鳝actin引物和18S r RNA基因引物扩增之后检测到了粘液中的较亮的DNA片段。为了进一步确认检测到的是黄鳝的DNA,我们又对得到的DNA进行了扩增,并得到的产物进行了克隆和测序。结果表明,粘液中提取到的DNA是黄鳝的基因组DNA。随后又选择了60尾黄鳝,提取其粘液之后对其成活率进行统计,养殖1个月后黄鳝的成活率都在95%以上。此黄鳝体表粘液提取DNA方法能提取高质量的DNA,可用于用于PCR鉴定。  相似文献