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31.
Extracellular products(ECP)produced by Vibrio anguillarum strain M3 originally isolated from diseased flounder(Paralichthys olivaceus)were prepared.ECP of M3 showed gelatinase,casinase,amylase and haemolytic activity on agarose plates.High protease activity against azocasin was detected.Bacterium M3 showed highest growth and protease activity at 25℃。The Protease present in ECP showed maximal activity at pH 8 and 55℃；was completely inactivated by application of 80℃ heat for 30 min;was completely inhibited by EDTA and HgCl2,and was partially inhibited by PMSF,SDS,MnCl2 and iodoacetic acid;but not inhibited by CaCl2 and MgCl2.The ECP was toxic to flounder fish at LD50 values of 3.1 μg protein/g body weight.The addition of HgCl2 and application of heat at 50℃ decreased the lethal toxicity of ECP.When heated at 100℃,ECP lethality to flounder was completely in-hibited.After intramuscular injection of ECP into flounder,it showed evident histopathological changes including necrosis of muscle,extensive deposition of haemosiderin in the spleen,dilated bolld vessels congested with numerous lymphocytes in the liver.These results showed that ECP protease was a lethal factor produced by the baterium V.anguillarum M3. 相似文献
32.
本研究将新发现的鳗弧菌毒力相关基因mltD(膜绑定溶胞壁质转糖酶基因)克隆于表达载体pET-32a(+),在大肠杆菌BL21(DE3)中诱导表达,并以镍琼脂糖亲和层析柱纯化表达的融合蛋白;经SDS-PAGE分析,纯化的融合蛋白为单一条带,分子量约为70.2 kDa,与理论预测值相符。生物信息学分析表明,鳗弧菌MltD由523个氨基酸组成,与其他弧菌的MltD氨基酸序列有较高的相似性;二级结构以α螺旋和无规则卷曲为主;功能区包括N端的信号肽区域、1个糖基转移酶结构域和C端的3个溶解酶结构域;蛋白的N端具有一段脂蛋白信号肽,信号肽外的其他部分为非细胞质蛋白。鳗弧菌MltD蛋白为亲水性;不稳定系数为27.40,属于稳定性蛋白;整个蛋白分子共有24个可能的抗原决定簇,具有较强的抗原性。推断MltD蛋白为1种外周膜脂蛋白,在弧菌中相对保守,抗原性强,可应用于抗弧菌病疫苗的开发研制。 相似文献
33.
Distribution of injected Vibrio anguillarum in body of P,,naeus monodon was studied with immunohistochemical method. Bacteria could be detected throughout the experiment in some individuals; however in lymphoid tissue, gill, heart and haemolymph of all vibrio injected shrimp, the bacteria could be observed only 5 min after injection. The bacteria density in haemolymph, haemolymph of the hepatopancreas and gills decreased with time. In the lymphoid organ and heart, the bacteria density was the highest 48 h after injection, then decreased. Nodules could be formed in the heart, lymphoid organ and injection site. 相似文献
34.
为了研究大菱鲆(Scophthalmus maximus)中趋化因子(Chenokine)的特征及其在大菱鲆免疫过程中发挥的作用,设计了本文中的实验。本实验从大菱鲆基因组和转录组数据库中鉴定了一个硬骨鱼特有的趋化因子CC亚家族成员—CCL34,并选取大菱鲆的8个健康组织以及2种细菌感染后的肠道和皮肤组织,采用实时荧光定量PCR(qRT-PCR)对其表达特征进行研究。序列分析结果显示该趋化因子全长mRNA包含1个327 bp的5’非编码区(UTR),1个246 bp的3’非编码区,和1个编码103个氨基酸残基长度为312 bp的开放阅读框(ORF)。此CCL34基因含有4个外显子和3个内含子。通过系统发育分析、共线性分析,将该大菱鲆趋化因子命名为CCL34。实时荧光定量PCR表明CCL34在大菱鲆健康组织中普遍表达,尤其在肾脏、肝脏、皮肤中有高水平表达。鳗弧菌(Vibrio anguillarum)和无乳链球菌(Streptococcus agalactiae)感染后,肠道组织CCL34基因表达量较对照组显著性上调(p<0.05),这表明CCL34可能在大菱鲆的肠道粘膜免疫中起重要作用。本研究为趋化因子家族功能的研究奠定了基础,为增强鱼类免疫力和抗病能力提供了理论依据。 相似文献
35.
We cloned and sequenced a prtV-like gene from Vibrio anguillarum M3 strain.This prtV gene encodes a putative protein of 918 amino acids,and is highly homologous to the V.cholerae prtV gene.We found that a prtV insertion mutant strain displayed lower gelatinase activity on gelatin agar,lower protease activity against azocasein,and lower activity for four glycosidases.This prtV mutant strain also had increased activity for two esterases in its extracellular products,as analyzed by the API ZYM system.In additi... 相似文献
36.
Shuo Li Chunmei Li Xubo Wang Yanan Wang Zhipeng Liu Teng Zhai Quanqi Zhang 《中国海洋大学学报(英文版)》2010,9(1):59-64
A novel immune-related gene was expressed in Japanese flounder (Paralichthys olivaceus) injected with Vibrio anguillarum. The complete cDNA contained a 169 bp 5’UTR, a 336 bp open reading frame (ORF) encoding 111 amino acids and a 556bp 3’UTR.
Six exons and five introns were identified in the PoIR2 gene. Blastp similarity comparison showed its encoding protein had 50% similarity to Danio rerio neuromedin S (NMS), but further alignment indicated they did not have NMS C-terminal conservational signature domain. So
it was not defined as an NMS homologue. Protein structure analysis indicated it had a 26aa signal peptide and was a secretory
pathway protein. RT-PCR demonstrated that the expression of PoIR2 was quickly induced and drastically increased in liver, kidney, spleen, gills, intestine, heart, and skeletal muscle after
infected with V. anguillarum. These results indicated that the PoIR2 might play some important role in Japanese flounder immune response system. This gene was named PoIR2 (P.olivaceus immune-related gene 2, GenBank accession number: EU224372). The mature PoIR2 peptide was expressed in BL21(DE3) pLysS using pET-32a(+) vector and a great part of the recombinant mature peptide existed
as soluble type. 相似文献
37.
基于16S rRNA和recA香鱼鳗利斯顿氏菌的分离鉴定 总被引：2，自引：0，他引：2
为研究引起香鱼(Plecoglossus altivelis)出血溃烂症病因及致病菌系统发育地位,本研究从患病香鱼的肝脏、肾脏及体表分离到11株病原菌(编号:X0901-X0911),运用常规细菌生理生化方法鉴定致病菌所属种类;运用16SrRNA基因、recA基因序列分析方法研究致病菌的系统发育地位。细菌生理生化鉴定结果表明:致病菌为鳗利斯顿氏菌,11株细菌生理生化结果相同,均为革兰氏阴性杆菌、氧化酶阳性、接触酶阳性、吲哚阳性、精氨酸脱羧酶阳性、精氨酸双水解酶阳性、硝酸盐还原阳性、甘露醇阳性、MR测定阳性;H2S阴性、V-P测定阴性等。根据16SrRNA基因、recA基因序列分别构建弧菌科常见细菌系统进化树,结果表明:致病菌与鳗利斯顿氏菌(Listonella anguillarum)均聚为一枝,聚类结果与细菌生理生化鉴定结果相符。致病菌与鳗利斯顿氏菌16SrRNA基因、recA基因的同源性分别为99.9%、99.8%。以recA基因构建的系统进化树的拓扑学结构与16SrRNA基因建树结果相类似。此外,与16SrRNA基因相比,recA基因在不同物种之间具有更高的鉴别能力,本研究表明recA基因适合作为弧菌科常见细菌物种间进化关系研究的标记。 相似文献
38.
2008年12月,江苏赣榆某渔场养殖半滑舌鳎出现大量死亡,症状主要表现为:头部、鳃盖及鳍基出血,尾鳍腐烂,腹腔膨胀并积有大量腹水,部分病鱼肠管脱出肛外。从病鱼肝脏、腹水中分离出大量优势生长的细菌,人工感染试验证明其对半滑舌鳎有较强的致病性。对3株分离菌进行了形态特征、理化特性、胞外酶及溶血素活性等表型生物学性状检验;测定了代表菌株的16S rRNA和gyrB基因序列,分析了16S rRNA和gyrB两种基因序列的同源性,并构建了系统发生树,比较了两种基因对分离菌的鉴定能力;结果表明gyrB基因用于细菌种间鉴定更具优越性;基于16S rRNA和gyrB基因的系统发育学分析表明分离菌与鳗利斯顿氏菌具有高同源性,根据分离菌的表型及分子特征,判定分离菌为利斯顿氏菌属(Listonella MacDonell and Colwell 1986)的鳗利斯顿氏菌[Listonella anguillarum (Bergeman 1909) MacDonell and Colwell 1986]。胞外酶活性及溶血活性检测表明3株分离菌均具有淀粉酶、蛋白酶、卵磷脂酶等胞外酶活性,在含7%家兔脱纤血液营养琼脂培养基上呈β型溶血,PCR检测3株供试菌均可扩增出大小约493 bp的溶血素基因和大小约248bp的金属蛋白酶基因。分离菌的耐药谱测定结果显示,对供试49种抗菌药物中的青霉素G等13种药物耐药,对羧苄青霉素等6种药物存在敏感与耐药的株间差异。 相似文献
39.
鳗弧菌(Vibrio anguillarum)是一种水产动物的重要致病菌,鳗弧菌分泌的胞外蛋白酶是该菌的致病因子之一.研究了环境因子对鳗弧菌W-1生长及胞外蛋白酶表达的影响.结果表明,鳗弧菌的最适生长温度为28℃,菌体生长量在21 h达到最高,胞外蛋白酶的活力在24 h最高.在37℃培养时,菌体生长和酶活力显著降低;添加葡萄糖、蔗糖、麦芽糖等碳源物质均能显著促进菌体的生长,但对于胞外蛋白酶表达有明显抑制作用;1 mmol/L EGTA对茵体的生长和胞外蛋白酶表达没有明显影响,而1 mmol/L的EDTA强烈抑制鳗弧菌的生长及其胞外蛋白酶的表达.添加不同浓度的氨苄青霉素对菌体的生长及其胞外蛋白酶表达有明显的抑制作用. 相似文献