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《中国动物传染病学报》2016,(5)
通过生物信息学方法预测2-型猪链球菌(Streptococcus suis serotype 2,SS2)保护性抗原ESA(epidermal surface antigen,ESA)在SS 2中的分布和B细胞表位,为研究SS2的多表位疫苗奠定基础。以ESA推定的氨基酸序列为基础,通过生物信息学软件对ESA进行信号肽、跨膜序列等分析,然后分别以Kyte-Doolittle法、Emini法和Jameson-Wolf法分析蛋白的亲水性、表面可能性以及抗原指数;辅以Garnier-Robson法、Chou-Fasman法和Karplus-Schulz法分析蛋白二级结构中柔性区域,进而预测ESA的B细胞表位。结果表明,ESA为跨膜蛋白,B细胞表位位于N-端第30~35、59~64、178~185、245~253区域。多参数或多个软件联合使用能预测SS2保护性抗原ESA的B细胞表位和细胞定位,为实验研究提供帮助,大大缩减实验时间。 相似文献
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以鸭肝炎病毒(DHV)SN株VP1基因序列为基础,运用Garnier-Robson、Chou-Fasman和Karplus-Schulz方法预测VP1蛋白的二级结构,并分别运用Kyte-Doolittle方法、Jameson-Wolf方法和Emini方法预测蛋白的亲水性、抗原指数和表面可及性,最后结合吴玉章的方法综合分析预测其B细胞抗原表位.结果显示,VP1蛋白C端第132～137、177～186、209～219区段有较好的亲水性、表面可及性和较高的抗原指数,并且在二级结构上含有易形成抗原表位的无规则卷曲和β-转角,可能是VP1蛋白的B细胞抗原优势表位;DHV SN株与弱毒疫苗株在推测的VP1 B细胞抗原表位177～186和209～219两个区域氨基酸序列变异较大. 相似文献
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为预测番鸭呼肠孤病毒(DRV)YB株σC蛋白二级结构和B细胞抗原表位,对DRV YB株的σC蛋白氨基酸序列与参考DRV相应序列以DNAStar软件进行同源性和遗传进化树分析;采用SOPMA法、Gamier-Robson法、Chou-Fasman法和Karplus-Schultz法方法预测其二级结构;用Hopp-Woods法、Emini法、Jameson-Wolf法和吴玉章等建立的方法分别预测蛋白的亲水性、表面可能性和抗原指数,然后综合分析其B细胞抗原表位.结果显示,DRV YB与国内DRV相应序列同源性在93.7％～99.6％,代表性较强;B细胞表位可能在15～27、36～49、40～49及90～98区段或其附近,这4个被预测的表位均含有β-转角和无规卷曲结构.本研究为进一步确定σC蛋白的B细胞表位和反向()计提供了理论基础. 相似文献
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为了对异源抗体对接方法预测流感病毒血凝素B细胞构象表位的可行性进行分析,本研究基于蛋白质相互作用界面的形状互补理论,使用异源抗体(来自于抗原抗体复合物1G9M、1RVF、1TPX、1NCA、1A2Y)的晶体结构同流感病毒血凝素(A/Aichi/2/68(H3N2))的晶体结构进行刚性分子对接,通过计算B细胞表位的预测表面和实际表面C-α原子之间的RSMD以及表位氨基酸的准确率,分析用异源抗体探测出血凝素B细胞表位的可能性。结果表明,不同的异源抗体均可结合到A/Aichi/2/68(H3N2)的已知B细胞表位,最佳的对接构象与原晶体结构重叠后表位残基C-α原子的均方根偏差均小于0.6埃,表位氨基酸预测的准确性大于60%,说明异源抗体对接方法可以尝试用于B细胞构象表位的预测。 相似文献
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运用ANTHEPROT 5.0综合分析二级结构、亲水性、表面可及性与抗原性指数,预测MRP的B细胞优势袁位簇;PCR扩增出表达预测的抗原肽段的基因(mrp-1),构建重组原核表达载体pET32a-mrp-1,IPTG诱导表达,以渗透性休克法纯化重组蛋白;使用重组蛋白免疫血清和灭活猪链球菌2型SS2免疫血清Western blotting检测重组蛋白的免疫原性及其与SS2免疫血清的反应性;纯化的重组蛋白免疫雌性SPF昆明小鼠,SS2攻毒受试动物评估重组蛋白对小鼠的免疫保护力.Western blotting显示预测的950～1 210 aa肽段与PET32a重组表达的蛋白(MRP)具有良好的免疫原性和反应性;攻毒试验结果表明重组蛋白对小鼠的免疫保护率为菌体免疫所提供保护率的66.7％.研究为利用该细胞表位簇作为高效免疫制剂奠定了基础. 相似文献
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根据前期试验中测得的蓝舌病病毒VP7蛋白的基因序列和推导的氨基酸序列,利用DNAStar软件和Biosun软件进行生物信息学预测,以单参数(亲水性、可及性、柔韧性、抗原性)预测为基础,结合二级结构预测来综合分析蓝舌病病毒VP7蛋白的B细胞表位.比较两种软件的预测结果发现,在蓝舌病病毒VP7蛋白的381个氨基酸序列中,第81～85、198～202、235～239、253～257区域有较好的亲水性、表面可及性和较高的抗原指数,并且在二级结构上含有易形成抗原表位的转角和无规则卷曲,最有可能为蓝舌病病毒VP7蛋白的B细胞线性优势表位.上述预测和判定结果表明,在蓝舌病病毒VP7蛋白序列中存在优势B细胞线型表位,为进一步合成多肽并分析已获得的VP7蛋白单抗的结合表位奠定了基础. 相似文献
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用生物信息学方法比较了GenBank中9株Ⅰ型IBDV与2株Ⅱ型IBDV基因片段A前体多聚蛋白的推导氨基酸(aa)序列,预测并分析了上述两型IBDV毒株前体多聚蛋白的线性B细胞抗原表位.结果表明,两型IBDV毒株的前体多聚蛋白有75个aa残基的差异,其上大多数预测B细胞抗原表位的位置及其指数值均无明显差异;但两型IBDV在第3～10位中的aa残基差异影响了该序列肽的预测指数和结构,以及与Ⅰ型IBDV多抗的反应性. 相似文献
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对2005年四川资阳脑膜炎病例猪链球菌2型分离株ZYH33溶血素编码蛋白中包含多个抗原表位的第230～593氨基酸残基区域的基因片段进行扩增并克隆.基因片段经酶切处理后插入表达载体pQE-30的BamH Ⅰ和Sal Ⅰ位点之阃,构建融合表达质粒.转化宿主菌TG1经IPTG诱导后融合基因得到了表达,用猪链球菌2型菌体抗血清对表达的融合蛋白进行免疫印迹试验,分析融合蛋白的免疫反应性.试验结果提示该溶血素蛋白第230～593氨基酸残基区域可作为猪链球菌的诊断抗原,为基因工程疫苗的研制奠定基础. 相似文献
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致病性猪链球菌2型的病原分离鉴定及毒力因子的PCR检测 总被引：6，自引：3，他引：3
通过对广西某猪场急性死亡的猪进行病原分离,分离到革兰氏阳性球菌,用链球菌快速鉴定试剂条Rapid ID 32 Strep鉴定为猪链球菌2型;并对分离菌进行猪链球菌2型荚膜多糖抗原（cps2J）及其重要毒力因子溶菌酶释放蛋白（MRP）、细胞外蛋白因子（EF）和溶血素（SLY）的多重PCR检测,同时对cps2J基因进行序列测定,与GenBank发表的猪链球菌2型相比,同源性为98.8%,毒力因子MRP、EF和SLY检测均为阳性,证实广西某猪场急性死亡的猪为高毒力猪链球菌2型感染所致。动物试验中,该菌可引起小白鼠部分死亡,家兔体温最高升至40.3℃,最后败血而死。 相似文献
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Kim D Han K Oh Y Kim CH Kang I Lee J Gottschalk M Chae C 《Canadian journal of veterinary research》2010,74(4):314-316
The objective of this study was to determine the capsular serotypes and potential virulence factors of Streptococcus suis isolated from pigs with polyserositis. Among the 24 isolates evaluated, serotype 3 [7 (29%) of the isolates] and serotype 4 [5 (21%)] were the most common. The isolates were also studied for the presence of the genes mrp, epf, and sly, which encode muramidase-released protein (MRP), extracellular factor (EF), and suilysin (SLY), respectively. Of the 24 isolates, 8 carried mrp: 4 of serotype 3, 2 of serotype 2, and 2 of serotype 4. One mrp(+) isolate (serotype 2) also carried the epf gene. All 24 isolates carried the sly gene. The serotype and genotype distribution greatly differed from that reported for isolates from pigs with other clinical manifestations of S. suis infection in other countries. 相似文献
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Characterization of the invasion of porcine endothelial cells by Streptococcus suis serotype 2 
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下载免费PDF全文 Ghyslaine Vanier Mariela Segura Marcelo Gottschalk 《Canadian journal of veterinary research》2007,71(2):81-89
Streptococcus suis serotype 2 is an important swine pathogen associated mainly with meningitis. In a previous study, we demonstrated the ability of S. suis serotype 2 to adhere to and invade immortalized porcine brain microvascular endothelial cells (PBMECs) forming the blood-brain barrier. The aim of the current work was to further characterize the mechanism(s) by which S. suis invades porcine endothelial cells. The ability of several S. suis strains to interact with PBMECs was not found to correlate with their geographic origin, virulence, host of origin, or suilysin production. Characterization studies demonstrated that proteinaceous adhesins/invasins, cell wall components, lipoteichoic acid, and serum components (including fibronectin) were involved in interactions between S. suis and PBMECs. In addition to PBMECs, S. suis was able to adhere to and invade 2 porcine aortic endothelial cell lines and primary PBMECs. 相似文献
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为了探讨猪链球菌2型(Streptococcus suis serotype 2,SS2)的Ⅲ型溶血素是否具有溶血活性以及Ⅲ型溶血素在SS2致病过程中的作用,本研究利用同源重组基因敲除法成功构建了SS205ZY的Ⅲ型溶血素(slyrp)基因缺失突变菌株△slyrp及双基因缺失突变菌株△sly/△slyrp,并比较了野生菌株和基因缺失突变菌株的溶血能力以及对小鼠的致病力.结果表明,slyrp基因敲除后可导致SS2裂解红细胞的能力有所下降,而双基因缺失突变菌株△sly/△slyrp的溶血能力完全丧失;slyrp基因敲除后对小鼠的致病力没有影响.结果提示猪链球菌2型Ⅲ型溶血素具有一定的溶血能力,该Ⅲ型溶血素在SS2感染过程中,对溶血素(sly)起协同作用,不是SS2主要的毒力相关基因. 相似文献
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Knowledge of virulence factors of Streptococcus suis is limited. Several virulence factor candidates have been proposed, among them suilysin, which is responsible for a toxic effect on epithelial cells. The aim of this study was to detect the suilysin gene sequence in Streptococcus suis strains of various origin. In total 63 Streptococcus suis isolates were investigated. Forty four of them originated from tissues of streptococcosis affected animals. The remaining 19 strains were isolated from tonsils of healthy carrier pigs. Suilysin gene specific sequence was detected in 79% of the strains tested. In isolates obtained from pigs with signs of streptococcosis this gene sequence was recorded in 85% of cases. In Streptococcus suis strains isolated from healthy carrier pigs the suilysin gene was detected in 63% of the isolates. It seems that suilysin toxic activity is only one of the many steps involved in the pathogenesis of Streptococcus suis infection and that strain's virulence cannot be stated only on the basis of suilysin gene sequence presence. 相似文献
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Putative mannose-specific phosphotransferase system component IID represses expression of suilysin in serotype 2 Streptococcus suis 总被引：2，自引：0，他引：2
In this study, we generated a genomic mutant library from a North American strain of serotype 2 Streptococcus suis using the pGh9:ISS1 transposition vector. Suilysin is the hemolysin made by S. suis. A hyper-hemolytic mutant was identified by screening for hemolytic phenotype using media with human blood. The hyper-hemolytic phenotype was characterised by a quantitative hemolysis microplate method. The use of green fluorescent protein (GFP) as a reporter also showed that suilysin gene expression was greater in the mutant. DNA sequence analysis of 3.8 kb surrounding the ISS1 insertion site revealed four open reading frames (ORFs) with three consecutive ORFs that belong to a putative mannose-specific phosphotransferase system (PTS). The S. suis gene homologous to mannose permease IID, manN, was interrupted by the transposon. A complementation test showed that manN repressed the expression of suilysin and the absence of manN was responsible for the hyper-hemolytic phenotype. However, both wild type and isogenic hyper-hemolytic mutant S. suis fermented mannose, glucose and lactose. Thus, despite its potential roles in carbohydrate transport, phosphorylation and metabolism, the manN homologue in the putative mannose-specific PTS regulates gene expression in S. suis. 相似文献
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Streptococcus suis is a major cause of meningitis, sepsis and arthritis in piglets and a zoonotic agent. Survival in the blood circulation system represents a major step in pathogenesis of S. suis infections. To get further insights into the mechanisms of S. suis survival in the host, we compared a highly virulent S. suis serotype 2 strain with its non-encapsulated and suilysin-deficient mutants in their abilities to resist phagocytosis and killing by polymorphonuclear neutrophils (PMNs) and mononuclear cells. PMNs displayed a higher capacity to take up encapsulated bacteria than mononuclear cells, whereas both cell types internalized efficiently non-encapsulated S. suis. Differentiation of extracellular and intracellular survival of the WT strain revealed that in PMNs the majority of the cell-associated streptococci were intracellular, whereas in mononuclear cells the majority remained attached to the cell surface. S. suis survived mainly extracellularly, since both cells killed intracellular bacteria to a similar extent. As a consequence of different resistance to phagocytosis, only the encapsulated S. suis strains survived co-cultivation with PMNs. Comparison of the WT strain with its encapsulated suilysin-deficient mutant revealed reduced survival of the mutant after co-cultivation with PMNs. Involvement of suilysin in inhibition of phagocytosis was further confirmed by the use of anti-suilysin antibodies and recombinant suilysin. Kinetic experiments with PMNs suggested that reduced survival of the mutant strain was mainly associated with an increased uptake, whilst both strains adhered similarly. Concluding, our results indicate that the capsule and the suilysin play important roles in S. suis survival in the host by interfering with phagocytic uptake. 相似文献
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为验证猪链球菌2型荧光PCR检测方法对临床样品检测的敏感性和适用性以及该方法所拥有的独特优点,分别用荧光PCR法、常规PCR法和细菌分离法对人工感染猪链球菌2型的小鼠肝脏和发病猪的心、肝、脾、肾等实质器官和血液、喉拭子进行抗原检测。结果显示,荧光PCR法检出率为70.8%,明显高于普通PCR法（检出率为20.8%）,也高于常规细菌分离法（检出率为45.8%）。由于临床样品常常会被其他细菌污染,细菌分离法很难准确分离到链球菌。但荧光PCR法不受其他细菌污染的影响,对实验室培养的猪链球菌2型菌液,该方法检测滴度可达10-6/0.1mL（42～52 CFU/0.1 mL）,而普通PCR方法检测滴度仅为10-4。 相似文献
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利用荧光标记技术,采用2对引物初步建立检测猪链球菌荧光DNA扩增片段长度多态性方法,结果表明12株猪链球菌扩增的多态性位点数从51～98条不等,该方法能够检测猪链球菌的多态性,区分不同血清型以及同一血清型不同特性的菌株,可用于菌株鉴定及流行病学研究中细菌源的追踪。 相似文献