首页 | 本学科首页   官方微博 | 高级检索  
相似文献
 共查询到20条相似文献,搜索用时 140 毫秒
1.
应用DNA芯片研究禽致病性大肠杆菌可能致病基因的表达.构建禽致病性大肠杆菌毒力基因、潜在毒力基因的DNA芯片,应用基因芯片技术对同属O2血清型的禽高致病性大肠杆菌E058株和低致病性大肠杆菌E526株在体外LB培养基和鸡血清培养状态下进行差异表达分析.结果:在体外LB静置培养状态下,低致病株E526与高致病株E058相比共有16个差异基因,均为下调基因.在鸡血清静置培养中,E526与E058相比共有15个差异基因,均为下调基因.应用基因芯片成功筛选了禽致病性大肠杆菌在体外不同条件下的毒力基因及可能毒力基因中差异表达基因,表明一些铁摄取系统相关基因对APEC的毒力较重要,同时也筛选出了一些新的可能致病基因aes-1,aes-2,aes-3,aes-4,aes-6,aes-8,aes-10,aes-13,aes-15,aes-31等.  相似文献   

2.
为了克隆与分析霍乱弧菌O1E1Tor流行株与非流行株两类菌株基因组差异片段 ,采用抑制差减杂交技术 (suppressionsubtractivehybridization ,SSH)分别以国内保留的流行株Wujiang 2及非流行株Js 32一株作为被检菌 ,另一株作为参考菌进行基因组差异研究 .在进行的差减杂交实验中 ,流行株Wujiang 2共检出 34个特异差异片段 ,经同源检索共代表 35个基因片段 ,其中包括许多重要的霍乱弧菌毒力相关基因如CTX遗传单元、TLC因子及可移动外来成分如霍乱弧菌整合子RVC序列及Tn10转位酶 .非流行株Js 32共检出 14个特异差异片段 ,经同源检索未能检出同源片段 ,可能代表新基因序列 .研究表明 ,霍乱弧菌流行株与非流行株基因组存在较多差异基因 ,表现在毒力、毒力相关基因、代谢以及其它噬菌体等可转移的基因成分  相似文献   

3.
[目的]探究毒力基因aerobactin与sit操纵子与禽致病性大肠杆菌E058株致病作用的相关性.[方法]利用Red同源重组方法,构建APEC E058株aerobactin与sit操纵子基因缺失株E058Δvir,并通过一系列的体内及体外试验对其生物学特性进行研究.[结果]生长曲线测定、细菌侵袭试验及体外竞争等试验结果表明,突变株与亲本株差异不显著;体内动态分布试验结果显示,突变株E058Δvir在5个被检脏器中均极显著地低于亲本株(P<0.001).[结论]aerobactin与sit操纵子与禽致病性大肠杆菌E058株的致病性相关,是其重要的致病因子.  相似文献   

4.
利用高通量测序获取1株抗原式为3,10∶a,r,z6的沙门氏菌(Salmonella)GX150603的全基因组序列.根据鞭毛抗原序列、致病性和抗性基因预测GX150603的血清型,利用分子生物学软件分析基因组岛和前噬菌体,并与其他菌株进行全基因组系统发育分析.经鉴定GX150603为韦太夫雷登沙门氏菌(Salmonella enteric asubsp.enterica serovar Weltevreden),并推测由于鞭毛抗原的氨基酸变异导致该菌株与H∶a血清产生了交叉反应.GX150603基因组携带148个毒力相关基因,8个耐药相关基因,27个基因组岛和3个前噬菌体.基因组岛包括沙门氏菌毒力岛SPI-1/2/3/4/5/6/9/11/13/14/19,centisome 63,CS54和3个该血清型特有的基因组岛.与7个韦太夫雷登沙门氏菌欧洲分离株比较,GX150603有4个共线性片段重排,4个共线性片段缺失和1个共线性片段存在较大差异,进化分析显示,同一血清型仍然表现出最近的亲缘关系.  相似文献   

5.
目的为测定小肠结肠炎耶尔森菌噬菌体phiYe-F10的裂解谱,分析噬菌体phiYe-F10裂解能力与宿主毒力基因间的关系。方法采用双层平板法观察噬菌体phiYe-F10对213株小肠结肠炎耶尔森菌,36株假结核耶尔森菌和1株鼠疫耶尔森菌的裂解能力;同时对不同来源不同地区的小肠耶尔森菌进行黏附侵袭位点基因(ail)、肠毒素基因(ystA、ystB)、黏附素基因A(yadA)、毒力因子基因F(virF)、O∶3血清型特异性基因(rfbc)检测和血清型别鉴定。结果表明213株小肠结肠炎耶尔森菌包括O∶3血清型84株(其中rfbc+78株),O∶5血清型10株,O∶8血清型13株,O∶9血清型34株,其它及未分型的共72株。携带毒力质粒的典型致病性小肠耶尔森菌(ail+,ystA+,ystB-,yadA+,virF+)77株,毒力质粒丢失的致病性小肠结肠炎耶尔森菌(ail+、ystA+、ystB-、yadA-、virF-)15株,其它非致病基因型121株。噬菌体phiYe-F10裂解71株O∶3小肠结肠炎耶尔森氏菌,其中致病性小肠结肠炎耶尔森菌52株,非致病性小肠结肠炎耶尔森菌19株。对O∶5,O∶9血清型和其它菌株均不裂解,对O∶3的裂解率高达84.5%(71/84)。噬菌体phiYe-F10对假结核耶尔森菌和鼠疫耶尔森氏菌均不裂解。结论:phiYe-F10噬菌体是一株小肠结肠炎耶尔森菌的裂性噬菌体,在25℃时表现出一个典型的窄裂解谱,高度专一性裂解O∶3血清型小肠结肠炎耶尔森菌。phiYe-F10对典型致病性小肠结肠炎耶尔森菌的裂解能力明显高于非致病性的小肠结肠炎耶尔森菌  相似文献   

6.
【目的】为探究脂多糖对O1、O78血清型禽致病性大肠杆菌(Avian pathogenic Escherichia coli,APEC)致病作用的影响。【方法】选取负责脂质A生物合成相关基因lpx L和lpx M,利用λ噬菌体的Red同源重组系统分别构建APECE516(O1血清型)和APECE522(O78血清型)缺失株E516Δlpx L、E516Δlpx M、E516Δlpx LΔlpx M、E522Δlpx L、E522Δlpx M和E522Δlpx LΔlpx M,并通过体内外试验对其生物学特性及致病性进行研究。【结果】各菌株生长速度基本一致。E516Δlpx L、E516Δlpx LΔlpx M、E522Δlpx M和E522Δlpx LΔlpx M的抗血清补体杀菌能力和抗鸡巨噬细胞HD-11吞噬能力较野生株显著下降,而缺失株E516Δlpx M、E522Δlpx L与野生株相比无明显差异;半数致死剂量测定结果显示,除E516Δlpx M、E522Δlpx L外,各缺失株毒力降低1000倍左右;SPF鸡体内动态分布试验结果显示,各缺失株在鸡体内定殖能力较野生株显著下降,但回补株的毒力未能恢复至野生株水平。【结论】lpx L和lpx M基因与O1血清型APECE516株和O78血清型APECE522株的毒力有关,但是lpx L和lpx M基因对E516和E522菌株毒力的影响存在差异。  相似文献   

7.
选择性捕获禽病原性大肠杆菌体内转录序列   总被引:5,自引:1,他引:4  
采用选择性捕获转录序列(SCOTS)方法鉴定禽病原性大肠杆菌E037株(血清型O78)在感染SPF鸡过程中的转录表达基因。通过总RNA分离、cDNAs合成、PCR扩增和SCOTS对cDNAs选择和致病性特异转录序列的富集,致病性特异的cDNAs被分离鉴定,共获得31个转录序列(命名为aec),其中分别有2、1、4、14、2和8个aec序列与黏附素、LPS的合成、铁的摄取系统、质粒编码基因、噬菌体编码基因和一些其它功能基因相关;从气囊中分离到16个aec序列,心包膜中分离到15个aec序列;有3种与质粒编码基因相关序列在气囊和心包膜中都被分离到。结果显示APEC致病性特异序列包括黏附素、LPS的合成、铁的转运、质粒编码基因、噬菌体编码基因和一些其它功能基因等。通过SCOTS方法建立了一种体内表达致病性特异基因的方法和APEC在自然宿主感染模型中致病性相关基因的表达谱的筛选方法。  相似文献   

8.
高清清  夏乐  刘娟华  高崧  刘秀梵 《微生物学报》2016,56(10):1571-1582
【目的】探究荚膜对肠道外致病性大肠杆菌致病作用的影响。【方法】选取负责荚膜多糖转运的基因kpsE和kpsD,利用λRed重组系统构建APEC E058和UPEC U17荚膜缺失株E058ΔkpsED和U17ΔkpsED,并通过一系列的体内及体外试验对其生物学特性及致病性进行研究。【结果】双基因缺失株的生长速度较野生株没有明显差异,但缺失株抗血清补体杀菌能力和抗鸡巨噬细胞HD-11细胞吞噬能力显著下降。1日龄雏鸡LD50致病性试验结果显示,缺失株E058ΔkpsED和U17ΔkpsED对鸡失去致病力,而回复株毒力恢复至野生株水平;35日龄SPF鸡体内动态分布和竞争试验显示ΔkpsED缺失株在鸡体内定殖能力和竞争性生长能力显著下降,表明kpsED双基因的缺失能显著降低APEC E058和UPEC U17的致病力。【结论】荚膜与肠道外致病性大肠杆菌的致病性相关,是其重要的毒力因子。  相似文献   

9.
[目的]为了研究噬菌体整合酶基因在猪链球菌2型(Streptococcus suis type 2,SS2)中的分布情况.[方法]根据噬菌体整合酶基因设计引物,建立了PCR方法,并对扩增产物进行测序.[结果]结果显示,25株SS2致病菌株均扩增出目的片段,非毒力株T15、5株其它血清型猪链球菌及兰氏C群猪源链球菌未扩增出目的片段.经丝裂霉素C诱导后,SS2致病菌株出现完全的细胞溶解,而非毒力株T15未出现溶解.SS2致病株HA9801和ZY05719诱导均产生溶原性噬菌体,分别命名为SS2-HA和SS2-ZY,电镜观察,二者均头部呈正六边形,无尾部,其核酸类型为dsDNA,可鉴定为复层噬菌体科(Tectiviridae)的成员.噬菌体SS2-HA和SS2-ZY整合酶基因序列与已报道的SS2噬菌体整合酶基因序列高度同源,显示SS2噬菌体整合酶具有较高的特异性.[结论]从SS2致病株中检出溶原性噬菌体和噬菌体整合酶基因,且噬菌体整合酶基因与SS2溶菌酶释放蛋白(mrp)等7种毒力相关基因有相关性,表明SS2的溶原性噬菌体可能与其致病性有关.  相似文献   

10.
利用大肠杆菌K12MG1655株全基因组ORFs和痢疾志贺氏菌A1型Sd51197株特异性ORFs探针制备的芯片,研究了痢疾志贺氏菌13个血清型代表株的基因组组成.结果显示,该血清群成员的基因组中包含有2654个保守的源于大肠杆菌ORFs;共同缺失了219个涉及前噬菌体基因、分子伴侣、特异性O抗原合成等大肠杆菌原有的基因;并通过水平转移获得了一些特异性基因,如Ⅱ型分泌系统相关组分、铁转运相关因子等.根据基因组组成所作的进化树,发现A1,A2,A8和A10这四型菌与其他痢疾志贺氏菌亲源关系较远.研究所得结果为进一步深入探索痢疾志贺氏菌的生理过程、致病性和进化奠定了基础.  相似文献   

11.
Genomes of prokaryotes differ significantly in size and DNA composition. Escherichia coli is considered a model organism to analyze the processes involved in bacterial genome evolution, as the species comprises numerous pathogenic and commensal variants. Pathogenic and nonpathogenic E. coli strains differ in the presence and absence of additional DNA elements contributing to specific virulence traits and also in the presence and absence of additional genetic information. To analyze the genetic diversity of pathogenic and commensal E. coli isolates, a whole-genome approach was applied. Using DNA arrays, the presence of all translatable open reading frames (ORFs) of nonpathogenic E. coli K-12 strain MG1655 was investigated in 26 E. coli isolates, including various extraintestinal and intestinal pathogenic E. coli isolates, 3 pathogenicity island deletion mutants, and commensal and laboratory strains. Additionally, the presence of virulence-associated genes of E. coli was determined using a DNA "pathoarray" developed in our laboratory. The frequency and distributional pattern of genomic variations vary widely in different E. coli strains. Up to 10% of the E. coli K-12-specific ORFs were not detectable in the genomes of the different strains. DNA sequences described for extraintestinal or intestinal pathogenic E. coli are more frequently detectable in isolates of the same origin than in other pathotypes. Several genes coding for virulence or fitness factors are also present in commensal E. coli isolates. Based on these results, the conserved E. coli core genome is estimated to consist of at least 3,100 translatable ORFs. The absence of K-12-specific ORFs was detectable in all chromosomal regions. These data demonstrate the great genome heterogeneity and genetic diversity among E. coli strains and underline the fact that both the acquisition and deletion of DNA elements are important processes involved in the evolution of prokaryotes.  相似文献   

12.
Morris RT  Drouin G 《Genomics》2008,92(3):168-172
We previously showed that gene conversions were more frequent in the genomes of three Escherichia coli pathogenic strains than in the genome of the nonpathogenic K-12 E. coli strain. However, that study did not address whether the more frequent conversions observed in the genes of pathogenic strains occurred between the backbone genes common to these four strains or in the numerous horizontally transferred genes found only in pathogenic strains. Here, we show that ectopic gene conversions are equally frequent in the backbone genes of pathogenic and nonpathogenic strains, that most of these conversions are short, and that the nucleotide changes they generate are probably selectively neutral. Backbone genes are therefore under similar selective constraints in both pathogenic and nonpathogenic E. coli strains. The higher frequency of gene conversions we previously observed in pathogenic strains is therefore due to higher conversion frequencies between the numerous horizontally transferred genes found only in pathogenic strains.  相似文献   

13.
Nonpathogenic Escherichia coli strain Nissle 1917 (O6:K5:H1) is used as a probiotic agent in medicine, mainly for the treatment of various gastroenterological diseases. To gain insight on the genetic level into its properties of colonization and commensalism, this strain's genome structure has been analyzed by three approaches: (i) sequence context screening of tRNA genes as a potential indication of chromosomal integration of horizontally acquired DNA, (ii) sequence analysis of 280 kb of genomic islands (GEIs) coding for important fitness factors, and (iii) comparison of Nissle 1917 genome content with that of other E. coli strains by DNA-DNA hybridization. PCR-based screening of 324 nonpathogenic and pathogenic E. coli isolates of different origins revealed that some chromosomal regions are frequently detectable in nonpathogenic E. coli and also among extraintestinal and intestinal pathogenic strains. Many known fitness factor determinants of strain Nissle 1917 are localized on four GEIs which have been partially sequenced and analyzed. Comparison of these data with the available knowledge of the genome structure of E. coli K-12 strain MG1655 and of uropathogenic E. coli O6 strains CFT073 and 536 revealed structural similarities on the genomic level, especially between the E. coli O6 strains. The lack of defined virulence factors (i.e., alpha-hemolysin, P-fimbrial adhesins, and the semirough lipopolysaccharide phenotype) combined with the expression of fitness factors such as microcins, different iron uptake systems, adhesins, and proteases, which may support its survival and successful colonization of the human gut, most likely contributes to the probiotic character of E. coli strain Nissle 1917.  相似文献   

14.
To identify novel virulence-associated genes in uropathogenic Escherichia coli (UPEC) strains, a suppression subtractive hybridization strategy was applied to genomic DNA of four clinical UPEC isolates from patients suffering from cystitis or pyelonephritis. The genomic DNA of four isolates (tester strains) was subtracted from the DNA of two different driver strains, the well characterized UPEC strain CFT073 and the non-pathogenic E. coli K-12 strain MG1655. We determined the sequence of 172 tester strain-specific DNA fragments, 86 of which revealed only low or no homology to nucleotide sequences of public databases. We further determined the virulence association of the 86 novel DNA fragments using each DNA fragment as a probe in Southern hybridizations of a reference strain collection consisting of 60 extraintestinal pathogenic E. coli isolates, and 40 non-virulent E. coli strains from stool samples. From this, 19 novel DNA fragments were demonstrated to be significantly associated with virulent strains and thus may represent new virulence traits. Our results support the idea of a considerable genetic variability among UPEC strains and suggest that novel genomic determinants might contribute to virulence of UPEC.  相似文献   

15.
The extent and nature of DNA polymorphism in the mutS-rpoS region of the Escherichia coli genome were assessed in 21 strains of enteropathogenic E. coli (EPEC) and enterohemorrhagic E. coli (EHEC) and in 6 strains originally isolated from natural populations. The intervening region between mutS and rpoS was amplified by long-range PCR, and the resulting amplicons varied substantially in length (7.8 to 14.2 kb) among pathogenic groups. Restriction maps based on five enzymes and sequence analysis showed that strains of the EPEC 1, EPEC 2, and EHEC 2 groups have a long mutS-rpoS region composed of a approximately 6.0-kb DNA segment found in strain K-12 and a novel DNA segment ( approximately 2.9 kb) located at the 3' end of rpoS. The novel segment contains three genes (yclC, pad1, and slyA) that occur in E. coli O157:H7 and related strains but are not found in K-12 or members of the ECOR group A. Phylogenetic analysis of the common sequences indicates that the long intergenic region is ancestral and at least two separate deletion events gave rise to the shorter regions characteristic of the E. coli O157:H7 and K-12 lineages.  相似文献   

16.
We have investigated the use of a top-down liquid chromatography/mass spectrometric (LC/MS) approach for the identification of specific protein biomarkers useful for differentiation of closely related strains of bacteria. The sequence information derived from the protein biomarker was then used to develop specific polymerase chain reaction primers useful for rapid identification of the strains. Shiga-toxigenic Escherichia coli (STEC) strains were used for this evaluation. The expressed protein profiles of two closely related serotype 0157:H7 strains, the predominant strain implicated in illness worldwide, and the nonpathogenic E. coli K-12 strain were compared with each other in an attempt to identify new protein markers that could be used to distinguish the 0157:H7 strains from each other and from the E. coli K-12 strain. Sequencing of a single protein unique to one of the 0157:H7 strains identified it as a cytolethal distending toxin, a potential virulence marker. The protein sequence information enabled the derivation of genetic sequence information for this toxin, thus allowing the development of specific polymerase chain reaction primers for its detection. In addition, the top-down LC/MS technique was able to identify other unique biomarkers and differentiate nearly identical 0157:H7 strains, which exhibited identical phenotypic, serologic, and genetic traits. The results of these studies demonstrate that this approach can be expanded to other serotypes of interest and provide a rational approach to identifying new molecular targets for detection.  相似文献   

17.
Escherichia coli, including the closely related genus Shigella, is a highly diverse species in terms of genome structure. Comparative genomic hybridization (CGH) microarray analysis was used to compare the gene content of E. coli K-12 with the gene contents of pathogenic strains. Missing genes in a pathogen were detected on a microarray slide spotted with 4,071 open reading frames (ORFs) of W3110, a commonly used wild-type K-12 strain. For 22 strains subjected to the CGH microarray analyses 1,424 ORFs were found to be absent in at least one strain. The common backbone of the E. coli genome was estimated to contain about 2,800 ORFs. The mosaic distribution of absent regions indicated that the genomes of pathogenic strains were highly diversified because of insertions and deletions. Prophages, cell envelope genes, transporter genes, and regulator genes in the K-12 genome often were not present in pathogens. The gene contents of the strains tested were recognized as a matrix for a neighbor-joining analysis. The phylogenic tree obtained was consistent with the results of previous studies. However, unique relationships between enteroinvasive strains and Shigella, uropathogenic, and some enteropathogenic strains were suggested by the results of this study. The data demonstrated that the CGH microarray technique is useful not only for genomic comparisons but also for phylogenic analysis of E. coli at the strain level.  相似文献   

18.
19.
To determine virulence-related genes in uropathogenic Escherichia coli (UPEC) showing invasiveness to T-24 bladder cancer cells, genomic subtractive hybridization was performed between a highly invasive and a less invasive strain. Forty-nine DNA fragments were isolated from the invasive strain. One of them showed homology with Salmonella invA gene. By chromosomal walking of the strain, a type III secretion system that has been described in E. coli O157:H7 was identified on the genome of the invasive strains. Three strains out of 100 UPEC isolates had a type III secretion system inserted at 64 min of the chromosome, corresponding to E. coli K-12 MG1655. This finding suggested that the type III secretion system could play a part in uropathogenicity of UPEC.  相似文献   

20.
Sequences encoding the production of a cytotoxin (VT) active on Vero cells were cloned in Escherichia coli K12 from a VT-determining phage that originated in E. coli strain H19 of serotype O26.H11. Subcloning resulted in the identification of a 2.5 kb fragment that still coded for VT production. Mutagenesis with transposon Tn1000 was used to map VT sequences and a 0.75 kb probe was developed. In colony hybridization tests with strains isolated from patients with haemolytic uraemic syndrome or diarrhoea, this probe derived from the H19 VT genes detected only some of the VT+ strains belonging to serogroup 0157. A VT+ strain, E32511, serotype 0157.H-, which was negative in colony hybridization was the source of another VT-determining phage from which VT sequences were cloned. Southern hybridization of the VT genes from E32511 with the H19 probe was negative under stringent conditions but there was weak homology under conditions of low stringency. These results indicate that there are differences in the VT genes of pathogenic E. coli.  相似文献   

设为首页 | 免责声明 | 关于勤云 | 加入收藏

Copyright©北京勤云科技发展有限公司  京ICP备09084417号