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1.
致断奶仔猪腹泻、水肿病大肠杆菌F18菌毛a因子单克隆抗体的研制及其初步应用 总被引：2，自引：0，他引：2
以表达 F18ac菌毛参考菌株 2 134(O15 7∶ H19)为研究对象 ,摸索出了 F18ac菌毛表达的最佳条件 ,采用热洗脱法获得了较纯的 F18ac菌毛 ,并以之免疫小鼠 ,利用淋巴细胞杂交瘤技术 ,用直接玻板凝集法筛选出 1株杂交瘤细胞株 19B4 ,能稳定分泌针对 F18ac菌毛和 F18ab菌毛共同抗原表位 a因子的单克隆抗体。细胞培养上清对 F18ac菌毛参考菌株 2 134和 F18ab菌毛参考菌株 10 7/ 86的直接凝集价均可达 1∶ 8,腹水单抗直接凝集效价可达 1∶ 10 2 4 ,而与猪源产 F4、F5、F6和 F4 1粘附素大肠杆菌菌株、鸡源产 型菌毛大肠杆菌菌株及沙门氏菌不发生凝集反应。免疫印迹试验结果显示 ,腹水单抗可同时特异识别 F18ac菌毛 170 0 0和 F18ab菌毛 15 0 0 0左右的主要亚单位多肽。应用所研制的单抗对 2 15株断奶仔猪腹泻大肠杆菌分离株进行了检测 ,结果上述分离株 TSB培养物的 F18菌毛的检出率为13.4 % ,TSA培养物的检出率为 8.4 %。F18菌毛阳性分离株的 O血清型除了常见血清型 O139、O14 1外 ,还有非常见血清型 O10 7、O131和 O9等。TSB培养物的 F18菌毛检出率明显高于 TSA培养物的检出率 ,表明 F18菌毛在 TSB培养基中比在 TSA培养基上更容易表达 ,TSB培养基是适合 F18菌毛检测、流行病学调查的培养基 相似文献
2.
从南阳地区各养鸡场的发病鸡中分离出大肠杆菌,经常规方法培养、纯化、镜检及生理生化鉴定。分离出29株鸡致病性大肠杆菌。有14株分离株被鉴定出血清型,其中O1为4株,O2为3株,O78为7株,O1,O2和O78血清型占分离株总数的48．3％。未定型15株,用29株分离菌株分别接种供试雏鸡后,发病症状及剖检病变均符合本病特征,其中高致病性菌株有13株,中度致病菌株有9株,低致病性菌株有7株,分别占试验菌株总数的44．8％、31％和24．2％。采用22种药敏纸片对29株分离菌进行药敏试验,结果表明,分离出的大肠杆菌耐药性相当普遍,仅对头孢噻肟、丁胺卡那霉素、壮观霉素及新霉素等药物敏感性较高。 相似文献
3.
对鸡大肠杆菌进行分离鉴定，分离出的56株大肠杆菌中，共鉴定出47株菌株的血清型，分属O1、O2、O5、O35、O78、O1116种不同O抗原血清型，占分离菌株的83．9％。其中O1、O2、O78、O111最常见，占已定型菌株83％。同时，进行了药敏试验，为药物防治提供依据。 相似文献
4.
5.
F107(F18ab)菌毛单抗的制备、鉴定与初步应用 总被引：9，自引：2，他引：7
将大肠杆菌107／86和水肿病菌株在TSB中培养，鉴定其表达F107菌毛，前者作为免疫原免疫BABL／C小鼠，后者作为检测抗原包装酶标板。细胞融合后经ELISA和玻板凝集检测，得到一株阳性杂交瘤细胞株，命名为F7－1。该细胞株制备的腹水ELISA效价为：2^16，试管凝集价为：1：5120，在免疫印迹中可与提取的F107菌毛特异性条带发生反应。利用此单抗检测表明，91％的水肿病菌株表达F107菌毛。 相似文献
6.
鸡大肠杆菌多价1型菌毛亚单位疫苗在鸡体内的免疫原性研究 总被引：4，自引：0，他引：4
用1日龄雏鸡免疫评价了鸡大肠杆菌多价1型菌毛油乳剂苗在鸡体内的免疫原性。用含1型菌毛的3个不同血清型（O1、O78及O88）菌株,大容量培养后提取菌毛制备多价菌毛油乳剂苗。接种多价菌毛油乳剂苗的鸡用同源菌株攻毒后出现10％～11．2％的死亡率,阳性对照组鸡攻毒后出现70％～80％的死亡率;用异源菌株（O36）攻毒后,免疫鸡出现33．3％的死亡率,而未免疫阳性对照组鸡死亡率达70％。免疫鸡在气囊,心包及肝脏的病变非常轻微,且攻毒后非免疫鸡能更有效地清除侵入气囊的大肠杆菌。 相似文献
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8.
禽病原性大肠杆菌Ⅰ型菌毛单克隆抗体的研制及其对分离株的检测 总被引：2，自引：0，他引：2
从禽病原性大肠杆菌分离株 TK3 ( O1 )提取 型菌毛免疫 BALB/c小鼠 ,取其脾细胞与小鼠骨髓瘤细胞融合 ,获得 4株能稳定分泌针对 型菌毛的单克隆抗体杂交瘤细胞株 ,分别命名为 a B6 、b G5 、c F3和 c G3。单抗阻断甘露糖敏感血凝试验、免疫胶体金和 Western blot试验证明 ,单抗是 型菌毛特异的。这些单抗培养上清及腹水的 ELISA效价为分别为 1 0 - 2～ 1 0 - 3和 1 0 - 5～ 1 0 - 6 ;腹水的平板凝集价为 1∶ 1 2 8～ 1∶ 2 56。上述4株单抗对 77株带 型菌毛的禽病原性大肠杆菌优势血清型分离株进行了检测 :O1 8分离株阳性率为72 %,O78分离株阳性率为 1 7%,O2分离株阳性率为 3 3 %,O88分离株阳性率 89%,O1 1分离株阳性率为60 %,O2 6分离株阳性率为 3 3 %。表明研制的 型菌毛单抗能检出大多数 O1 8、O88、O1 1分离株的 型菌毛 ,而对 O78、O2、O2 6分离株相应菌毛的检出率较低 相似文献
9.
猪水肿病大肠杆菌分离、鉴定及药敏试验 总被引：6，自引：3，他引：3
本实验从疑似猪水肿病的病例分离到5株大肠杆菌,O抗原鉴定结果表明所有菌株均为O139血清型;应用F18ab菌毛单克隆抗体对这5株大肠杆菌能否表达F18ab菌毛进行了鉴定,结果表明其中2个菌株能表达F18ab菌毛;利用聚合酶链式反应(PCR)对志贺氏菌样毒素Ⅱ型变异体(SLT-Ⅱe)操纵子基因保守区进行了扩增,结果发现在能表达F18ab菌毛2个菌株中可扩增一段特异性序列。以上数据表明这2株大肠杆菌为致仔猪水肿病大肠杆菌。药敏试验表明这两株菌株均对氟哌酸、妥布霉素、庆大霉素、利福平等抗生素高度敏感。 相似文献
10.
致猪水肿病大肠杆菌F18ab菌毛单克隆抗体的研制及其初步应用 总被引：4，自引：0，他引：4
将致猪水肿病大肠杆菌 F18ab菌毛提纯 ,以其免疫 BAL B/ c小鼠 ,利用淋巴细胞杂交瘤技术 ,经 3次融合 ,共筛选出 4株能稳定分泌针对 F18ab菌毛的特异单克隆抗体的杂交瘤细胞株 4 G8、4 H11、4 A2和 3A12 ,腹水单抗的直接凝集价最高可达 1∶ 32 0 0。免疫印迹试验表明 ,4株腹水单抗均可特异识别 F18ab菌毛 Mr 15 0 0 0左右的主要亚单位多肽。应用所研制的单抗对 11株猪水肿病大肠杆菌分离株 F18ab菌毛表达情况进行了检测 ,结果显示 ,上述分离株中F18ab菌毛的出现率为 72 .7% (8/ 11) ,检出的 F18ab菌毛阳性分离株的 O血清型均为 O1 39。 相似文献
11.
Prevalence of serogroups and virulence factors of Escherichia coli strains isolated from pigs with postweaning diarrhoea in eastern China 总被引：5，自引：0，他引：5
This study was undertaken to determine the present distribution of serogroups, hemolytic activity and virulence factors among Escherichia coli strains isolated from pigs with postweaning diarrhoea from eight provinces in eastern China. Two hundred and fifteen E. coli isolates were serogrouped with O-antisera, investigated for hemolytic activity, assessed for F4, F5, F6, F18 and F41 fimbrial antigens by monoclonal antibodies and detected for genes of enterotoxins and shiga-toxin-two-variant (Stx2e) by a multiplex polymerase chain reaction (PCR). Among these E. coli isolates, 140 were determined to be placed in serogroups, 52 were unable to be serogrouped and the rest 23 auto-agglutinated. These isolates distributed in 45 serogroups and 64.3% (90/140) belonged to 12 O serogroups: O8, O9, O11, O20, O32, O91, O93, O101, O107, O115, O116 and O131. Hemolytic activity was detected in 11.6% (25/215) of all isolates. Several uncommon O serogroups were discovered in this study. Agglutination tests showed that 50.2% (108/215) of these isolates were positive for one or more of the five fimbrial antigens. Seventy-two E. coli strains expressed single fimbria and 36 strains expressed two or more fimbriae. Among these 215 E. coli isolates, strains expressing F18, F4, F6, F6 + F18 or F5 + F41 occurred more frequently. PCR analysis showed that 60.5% (130/215) of the isolates only harboured the gene of estI (STI) while 6.0% (13/215) strains possessed the genes of stx2e, estI and estII and 5.6% (12/215) of strains had the genes of estI/estII. Of all these isolates, 107 (49.8%) were negative for the fimbrial antigens examined. The fimbria-negative isolates usually possessed genetic determinant of estI (78, 72.9%). 相似文献
12.
L H Arp 《American journal of veterinary research》1985,46(12):2644-2647
Young turkeys (n = 20) were inoculated IV with fimbriated, virulent Escherichia coli ECl (O78:K80: H9:F1). Blood samples were collected for bacterial quantitation at postinoculation minutes (PIM) 10, 20, 30, 40, 50, and 60. Immediately after the PIM 30 sampling, the turkeys were allotted into 4 groups (5 turkeys/group) and were injected IV with 1 of the following antisera: group 1, antibodies to F1 fimbriae (AF); group 2, antibodies to E coli O78 antigen (AO); group 3, antibodies to live, fimbriated (F1+) homologous E coli (ALEC); or group 4, normal turkey serum (NTS) collected from a healthy turkey. Compared with NTS, ALEC and AO caused a significant reduction in blood-borne E coli, whereas AF did not reduce bacterial numbers. In addition, 2 groups of 10 turkeys were inoculated IV with live, F1+ or nonfimbriated (F1-) E coli ECl. Numbers of viable bacteria were determined in blood samples and liver specimens collected 2 minutes after inoculation. Compared with F1- bacteria, significantly more F1+ bacteria were found in liver specimens and significantly fewer F1+ bacteria were found in blood samples. Results indicated that antibodies to F1 fimbriae do not enhance clearance of F1+ E coli from the bloodstream of turkeys probably because F1+ bacteria are selectively cleared by the liver, even without antibody. 相似文献
13.
利用F18菌毛a因子单克降抗体以及已建立的鉴定F18菌毛及其亚型的双重PCR法,对来自断奶仔猪水肿病和／或腹泻病例的60株VTEC、24株VTEC／ETEC以及24株ETEC的进行了F18菌毛检测,以了解F18ab^＋和F18ac^＋大肠杆菌在江苏省断奶仔猪群的分子流行病学。结果表明：通过F18菌毛a因子单克隆抗体,可检测出52株大肠杆菌为F18^＋,检出率为48．15％;而通过双重PCIL方法,共检测出63株大肠杆菌为F18^＋,检出率为58．33％,其中53株（49．07％）为F18ab^＋10株（92．6％）为F18ac^＋。另外还发现：在VTEC、VTEC／ETEC以及ETEC的菌株之间,这2种F18菌毛亚型的分子流行病学是不同的。在VTEC中,F18ab^＋,菌株37株（61．67％）,未发现F18ac^＋菌株;在VTEC／ETEC中,F18ab^＋菌株15株（62．50%）,F18ac^＋菌株8株（33．33％）;而在ETEC中F18ab^＋菌株只有1株（4．17％）,F18ac^＋菌株只有2株（8.33％）。以上数据表明：④PCR法检测F18菌毛优于单抗法;②F18菌毛是VTEC／ETEC、VTEC的重要致病因子,而在ETEC中则明显低于VTEC／ETEC和VTEC;⑧F18ab^＋菌株一般为SLT-IIe^＋,而F8ac^＋菌株一般为STI^＋。 相似文献
14.
The P fimbriae F11 and F165 that have been demonstrated on Escherichia coli septicaemic strains in poultry and calves, respectively, possess a nearly identical major subunit that demonstrates a serological cross-reaction. A polyclonal antibody-based sandwich ELISA (sELISA) that was specific for both F11 and F165 fimbriated strains was compared with a PCR method to detect F11/F165 fimbriated strains, in a collection of E. coli strains isolated from diseased animals. Of 298 isolates tested, 36 were positive by PCR of which only 14 were sELISA positive. There were no sELISA positive but PCR negative results. The 36 PCR positive isolates comprised 11 avian strains of which 10 were sELISA positive, 20 bovine strains of which 4 were sELISA positive and 3 ovine strains, 1 porcine strain and 1 equine strain all of which were sELISA negative. The F11/F165 incidence of 10.7% in 103 poultry and 18.3% in 109 bovine isolates demonstrates a moderate level of these factors in E. coli septicaemic cases in Northern Ireland. 相似文献
15.
A total of 720 Escherichia coli strains isolated from diarrheic piglets on 756 swine farms were screened for the presence of the enteroaggregative E. coli heat-stable enterotoxin 1 (EAST1) gene by polymerase chain reaction (PCR). Escherichia coli strains that carried EAST1 genes were also tested by PCR for the presence of 4 fimbriae (F4, F5, F6, F41), 2 heat-stable enterotoxins (STa and STb), and 1 heat-labile enterotoxin (LT) gene. One hundred sixty-four (22.7%) of the 720 E. coli isolates carried genes for EAST1. Of these 164 isolates, 62 (37.8%) carried EAST1 genes only, 11 (6.7%) carried genes for at least 1 of the fimbrial adhesins, 51 (31.1%) carried genes for at least 1 of the enterotoxins, and 40 (23.8%) carried genes for at least 1 of the fimbrial adhesins and enterotoxins. Forty-six percent of strains that carried EAST1 genes carried STa genes, and 16% of strains that carried EAST1 genes carried F4. The isolation rate of enterotoxigenic E. coli strains carrying genes for EAST1 gene was 63%. The 6 major genotypes observed in this study (in decreasing order) were EAST1+, EAST1+STa+, EAST1+STa+STb+, EAST1+STa+F5+, EAST1+STa+F4+, and EAST1+STb+F4+. EAST1 is widely prevalent among diarrheagenic strains of E. coli and may represent an important virulence determinant in the pathogenesis of enteric colibacillosis of preweaned pigs. 相似文献
16.
Polymerase chain reaction for 4 fimbriae (F4, F5, F6, F41), 2 heat-stable enterotoxins (STa, STb), and 1 heat-labile enterotoxin (LT) were performed on 400 Escherichia coli isolates to determine their genotype prevalence among enterotoxigenic E. coli isolates from preweaned pigs with diarrhea in the Republic of Korea. A total of 200 of the 400 E. coli isolates were also selected for characterization of the O serogroup. Of these 200 isolates, serogroup could be determined in 139 (69.5%) but not in 61 isolates (30.5%). Isolates of serogroup O101 were the most common, followed in descending order by 08, 020, 0162, 0141, and 0149. Ninety-seven (24.3%) of the 400 E. coli isolates carried genes for at least 1 of the entertoxins or fimbrial adhesins. Of these 97 isolates, 27 carried genes for at least 1 of the fimbrial adhesins and entertoxins. Sixty-six percent of the isolates that carried fimbrial adhesin genes carried genes for at least 1 of the enterotoxins, and 71% of the isolates that carried enterotoxin genes carried genes for at least 1 of the fimbrial adhesins. Genes for the F6 fimbriae were detected in 6% of the E. coli isolates, and F4+, F41+, and F5+ genes were detected in 4.3%, 3.3%, and 2% of the isolates, respectively. Genes for STa, STb, and LT were detected in 10%, 8.5%, and 4.3% of the isolates, respectively. The 6 major genotypes observed in this study (in decreasing order) were F6+, STb+, F41+, STa+STb+, F6+STa+, and STa+. 相似文献
17.
Seventy-six Escherichia coli serotypes possessing the ipaH gene typical of enteroinvasive E. coli (EIEC) strains were characterized. Biochemical identification of our strains shows positive reactions for lactose fermentation (100% of strains), lysine decarboxylase (98.7% of strains) and motility (67.1% of strains), properties that do not correspond with those described to the EIEC group. The serotypes agree with an initial classification. In this, some common O antigens identified among ipaH+ strains were O2 (n=20), OR (n=11) and non-determined O? (n=10). The O2:NM serotype was the most common. Sixty-six percent (n=50) of the ipaH+ E. coli strains were colicin producers, of them, 26 (34%) produced Col V and other colicins, 13 (17%) produced colicins other than Col V, and 11 (14.5%) produced Col V only. Trimethoprim/Sulfa (72%), ampicillin (64.5%), enrofloxacin (55.3%), and ciprofloxacin (47.4%) were the major antimicrobial resistance frequencies observed. Twenty-five different multiresistance patterns were observed, where sixty-six strains (86.8%) were included. A MIC test showed that most of the strains were sensitive to low gentamicin and kanamycin concentrations, whereas most of the strains were resistant to tetracycline. An invasiveness assay showed that the predominant alterations caused to HEp-2 cells were changes in shape and staining, and in most of the specimens, a partial monolayer detachment was also seen. Fifteen strains invaded more than 30% of the monolayer cells, causing the formation of intercellular bridges or filipoidal-like protrusions. The results suggest the existence of specific clone complexes derived from EIEC strains adapted to the avian host. To our knowledge, this is the first study that demonstrates the presence of extraintestinal invasive E. coli (ExIEC) strains. 相似文献
18.
Inhibition of adhesion of F18+ Escherichia coli to piglet intestinal villous enterocytes by monoclonal antibody against blood group H-2 antigen 总被引：3，自引：0，他引：3
Enterotoxigenic and verotoxigenic F18+ Escherichia coli colonising the pig small intestine, adhere to receptors on intestinal villous enterocytes by F18 fimbriae. The aim of the present study was to define the F18R nature. The knowledge on the nature of this receptor could be important for the development of receptor-based treatments against F18+ E. coli-induced disease. The adhesion of F18+ E. coli to pig intestinal villous enterocytes was analysed in an in vitro assay. The adhesion of F18+ E. coli but not of F4ac+ E. coli was strongly inhibited by monoclonal antibodies (mAb) with blood group H-2 specificity. Conversely, blood group H-1 specific mAb could not inhibit the adhesion of F18+ E. coli nor F4ac+ E. coli. Moreover, the blood group H-2 trisaccharide strongly inhibited the adhesion of F18+ E. coli, but only partially the adhesion of F4ac+ E. coli. These data demonstrate that the F18 receptor contains the blood group antigen H-2 (-fuc-(1-2)-β-Gal-(1-4)-GlcNAc) as major carbohydrate. 相似文献
19.
Three hundred and twenty-four strains of Escherichia coli isolated from weaned pigs with diarrhoea or oedema disease in Eastern China were screened by multiplex PCR for the presence of the gene encoding adhesin involved in diffuse adhesion I (AIDA-I). Two AIDA-I positive strains were subjected to analysis of the nucleotide sequence of the complete orfA and orfB of the AIDA gene. The AIDA-I positive E. coli isolates were also assessed for five fimbriae (F4, F5, F6, F18 and F41) by monoclonal antibodies and for toxin genes (STa, STb, LT, EAST1, Stx2e) by PCR. Twenty-one (6.5%) of the isolates possessed AIDA-I genes. Of these isolates, two carried AIDA-I genes as the only demonstrated virulence factors, and the remaining isolates carried other virulence factor genes. Comparing the AIDA-I sequence from porcine and human sources, a high homology of orfA both in porcine E. coli and human E. coli was observed. However, each orfB of the two porcine E. coli isolates was 3864 nucleotides long compared with 3861 for the E. coli 2787 orfB, and showed 96.5% homology to E. coli 2787. The data indicated (1) that AIDA-I may be an occasional virulence factor in post-weaning diarrhoea and oedema disease in pigs, (2) that it has the potential to transfer between porcine and human E. coli, and (3) that there is a genetic diversity in orfB between human and porcine E. coli. 相似文献
20.
对江苏省海安县2003年5月～2004年5月各乡镇猪水肿病的发病和死亡情况进行了流行病学调查，发现猪水肿病在海安地区呈零星散发性发生，发病率为0.31％～1．84％，死亡率为0．07％—0．56％，病死率为16．0％～35．0％。水肿病主要侵害仔猪，育成猪也偶有发生，一般以气温比较高的季节多发。同时从疑似猪水肿病的病料中分离到12株大肠杆菌，经O血清型鉴定，5株为0139，2株为O45，O55、O93，O9、O101、O119各1株。经多重PCR检测毒素基因(STa，STb、LT、SLT-2e)和大肠杆菌粘附素单抗(F4、F5、F6、F41、F18)检测茵毛，共6个分离株带有SLT-2e基因，其中O139分离株5株，055分离株1株，其余6株均带STb基因。12个分离株中，单独表达F18菌毛的4株(O1392株，O45、O119各1株)，F5 F411株(O93)，F41 F181株(O139)，F5 F41 F182株(均为O139)。经药敏试验，多数分离株对壮观霉素和新霉素高度敏感。 相似文献