猪源大肠杆菌F18菌毛的体外表达和抗原特性

Antigenic determinants analysis and detection of virulence factors in F18 fimbriae Escherichia coli strains isolated from pigs

摘要:

摘要:【目的】揭示从仔猪腹泻和/或水肿病猪体内分离到的fedA + 大肠杆菌所携带的毒力因子、F18菌毛在体外表达及其抗原变异情况。【方法】利用凝集试验测定O 血清型，PCR方法检测毒力基因，单克隆抗体分析F18菌毛抗原特性。【结果】在75个fedA + 分离株中，有62株测定出其O血清型，覆盖8种血清型，以O107和O139为主(74.2%) ;estI、estII、elt、stx-2e、astA、orfA、irp2、fyuA、ler和eaeA基因在这75个菌株中的检出率分别为64.0%、46.7%、28.0%、62.7%、26.7%、9.3%、9.3%、9.3%、1.3%和1.3%，其中仅拥有stx-2e基因的菌株有19株，同时拥有estI/estII/stx-2e基因的菌株有20株。单抗鉴定结果显示，在33株体外表达F18菌毛的菌株中，21株(63.6%)被鉴定为F18ac变体，2株(6.1%) 被鉴定为F18ab变体，其余10株(30.3%)仅跟F18“a”因子单抗反应，而不跟F18“b”、“c”因子单抗反应。间接ELISA显示，11株单抗至 少识别F18菌毛的6个表位，其中“a”因子至少有3个表位，“b”因子至少有2个表位，“c”因子至少有1个表位。【结论】在猪源菌株中，F18ab菌毛在体外表达率较低;F18ac菌毛在体外表达率较高，主要与肠毒素和O107血清型相关，同时我国存在F18菌毛的抗原变异。

Abstract:

Abstract:［Objective］We determined the present distribution of serogroups and virulence factors among F18 Escherichia coli isolates from pigs with diarrhea and/or edema disease during 1998 to 2006．Epitope of F18 fimbriae was also analyzed．［Methods］A total of 75 E.coli isolates harbored fedA gene coding the major subunit of F18 fimbriae were identified by biochemical procedures and serological techniques． PCR was used to detect the virulence-related genes．An indirect ELISA was also used to analyze the antigen patterns of 33 F18 bearing E．coli strains． ［Results］Among these 75 isolates，62 were determined to be placed in total 8 serogroups，and O107 and O139 were main serogroups (61.3%) of fedA-positive isolates．The percentage of the detection of the estI，estII，elt，stx-2e，astA，orfA，irp2，fyuA，ler and eaeA among 75 strains was 64.0%，46.7%，28.0%，62.7%，26.7%，9.3%，9.3%，9.3%，1.3% and 1.3%，respectively．of the 75 strains 19 were positive for stx-2e gene only，and 20 for estI/estII/stx-2e．The 11 MAbs against F18“a”，“b”and“c”had been used in indirect ELISA to detect F18 pili antigen，44.0% (33 /75) of these isolates were F18 + when cultured in TSB cultures．Among 33 isolates expressed F18 fimbriae，21 isolates were identified as the “ac”variant，2 were identified as the“ab”variant and 10 reacted only with F18“a”specific MAbs while not with F18“b”，“c”specific MAbs． The results also revealed that the 11 MAbs recognized at least 6 epitopes including at least three common typespecific antigenic determinants“a”and two“b”antigen determinants and one“c”antigen determinant.［Conclusion］The antigenic variants of F18 fimbriae are biologically distinct．F18ab fimbriae are expressed poorly in vitro，while F18ac are more efficiently expressed in vitro and most often are linked with enterotoxin (STa and/or STb) production and serogroups O107．Some F18 fimbrial antigens experienced some variations comparing with those of F18ab and F18ac strains.