禽致病性大肠杆菌gspL基因缺失株构建及生物学特性

Construction and characterization of a gspL mutant of avian pathogenic Escherichia coli

摘要:

摘要:【目的】研究gspL基因缺失对禽致病性大肠杆菌(Avian pathogenic Escherichia coli，APEC)生物学特性的影响。【方法】利用Red重组方法构建禽致病性大肠杆菌DE17株的gspL缺失株;分析野生株与缺失株的生长特性、黏附和入侵DF1细胞的差异;采用荧光定量PCR的方法比较野生株和缺失株毒力基因转录水平的变化;比较野生株与缺失株的半数致死量(LD50)差异。【结果】gspL缺失不影响DE17的生长特性，但其黏附和入侵DF1细胞能力显著下调。荧光定量PCR检测结果表明，缺失株毒力基因luxS，pfs，fyuA和iss转录水平明显上调，tsh的转录水平明显下调，而vat，ibeA，stx2f和ompA的转录水平无显著变化;LD50检测结果表明，缺失株比野生株毒力增强了12倍。【结论】gspL基因的缺失不影响禽致病性大肠杆菌的生长特性，但能减弱其黏附和入侵能力，且可以正调控禽致病性大肠杆菌部分毒力基因的转录水平，推测gspL基因可能与APEC对宿主的致病性有关。

Abstract:

Abstract:［Objective］To study the role of gspL gene in avian pathogenic Escherichia coli．［Methods］The gspL mutant of Avian pathogenic Escherichia coli (APEC) was constructed by homologous recombination assay．The growth characteristics，the ability of adhesion and invasion to DF1 cells，the virulence genes transcription level and median lethal dose (LD50) were analyzed between the gspL mutant strain and the wild strain．［Results］Compared with the wild strain，the mutant strain had no significant difference in the growth status．However，its ability of adhesion and invasion was significantly lower．The transcription of genes pfs，fyuA，iss and vat increased obviously，the tsh decreased and the transcription level of luxS，ibeA，stx2f and ompA had no significant change． LD50 showed that the gspL mutant strain had 12-fold increase in virulence．［Conclusion］The deletion of gspL gene could abate the ability of adhesion and invasion，regulate and control some virulence gene transcription level，enhance the virulence of APEC． The results show that the gspL gene play roles in pathogenicity of APEC．