Roles Of Iron Acquisition-related Regulation Genes Rsta And Fur In Virulence Of APEC And UPEC

Escherichia coli commonly colonize the digestive tract or other mucosal surfaces of newly born mammalian and homothermal animals for a lifetime and become a normal flora in tract. While most of these strains are nonpathogenic certain few pathogenic strains can become virulent in some circumstances. It can mainly cause intestinal or extraintestinal infection. Avian pathogenic E.coli (APEC) and uropathogenic E.coli (UPEC) are both found in extraintestinal infections, and then regarded as extraintestinal pathogenic E.coli (ExPEC), APEC can cause systemic infection in poultry while UPEC can cause infection in human urethra tact. Some studies have shown that these two pathotypes own some similar virulence determinants and iron acquisition system is included. Also there is some hypothesis saying that there might be some close Homologous relationship between APEC and UPEC which we need more studies to clarify.Two-component system is often used by bacteria to adapt response in changing environment for survival. RstA/rstB is constituted of protein membrane senor protein RstB and responsor regulator, RstA. The protein RstA which coded by gene rstA is responsible to an unknown effect on regulation of iron acquisition system. And fur is a kind of protein which plays a pivotal role in iron acquisition system. It is confirmed that Fur is a negative regulation protein that could prevent bacteria from iron toxicity by taking iron overload. Thus, the Fur is an important role in keeping equilibrium concentration of iron. After all, RstA and Fur are both significant regulators in the pathway of iron acquisition.In this study, the Lamda red-combination system is used to construct mutants such as E058△fur, E058ArstA, E058AfurArstA, U17△fur, U17△rstA and U17△fur△rstA. And the plasmid pM18was electroporated into mutants above to generate the revertants of them. And then, the research of biological characteristics and evaluation of pathogenic on experimental animals was carried on to compare the differences of the virulence between APEC and UPEC and also could find the similar of the virulent determinants. The data of this study might offer some important information to further research of the pathogenesis and specific prevention measures for APEC and UPEC.The experimental results show that there was no obvious difference between mutants and wild type strains in vitro, such as the growth curves which realized that there were very few changes between mutants and wild type strains while cultured in Iron deficiency medium. However, the growth rate of U17△rstA and E058ArstA is a slight decline in LB broth while the one of U17△fur and E058△fur is much lower than the wild type strains. What’s more, there were also no significant differences between the mutants and the wild-type strains in resistance to serum, cellular ingestion and intracellular survival in HD-11. Results indicated that APEC and UPEC utilize similar iron acquisition mechanisms in chickens. And, the lethal test in1-day old SPF chicken and the body dynamic distribution in test35days of age chicken consistently show that the deletion of rstA leads to attenuation of virulence and decreased colonization than wild types. In consequence, the gene rstA of two-component system RstA/RstB might be more strong correlation than fur, and there might be some relationship between these two genes. The slight changes on other phenotypic tests between the mutants and the wild-type strains suggest the existence of other virulence compensatory mechanisms.