Abstract
Escherichia coli is one of the major pathogens causing mastitis in dairy cattle. Yet, the factors which mediate the ability for E. coli to develop in the bovine mammary gland remain poorly elucidated. In a mouse model, infections induced by the reference mastitis E. coli P4 showed a strong colonisation of the mammary gland, while this strain had a low stimulating power on cells of the PS bovine mammary epithelial cell line. In order to understand if such a reduced response contributes to the severity of infection, a library of random mutants of P4 strain was screened to identify mutants inducing stronger response of PS cells. Among hyper-stimulating mutants, six were altered in genes involved in biosynthesis of lipopolysaccharide (LPS) and had lost their O-polysaccharide region, suggesting that the presence of O-antigen impairs the response of PS cells to LPS. Using purified smooth (S) and rough (R) fractions of LPS, we showed that the R-LPS fraction induced a stronger response from PS cells than the smooth LPS fraction. Biological activity of the S-LPS fraction could be restored by the addition of recombinant bovine CD14 (rbCD14), indicating a crucial role of CD14 in the recognition of S-LPS by Mammary Epithelial Cells (MEC). When S-LPS and R-LPS were injected in udder quarters of healthy lactating cows, an inflammation developed in all infused quarters, but the S-LPS induced a more intense pro-inflammatory response, possibly in relation to sizeable concentrations of CD14 in milk. Altogether, our results demonstrate that the O-antigen modulates the pro-inflammatory response of MEC to LPS, that S-LPS and R-LPS trigger different responses of MEC and that these responses depend on the presence of CD14.
Highlights
Bovine mastitis is defined as an inflammation of the mammary gland in cows
Because our previous studies had indicated that CD14 was important for the recognition of LPS by PS cells [23], the ability of these mutants to stimulate PS cells was performed in the presence of purified recombinant bovine CD14 (Fig 1B)
By screening of a library of random transposome mutants of E. coli P4, we demonstrate that the presence of an O-antigen decreases the ability of this strain to stimulate a pro-inflammatory response by Mammary Epithelial Cells (MEC) (Fig 1)
Summary
Bovine mastitis is defined as an inflammation of the mammary gland in cows. This disease remains an important issue with a major economic impact for the dairy farmers [1, 2]. O-antigen and response of mammary epithelial cells analysis, decision to publish, or preparation of the manuscript
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