Lipopolysaccharide O antigen status of Yersinia enterocolitica O:8 is essential for virulence and absence of O antigen affects the expression of other Yersinia virulence factors

Abstract

Lipopolysaccharide (LPS) is the major component of the outer membrane of Gram-negative bacteria. Although much attention has been given to the biological effects of its lipid A portion, a great body of evidence indicates that its O chain polysaccharide (O antigen) portion plays an important role in the bacterium-host interplay. In this work we have studied in-depth the role of the O antigen in Yersinia enterocolitica serotype O:8 pathogenesis. We made a detailed virulence analysis of three mutants having different O antigen phenotypes: (i) LPS with no O antigen (rough mutant); (ii) LPS with one O unit (semirough mutant) and (iii) LPS with random distribution of O antigen chain lengths. We demonstrated that these LPS O antigen mutants were attenuated in virulence regardless of the infection route used. Co-infection experiments revealed that the rough and semirough mutants were severely impaired in their ability to colonize the Peyer's patches and in contrast to the wild-type strain they did not colonize spleen and liver. The mutant with random distribution of O antigen chain lengths, however, survived better but started to be cleared from mouse organs after 8 days. As an explanation to this attenuation we present here evidence that other Yersinia virulence factors depend on the presence of O antigen for their proper function and/or expression. We demonstrated that in the rough mutant: (i) the YadA function but not its expression was altered; (ii) Ail was not expressed and (iii) inv expression was downregulated. On the other hand, expression of flhDC, the flagellar master regulatory operon, was upregulated in this mutant with a concomitant increase in the production of flagellins. Finally, expression of yplA, encoding for the Yersinia phospholipase A, was also upregulated accompanied by an increased flagellar type III secretion system mediated secretion of YplA to culture medium. Together these findings suggest that the absence of O antigen in the outer membrane of Yersinia either directly or indirectly, for example through a cellular or membrane stress, could act as a regulatory signal.

Expression of the Yersinia enterocolitica O:3 LPS O-antigen and outer core gene clusters is RfaH-dependent

“…Whilst a rough Y. enterocolitica O:8 mutant exhibited alterations in YadA function, downregulation of inv and ail, and hypermotility due to upregulation of flhDC (Bengoechea et al, 2004), no such changes were observed in the YeO3-DrfaH strain. These discrepancies may be explained by differences between the serotypes.…”

An aflagellate mutant Yersinia enterocolitica biotype 1A strain displays altered invasion of epithelial cells, persistence in macrophages, and cytokine secretion profiles in vitro

“…All of these virulence determinants are present in BT1b and BT2-5 strains, but absent in BT1A strains (Bottone, 1999). In addition, several key publications have highlighted the key role of LPS and O-antigen variation in Y. enterocolitica pathogenesis (Bengoechea et al, 2003(Bengoechea et al, , 2004Rebeil et al, 2004).…”

Unique Virulence Properties of Yersinia enterocolitica O:3

“…An interesting feature of Y. enterocolitica O antigen is that its production is tightly controlled in response to temperature. Y. enterocolitica produce O-antigen in abundance at 22-25°C, but only very low amounts can be detected at 37°C (Bengoechea et al, 2004;2002). This led to speculations, that the function of YeO:3 adhesion factors might be inhibited by the presence of the distinct YeO:3 O antigen.…”

“…In contrast, O-antigen of YeO:3 is composed of a homopolymer of 6-deoxy-L-altrose linked by α-1,2 glycosidic bonds. The O-antigen and the outer core are both connected to the inner core, resulting in the formation of a branched LPS molecule (Al-Hendy et al, 1991;Bengoechea et al, 2004;Brubaker, 1991). An interesting feature of Y. enterocolitica O antigen is that its production is tightly controlled in response to temperature.…”