Abstract
Lawsonia intracellularis is an obligately intracellular pathogen which is the cause of the
disease complex known as proliferative enteropathy (PE) or ileitis. This bacterium is
pathogenic in a broad range of animal hosts, disease being most notable in pigs. L.
intracellularis has a tropism for immature (crypt) epithelial cells and disease is
characterised by epithelial hyperplasia in infected crypts. This pathology presumably
reflects expression of novel virulence factors during infection. To date few genes have
been identified and of these only IsaA, a tlyA homologue, has any function ascribed.
TlyA proteins of bacteria belong to a novel family identified across a phylogenetically
diverse range of bacteria. These include several gastrointestinal pathogens such as
Helicobacter pylori, Campylobacterjejuni, Brachyspira hyodysenteriae and Lawsonia
intracellularis. These proteins have been identified mainly through genomic sequencing
and their expression and role(s) during infection remain to be fully defined. TlyA
deletion mutants in H. pylori and B. hyodysenteriae are attenuated, suggesting that these
proteins perform important roles during infection. LsaA (lawsonia surface antigen) the
L. intracellularis orthologue, is expressed during infection in vitro and in vivo, which
suggests that this factor is involved during adherence and/or invasion of intestinal
epithelial cells. The principal aim was to characterise function(s) of LsaA. Specifically
the putative function as an adhesin was investigated further using a combination of
biochemical and molecular approaches (including affinity purification and yeast 2-
hybrid analysis) to elucidate possible receptor(s). However, no consistent partner was
evident therefore mammalian epithelial cell receptors could not be defined using this
range of approaches. It is possible that LsaA's role in adherence is adventitious - for
example, it has been proposed that the TlyA family of proteins possess a regulatory role
in bacterial colonisation as opposed to a direct involvement in bacterial adherence. The
existence of two conserved putative functional domains, S4 RNA binding and
methyltransferase motifs have been noted in all members of the TlyA family examined
to date. These domains are found separately in several protein families known to be
involved in gene regulation. Since no system has been developed for mutating genes in
L. intracellularis the proteome of a TlyA deletion mutant of H. pylori was compared to its parent to further this potentially new and interesting function of TlyA family proteins
Notably, flagellin B and catalase were absent in the tlyA mutant. Since deletion of tlyA
corresponds with changes in expression of several H. pylori genes, it can be concluded
that reduced colonisation of H. pylori tlyA mutant is likely to be as a result of effects on
expression of virulence genes rather than a direct role in adherence.
