Enteric Diseases of Food Animals: Enhanced Prevention, Control and Food Safety

Non-Technical Summary:<br/>
Campylobacter jejuni is a major cause of gastroenteritis with vomiting and diarrhea. It causes disease and damage to the colon after ingestion, but most individuals recover after 7-10 days. However, some people infected with C. jejuni develop autoimmune diseases like inflammatory bowel disease (IBD) and peripheral nerve disease like Guillain Barre syndrome. We will study how C. jejuni adapts to its host because this process leads to the most severe manifestations of the pathogen. To do this we will sequence genes and examine gene expression both before and after passage of the bacterium in the model host, the mouse. We will focus especially on contingency genes, which are those that can change rapidly by an evolutionary process. Some of these rapid changes have been correlated to more severe disease in the mouse model. We plan to figure out the mechanism of how contingency genes and other genes lead to enhanced virulence. This outcome will be measured by comparing genes that are present/absent or up or down regulated in the adapted compared to the nonadapted C. jejuni strain. We will publish these outcomes and this work will inform development of vaccines and treatments for this bacterium.
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Approach:<br/>
1) Host adaptation of C. jejuni strains. C. jejuni adapts quickly to its host environment, gaining virulence in serial transfers through a mouse model of campylobacteriosis. Gain of virulence appears to be entirely by mutations in contingency loci, homopolymeric tracts (HTs) that experience frequent base insertion and deletions that alter gene sequence and expression. We used population-level re-sequencing to show that these were the only genomic differences between a pre- and post passage population, supporting the hypothesis that contingency loci generate genomic diversity that enables C. jejuni to rapidly evolve to exploit new host environments. We also expect that within host variation in contingency genes may account for variability of expression of paralysis in humans infected with C. jejuni class A lipo-oligosaccharide (LOS) strains. We will sequence other replicate C. jejuni lineages, already passaged through mice to determine where or not the same contingency loci changes are responsible for adaptation in these populations. We will use breseq software to predict mutations in population level bacterial re-sequencing data, analyze contingency loci evolution in lab-evolved C. jejuni lineages with relaxed selection, and initiate an effort to quantitatively analyze the evolution of contingency loci in C. jejuni using a quasispecies model.
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2) Defining C. jejuni genes encoding surface structures that contribute to development of autoimmunity. Presence of certain chemical variants of the LOS surface molecule of C. jejuni was correlated with development of an ascending paralysis (Guillain Barre Syndrome)in patients previously infected with C. jejuni. Particular LOS variants were correlated with changes in particular C. jejuni genes in patient strains. It is likely that LOS variants mimic gangliosides on human peripheral nerves and trigger autoantibodies that cross-react with nerves inducing immune system attack, resulting in nerve damage and paralysis. Some of these genes contain HTs, subject to high rates of slip-strand mutagenesis, which alters the length of the tract. Such mutations often truncate and inactivate the encoded proteins, thereby altering the chemical structure of LOS. We demonstrated variation in HT lengths in contingency genes during passage of C. jejuni strain 11168 in mice such that C. jejuni populations recovered from mice after passage contained different proportions of tract lengths from those in the inoculum. We will use stored isolates of C. jejuni 260.94, a sequenced GBS-associated human clinical strain, recovered from infected NOD WT, NOD IL-10 deficient, NOD B7-2 deficient, and C57BL/6 WT mice after infection. Some mice of each genotype had autoantibodies and/or neurological deficits. We will examine the distribution of HT lengths in three contingency genes in the LOS locus of C. jejuni 260.94 (neuA, neuB, and hldE) pre- and post passage in the mice using PCR-fragment length method. We expect to find shifts in proportions of tract lengths in one or more of these genes; we will then correlate such shifts with the clinical outcome and/or development of autoantibodies in individual mice.