Passage of Campylobacter jejuni through the chicken reservoir or mice promotes phase variation in contingency genes Cj0045 and Cj0170 that strongly associates with colonization and disease in a mouse model

Abstract

Human illness due to Camplyobacter jejuni infection is closely associated with consumption of poultry products. We previously demonstrated a 50 % shift in allele frequency (phase variation) in contingency gene Cj1139 (wlaN) during passage of C. jejuni NCTC11168 populations through Ross 308 broiler chickens. We hypothesized that phase variation in contingency genes during chicken passage could promote subsequent colonization and disease in humans. To test this hypothesis, we passaged C. jejuni strains NCTC11168, 33292, 81-176, KanR4 and CamR2 through broiler chickens and analysed the ability of passaged and non-passaged populations to colonize C57BL6 IL-10-deficient mice, our model for human colonization and disease. We utilized fragment analysis and nucleotide sequence analysis to measure phase variation in contingency genes. Passage through the chicken reservoir promoted phase variation in five specific contingency genes, and these 'successful' populations colonized mice. When phase variation did not occur in these same five contingency genes during chicken passage, these 'unsuccessful' populations failed to colonize mice. Phase variation during chicken passage generated small insertions or deletions (indels) in the homopolymeric tract (HT) in contingency genes. Singlecolony isolates of C. jejuni strain KanR4 carrying an allele of contingency gene Cj0170 with a 3These authors contributed equally to this work. Two supplementary tables, listing PCR primer sequences for fragment and sequence analysis used in this study, and results of sequence and fragment analysis to confirm large shifts in allele frequency, with a supplementary reference, are available with the online version of this paper. 10G HT colonized mice at high frequency and caused disease symptoms, whereas single-colony isolates carrying the 9G allele failed to colonize mice. Supporting results were observed for the successful 9G allele of Cj0045 in strain 33292. These data suggest that phase variation in Cj0170 and Cj0045 is strongly associated with mouse colonization and disease, and that the chicken reservoir can play an active role in natural selection, phase variation and disease.

Genomic variations leading to alterations in cell morphology of Campylobacter spp

“…In C. jejuni , PVRs have been identified in capsule, LOS and flagellin glycosylation loci as well as a range of loci of unknown function4272845. Most identified PVRs in C. jejuni are within polyG:C tracts of seven or more nucleotides in length262745.…”

Coadministration of the Campylobacter jejuni N-Glycan-Based Vaccine with Probiotics Improves Vaccine Performance in Broiler Chickens

“…Passage through chickens can also promote phase variation in certain contingency genes (71,72), including Cj0170, which is involved in regulating motility, and Cj0045, which is associated with colonization and disease in mice (73). Here, we compared average phase-variable expression states for 19 loci by analysis of 30 colonies isolated from each C. jejuni-positive bird versus 30 colonies obtained from the input culture.…”

Whole-Genome Sequencing in Epidemiology of Campylobacter jejuni Infections

“…The genomic changes that occurred during host passage (either human or animal) were small and, except for rare single nucleotide variants (SNVs) at one or two loci, limited to indels in homopolymeric tracts in contingency loci. These frameshift mutations were generally found in genes regulating phase variations of surface structures and therefore are likely to play an important role in host adaptation (11)(12)(13)(14). Furthermore, these mutations accumulated rapidly in the C. jejuni population infecting a single individual, even in the absence of selective pressure, demonstrating mutation rates between 10 and 100 times faster than mutation rates in other parts of the genome (10).…”

“…The fixation of such mutations is determined by evolutionary processes such as genetic drift, the bottleneck effect, and selection and can be observed due to host adaption (10). WGS has been applied to study the genomic diversity and changes that can accumulate during human infection (11,12) and animal colonization (13,14). The genomic changes that occurred during host passage (either human or animal) were small and, except for rare single nucleotide variants (SNVs) at one or two loci, limited to indels in homopolymeric tracts in contingency loci.…”