High-Frequency Variation of Purine Biosynthesis Genes Is a Mechanism of Success in Campylobacter jejuni
Authors: Cameron Andrew , Huynh Steven , Scott Nichollas E. , Frirdich Emilisa , Apel Dmitry , Foster Leonard J. , Parker Craig T. , Gaynor Erin C. 
Published: September 2015

Phenotypic variation is prevalent in the zoonotic pathogen
Campylobacter jejuni
, the leading agent of enterocolitis in the developed world. Heterogeneity enhances the survival and adaptive malleability of bacterial populations because variable phenotypes may allow some cells to be protected against future stress. Exposure to hyperosmotic stress previously revealed prevalent differences in growth between
C. jejuni
strain 81-176 colonies due to resistant or sensitive phenotypes, and these isolated colonies continued to produce progeny with differential phenotypes. In this study, whole-genome sequencing of isolated colonies identified allelic variants of two purine biosynthesis genes,
purF
and
apt
, encoding phosphoribosyltransferases that utilize a shared substrate. Genetic analyses determined that
purF
was essential for fitness, while
apt
was critical. Traditional and high-depth amplicon-sequencing analyses confirmed extensive intrapopulation genetic variation of
purF
and
apt
that resulted in viable strains bearing alleles with in-frame insertion duplications, deletions, or missense polymorphisms. Different
purF
and
apt
alleles were associated with various stress survival capabilities under several niche-relevant conditions and contributed to differential intracellular survival in an epithelial cell infection model. Amplicon sequencing revealed that intracellular survival selected for stress-fit
purF
and
apt
alleles, as did exposure to oxygen and hyperosmotic stress. Putative protein recognition direct repeat sequences were identified in
purF
and
apt
, and a DNA-protein affinity screen captured a predicted exonuclease that promoted the global spontaneous mutation rate. This work illustrates the adaptive properties of high-frequency genetic variation in two housekeeping genes, which influences
C. jejuni
survival under stress and promotes its success as a pathogen.

IMPORTANCE
C. jejuni
is an important cause of bacterial diarrheal illness. Bacterial populations have many strategies for stress survival, but phenotypic variation due to genetic diversity has a powerful advantage: no matter how swift the change in environment, a fraction of the population already expresses the survival trait. Nonclonality is thus increasingly viewed as a mechanism of population success. Our previous work identified prominent resistant/sensitive colonial variation in
C. jejuni
bacteria in response to hyperosmotic stress; in the work presented here, we attribute that to high-frequency genetic variation in two purine biosynthesis genes,
purF
and
apt
. We demonstrated selective pressure for nonlethal mutant alleles of both genes, showed that single-cell variants had the capacity to give rise to diverse
purF
and
apt
populations, and determined that stress exposure selected for desirable alleles. Thus, a novel
C. jejuni
adaptive strategy was identified, which was, unusually, reliant on prevalent genetic variation in two housekeeping genes.