Peptidoglycan-Modifying Enzyme Pgp1 Is Required for Helical Cell Shape and Pathogenicity Traits in Campylobacter jejuni

Emilisa Frirdich,Jeremy Ellermeier,Erin C Gaynor,Calvin Adams,Waldemar Vollmer,Jooeun Lee,Jacob Biboy,Lindsay Davis Gielda,Stephen E Girardin,Victor J Dirita,Steven R Blanke

doi:10.1371/journal.ppat.1002602

Emilisa Frirdich, Jeremy Ellermeier + Show 9 more

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https://doi.org/10.1371/journal.ppat.1002602

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Abstract

The impact of bacterial morphology on virulence and transmission attributes of pathogens is poorly understood. The prevalent enteric pathogen Campylobacter jejuni displays a helical shape postulated as important for colonization and host interactions. However, this had not previously been demonstrated experimentally. C. jejuni is thus a good organism for exploring the role of factors modulating helical morphology on pathogenesis. We identified an uncharacterized gene, designated pgp1 (peptidoglycan peptidase 1), in a calcofluor white-based screen to explore cell envelope properties important for C. jejuni virulence and stress survival. Bioinformatics showed that Pgp1 is conserved primarily in curved and helical bacteria. Deletion of pgp1 resulted in a striking, rod-shaped morphology, making pgp1 the first C. jejuni gene shown to be involved in maintenance of C. jejuni cell shape. Pgp1 contributes to key pathogenic and cell envelope phenotypes. In comparison to wild type, the rod-shaped pgp1 mutant was deficient in chick colonization by over three orders of magnitude and elicited enhanced secretion of the chemokine IL-8 in epithelial cell infections. Both the pgp1 mutant and a pgp1 overexpressing strain – which similarly produced straight or kinked cells – exhibited biofilm and motility defects. Detailed peptidoglycan analyses via HPLC and mass spectrometry, as well as Pgp1 enzyme assays, confirmed Pgp1 as a novel peptidoglycan DL-carboxypeptidase cleaving monomeric tripeptides to dipeptides. Peptidoglycan from the pgp1 mutant activated the host cell receptor Nod1 to a greater extent than did that of wild type. This work provides the first link between a C. jejuni gene and morphology, peptidoglycan biosynthesis, and key host- and transmission-related characteristics.

Highlights

Campylobacter jejuni is a helical, highly motile, Gram-negative eProteobacterium and a prevalent zoonotic organism existing asymptomatically in the intestinal tract of birds and other animal species [1,2,3]
We found that C. jejuni strain 81-176 binds calcofluor white (CFW), a compound that reacts with b1–3 and b1–4 carbohydrate linkages and fluoresces under long wave UV light [8,9]
Bacterial cell shape is dictated by the composition of the cell envelope component peptidoglycan

Summary

Introduction

Campylobacter jejuni is a helical, highly motile, Gram-negative eProteobacterium and a prevalent zoonotic organism existing asymptomatically in the intestinal tract of birds and other animal species [1,2,3]. C. jejuni lacks many of the frequently identified virulence factors encoded by other enteric pathogens such as pili, enterotoxins, and specialized secretion mechanisms [6,7]. Genes affecting fundamental aspects of C. jejuni biology in hosts other than humans, such as stress survival, transmission, and asymptomatic colonization, affect virulence in disease models. The carbohydrate responsible for CFW reactivity in C. jejuni has not yet been identified, it was previously shown not to be one of the well-characterized surface polysaccharides expressed by C. jejuni: the capsular polysaccharide, lipooligosaccharide (LOS), N-linked glycoproteins, or O-linked flagellar glycoproteins [10]. C. jejuni mutants with altered CFW reactivity can be readily identified in screens. All C. jejuni CFW hyper- or hyporeactive mutants characterized to date exhibit changes in pathogenesis, virulence, fundamental, and/or stress survival phenotypes

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