Identification of Novel Phage Resistance Mechanisms in Campylobacter jejuni by Comparative Genomics

Martine C H Sørensen; Yilmaz Emre Gencay; Florian Fanger; Mariana A T Chichkova; Mária Mazúrová; Jochen Klumpp; Eva M Nielsen; Lone Brøndsted

doi:10.3389/fmicb.2021.780559

Identification of Novel Phage Resistance Mechanisms in Campylobacter jejuni by Comparative Genomics

Front Microbiol. 2021 Dec 14:12:780559. doi: 10.3389/fmicb.2021.780559. eCollection 2021.

Authors

Martine C H Sørensen¹, Yilmaz Emre Gencay¹, Florian Fanger¹, Mariana A T Chichkova¹, Mária Mazúrová¹, Jochen Klumpp², Eva M Nielsen³, Lone Brøndsted¹

Affiliations

¹ Food Safety and Zoonoses, Department of Veterinary and Animal Sciences, University of Copenhagen, Frederiksberg, Denmark.
² Institute for Food, Nutrition and Health, ETH Zürich, Zurich, Switzerland.
³ Foodborne Infections, Department of Bacteria, Parasites & Fungi, Statens Serum Institut, Copenhagen, Denmark.

Abstract

Phages infecting Campylobacter jejuni are considered a promising intervention strategy at broiler farms, yet phage sensitivity of naturally occurring poultry isolates is not well studied. Here, we investigated phage sensitivity and identified resistance mechanisms of C. jejuni strains originating from Danish broilers belonging to the most prevalent MLST (ST) types. Determining plaque formation of 51 phages belonging to Fletchervirus or Firehammervirus showed that 21 out of 31 C. jejuni strains were susceptible to at least one phage. While C. jejuni ST-21 strains encoded the common phase variable O-methyl phosphoramidate (MeOPN) receptor of the Fletchervirus and were only infected by these phages, ST-45 strains did not encode this receptor and were exclusively infected by Firehammervirus phages. To identify internal phage resistance mechanism in ST-21 strains, we performed comparative genomics of two strains, CAMSA2002 sensitive to almost all Fletchervirus phages and CAMSA2038, resistant to all 51 phages. The strains encoded diverse clustered regularly interspaced short palindromic repeats (CRISPR) spacers but none matched the tested phages. Sequence divergence was also observed in a predicted SspE homolog and putative restriction modification systems including a methyl-specific McrBC endonuclease. Furthermore, when mcrB was deleted, CAMSA2038 became sensitive to 17 out of 43 phages, three being Firehammervirus phages that otherwise did not infect any ST-21 strains. Yet, 16 phages demonstrated significantly lower efficiencies of plating on the mcrB mutant suggesting additional resistance mechanism still restricting phage propagation in CAMSA2038. Thus, our work demonstrates that C. jejuni isolates originating from broilers may have acquired several resistance mechanisms to successfully prevent phage infection in their natural habitat.

Keywords: Campylobacter jejuni; MLST (multilocus sequence typing); McrBC; bacteriophage; comparative genomics; phage resistance; phage sensitivity; restriction modification system.