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

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.

A hybrid receptor binding protein enables phage F341 infection of Campylobacter by binding to flagella and lipooligosaccharides

“…It is therefore unclear how phage F341 may have acquired the hybrid receptor binding protein and what environmental pressures may have selected for this unique phage. We previously also observed other genes that are shared among C. jejuni prophages and lytic phages although only annotated as hypothetical proteins ( Sørensen et al, 2021b ). This suggests that acquisition or exchange of genetic content between these groups of phages may be a more common phenomenon.…”

Campylobacter prophage diversity reveals pervasive recombination between prophages from different Campylobacter species

“…One of the most relevant findings of our study is that we uncovered a previously undescribed prophage population that comprised of 318 bacteriophage species and 154 genera. Few studies have described a part of the prophage population only for C. jejuni and C. coli species ( 21 , 22 ), resulting in the already-known CJIE-like prophages ( 50 ). However, this type of prophage is only a tiny part ( Fig.…”

“…Campylobacter phages have been studied mainly from epidemiological aspects, where most reported phages mainly infect C. jejuni and Campylobacter coli ( 21 , 22 ). In addition, the classification of Campylobacter phages has not been fully explored and standardized.…”

“…For example, lytic Campylobacter phages are classified into three groups according to their genome size ( 22 , 23 ). Group I contained uncharacterized phages with large genomes (320–425 kbs), whereas groups II and III included phages with genomes between 175 and 183 kbs ( Firehammerviruses ) and 131–135 kbs, respectively ( Fletcherviruses ) ( 21 ). Additionally, according to the National Center for Biotechnology Information (NCBI) database, there are 11 families and 11 genera of Campylobacter phages, of which only 59 phage genomes are complete (accessed in late February 2023).…”

The rise of antibiotic resistance in Campylobacter

“…C. jejuni isolates from broilers may have acquired several resistance mechanisms to prevent phage infection in their natural habitats. The restriction modification system of Campylobacter may play a protective role in avoiding phage clearance when mcrB , a methyl-specific McrBC endonuclease, is deleted, which could enhance sensitivity to CAMSA2038 [63]. Phage therapies are an exciting challenge in the fight to control the Campylobacter epidemic.…”