Campylobacter bacteriophage DA10: an excised temperate bacteriophage targeted by CRISPR-cas

Abstract

Background Lytic bacteriophages that infect Campylobacter spp. have been utilized to develop therapeutic/decontamination techniques. However, the association of Campylobacter spp. and bacteriophages has been the focus of several strands of research aimed at understanding the complex relationships that have developed between predators and prey over evolutionary time. The activities of endogenous temperate bacteriophages have been used to evaluate genomic rearrangements and differential protein expression in host cells, and mechanisms of resistance to bacteriophage infection in campylobacters such as phase variation and CRISPR-mediated immunity. Results Temperate bacteriophage DA10 represents a novel excised and infective virus capable of replication in a restricted set of C. jejuni and C. coli hosts. Whole genome sequencing reveals that DA10 (35,379 bp) forms part of a novel group of temperate bacteriophages that have limited distribution among database host genome sequences. Analysis of potential host genomes reveals a robust response against DA10 and DA10-like bacteriophages is driven by CRISPR-mediated immunity with 75% of DA10 ORFs represented as ~ 30 bp spacer sequences in numerous Campylobacter Type II-C CRISPR arrays. Several DA10-like homologues have been identified in a small sub-set of C. jejuni and C. coli genome sequences (ranging from near complete integrated prophage sequences to fragments recognisable in the sequence read archive). Conclusions A complete intact DA10-like prophage in C. jejuni CJ677CC520 provides evidence that the associations between host and DA10-like bacteriophages are long-standing in evolutionary timescales. Extensive nucleotide substitution and loss can be observed in the integrated DA10-like prophage of CJ677CC520 compared to other relatives as observed through pairwise genome comparisons. Examining factors that have limited the population expansion of the prophage, while others appear to have thrived and prospered (Mu-like, CJIE-like, and lytic Campylobacter bacteriophages) will assist in identifying the underlying evolutionary processes in the natural environment.

A Campylobacter integrative and conjugative element with a CRISPR-Cas9 system targeting competing plasmids: a history of plasmid warfare?

“…jejuni and C. coli and most spacers cannot (yet) be linked to mobile elements or phages [34, 59–61], the chromosomal copy of Cas9 may perform additional or alternative functions in C. jejuni , such as control or activity in virulence [62–66].…”

“…We have previously shown that ~98 % of all C. jejuni genomes have a CRISPR-Cas system, while this is more limited in C. coli, where only ~10 % of C. coli genomes have a CRISPR-Cas system [34]. Since the diversity in CRISPR spacers is also low in the chromosomal version of CRISPR-Cas of C. jejuni and C. coli and most spacers cannot (yet) be linked to mobile elements or phages [34,[59][60][61], the chromosomal copy of Cas9 may perform additional or alternative functions in C. jejuni, such as control or activity in virulence [62][63][64][65][66]. However, this is not the case for the CampyICE1 CRISPR-Cas9 system, as a majority of spacers can be linked to the three main families of plasmids in C. jejuni and C. coli: pTet, pVir and pCC42.…”

Distribution of CRISPR Types in Fluoroquinolone-Resistant Campylobacter jejuni Isolates

“…In general, phages within a group share high genomic homology, but the overall homology among the different groups may be quite low [ 48 ]. Even though a few open reading frames (ORFs) of phage DA10 (a novel class of Campylobacter phage) have homologs in other Campylobacter phages, including CP39 (a class III Campylobacter phage) and the prophage CJIE4-5 [ 49 ], the likelihood of these shared ORFs in the induction of cross-immunity among different phage groups is expected to be minimal. Furthermore, these data showed that the CRISPR region of most of the FQ-resistant C. jejuni strains tested in this study contained nucleic acids derived not only from phages but also from plasmid or other sources.…”

A Campylobacter integrative and conjugative element with a CRISPR-Cas9 system targeting competing plasmids: a history of plasmid warfare?

“…We have previously shown that ~98% of all C. jejuni genomes have a CRISPR-Cas system, while this is more limited in C. coli, where only ~10% of C. coli genomes have a CRISPR-Cas system [34]. Since the diversity in CRISPR spacers is also low in the chromosomal version of CRISPR-Cas of C. jejuni and C. coli and most spacers cannot (yet) be linked to mobile elements or phages [34,[57][58][59], it may represent additional or alternative functions for Cas9 in C. jejuni, such as control or activity in virulence [60][61][62][63][64]. However, this is not the case for the CampyICE1 CRISPR-Cas9 system, as a majority of spacers can be linked to the three main families of plasmids in C. jejuni and C. coli: pTet, pVir and pCC42.…”