The Bacteriophage Carrier State of Campylobacter jejuni Features Changes in Host Non-coding RNAs and the Acquisition of New Host-derived CRISPR Spacer Sequences

Abstract

Incorporation of self-derived CRISPR DNA protospacers in Campylobacter jejuni PT14 occurs in the presence of bacteriophages encoding a CRISPR-like Cas4 protein. This phenomenon was evident in carrier state infections where both bacteriophages and host are maintained for seemingly indefinite periods as stable populations following serial passage. Carrier state cultures of C. jejuni PT14 have greater aerotolerance in nutrient limited conditions, and may have arisen as an evolutionary response to selective pressures imposed during periods in the extra-intestinal environment. A consequence of this is that bacteriophage and host remain associated and able to survive transition periods where the chances of replicative success are greatly diminished. The majority of the bacteriophage population do not commit to lytic infection, and conversely the bacterial population tolerates low-level bacteriophage replication. We recently examined the effects of Campylobacter bacteriophage/C. jejuni PT14 CRISPR spacer acquisition using deep sequencing strategies of DNA and RNA-Seq to analyze carrier state cultures. This approach identified de novo spacer acquisition in C. jejuni PT14 associated with Class III Campylobacter phages CP8/CP30A but spacer acquisition was oriented toward the capture of host DNA. In the absence of bacteriophage predation the CRISPR spacers in uninfected C. jejuni PT14 cultures remain unchanged. A distinct preference was observed for incorporation of self-derived protospacers into the third spacer position of the C. jejuni PT14 CRISPR array, with the first and second spacers remaining fixed. RNA-Seq also revealed the variation in the synthesis of non-coding RNAs with the potential to bind bacteriophage genes and/or transcript sequences.

The CRISPR Spacer Space Is Dominated by Sequences from Species-Specific Mobilomes

“…Complete reannotation of the available prokaryotic genomes is a demanding project outside the scope of the present work but, with this caveat, only a small fraction of the detected protospacers could be traced to sequences demonstrably not originating from viruses or other mobile genetic elements. Previous analyses of CRISPR arrays from individual bacterial and archaeal genomes have found widely different fractions of self-matching spacers (1,(40)(41)(42). Our current, comprehensive analysis indicates that the overwhelming majority of the spacers that persist long enough to be detected are derived from viruses and other mobile elements (collectively known as the mobilome [59]), apparently indicating strong selection against self-targeting spacers.…”

“…However, for type II systems, it has been shown that when the effector nuclease Cas9 is inactivated, preventing target cleavage, the great majority of the incorporated spacers originate from the host DNA (39). Moreover, self-derived spacers have been detected also in some microbial genomes (40)(41)(42). Thus, the specificity of the spacers toward foreign genomes could be caused primarily by selection against cells with incorporated self-targeting spacers, and the efficacy of such selection is likely to differ among microbes.…”

Review article: bacteriophages in gastroenterology—from biology to clinical applications

“…Another interaction observed between phages and bacteria is a stable equilibrium, where neither lysogeny nor lysis takes place, known as a carrier state or pseudolysogeny. This carrier state has been described in several bacteria, such as Campylobacter spp., Shigella dysenteriae , Brucella abortus and Proteus mirabilis …”

The Minor Flagellin of Campylobacter jejuni (FlaB) Confers Defensive Properties against Bacteriophage Infection

“…Moreover, Campylobacter bacteriophage carrier state cultures also escape phage lysis by undergoing growth phase dependent motility attenuation as a response to phage association (Siringan et al, 2014). Whereas, flaA gene transcription is down-regulated in carrier state cultures, which likely accounts for their impaired mobility, flaB transcription is up-regulated, which based on the current data could be an adaption to limit phage infection leading to cell lysis (Brathwaite et al, 2015; Hooton et al, 2016). It is recognized that σ 54 is part of a group of transcriptional regulators termed class 1 genes that demonstrate hierarchical regulation of class 2 genes, which are fundamental for the formation of the flagellar secretory apparatus and code for the structural components of the basal body of the flagellum (Hendrixson and DiRita, 2003; Lertsethtakarn et al, 2011).…”