Genetic recombination-mediated evolutionary interactions between phages of potential industrial importance and prophages of their hosts within or across the domains of Escherichia, Listeria, Salmonella, Campylobacter, and Staphylococcus.

BACKGROUND: The in-depth understanding of the role of lateral genetic transfer (LGT) in phage-prophage interactions is essential to rationalizing phage applications for human and animal therapy, as well as for food and environmental safety. This in silico study aimed to detect LGT between phages of potential industrial importance and their hosts. METHODS: A large array of genetic recombination detection algorithms, implemented in SplitsTree and RDP4, was applied to detect LGT between various Escherichia, Listeria, Salmonella, Campylobacter, Staphylococcus, Pseudomonas, and Vibrio phages and their hosts. PHASTER and RAST were employed respectively to identify prophages across the host genome and to annotate LGT-affected genes with unknown functions. PhageAI was used to gain deeper insights into the life cycle history of recombined phages. RESULTS: The split decomposition inferences (bootstrap values: 91.3-100; fit: 91.433-100), coupled with the Phi (0.0-2.836E-12) and RDP4 (P being well below 0.05) statistics, provided strong evidence for LGT between certain Escherichia, Listeria, Salmonella, and Campylobacter virulent phages and prophages of their hosts. The LGT events entailed mainly the phage genes encoding for hypothetical proteins, while some of these genetic loci appeared to have been affected even by intergeneric recombination in specific E. coli and S. enterica virulent phages when interacting with their host prophages. Moreover, it is shown that certain L. monocytogenes virulent phages could serve at least as the donors of the gene loci, involved in encoding for the basal promoter specificity factor, for L. monocytogenes. In contrast, the large genetic clusters were determined to have been simultaneously exchanged by many S. aureus prophages and some Staphylococcus temperate phages proposed earlier as potential therapeutic candidates (in their native or modified state). The above genetic clusters were found to encompass multiple genes encoding for various proteins, such as e.g., phage tail proteins, the capsid and scaffold proteins, holins, and transcriptional terminator proteins. CONCLUSIONS: It is suggested that phage-prophage interactions, mediated by LGT (including intergeneric recombination), can have a far-reaching impact on the co-evolutionary trajectories of industrial phages and their hosts especially when excessively present across microbially rich environments.

Isolation, host specificity and genetic characterization of Campylobacter specific bacteriophages from poultry and swine sources.

The isolation and characterization of 304 Campylobacter specific bacteriophage isolates from broiler and swine sources is reported in this study. Genome size characterization determined by PFGE classified these isolates,called CAM1-CAM304, within the campylophages group II (n = 18) and group III (n = 286). Host range analyses showed a high host specificity and similar lytic spectrum among isolates of the same group. Campylophages of group II infected C. jejuni, C. coli and even a C. fetus strain whereas those of group III only infected C. jejuni strains. The most promising 59 campylophage candidates were selected according to their lytic activity and their genetic diversity was analyzed by RFLP using SmiI and HhaI endonucleases for group II and III campylophages, respectively. Moreover, RAPD-PCR technique was for the first time assessed in the genetic characterization of campylophages and it was shown to be effective only for those of group II. Bacteriophage isolates grouped in a same genotype displayed different host ranges, therefore, 13 campylophages of group II and eight of group III were differentiated considering all the approaches assayed. An in-depth analysis of these bacteriophages will be performed to confirm their promising potential for the biocontrol of Campylobacter within the farm to fork process.

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

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.

Efficient isolation of Campylobacter bacteriophages from chicken skin, analysis of several isolation protocols.

The application of Campylobacter specific bacteriophages appears as a promising food safety tool for the biocontrol of this pathogen in the poultry meat production chain. However, their isolation is a complicated challenge since their occurrence appears to be low. This work assessed the efficiency of seven protocols for recovering Campylobacter phages from chicken skin samples inoculated at phage loads from 5.0 × 101 to 5.0 × 106 PFU/g. The enrichment of chicken skin in selective Bolton broth containing target isolates was the most efficient procedure, showing a low detection limit of 5.0 × 101 PFU/g and high recovery rates of up to 560%. This method's effectiveness increased as phage concentration decreased, showing its suitability for phage isolation. When this method was applied to isolate new Campylobacter phages from retail chicken skin, a total of 280 phages were recovered achieving an isolation success rate of 257%. From the 109 samples 68 resulted phage positive (62%). Chicken skin could be, therefore, considered a rich source in Campylobacter phages. This method is a simple, reproducible and efficient approach for the successful isolation of both group II and III Campylobacter specific bacteriophages, which could be helpful for the enhancement of food safety by reducing this pathogen contamination in broiler meat.

Comparative genomics of Cp8viruses with special reference to Campylobacter phage vB_CjeM_los1, isolated from a slaughterhouse in Ireland.

Campylobacter phage vB_CjeM_Los1 was recently isolated from a slaughterhouse in the Republic of Ireland using the host Campylobacter jejuni subsp. jejuni PT14, and full-genome sequencing and annotation were performed. The genome was found to be 134,073 bp in length and to contain 169 predicted open reading frames. Transmission electron microscopy images of vB_CjeM_Los1 revealed that it belongs to the family Myoviridae, with tail fibres observed in both extended and folded conformations, as seen in T4. The genome size and morphology of vB_CjeM_Los1 suggest that it belongs to the genus Cp8virus, and seven other Campylobacter phages with similar size characteristics have also been fully sequenced. In this work, comparative studies were performed in relation to genomic rearrangements and conservation within each of the eight genomes. None of the eight genomes were found to have undergone internal rearrangements, and their sequences retained more than 98% identity with one another despite the widespread geographical distribution of each phage. Whole-genome phylogenetics were also performed, and clades were shown to be representative of the differing number of tRNAs present in each phage. This may be an indication of lineages within the genus, despite their striking homology.

Salmonella and Campylobacter: Antimicrobial resistance and bacteriophage control in poultry.

Salmonella and Campylobacter are major causes of foodborne related illness and are traditionally associated with consuming undercooked poultry and/or consuming products that have been cross contaminated with raw poultry. Many of the isolated Salmonella and Campylobacter that can cause disease have displayed antimicrobial resistance phenotypes. Although poultry producers have reduced on-the-farm overuse of antimicrobials, antimicrobial resistant Salmonella and Campylobacter strains still persist. One method of bio-control, that is producing promising results, is the use of lytic bacteriophages. This review will highlight the current emergence and persistence of antimicrobial resistant Salmonella and Campylobacter recovered from poultry as well as bacteriophage research interventions and limitations.

Campylobacter group II phage CP21 is the prototype of a new subgroup revealing a distinct modular genome organization and host specificity.

BACKGROUND: The application of phages is a promising tool to reduce the number of Campylobacter along the food chain. Besides the efficacy against a broad range of strains, phages have to be safe in terms of their genomes. Thus far, no genes with pathogenic potential (e.g., genes encoding virulence factors) have been detected in Campylobacter phages. However, preliminary studies suggested that the genomes of group II phages may be diverse and prone to genomic rearrangements. RESULTS: We determined and analysed the genomic sequence (182,761 bp) of group II phage CP21 that is closely related to the already characterized group II phages CP220 and CPt10. The genomes of these phages are comprised of four modules separated by very similar repeat regions, some of which harbouring open reading frames (ORFs). Though, the arrangement of the modules and the location of some ORFs on the genomes are different in CP21 and in CP220/CPt10. In this work, a PCR system was established to study the modular genome organization of other group II phages demonstrating that they belong to different subgroups of the CP220-like virus genus, the prototypes of which are CP21 and CP220. The subgroups revealed different restriction patterns and, interestingly enough, also distinct host specificities, tail fiber proteins and tRNA genes. We additionally analysed the genome of group II phage vB_CcoM-IBB_35 (IBB_35) for which to date only five individual contigs could be determined. We show that the contigs represent modules linked by long repeat regions enclosing some yet not identified ORFs (e.g., for a head completion protein). The data suggest that IBB_35 is a member of the CP220 subgroup. CONCLUSION: Campylobacter group II phages are diverse regarding their genome organization. Since all hitherto characterized group II phages contain numerous genes for transposases and homing endonucleases as well as similar repeat regions, it cannot be excluded that these phages are genetically unstable. To answer this question, further experiments and sequencing of more group II phages should be performed.

Phage-bacteria network analysis and its implication for the understanding of coral disease.

Multiple studies have explored microbial shifts in diseased or stressed corals; however, little is known about bacteriophage interactions with microbes in this context. This study characterized microbial 16S rRNA amplicons and phage metagenomes associated with Montastraea annularis corals during a concurrent white plague disease outbreak and bleaching event. Phage consortia differed between bleached and diseased tissues. Phages in the family Inoviridae were elevated in diseased or healthy tissues compared with bleached portions of diseased tissues. Microbial communities also differed between diseased and bleached corals. Bacteria in the orders Rhodobacterales and Campylobacterales were increased while Kiloniellales was decreased in diseased compared with other tissues. A network of phage-bacteria interactions was constructed of all phage strains and 11 bacterial genera that differed across health states. Phage-bacteria interactions varied in specificity: phages interacted with one to eight bacterial hosts while bacteria interacted with up to 59 phages. Six phages were identified that interacted exclusively with Rhodobacterales and Campylobacterales. These results suggest that phages have a role in controlling stress-associated bacteria, and that networks can be utilized to select potential phages for mitigating detrimental bacterial growth in phage therapy applications.

A suggested classification for two groups of Campylobacter myoviruses.

Most Campylobacter bacteriophages isolated to date have long contractile tails and belong to the family Myoviridae. Based on their morphology, genome size and endonuclease restriction profile, Campylobacter phages were originally divided into three groups. The recent genome sequencing of seven virulent campylophages reveal further details of the relationships between these phages at the genome organization level. This article details the morphological and genomic features among the campylophages, investigates their taxonomic position, and proposes the creation of two new genera, the "Cp220likevirus" and "Cp8unalikevirus" within a proposed subfamily, the "Eucampyvirinae"

Combined application of bacteriophages with a competitive exclusion culture and carvacrol with organic acids can reduce Campylobacter in primary broiler production.

For reducing Campylobacter (C.) in the food production chain and thus the risk to the consumer, the combined application of different measures as a multiple-hurdle approach is currently under discussion. This is the first study to investigate possible synergistic activities in vivo, aiming at reducing intestinal C. jejuni counts by administering (i) bacteriophages (phages) in combination with a competitive exclusion (CE) product and (ii) carvacrol combined with organic acids. The combined application of the two selected phages (Fletchervirus phage NCTC 12673 and Firehammervirus phage vB_CcM-LmqsCPL1/1) and the CE product significantly reduced C. jejuni loads by 1.0 log10 in cecal and colonic contents as well as in cloacal swabs at the end of the trial (33 and 34 days post hatch). The proportion of bacterial isolates showing reduced phage susceptibility ranged from 10.9% (isolates from cecal content) to 47.8% (isolates from cloacal swabs 32 days post hatch) for the Fletchervirus phage, while all tested isolates remained susceptible to the Firehammervirus phage. The use of carvacrol combined with an organic acid blend (sorbic acid, benzoic acid, propionic acid, and acetic acid) significantly reduced Campylobacter counts by 1.0 log10 in cloacal swabs on day 30 only.

The complete genome sequence of bacteriophage CP21 reveals modular shuffling in Campylobacter group II phages.

Campylobacter group II phages described so far share a high degree of sequence similarity. We report the 182,833-bp genomic sequence of the closely related group II phage CP21 and show that it has a completely different genomic organization. As in other group II phages, the CP21 genome is composed of large modules separated by long DNA repeat regions which obviously trigger recombination and modular shuffling.

Campylobacter bacteriophages and bacteriophage therapy.

Members of the genus Campylobacter are frequently responsible for human enteric disease with occasionally very serious outcomes. Much of this disease burden is thought to arise from consumption of contaminated poultry products. More than 80% of poultry in the UK harbour Campylobacter as a part of their intestinal flora. To address this unacceptably high prevalence, various interventions have been suggested and evaluated. Among these is the novel approach of using Campylobacter-specific bacteriophages, which are natural predators of the pathogen. To optimize their use as therapeutic agents, it is important to have a comprehensive understanding of the bacteriophages that infect Campylobacter, and how they can affect their host bacteria. This review will focus on many aspects of Campylobacter-specific bacteriophages including: their first isolation in the 1960s, their use in bacteriophage typing schemes, their isolation from the different biological sources and genomic characterization. As well as their use as therapeutic agents to reduce Campylobacter in poultry their future potential, including their use in bio-sanitization of food, will be explored. The evolutionary consequences of naturally occurring bacteriophage infection that have come to light through investigations of bacteriophages in the poultry ecosystem will also be discussed.

Evidence for a lineage of virulent bacteriophages that target Campylobacter.

BACKGROUND: Our understanding of the dynamics of genome stability versus gene flux within bacteriophage lineages is limited. Recently, there has been a renewed interest in the use of bacteriophages as 'therapeutic' agents; a prerequisite for their use in such therapies is a thorough understanding of their genetic complement, genome stability and their ecology to avoid the dissemination or mobilisation of phage or bacterial virulence and toxin genes. Campylobacter, a food-borne pathogen, is one of the organisms for which the use of bacteriophage is being considered to reduce human exposure to this organism. RESULTS: Sequencing and genome analysis was performed for two Campylobacter bacteriophages. The genomes were extremely similar at the nucleotide level (> or = 96%) with most differences accounted for by novel insertion sequences, DNA methylases and an approximately 10 kb contiguous region of metabolic genes that were dissimilar at the sequence level but similar in gene function between the two phages. Both bacteriophages contained a large number of radical S-adenosylmethionine (SAM) genes, presumably involved in boosting host metabolism during infection, as well as evidence that many genes had been acquired from a wide range of bacterial species. Further bacteriophages, from the UK Campylobacter typing set, were screened for the presence of bacteriophage structural genes, DNA methylases, mobile genetic elements and regulatory genes identified from the genome sequences. The results indicate that many of these bacteriophages are related, with 10 out of 15 showing some relationship to the sequenced genomes. CONCLUSIONS: Two large virulent Campylobacter bacteriophages were found to show very high levels of sequence conservation despite separation in time and place of isolation. The bacteriophages show adaptations to their host and possess genes that may enhance Campylobacter metabolism, potentially advantaging both the bacteriophage and its host. Genetic conservation has been shown to extend to other Campylobacter bacteriophages, forming a highly conserved lineage of bacteriophages that predate upon campylobacters and indicating that highly adapted bacteriophage genomes can be stable over prolonged periods of time.

Trileucine and Pullulan Improve Anti-Campylobacter Bacteriophage Stability in Engineered Spray-Dried Microparticles.

Spray drying biologics into a powder can increase thermal stability and shelf-life relative to liquid formulations, potentially eliminating the need for cold chain infrastructure for distribution in developing countries. In this study, process modelling, microparticle engineering, and a supplemented phase diagram were used to design physically stable fully amorphous spray-dried powder capable of stabilizing biological material. A greater proportion of anti-Campylobacter bacteriophage CP30A remained biologically active after spray drying using excipient formulations containing trehalose and a high glass transition temperature amorphous shell former, either trileucine or pullulan, as compared to the commonly used crystalline shell former, leucine, or a low glass transition temperature amorphous shell former, pluronic F-68. Particle formation models suggest that the stabilization was achieved by protecting the bacteriophages against the main inactivating stress, desiccation, at the surface. The most promising formulation contained a combination of trileucine and trehalose for which the combined effects of feedstock preparation, spray drying, and 1-month dry room temperature storage resulted in a titer reduction of only 0.6 ± 0.1 log10(PFU mL-1). The proposed high glass transition temperature amorphous formulation platform may be advantageous for stabilizing biologics in other spray drying applications in the biomedical engineering industry.

Two Distinct Modes of Lysis Regulation in Campylobacter Fletchervirus and Firehammervirus Phages.

Campylobacter phages are divided into two genera; Fletchervirus and Firehammervirus, showing only limited intergenus homology. Here, we aim to identify the lytic genes of both genera using two representative phages (F352 and F379) from our collection. We performed a detailed in silico analysis searching for conserved protein domains and found that the predicted lytic genes are not organized into lysis cassettes but are conserved within each genus. To verify the function of selected lytic genes, the proteins were expressed in E. coli, followed by lytic assays. Our results show that Fletchervirus phages encode a typical signal peptide (SP) endolysin dependent on the Sec-pathway for translocation and a holin for activation. In contrast, Firehammervirus phages encode a novel endolysin that does not belong to currently described endolysin groups. This endolysin also uses the Sec-pathway for translocation but induces lysis of E. coli after overexpression. Interestingly, co-expression of this endolysin with an overlapping gene delayed and limited cell lysis, suggesting that this gene functions as a lysis inhibitor. These results indicate that Firehammervirus phages regulate lysis timing by a yet undescribed mechanism. In conclusion, we found that the two Campylobacter phage genera control lysis by two distinct mechanisms.

Campylobacters and bacteriophages in the surface waters of Canterbury (New Zealand).

AIM: To determine the relationship between the presence of thermotolerant campylobacters and their bacteriophages (phages) in surface waters for the potential to use phages as an indicator of Campylobacter spp. METHODS AND RESULTS: Thermotolerant campylobacters were enumerated in 53 water samples using a three tube most probable number (MPN) series in m-Exeter broth. The presence of phages in the same samples was tested using two approaches: qualitative enrichment with five different Campylobacter hosts and a quantitative membrane concentration method. Phages infecting an Escherichia coli O157:H7 isolate were also enumerated by the membrane concentration method. Campylobacter spp. were isolated from 45/53 (85%) of the samples at 0.4-110 MPN 100 ml(-1). No Campylobacter phages were isolated, but coliphages were present in 43/46 (93%) of samples. CONCLUSIONS: The membrane concentration method recovered >80% of Campylobacter phages from spiked samples. The absence of Campylobacter phages in environmental samples, from both enrichment and concentration methods, suggests that, if present, they are at very low titres. SIGNIFICANCE AND IMPACT OF THE STUDY: Testing for Campylobacter phages as an indicator of Campylobacter spp. presence is not effective. The quantitative data for Campylobacter spp. will be useful for risk assessment purposes.

Spray-dried anti-Campylobacter bacteriophage CP30A powder suitable for global distribution without cold chain infrastructure.

Campylobacter jejuni is a leading cause of foodborne illness globally. In this study, a spray drying and packaging process was developed to produce a thermally-stable dry powder containing bacteriophages that retains biological activity against C. jejuni after long distance shipping at ambient temperature. Spray drying using a twin-fluid atomizer resulted in significantly less (p < 0.05) titer reduction than spray drying using a vibrating mesh nebulizer. The use of centrifugation and dilution of filtered bacteriophage lysate in the formulation step resulted in a significantly greater (p < 0.05) proportion of bacteriophages remaining active relative to use of no centrifugation and dilution. The spray-dried bacteriophage powder generated using leucine and trehalose as excipients was flowable, non-cohesive, and exhibited a high manufacturing yield. The powder retained its titer with no significant differences (p > 0.05) in biological activity after storage in suitable packaging for at least 3 weeks at room temperature and after ambient temperature shipping a total distance of approximately 19,800 km, including with a 38 °C temperature excursion. The bacteriophage powder therefore appears suitable for global distribution without the need for cold chain infrastructure.

The applications of bacteriophages and their lysins as biocontrol agents against the foodborne pathogens Listeria monocytogenes and Campylobacter: An updated look.

Listeria monocytogenes and Campylobacter spp. are foodborne pathogens responsible for outbreaks and disease in humans. The emerging problem of bacterial antibiotic resistance and the persistence of pathogens in the environment, especially where foods are processed, are some of the reasons that have led to a re­emerging interest in bacteriophages and their lysins as potential candidates for bio­control. This review focuses on the use of bacteriophages and their lysins as alternative strategies for controlling the foodborne pathogens L. monocytogenes and Campylobacter spp. In addition, the application of bacteriophages and their lysins in food safety and animal health, as well as phage­resistance development, legislation, and future prospects were discussed.

Characterization of Campylobacter phages including analysis of host range by selected Campylobacter Penner serotypes.

BACKGROUND: The predominant food borne pathogen in the western world today is Campylobacter. Campylobacter specific bacteriophages (phages) have been proposed as an alternative agent for reducing the burden of Campylobacter in broilers. One concern in relation to phage biocontrol is the narrow host range often displayed by phages. To identify the potential of phages as a Campylobacter reducing agent we needed to determine their infectivity on a panel of isolates representing the Campylobacter strains found in broilers as well as humans. RESULTS: In this study, Campylobacter phages were isolated from the intestines of broilers and ducks and from abattoir sewage. Twelve phages were investigated to determine their ability to infect the Campylobacter Penner serotypes commonly present in Danish poultry and patients with campylobacteriosis. A total of 89% of the Campylobacter jejuni strains and 14% of the Campylobacter coli strains could be infected by at least one of the bacteriophages. The majority of the phages infected the most common serotypes in Danish broilers (O:1,44; O:2; O:4-complex), but showed limited ability to infect 21 of the less frequent Campylobacter serotypes. Pulse field gel electrophoresis (PFGE) and restriction endonuclease analysis (REA) were used to characterize the phage genomes. Three categories of bacteriophages were observed. I: a genome size of approximately 194 kb and refractory to digestion with HhaI; II: a genome size of approximately 140 kb and digestible by HhaI; and III: a genome size undeterminable in PFGE. The categorization of the phages correlated with the host range patterns displayed by the phages. Six phages were subjected to transmission electron microscopy (TEM). They all belonged to the family of Myoviridae. CONCLUSION: We have characterized and identified the host range of 12 Danish Campylobacter phages. Due to their ability to infect the majority of the common serotypes in Denmark we suggest the phages can become an effective agent in the effort to reduce the incidence of campylobacteriosis in Denmark. This study provides the basis for future experiments in Campylobacter phages and knowledge for the selection of Campylobacter phages for biocontrol in broilers.

Campylobacters and their bacteriophages from chicken liver: The prospect for phage biocontrol.

Consumption of foods containing chicken liver has been associated with Campylobacter enteritis. Campylobacters can contaminate the surface of livers post-mortem but can also arise through systemic infection of colonising bacteria in live birds. The use of bacteriophage to reduce levels of Campylobacter entering the food chain is a promising intervention approach but most phages have been isolated from chicken excreta. This study examined the incidence and contamination levels of Campylobacter and their bacteriophage in UK retail chicken liver. Using enrichment procedures, 87% of 109 chicken livers were surface contaminated with Campylobacter and 83% contaminated within internal tissues. Direct plating on selective agar allowed enumeration of viable bacteria from 43% of liver samples with counts ranging from 1.8->3.8log10CFU/cm2 for surface samples, and 3.0->3.8log10CFU/g for internal tissue samples. Three C. jejuni isolates recovered from internal liver tissues were assessed for their ability to colonise the intestines and extra-intestinal organs of broiler chickens following oral infection. All isolates efficiently colonised the chicken intestines but were variable in their abilities to colonise extra-intestinal organs. One isolate, CLB104, could be recovered by enrichment from the livers and kidneys of three of seven chickens. Campylobacter isolates remained viable within fresh livers stored at 4°C over 72h and frozen livers stored at -20°C over 7days in atmospheric oxygen, and therefore constitute a risk to human health. Only three Campylobacter-specific bacteriophages were isolated, and these exhibited a limited host range against the Camplylobacter chicken liver isolates. All were identified as group III virulent bacteriophage based on their genome size of 140kb. The application of broad host range group II virulent phages (8log10PFU/g) to liver homogenates containing C. jejuni strains of diverse origin at 4°C resulted in modest but significant reductions in the viable counts ranging from 0.2 to 0.7log10CFU/g.