Phenotypic characterization and analysis of complete genomes of two distinct strains of the proposed species "L. swaminathanii".

Recently, a new Listeria species, "Listeria swaminathanii", was proposed. Here, we phenotypically and genotypically characterize two additional strains that were previously obtained from soil samples and compare the results to the type strain. Complete genomes for both strains were assembled from hybrid Illumina and Nanopore sequencing reads and annotated. Further genomic analysis including average nucleotide identity (ANI) and detection of mobile genetic elements and genes of interest (e.g., virulence-associated) were conducted. The strains showed 98.7-98.8% ANI with the type strain. The UTK C1-0015 genome contained a partial monocin locus and a plasmid, while the UTK C1-0024 genome contained a full monocin locus and a prophage. Phenotypic characterization consistent with those performed on the proposed type strain was conducted to assess consistency of phenotypes across a greater diversity of the proposed species (n = 3 instead of n = 1). Only a few findings were notably different from those of the type strain, such as catalase activity, glycerol metabolism, starch metabolism, and growth at 41 °C. This study further expands our understanding of this newly proposed sensu stricto Listeria species.

Characterization of a Novel Group of Listeria Phages That Target Serotype 4b Listeria monocytogenes.

Listeria monocytogenes serotype 4b strains are the most prevalent clinical isolates and are widely found in food processing environments. Bacteriophages are natural viral predators of bacteria and are a promising biocontrol agent for L. monocytogenes. The aims of this study were to characterize phages that specifically infect serotype 4b strains and to assess their ability to inhibit the growth of serotype 4b strains. Out of 120 wild Listeria phages, nine phages were selected based on their strong lytic activity against the model serotype 4b strain F2365. These nine phages can be divided into two groups based on their morphological characteristics and host range. Comparison to previously characterized phage genomes revealed one of these groups qualifies to be defined as a novel species. Phages LP-020, LP-027, and LP-094 were selected as representatives of these two groups of phages for further characterization through one-step growth curve and inhibition of serotype 4b L. monocytogenes experiments. Listeria phages that target serotype 4b showed an inhibitory effect on the growth of F2365 and other serotype 4 strains and may be useful for biocontrol of L.monocytogenes in food processing environments.

Complete Genome Sequence of a Serotype 7 Listeria monocytogenes Strain, FSL R9-0915.

Listeria monocytogenes serotype 7 lacks glycosidic constituents in wall teichoic acids. Here, we present the complete genome sequence of L. monocytogenes serotype 7 strain FSL R9-0915 and an analysis of genes known to affect L. monocytogenes antigenicity. This strain is used as a control strain in Listeria phage host range analyses.

Complete Genome Sequences of Three Listeria monocytogenes Bacteriophage Propagation Strains.

Bacteriophages can be used as a biocontrol for the foodborne pathogen Listeria monocytogenes Propagation of phages is a necessary step for their use in experimental studies and biocontrol applications. Here, we present the complete genomes of three Listeria monocytogenes strains commonly used as propagation hosts for Listeria phages.

Mutant and Recombinant Phages Selected from In Vitro Coevolution Conditions Overcome Phage-Resistant Listeria monocytogenes.

Bacteriophages (phages) are currently available for use by the food industry to control the foodborne pathogen Listeria monocytogenes Although phage biocontrols are effective under specific conditions, their use can select for phage-resistant bacteria that repopulate phage-treated environments. Here, we performed short-term coevolution experiments to investigate the impact of single phages and a two-phage cocktail on the regrowth of phage-resistant L. monocytogenes and the adaptation of the phages to overcome this resistance. We used whole-genome sequencing to identify mutations in the target host that confer phage resistance and in the phages that alter host range. We found that infections with Listeria phages LP-048, LP-125, or a combination of both select for different populations of phage-resistant L. monocytogenes bacteria with different regrowth times. Phages isolated from the end of the coevolution experiments were found to have gained the ability to infect phage-resistant mutants of L. monocytogenes and L. monocytogenes strains previously found to be broadly resistant to phage infection. Phages isolated from coinfected cultures were identified as recombinants of LP-048 and LP-125. Interestingly, recombination events occurred twice independently in a locus encoding two proteins putatively involved in DNA binding. We show that short-term coevolution of phages and their hosts can be utilized to obtain mutant and recombinant phages with adapted host ranges. These laboratory-evolved phages may be useful for limiting the emergence of phage resistance and for targeting strains that show general resistance to wild-type (WT) phages.IMPORTANCEListeria monocytogenes is a life-threatening bacterial foodborne pathogen that can persist in food processing facilities for years. Phages can be used to control L. monocytogenes in food production, but phage-resistant bacterial subpopulations can regrow in phage-treated environments. Coevolution experiments were conducted on a Listeria phage-host system to provide insight into the genetic variation that emerges in both the phage and bacterial host under reciprocal selective pressure. As expected, mutations were identified in both phage and host, but additionally, recombination events were shown to have repeatedly occurred between closely related phages that coinfected L. monocytogenes This study demonstrates that in vitro evolution of phages can be utilized to expand the host range and improve the long-term efficacy of phage-based control of L. monocytogenes This approach may also be applied to other phage-host systems for applications in biocontrol, detection, and phage therapy.

Homburgvirus LP-018 Has a Unique Ability to Infect Phage-Resistant Listeria monocytogenes.

Listeria phage LP-018 is the only phage from a diverse collection of 120 phages able to form plaques on a phage-resistant Listeria monocytogenes strain lacking rhamnose in its cell wall teichoic acids. The aim of this study was to characterize phage LP-018 and to identify what types of mutations can confer resistance to LP-018. Whole genome sequencing and transmission electron microscopy revealed LP-018 to be a member of the Homburgvirus genus. One-step-growth curve analysis of LP-018 revealed an eclipse period of ~60-90 min and a burst size of ~2 PFU per infected cell. Despite slow growth and small burst size, LP-018 can inhibit the growth of Listeria monocytogenes at a high multiplicity of infection. Ten distinct LP-018-resistant mutants were isolated from infected Listeria monocytogenes 10403S and characterized by whole genome sequencing. In each mutant, a single mutation was identified in either the LMRG_00278 or LMRG_01613 encoding genes. Interesting, LP-018 was able to bind to a representative phage-resistant mutant with a mutation in each gene, suggesting these mutations confer resistance through a mechanism independent of adsorption inhibition. Despite forming plaques on the rhamnose deficient 10403S mutant, LP-018 showed reduced binding efficiency, and we did not observe inhibition of the strain under the conditions tested. Two mutants of LP-018 were also isolated and characterized, one with a single SNP in a gene encoding a BppU domain protein that likely alters its host range. LP-018 is shown to be a unique Listeria phage that, with additional evaluation, may be useful in biocontrol applications that aim to reduce the emergence of phage resistance.

Complete Genome Sequences of Two Listeria Phages of the Genus Pecentumvirus.

Bacteriophages isolated from environmental sources can be used as a biocontrol against the foodborne pathogen Listeria monocytogenes Here, we present the complete genomes of LP-039 and LP-066, two Pecentumvirus bacteriophages that infect L. monocytogenes The genome sizes of LP-039 and LP-066 are 136.2 kb and 139.0 kb, respectively.

Complete Genome Sequences and Transmission Electron Micrographs of Listeria Phages of the Genus Homburgvirus.

Bacteriophages that infect the foodborne pathogen Listeria monocytogenes were previously isolated from New York dairy farms. The complete genome sequences for three of these Listeria phages, with genome sizes of 64.6 to 65.7 kb, are presented here. Listeria phages LP-010, LP-013, and LP-031-2 are siphoviruses that belong to the genus Homburgvirus.

Use of Bacteriophages to Control Escherichia coli O157:H7 in Domestic Ruminants, Meat Products, and Fruits and Vegetables.

Escherichia coli O157:H7 is an important foodborne pathogen that causes severe bloody diarrhea, hemorrhagic colitis, and hemolytic uremic syndrome. Ruminant manure is a primary source of E. coli O157:H7 contaminating the environment and food sources. Therefore, effective interventions targeted at reducing the prevalence of fecal excretion of E. coli O157:H7 by cattle and sheep and the elimination of E. coli O157:H7 contamination of meat products as well as fruits and vegetables are required. Bacteriophages offer the prospect of sustainable alternative approaches against bacterial pathogens with the flexibility of being applied therapeutically or for biological control purposes. This article reviews the use of phages administered orally or rectally to ruminants and by spraying or immersion of fruits and vegetables as an antimicrobial strategy for controlling E. coli O157:H7. The few reports available demonstrate the potential of phage therapy to reduce E. coli O157:H7 carriage in cattle and sheep, and preparation of commercial phage products was recently launched into commercial markets. However, a better ecological understanding of the phage E. coli O157:H7 will improve antimicrobial effectiveness of phages for elimination of E. coli O157:H7 in vivo.