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.

Soil Collected in the Great Smoky Mountains National Park Yielded a Novel Listeria sensu stricto Species, L. swaminathanii.

Soil samples collected in the Great Smoky Mountains National Park yielded a Listeria isolate that could not be classified to the species level. Whole-genome sequence-based average nucleotide identity BLAST and in silico DNA-DNA Hybridization analyses confirmed this isolate to be a novel Listeria sensu stricto species with the highest similarity to L. marthii (ANI = 93.9%, isDDH = 55.9%). Additional whole-genome-based analysis using the Genome Taxonomy Database Toolkit further supported delineation as a novel Listeria sensu stricto species, as this tool failed to assign a species identification. Phenotypic and genotypic characterization results indicate that this species is nonpathogenic. Specifically, the novel Listeria species described here is phenotypically (i) nonhemolytic and (ii) negative for phosphatidylinositol-specific phospholipase C activity; the draft genome lacks all virulence genes found in the Listeria pathogenicity islands 1, 2, 3, and 4 as well as the internalin genes inlA and inlB. While the type strain contains an apparently intact catalase gene (kat), this strain is phenotypically catalase-negative (an unusual characteristic for Listeria sensu stricto species). Additional analyses identified a nonsynonymous mutation in a conserved codon of kat that is likely linked to the catalase-negative phenotype. Rapid species identification systems, including two biochemical and one matrix-assisted laser desorption/ionization, misidentified this novel species as either L. monocytogenes, L. innocua, or L. marthii. We propose the name L. swaminathanii, and the type strain is FSL L7-0020T (=ATCC TSD-239T). IMPORTANCEL. swaminathanii is a novel sensu stricto species that originated from a US National Park and it will be the first Listeria identified to date without official standing in the nomenclature. Validation was impeded by the National Park's requirements for strain access, ultimately deemed too restrictive by the International Committee on Systematics of Prokaryotes. However, lack of valid status should not detract from the significance of adding a novel species to the Listeria sensu stricto clade. Notably, detection of non-monocytogenes sensu stricto species in a food processing environment indicate conditions that could facilitate the presence of the pathogen L. monocytogenes. If isolated, our data show a potential for L. swaminathanii to be misidentified as another sensu stricto, notably L. monocytogenes. Therefore, developers of Listeria spp. detection and identification methods, who historically only include validly published species in their validation studies, should include L. swaminathanii to ensure accurate results.

Novel Salmonella Phage, vB_Sen_STGO-35-1, Characterization and Evaluation in Chicken Meat.

Salmonellosis is one of the most frequently reported zoonotic foodborne diseases worldwide, and poultry is the most important reservoir of Salmonella enterica serovar Enteritidis. The use of lytic bacteriophages (phages) to reduce foodborne pathogens has emerged as a promising biocontrol intervention for Salmonella spp. Here, we describe and evaluate the newly isolated Salmonella phage STGO-35-1, including: (i) genomic and phenotypic characterization, (ii) an analysis of the reduction of Salmonella in chicken meat, and (iii) genome plasticity testing. Phage STGO-35-1 represents an unclassified siphovirus, with a length of 47,483 bp, a G + C content of 46.5%, a headful strategy of packaging, and a virulent lifestyle. Phage STGO-35-1 reduced S. Enteritidis counts in chicken meat by 2.5 orders of magnitude at 4 °C. We identified two receptor-binding proteins with affinity to LPS, and their encoding genes showed plasticity during an exposure assay. Phenotypic, proteomic, and genomic characteristics of STGO-35-1, as well as the Salmonella reduction in chicken meat, support the potential use of STGO-35-1 as a targeted biocontrol agent against S. Enteritidis in chicken meat. Additionally, computational analysis and a short exposure time assay allowed us to predict the plasticity of genes encoding putative receptor-binding proteins.

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.

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.