Degradation of Listeria monocytogenes biofilm by phages belonging to the genus Pecentumvirus.

Listeria monocytogenes is a pathogenic foodborne bacterium that is a significant cause of mortality associated with foodborne illness and causes many food recalls attributed to a bacteriological cause. Their ability to form biofilms contributes to the persistence of Listeria spp. in food processing environments. When growing as biofilms, L. monocytogenes are more resistant to sanitizers used in the food industry, such as benzalkonium chloride (BAC), as well as to physical stresses like desiccation and starvation. Lytic phages of Listeria are antagonistic to a broad range of Listeria spp. and may, therefore, have utility in reducing the occurrence of Listeria-associated food recalls by preventing food contamination. We screened nine closely related Listeria phages, including the commercially available Listex P100, for host range and ability to degrade microtiter plate biofilms of L. monocytogenes ATCC 19111 (serovar 1/2a). One phage, CKA15, was selected and shown to rapidly adsorb to its host under conditions relevant to applying the phage in dairy processing environments. Under simulated dairy processing conditions (SDPC), CKA15 caused a 2-log reduction in Lm19111 biofilm bacteria. This work supports the biosanitation potential of phage CKA15 and provides a basis for further investigation of phage-bacteria interactions in biofilms grown under SDPC. IMPORTANCE: Listeria monocytogenes is a pathogenic bacterium that is especially dangerous for children, the elderly, pregnant women, and immune-compromised people. Because of this, the food industry takes its presence in their plants seriously. Food recalls due to L. monocytogenes are common with a high associated economic cost. In food-processing plants, Listeria spp. typically reside in biofilms, which are structures produced by bacteria that shield them from environmental stressors and are often attached to surfaces. The significance of our work is that we show a bacteriophage-a virus-infecting bacteria-can reduce Listeria counts by two orders of magnitude when the bacterial biofilms were grown under simulated dairy processing conditions. This work provides insights into how phages may be tested and used to develop biosanitizers that are effective but are not harmful to the environment or human health.

Isolation, Characterization, and Genome Analysis of a Novel Bacteriophage, Escherichia Phage vB_EcoM-4HA13, Representing a New Phage Genus in the Novel Phage Family Chaseviridae.

Shiga toxin-producing Escherichia coli (STEC) is one of the leading causes of foodborne illnesses in North America and can lead to severe symptoms, with increased fatality risk for young children. While E. coli O157:H7 remains the dominant STEC serotype associated with foodborne outbreaks, there has been an increasing number of non-O157 STEC outbreaks in recent years. For the food industry, lytic bacteriophages offer an organic, self-limiting alternative to pathogen reduction-one that could replace or reduce the use of chemical and physical food processing methods. From EHEC-enriched sewage, we isolated a novel bacteriophage, vB_EcoM-4HA13 (4HA13). Phenotypic characterizations revealed 4HA13 to possess a myoviral morphotype, with a high specificity to non-motile O111 serotype, and a long latent period (90 min). Through genomic analyses, this 52,401-bp dsDNA phage was found to contain 81 CDS, but no detectable presence of antibiotic resistance, integrase, or virulence genes. A BLASTn search for each of the identified 81 CDS yielded homologues with low levels of similarity. Comparison of RNA polymerase and terminase large subunit amino acid sequences led to the proposal and acceptance of a new bacteriophage family, Chaseviridae, with 4HA13 representing a new species and genus. The discovery of this phage has broadened our current knowledge of bacteriophage diversity.

Complete Genome Sequence of a Jumbo Bacteriophage, Escherichia Phage vB_EcoM_EC001.

Here, we report the genome sequence of a jumbo Escherichia phage vB_EcoM_EC001, a myovirus isolated from primary sludge using enterohemorrhagic Escherichia coli O157:H7. The genome is 240,200 bp long and has 270 predicted coding sequences, including a tryptophanyl tRNA gene. It belongs to genus Seoulvirus.

PHIDA: A High Throughput Turbidimetric Data Analytic Tool to Compare Host Range Profiles of Bacteriophages Isolated Using Different Enrichment Methods.

Bacteriophages are viruses that infect bacteria and are present in niches where bacteria thrive. In recent years, the suggested application areas of lytic bacteriophage have been expanded to include therapy, biocontrol, detection, sanitation, and remediation. However, phage application is constrained by the phage's host range-the range of bacterial hosts sensitive to the phage and the degree of infection. Even though phage isolation and enrichment techniques are straightforward protocols, the correlation between the enrichment technique and host range profile has not been evaluated. Agar-based methods such as spotting assay and efficiency of plaquing (EOP) are the most used methods to determine the phage host range. These methods, aside from being labor intensive, can lead to subjective and incomplete results as they rely on qualitative observations of the lysis/plaques, do not reflect the lytic activity in liquid culture, and can overestimate the host range. In this study, phages against three bacterial genera were isolated using three different enrichment methods. Host range profiles of the isolated phages were quantitatively determined using a high throughput turbidimetric protocol and the data were analyzed with an accessible analytic tool "PHIDA". Using this tool, the host ranges of 9 Listeria, 14 Salmonella, and 20 Pseudomonas phages isolated with different enrichment methods were quantitatively compared. A high variability in the host range index (HRi) ranging from 0.86-0.63, 0.07-0.24, and 0.00-0.67 for Listeria, Salmonella, and Pseudomonas phages, respectively, was observed. Overall, no direct correlation was found between the phage host range breadth and the enrichment method in any of the three target bacterial genera. The high throughput method and analytics tool developed in this study can be easily adapted to any phage study and can provide a consensus for phage host range determination.