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.

Bacteriophage Taxonomy: A Continually Evolving Discipline.

While taxonomy is an often underappreciated branch of science, it serves very important roles. Bacteriophage taxonomy has evolved from a discipline based mainly on morphology, characterized by the work of David Bradley and Hans-Wolfgang Ackermann, to the sequence-based approach that is taken today. The Bacterial Viruses Subcommittee of the International Committee on Taxonomy of Viruses (ICTV) takes a holistic approach to classifying prokaryote viruses by measuring overall DNA and protein similarity and phylogeny before making decisions about the taxonomic position of a new virus. The huge number of complete genomes being deposited with the National Center for Biotechnology Information (NCBI) and other public databases has resulted in a reassessment of the taxonomy of many viruses, and the future will see the introduction of new viral families and higher orders.

Multireceptor phage cocktail against Salmonella enterica to circumvent phage resistance.

Non-Typhoidal Salmonella (NTS) is one of the most common food-borne pathogens worldwide, with poultry products being the major vehicle for pathogenesis in humans. The use of bacteriophage (phage) cocktails has recently emerged as a novel approach to enhancing food safety. Here, a multireceptor Salmonella phage cocktail of five phages was developed and characterized. The cocktail targets four receptors: O-antigen, BtuB, OmpC, and rough Salmonella strains. Structural analysis indicated that all five phages belong to unique families or subfamilies. Genome analysis of four of the phages showed they were devoid of known virulence or antimicrobial resistance factors, indicating enhanced safety. The phage cocktail broad antimicrobial spectrum against Salmonella, significantly inhibiting the growth of all 66 strains from 20 serovars tested in vitro. The average bacteriophage insensitive mutant (BIM) frequency against the cocktail was 6.22 × 10-6 in S. Enteritidis, significantly lower than that of each of the individual phages. The phage cocktail reduced the load of Salmonella in inoculated chicken skin by 3.5 log10 CFU/cm2 after 48 h at 25°C and 15°C, and 2.5 log10 CFU/cm2 at 4°C. A genome-wide transduction assay was used to investigate the transduction efficiency of the selected phage in the cocktail. Only one of the four phages tested could transduce the kanamycin resistance cassette at a low frequency comparable to that of phage P22. Overall, the results support the potential of cocktails of phage that each target different host receptors to achieve complementary infection and reduce the emergence of phage resistance during biocontrol applications.