Phenotypic Characterization And Comparative Genomics Analysis Of Listeriaphages
Listeria monocytogenes is a foodborne pathogen that is widespread in natural and urban environments. L. monocytogenes is also commonly found in food processing facilities where specific strains or subtypes have been shown to persist over time. Listeriaphages are phages that can specifically infect L. monocytogenes and other Listeria spp. Phages have been approved for use as a biocontrol agent for this pathogen; however knowledge on its efficacy against diverse L. monocytogenes, particularly from food associated environments, is still limited. We developed a collection of diverse phages for further studies which included (i) phenotypic and genotypic characterization of listeriaphages isolated from dairy farms, (ii) evaluation of phage susceptibility and development of phage cocktails against persistent subtypes from food-associated environments, and (iii) whole genome sequencing and comparative genomics of selected phages. Over 100 listeriaphages were isolated from silage samples collected over the course of 1.5 years on two dairy farms. Initial phenotypic and genotypic characterization of phages from this collection revealed considerable host range (9 lysis groups) and genomic diversity (genome sizes of 25- 140 kb). Among the 9 major Listeria serotypes used to determine the host range, the serotype 3c strain was found to be highly resistant to phages while serotype 4 strains were the most susceptible to phages. Variation in phage susceptibility (4.6-95.4%) was observed among different persistent isolates of multiple ribotypes from a food processing facility. While phage cocktails could temporally reduce the bacterial populations of some subtypes, others were unaffected by phage treatment. L. monocytogenes isolates also rapidly developed phage-resistance characteristics in laboratory challenge studies. Whole genome sequencing of 10 listeriaphages revealed considerable genomic diversity of listeriaphages on dairy farms. The phage genome sizes could be classified into 3 ranges: small (36-38 kb; n=3), mid-sized (64-67 kb; n=4), and large (133-135 kb; n=3). All genomes were found to be organized into 3 functional modules: (i) DNA packaging and structural proteins; (ii) cell lysis; (iii) DNA replication, modification, and metabolism. Genomes of six newly sequenced phages appear to resemble three previously described listeriaphages. However, four phages showed no sequence homology to any bacteriophages in the NCBI databases, suggesting they are novel listeriaphages. Our data provide valuable information for further development of effective and suitable phage-based biocontrol agents and other applications.
Listeriaphage; Food processing facility; Biocontrol
Boor, Kathryn Jean; Tauer, Loren William
Food Science and Technology
Ph. D., Food Science and Technology
Doctor of Philosophy
dissertation or thesis