A Novel Bacteriophage Targeting Cronobacter sakazakii Is a Potential Biocontrol Agent in Foods.

Cronobacter sakazakii is an important pathogen that causes high mortality in infants. Due to its occasional antibiotic resistance, a bacteriophage approach might be an alternative effective method for the control of this pathogen. To develop a novel biocontrol agent using bacteriophages, the C. sakazakii-infecting phage CR5 was newly isolated and characterized. Interestingly, this phage exhibited efficient and relatively durable host lysis activity. In addition, a specific gene knockout study and subsequent complementation experiment revealed that this phage infected the host strain using the bacterial flagella. The complete genome sequence analysis of phage CR5 showed that its genome contains 223,989 bp of DNA, including 231 predicted open reading frames (ORFs), and it has a G+C content of 50.06%. The annotated ORFs were classified into six functional groups (structure, packaging, host lysis, DNA manipulation, transcription, and additional functions); no gene was found to be related to virulence or toxin or lysogen formation, but >80% of the predicted ORFs are unknown. In addition, a phage proteomic analysis using SDS-PAGE and matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) revealed that seven phage structural proteins are indeed present, supporting the ORF predictions. To verify the potential of this phage as a biocontrol agent against C. sakazakii, it was added to infant formula milk contaminated with a C. sakazakii clinical isolate or food isolate, revealing complete growth inhibition of the isolates by the addition of phage CR5 when the multiplicity of infection (MOI) was 10(5).

Core lipopolysaccharide-specific phage SSU5 as an Auxiliary Component of a Phage Cocktail for Salmonella biocontrol.

Salmonella spp. are among the major food-borne pathogens that cause mild diarrhea to severe bacteremia. The use of bacteriophages to control various food-borne pathogens, including Salmonella, has emerged as a promising alternative to traditional chemotherapy. We isolated the Siphoviridae family phage SSU5, which can infect only rough strains of Salmonella. The blocking of SSU5 adsorption by periodate treatment of host Salmonella cells and spotting and adsorption assays with mutants that contain various truncations in their lipopolysaccharide (LPS) cores revealed that the outer core region of the LPS is a receptor of SSU5. SSU5 could infect O-antigen (O-Ag)-deficient Salmonella mutants that developed by challenging of O-Ag-specific phages, and consequently, it delayed the emergence of the phage-resistant Salmonella population in broth culture when treated together with phages using O-Ag as a receptor. Therefore, these results suggested that phage SSU5 would be a promising auxiliary component of a phage cocktail to control rough strains of Salmonella enterica serovar Typhimurium, which might emerge as resistant mutants upon infection by phages using O-Ag as a receptor.

A Nitrogen Metabolic Enzyme Provides Salmonella Fitness Advantage by Promoting Utilization of Microbiota-Derived Carbon Source.

Microbes support their growth in vertebrate hosts by exploiting a large variety of dietary components as nutrients, which determines the composition of gut microbiota. A pathogen Salmonella expands by utilizing 1,2-propanediol, a microbiota-fermented product, during mucosal inflammation. However, it remains largely unknown how the pathogen decides which nutrient to consume from the complex mixture in the gut. Here, we show that Salmonella enterica serovar Typhimurium utilizes 1,2-propanediol by EIIANtr (a nitrogen-metabolic PTS component implicated in virulence)-mediated regulation of the pdu operon, thereby expanding in the murine intestine. Propionyl-CoA, a metabolic intermediate produced by 1,2-propanediol catabolism, elevates EIIANtr protein amounts, entailing positive feedback, thereby boosting the 1,2-propanediol-utilization process. EIIANtr promotes pdu expression by enhancing glutathione synthesis. CRP (cAMP receptor protein) induces pdu genes by increasing EIIANtr expression in response to glucose availability. Notably, EIIANtr-mediated 1,2-propanediol-utilization conferred a growth benefit even under high glucose conditions which reduces CRP activity. The EIIANtr-mediated activation is likely conserved in pathogenic enterobacteria including Escherichia coli. Collectively, our findings suggest that Salmonella promotes its fitness by precisely modulating the utilization system for microbiota-derived carbon source. They also suggest that Salmonella may integrate signals, processed via EIIANtr, into its metabolic program as well as virulence circuit.