Natural Killers: Opportunities and Challenges for the Use of Bacteriophages in Microbial Food Safety from the One Health Perspective.

Ingestion of food or water contaminated with pathogenic bacteria may cause serious diseases. The One Health approach may help to ensure food safety by anticipating, preventing, detecting, and controlling diseases that spread between animals, humans, and the environment. This concept pays special attention to the increasing spread and dissemination of antibiotic-resistant bacteria, which are considered one of the most important environment-related human and animal health hazards. In this context, the development of innovative, versatile, and effective alternatives to control bacterial infections in order to assure comprehensive food microbial safety is becoming an urgent issue. Bacteriophages (phages), viruses of bacteria, have gained significance in the last years due to the request for new effective antimicrobials for the treatment of bacterial diseases, along with many other applications, including biotechnology and food safety. This manuscript reviews the application of phages in order to prevent food- and water-borne diseases from a One Health perspective. Regarding the necessary decrease in the use of antibiotics, results taken from the literature indicate that phages are also promising tools to help to address this issue. To assist future phage-based real applications, the pending issues and main challenges to be addressed shortly by future studies are also taken into account.

Campylobacter spp. prevalence and mitigation strategies in the broiler production chain.

This study aims to discuss the microbial ecology of the broiler gut environment, Campylobacter prevalence across the broiler production chain with a follow-up focus on a possible mitigation strategy, based on the use of bacteriophages. Scientific literature published from the last two decades was reviewed and data were collected to establish the ranges of Campylobacter loads from different samples. Results showed that the pathogen load in the sample is likely to increase from the different stages of the production chain. Contamination of water and feed represents the most notable source of contamination during the primary production, while cross-contamination of broiler carcasses, skin, and meat occurs during the slaughter, dressing, and processing via machinery, work surfaces, water, and air partially due to the leaking of contaminated feces from visceral rupture. Knowledge gaps were identified and included: a lack of studies detecting Campylobacter in broilers in most of the European countries over the last decade and a low number of studies determining the bacterial load in crates used to transport broilers to the slaughterhouse. Determining the prevalence of Campylobacter in the broiler industry will enable us to set critical control points to produce broiler flocks and meat products with a low risk of Campylobacter contamination.

Isolation, host specificity and genetic characterization of Campylobacter specific bacteriophages from poultry and swine sources.

The isolation and characterization of 304 Campylobacter specific bacteriophage isolates from broiler and swine sources is reported in this study. Genome size characterization determined by PFGE classified these isolates,called CAM1-CAM304, within the campylophages group II (n = 18) and group III (n = 286). Host range analyses showed a high host specificity and similar lytic spectrum among isolates of the same group. Campylophages of group II infected C. jejuni, C. coli and even a C. fetus strain whereas those of group III only infected C. jejuni strains. The most promising 59 campylophage candidates were selected according to their lytic activity and their genetic diversity was analyzed by RFLP using SmiI and HhaI endonucleases for group II and III campylophages, respectively. Moreover, RAPD-PCR technique was for the first time assessed in the genetic characterization of campylophages and it was shown to be effective only for those of group II. Bacteriophage isolates grouped in a same genotype displayed different host ranges, therefore, 13 campylophages of group II and eight of group III were differentiated considering all the approaches assayed. An in-depth analysis of these bacteriophages will be performed to confirm their promising potential for the biocontrol of Campylobacter within the farm to fork process.

Bacteriophage biocontrol to fight Listeria outbreaks in seafood.

Listeria monocytogenes is a well-known pathogen responsible for the severe foodborne disease listeriosis. The control of L. monocytogenes occurrence in seafood products and seafood processing environments is an important challenge for the seafood industry and the public health sector. However, bacteriophage biocontrol shows great potential to be used as safety control measure in seafood. This review provides an update on Listeria-specific bacteriophages, focusing on their application as a safe and natural strategy to prevent L. monocytogenes contamination and growth in seafood products and seafood processing environments. Furthermore, the main properties required from bacteriophages intended to be used as biocontrol tools are summarized and emerging strategies to overcome the current limitations are considered. Also, major aspects relevant for bacteriophage production at industrial scale, their access to the market, as well as the current regulatory status of bacteriophage-based solutions for Listeria biocontrol are discussed.

Efficient isolation of Campylobacter bacteriophages from chicken skin, analysis of several isolation protocols.

The application of Campylobacter specific bacteriophages appears as a promising food safety tool for the biocontrol of this pathogen in the poultry meat production chain. However, their isolation is a complicated challenge since their occurrence appears to be low. This work assessed the efficiency of seven protocols for recovering Campylobacter phages from chicken skin samples inoculated at phage loads from 5.0 × 101 to 5.0 × 106 PFU/g. The enrichment of chicken skin in selective Bolton broth containing target isolates was the most efficient procedure, showing a low detection limit of 5.0 × 101 PFU/g and high recovery rates of up to 560%. This method's effectiveness increased as phage concentration decreased, showing its suitability for phage isolation. When this method was applied to isolate new Campylobacter phages from retail chicken skin, a total of 280 phages were recovered achieving an isolation success rate of 257%. From the 109 samples 68 resulted phage positive (62%). Chicken skin could be, therefore, considered a rich source in Campylobacter phages. This method is a simple, reproducible and efficient approach for the successful isolation of both group II and III Campylobacter specific bacteriophages, which could be helpful for the enhancement of food safety by reducing this pathogen contamination in broiler meat.

Static and Continuous flow-through pulsed light technology for pesticide abatement in water.

Perspectives concerning pulsed light (PL) technology as a novel water decontamination treatment are overviewed in this work. Degradation of atrazine, malathion, chlorpyriphos-methyl and bromopropylate in aqueous solutions at different concentrations was performed employing static and continuous flow-through PL units. Results for both PL systems were compared in terms of efficacy of pesticides degradation and derived photoproducts formed. The pesticides degradation increased with the applied total fluence for both PL systems. In general, PL induced rapid degradation of all studied compounds although some differences were observed in the rate of degradation among them. The most intense treatment in the static unit (11J/cm2) yielded degradations between 60 and 85% whereas degradations between 74 and 93% were observed after exposure to slightly lower fluences (10J/cm2) in the dynamic unit. For both PL systems bromopropylate was the most affected compound. Photodegradation products arose from chemical reactions involving structural changes like dehalogenation, desulfuration, dealkylaytion and oxidation of the alkyl chains. Although further research is needed before an efficient application in full-scale systems, PL appears as a promising technology in order to reduce the presence of pesticides residues in process or waste waters, allowing thereby recirculation of these waters with the subsequent economic and environmental advantages.

Survival and growth of Listeria innocua treated by pulsed light technology: impact of post-treatment temperature and illumination conditions.

Inactivation of Listeria innocua by pulsed light (PL) was evaluated at different post-treatment temperature and illumination conditions. The impact of post-PL-treatment temperature on L. innocua culturability was evaluated for cells cultured at 37 °C (optimal growth temperature) and 4 °C (classical refrigerated food temperature). For both culture conditions, significant higher reductions (up to 3 log) were observed after post-PL-treatment temperature of 4 °C than of 37 °C. Contrarily, L. innocua culturability after PL treatment increased up to 2.2 log in presence of daylight illumination in comparison to dark storage. This photorepair mechanism was quickly activated reaching the maximum photoreactivation level after only 30 min of illumination. Moreover, photorepair capacity was rapidly reduced by increasing the time in darkness from PL treatment to samples illumination, being completely lost after time in darkness equal or greater than 5 h. According to these findings, the combination of PL with post-treatment temperature of 4 °C has a synergistic effect on the inactivation of L. innocua, whereas post-treatment daylight illumination has an antagonic effect on PL antimicrobial efficacy. Post-PL-treatment temperature and illumination conditions could be thereby considered important environmental factors to activate, inhibit or control the repair and/or growth of L. innocua survivors after PL treatment.

Sensitivity to pulsed light technology of several spoilage and pathogenic bacteria isolated from fish products.

The effect of pulsed light (PL) on the inactivation of six fish spoilage bacteria (Photobacterium phosphoreum, Serratia liquefaciens, Shewanella putrefaciens, Brochothrix thermosphacta, Pseudomonas group I, and Pseudomonas groups III and IV), six Listeria monocytogenes isolates from fish products, and one strain of Listeria innocua was evaluated. For all tested strains, grown at 4°C (temperature to process, store, and distribute convenient, lightly preserved fish products), the maximum detectable inactivation (6 to 6.5 log) was observed after treatments lower than 0.3 J/cm(2). The most sensitive bacteria were P. phosphoreum and S. liquefaciens, while both L. innocua and L. monocytogenes were the most resistant species to PL. Bacterial inactivation kinetics depended on their susceptibility to PL. Typically, sigmoidal curves were observed, with an initial shoulder followed by an exponential loss of cell culturability down to the maximum detectable inactivation. However, no initial shoulder was pointed out for the most sensitive species (P. phosphoreum and S. liquefaciens). Since the PL resistance of all tested Listeria strains (L. innocua and L. monocytogenes isolates) was similar, we propose the use of L. innocua as a surrogate for L. monocytogenes in PL technology validation.