Pathogens in the food chain - persistence, elimination and risk management

The project consists of several work packages with different activities. A number of bacterial strains have been adapted to food related stress. Survival of stressed pathogenic E. coli in fermented sausages and in digestion challenge models depended on recipe, strain and type of stress. Knowledge about factors which influence survival of human pathogens on leaf surfaces is important for developing strategies to prevent survival of the pathogens on fresh produce. Metagenomic studies on mapping of bacteria on lettuce leaves have been carried out. Survival of E. coli in biofilms with bacteria from lettuce leaves has been studied. E. coli was transported into the interior of leaves regardless of mildew infections. Gene expression studies have been performed under biofilm formation with and without E. coli present. The genomes of two biofilm formers have been sequenced. The CRISPR/Cas9 system for gene editing is under development for one of them (Pectobacterium). The use of UV light as a decontamination strategy in food production has been studied. Salmonella, E. coli and L. monocytogenes can be reduced when exposed to UV light on eggs and poultry. On poultry we have also shown decontamination and growth inhibition by use of organic acids and corresponding salts. Continuous UV irradiation, pulsed UV irradiation, organic salts and chlorine-based compounds have been shown to reduce L. monocytogenes on salmon. We also investigated alternative strategies such as use of steam and ozone for reduction of pathogens and microflora on meat. The food industry is under pressure to reducing the NaCl content in food. In vitro digestion using a digestion model indicated that broth with 5 versus 0.5 % NaCl significantly altered the survival of L. monocytogenes through the simulated human digestive system and by cell-culture assays simulating the intestinal epithelium. These experiments generate data for quantitative risk assessments and highlight potential effects of reducing the NaCl content in foods and its effect on L. monocytogenes survival in the digestive system. Visible light affects the life style of non-phototrophic bacteria. A global transcriptomic analysis revealed significant alteration in gene-expression of 82 % of the L. monocytogenes genome, encompassing 18 metabolic pathways. This study generates new hypotheses and emphasizes the importance of visible light as a potential bias factor when designing experiments. The study revealed inhibition of L. monocytogenes growth by light. Visible light might represent a novel strategy to reduce L. monocytogenes contamination in the food industry. A study on modelling of spreading of L. monocytogenes in products from salmon and poultry products has been carried out. A risk model based on realistic contamination, processing and storage conditions has been developed for calculating the number of cases of listeriosis from sushi. Growth models for Listeria in pasta salads have also been produced. Two studies on prevalence of shigatoxin 2a containing E. coli in cattle and in minced meat have been performed. Positive isolated have been further studied using whole genome sequencing. Studies on transfer of phages in food products during storage have been carried out. This increases knowledge about unwanted spreading in the environment. Close to 100 strains of E. coli O26:H11 have been sequenced to look at relatedness and possible correlations between uptake of Stx phages and pathogenicity. Both E. coli O103:H2 and E. coli O26:H11 were susceptible to shigatoxin phages and could develop into STEC. Investigations on survival of Stx phages on salad and in minced meat have been carried out. In addition, a new method using digital droplet PCR has been developed for the transfer studies. A study of prevalence of STEC in Norwegian mussels has been carried out, and one has also looked at different E. coli strains ability to form biofilm. We have also investigated to what extent enzyme-based washing solutions designed for removal of biofilm can contribute to a more efficient and environmentally friendly washing and disinfection in food production environments. The enzyme-based washing solutions were comparable to conventional chlor-alkali based washing solutions at removing of biofilms of L. monocytogenes from conveyor belts. Moreover, there have been performed studies on strategies for managing risk analysis and implementing food safety rules, especially regarding risk analyses of importance for trade. A study on the framework for risk analysis in Norway, Italy and Sweden has been carried out. Data for a comparative study on the strategies of Norwegian and Swedish food producing companies for reducing pathogens in food has been collected. Most important in this respect is the collection of information in Norway, Sweden and EU regarding management of antibiotic resistance. Some of the results were presented at the common arrangement Mattrygghetsdagen.

Forskningsprosjektet har bidratt til: Kompetanseoppbygging i institusjonene, økt samarbeid mellom institusjonene, økt samarbeid med eksterne samarbeidspartnere (forskning, forvaltning og industri), økt forståelsen rundt overlevelse og vekst av patogene mikroorganismer, utarbeidelse av risikomodeller, innføring av nye laboratorieteknikker og in vitro fordøyelsesmodeller, metoder for reduksjon av mikroorganismer i mat, publisering av vitenskapelige artikler, utdanning av PhD student og post doc, posterpris Food Micro 2018. For forvaltningen: Risikomodellene har betydning for risikovurderingene. Endrede retningslinjer for prøvetaking i Codex og EU. Kunnskap om regulative og organisatoriske rammeverk til å håndtere mattrygghetsutfordringene og utforming av organisatoriske rammeverk og forvaltningsstrategier for redusert antibiotikaresistens. For industri: Behandling av råvarer, prosessering og lagring for reduksjon av patogener i matproduksjon. Forbedrede kontrollrutiner for Listeria.

The food industry is constantly facing new microbiological challenges related to foodborne risks; foodborne illness, spoilage, withdrawals etc. More knowledge about food pathogens under real production conditions is needed, both to reduce the risk as much as possible, and to manage the residual risk. We want to address the behaviour of food pathogens during pretreatment, production and storage of foods by producing foods contaminated with food pathogens. Nofima is currently building a unique separate path ogen pilot plant unit where conventional and new food can be produced with pathogens in order to investigate the fate of pathogens under different production conditions. The investigation of behaviour of pathogens during food processing, packaging and sto rage will contribute to safer production both for small and large scale producers. We will primarily work with STEC and L. monocytogenes and food products of both animal and plant origin. Initially we will study the behaviour of STEC in fermented meats an d on leafy greens, and behaviour of L. monocytogenes during small scale cheese production. Other patogens and food products will be included later in the project after consultations with the reference group consisting of representatives from the food indu stries. Research questions to be answered in the project are listed above under objectives of project. The work will be organised in 9 workpackages (WPs). Development and characterization of stress adapted pathogenic strains are done in WP1. Strains inve stigated in WP1 will be used in the subsequent WPs. Intervention strategies to reduce survival and growth on raw materials (WP3), during food processing (WP4) and packaging and storage (WP5) will be studied. Mapping residual risk is done in (WP6) and chan ges in virulence through production is studied in WP7. Effects of sanitation processes on bacteria on production surfaces is studied in WP8, while managing strategies for reducing pathogens is the theme of WP9.