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There is limited knowledge of the characteristics, differences, and similarities of the bacterial composition in residential kitchens. Here, we report the microbiota of cleaning utensils (sponges and cloths) and five different surface samples in 74 households across five European countries.

To investigate the diversity, distribution, persistence, and prevalence of stress survival and resistance genes of Listeria monocytogenes clones dominating in food processing environments in Norway, genome sequences from 769 L. monocytogenes isolates from food industry environments, foods, and raw materials (512 of which were sequenced in the present study) were subjected to whole-genome multilocus sequence typing (wgMLST), single-nucleotide polymorphism (SNP), and comparative genomic analyses. The data set comprised isolates from nine meat and six salmon processing facilities in Norway collected over a period of three decades. The most prevalent clonal complex (CC) was CC121, found in 10 factories, followed by CC7, CC8, and CC9, found in 7 factories each. Overall, 72% of the isolates were classified as persistent, showing 20 or fewer wgMLST allelic differences toward an isolate found in the same factory in a different calendar year. Moreover, over half of the isolates (56%) showed this level of genetic similarity toward an isolate collected from a different food processing facility. These were designated as pervasive strains, defined as clusters with the same level of genetic similarity as persistent strains but isolated from different factories. The prevalence of genetic determinants associated with increased survival in food processing environments, including heavy metal and biocide resistance determinants, stress response genes, and

Whole genome sequencing (WGS) of foodborne pathogens such as Listeria monocytogenes is globally on the rise in the food industry. It provides an improvement for proactive surveillance and source-tracking and allows in-depth genetic characterization of the pathogen. In the present study, the virulence gene profile including 99 virulence genes of 767 L. monocytogenes isolates from the Norwegian meat and salmon processing industry was characterized. The isolate collection comprised 28 clonal complexes (CCs) that occur globally. We additionally determined the in vitro virulence potential for 13 major CCs in human intestinal epithelial Caco2 cells using cocktails of three to six representative isolates. Our aim was to test whether the virulence potential could be predicted from the virulence gene profiles to estimate the application potential of WGS in risk assessment in the food industry. The virulence gene profiles were highly conserved within the individual CCs and similar among phylogenetically closely related CCs. We observed a CC-associated distribution of accessory virulence genes in addition to different length polymorphisms. Furthermore, we detected different premature stop codons (PMSC) in the inlA gene, which were mainly present in CC9, CC121 and CC5 isolates. Accordingly, CC9 and CC5 were unable to invade Caco2 cells, whereas CC121 showed moderate virulence potential due to the presence of an isolate harboring full-length inlA. The highest invasion was observed for CC403 and CC415, potentially due to the presence of accessory virulence genes. We demonstrated that CC14, which harbored full-length inlA, was unable to invade Caco2 cells due to a low inlA gene expression. Reconstruction of inlA in CC9 and CC121 isolates showed that without the presence of InlA on the cell wall (as detected in the CC9 isolates), invasion into host cells failed. Our study showed that predicting the virulence potential based on genetic virulence profiles provides valuable information for risk assessment in the food industry but also has its limitations. The mere presence of a full-length inlA gene is not sufficient for virulence, but gene expression and the presence of the protein on the cell wall is required for the successful invasion of L. monocytogenes into host cells. Moreover, hypovirulent CCs like CC121 were among the most abundant human clinical isolates in Norway despite harboring a PMSC mutation in the inlA gene. In conclusion, our study highlights that combining genotypic and phenotypic data is of great importance to improve the informative value of applying WGS in the food industry.

To investigate the diversity, distribution, persistence, and prevalence of stress survival and resistance genes ofListeria monocytogenesclones dominating in food processing environments in Norway, genome sequences from 769L. monocytogenesisolates from food industry environments, foods, and raw materials (of which 512 were sequenced in the present study) were subjected to wgMLST, SNP, and comparative genomic analyses. The dataset comprised isolates from nine meat and six salmon processing facilities in Norway collected over a period of three decades. The most prevalent clonal complex (CC) was CC121, found in ten factories, followed by CC7, CC8, and CC9, found in seven factories each. Overall, 72% of the isolates were classified as persistent, showing 20 or fewer wgMLST allelic differences towards an isolate found in the same factory in a different calendar year. Moreover, over half of the isolates (56%) showed this level of genetic similarity towards an isolate collected from a different food processing facility. These were designated as pervasive strains, defined as clusters with the same level of genetic similarity as persistent strains but isolated from different factories. The prevalence of genetic determinants associated with increased survival in food processing environments, including heavy metal and biocide resistance determinants, stress response genes andinlAtruncation mutations, showed a highly significant increase among pervasive isolates, but not among persistent isolates. Furthermore, these genes were significantly more prevalent among the isolates from food processing environments compared to in isolates from natural and rural environments (n=218) and clinical isolates (n=111) from Norway.ImportanceListeria monocytogenescan persist in food processing environments for months to decades and spread through the food system by e.g., contaminated raw materials. Knowledge about the distribution and diversity ofL. monocytogenesis of importance in outbreak investigations and essential to effectively track and control this pathogen in the food system. The current study presents a comprehensive overview of the prevalence of persistent clones and of the diversity ofL. monocytogenesin Norwegian food processing facilities. The results demonstrate extensive spread of highly similar strains throughout the Norwegian food system, in that 56% of the 769 collected isolates from food processing factories belonged to clusters ofL. monocytogenesidentified in more than one facility. These strains were associated with an overall increase in the prevalence of plasmids and determinants of heavy metal and biocide resistance as well as other genetic elements associated with stress survival mechanisms and persistence.

Aims The purpose of the work was to investigate bacterial levels and diversity as well as survival of Salmonella in used dish washing sponges and brushes and identify consumer practices that can potentially explain bacterial status of these items. Methods and Results Used washing up utensils were collected from consumers. The bacterial numbers (TVC) were very variable with an extremely high median level (10.3 log cfu/item) in Portuguese sponges and lower levels in Norwegian items (7.3 and 7.0 cfu/item for sponges and brushes). No self-reported practices or household composition could explain differences found in TVC levels among the collected sponges. Lower mean TVC levels were found in unworn brushes and brushes regularly cleaned with soap, but the differences were modest (1.5 log or less). A common set of bacteria was found in brushes and sponges, dominated by Acinetobacter, Chryseobacterium, Enhydrobacter, Enterobacteriaceae and Pseudomonas. There was no difference in TVC or bacterial diversity between conventional and antimicrobial sponges containing silver after 4 weeks of use. For used brushes inoculated with Salmonella and allowed to dry overnight, a significant reduction in Salmonella numbers was observed. No reduction was observed for brushes stored in humid conditions (in a plastic bag) or for sponges regardless of storing conditions. Conclusions Overall, lower bacterial levels were observed in used brushes than in sponges, and Salmonella died more rapidly in brushes. A common set of non-pathogenic bacteria dominated in brushes and sponges. Significance and Impact of Study The study demonstrates that the use of brushes may be more hygienic than the use of sponges.

In this study, we addressed different aspects regarding the implementation of quasimetagenomic sequencing as a hybrid surveillance method in combination with enrichment for early detection of Listeria monocytogenes in the food industry. Different experimental enrichment cultures were used, comprising seven L. monocytogenes strains of different sequence types (STs), with and without a background microbiota community. To assess whether the proportions of the different STs changed over time during enrichment, the growth and population dynamics were assessed using dapE colony sequencing and dapE and 16S rRNA amplicon sequencing. There was a tendency of some STs to have a higher relative abundance during the late stage of enrichment when L. monocytogenes was enriched without background microbiota. When coenriched with background microbiota, the population dynamics of the different STs was more consistent over time. To evaluate the earliest possible time point during enrichment that allows the detection of L. monocytogenes and at the same time the generation of genetic information that enables an estimation regarding the strain diversity in a sample, quasimetagenomic sequencing was performed early during enrichment in the presence of the background microbiota using Oxford Nanopore Technologies Flongle and Illumina MiSeq sequencing. The application of multiple displacement amplification (MDA) enabled detection of L. monocytogenes (and the background microbiota) after only 4 h of enrichment using both applied sequencing approaches. The MiSeq sequencing data additionally enabled the prediction of cooccurring L. monocytogenes strains in the samples. IMPORTANCE We showed that a combination of a short primary enrichment combined with MDA and Nanopore sequencing can accelerate the traditional process of cultivation and identification of L. monocytogenes. The use of Illumina MiSeq sequencing additionally allowed us to predict the presence of cooccurring L. monocytogenes strains. Our results suggest quasimetagenomic sequencing is a valuable and promising hybrid surveillance tool for the food industry that enables faster identification of L. monocytogenes during early enrichment. Routine application of this approach could lead to more efficient and proactive actions in the food industry that prevent contamination and subsequent product recalls and food destruction, economic and reputational losses, and human listeriosis cases.

Pseudomonas is ubiquitous in nature and a predominant genus in many foods and food processing environments, where it primarily represents major food spoilage organisms. The food chain has also been reported to be a potential reservoir of antibiotic-resistant Pseudomonas. The purpose of the current study was to determine the occurrence of antibiotic resistance in psychrotrophic Pseudomonas spp. collected over a time span of 26 years from retail chicken in Norway and characterize their genetic diversity, phylogenetic distribution and resistance genes through whole-genome sequence analyses. Among the 325 confirmed Pseudomonas spp. isolates by 16S rRNA gene sequencing, antibiotic susceptibility profiles of 175 isolates to 12 antibiotics were determined. A subset of 31 isolates being resistant to &ge, 3 antibiotics were whole-genome sequenced. The isolates were dominated by species of the P. fluorescens lineage. Isolates susceptible to all antibiotics or resistant to &ge, 3 antibiotics comprised 20.6% and 24.1%, respectively. The most common resistance was to aztreonam (72.6%), colistin (30.2%), imipenem (25.6%) and meropenem (12.6%). Resistance properties appeared relatively stable over the 26-year study period but with taxa-specific differences. Whole-genome sequencing showed high genome variability, where isolates resistant to &ge, 3 antibiotics belonged to seven species. A single metallo-betalactmase gene (cphA) was detected, though intrinsic resistance determinants dominated, including resistance&ndash, nodulation (RND), ATP-binding cassette (ABC) and small multidrug resistance (Smr) efflux pumps. This study provides further knowledge on the distribution of psychrotrophic Pseudomonas spp. in chicken meat and their antibiotic resistance properties. Further monitoring should be encouraged to determine food as a source of antibiotic resistance and maintain the overall favorable situation with regard to antibiotic resistance in the Norwegian food chain.

Pseudomonas is ubiquitous in nature and a predominant genus in many foods and food processing environments, where it primarily represents major food spoilage organisms. The food chain has also been reported to be a potential reservoir of antibiotic-resistant Pseudomonas. The purpose of the current study was to determine the occurrence of antibiotic resistance in psychrotrophic Pseudomonas spp. collected over a time span of 26 years from retail chicken in Norway and characterize their genetic diversity, phylogenetic distribution and resistance genes through whole-genome sequence analyses. Among the 325 confirmed Pseudomonas spp. isolates by 16S rRNA gene sequencing, antibiotic susceptibility profiles of 175 isolates to 12 antibiotics were determined. A subset of 31 isolates being resistant to ≥3 antibiotics were whole-genome sequenced. The isolates were dominated by species of the P. fluorescens lineage. Isolates susceptible to all antibiotics or resistant to ≥3 antibiotics comprised 20.6% and 24.1%, respectively. The most common resistance was to aztreonam (72.6%), colistin (30.2%), imipenem (25.6%) and meropenem (12.6%). Resistance properties appeared relatively stable over the 26-year study period but with taxa-specific differences. Whole-genome sequencing showed high genome variability, where isolates resistant to ≥3 antibiotics belonged to seven species. A single metallo-betalactmase gene (cphA) was detected, though intrinsic resistance determinants dominated, including resistance–nodulation (RND), ATP-binding cassette (ABC) and small multidrug resistance (Smr) efflux pumps. This study provides further knowledge on the distribution of psychrotrophic Pseudomonas spp. in chicken meat and their antibiotic resistance properties. Further monitoring should be encouraged to determine food as a source of antibiotic resistance and maintain the overall favorable situation with regard to antibiotic resistance in the Norwegian food chain.

Sponges are frequently used in kitchens and have been shown to harbor large numbers of bacteria, occasionally also pathogens. Less is known about kitchen brushes regarding usage and presence of bacteria. In the present study, the use of sponges and brushes was studied in a survey among 9966 European consumers in ten countries, and growth and survival of bacteria in sponges and brushes were examined in laboratory experiments. Sponges were the preferred hand-cleaning utensils for washing-up in the majority of countries, while brushes were most frequently used in Denmark and Norway. Consumers mostly change their sponges at regular times, but also sensory cues (looks dirty, smelly, slimy) and usage occurrences such as wiping up meat juices may trigger replacement. Besides cleaning the dishes, over a quarter of the dish brush users also use it to clean a chopping board after soilage from chicken meat juices. The water uptake and drying rate varied considerably, both between different sponges and between brushes and sponges, where brushes dried fastest. Campylobacter survived one day in all sponges and Salmonella more than seven days in two of three types of sponges. In the type of sponge that dried slowest, Salmonella grew on the first day and was always found in higher levels than in the other sponges. Non-pathogenic bacteria grew in the sponges and reached levels around 9 log CFU/sponge. In brushes all types of bacteria died over time. Campylobacter and Salmonella were reduced by more than 2.5 log to below the detection limit after one and three days, respectively. Bacteriota studies revealed a tendency for a dominance by Gram-negative bacteria and a shift to high relative prevalence of Pseudomonas over time in sponges. Both enumeration by agar plating and bacteriota analysis confirmed that the pathogens were in a minority compared to the other bacteria. Treatments of sponges and brushes with chlorine, boiling or in the dishwasher were effective to reduce Salmonella. We conclude that brushes are more hygienic than sponges and that their use should be encouraged. Contaminated sponges or brushes should be replaced or cleaned when they may have been in contact with pathogenic microorganisms, e.g. used on raw food spills. Cleaning of sponges and brushes with chlorine, boiling or dishwasher may be a safe alternative to replacing them with new ones. Dishwashing sponges and brushes: Consumer practices and bacterial growth and survival

Fresh Atlantic salmon (Salmo salar) represents a healthy, nutritious food with global distribution and increasing consumption and economic value. Contaminating Listeria monocytogenes in fresh salmon represents a health hazard to consumers, is linked to extensive product recalls and is a major challenge for salmon processors. Verdad N6, a commercially available buffered vinegar, was evaluated as a treatment for raw salmon fillets either alone or in combination with the antimicrobial peptide nisin, with regard to anti-listerial effects under processing and storage, and influence on sensory quality and background microbiota. Salmon fillets were surface contaminated with L. monocytogenes and immersed in solutions of Verdad N6 or treated with nisin or a combination of these two treatments. Levels of L. monocytogenes were determined during vacuum-pack refrigerated storage. The use of Verdad N6 resulted in increased lag times and substantially reduced growth of L. monocytogenes. The inhibitory effects were dependent on Verdad N6 levels, immersion time, and storage time and temperature. A 5 s immersion in 10% Verdad N6 solution at 4 °C reduced growth of L. monocytogenes from log 2.8 to log 1 after 12 days of storage. Nisin (0.2–1 ppm) had listericidal effects up to 1 log but did not inhibit regrowth when used alone. Appropriate combinations of Verdad N6 and nisin led to L. monocytogenes levels no higher after 12 days of storage than the initial levels. The inhibitory effects were markedly lower at 7 °C than at 4 °C. Salmon with Verdad N6 showed reduced levels of total counts during storage indicating a longer shelf-life, and a shift in the dominating bacteria with reduced and increased relative levels of Enterobacteriaceae and lactic acid bacteria, respectively. Sensory analyses of raw and cooked Verdad N6 treated a non-treated salmon resulted in small differences. In summary, Verdad N6 is an option for production of high-quality raw salmon with increased shelf-life and enhanced food safety through its Listeria inhibiting effects. The application of Verdad N6 in combination with nisin treatment can further reduce the listeria-risks of these products.

Improved quality control and prolonged shelf life are important actions in preventing food waste. To get an overview of the bacterial diversity of fillets from live stored mature Atlantic cod, bacterial isolates were identified before and after storage (air and vacuum) and freezing/thawing. Based on the load of dominating bacteria, the effect of different packaging methods and a short freezing/thawing process on prolonged shelf-life was evaluated (total viable counts, bacteriota, sensory attributes, and volatile components). Hand filleted (strict hygiene) cod fillets had a low initial bacterial load dominated by the spoilage organism Photobacterium, whereas industrially produced fillets had higher bacterial loads and diversity (Pseudomonas, Arthrobacter, Psychrobacter, Shewanella). The identified bacteria after storage in vacuum or air were similar to the initially identified bacteria. Bacteriota analysis showed that a short time freezing/thawing process reduced Photobacterium while modified atmosphere packaging (MAP, 60%CO2/40%O2 or 60%CO2/40%N2) inhibited the growth of important spoilage bacteria (Photobacterium, Shewanella, Pseudomonas) and allowed the growth of Carnobacterium/Carnobacteriaceae and Acinetobacter. Despite being dominated by Photobacterium, fresh fillets stored in MAP 60%CO2/40%N2 demonstrated better sensory quality after 13 days of storage than fillets stored in MAP 60%CO2/40%O2 (dominated by Carnobacterium/Carnobacteriaceae). Carnobacterium spp. or other members of Carnobacteriaceae may therefore be potential spoilage organisms in cod when other spoilage bacteria are reduced or inhibited.

The processing environment of salmon processing plants represents a potential major source of bacteria causing spoilage of fresh salmon. In this study, we have identified major contamination routes of important spoilage associated species within the genera Pseudomonas, Shewanella and Photobacterium in pre-rigor processing of salmon. Bacterial counts and culture-independent 16S rRNA gene analysis on salmon fillet from seven processing plants showed higher levels of Pseudomonas spp. and Shewanella spp. in industrially processed fillets compared to salmon processed under strict hygienic conditions. Higher levels of Pseudomonas spp. and Shewanella spp. were found on fillets produced early on the production day compared to later processed fillets. The levels of Photobacterium spp. were not dependent on the processing method or time of processing. In follow-up studies of two plants, bacterial isolates (n=2101) from the in-plant processing environments (sanitized equipment/machines and seawater) and from salmon collected at different sites in the production were identified by partial 16S rRNA gene sequencing. Pseudomonas spp. dominated in equipment/machines after sanitation with 72 and 91% of samples from the two plants being Pseudomonas-positive. The phylogenetic analyses, based on partial 16S rRNA gene sequencing, showed 48 unique sequence profiles of Pseudomonas of which two were dominant. Only six profiles were found on both machines and in fillets in both plants. Shewanella spp. were found on machines after sanitation in the slaughter department while Photobacterium spp. were not detected after sanitation in any parts of the plants. Shewanella spp. and Photobacterium spp. were found on salmon in the slaughter departments. Shewanella was frequently present in seawater tanks used for bleeding/short term storage.In conclusion, this study provides new knowledge on the processing environment as a source of contamination of salmon fillets with Pseudomonas spp. and Shewanella spp., while Photobacterium spp. most likely originate from the live fish and seawater. The study show that strict hygiene during processing is a prerequisite for optimal shelf life of salmon fillets and that about 90% reductions in the initial levels of bacteria on salmon fillets can be obtained using optimal hygienic conditions.

The purpose of this study was to explore the microbiota of Norwegian fermented fish (rakfisk), a traditional product popular in the Norwegian market. Brine samples, collected from six producers during two subsequent years, were used. The producers applied different salt concentrations (between 3.8% and 7.2% NaCl), ripening temperatures (between 3.5 and 7.5 &deg, C), fish species (trout or char), and fish upbringing (wild trout, on-shore farmed trout or char, and off-shore farmed char). The microbiota in the brine during the ripening process was mainly characterized by DNA-based, culture-independent methods. In total, 1710 samples were processed and of these 1342 were used for the final analysis. The microbiota was dominated by Gammaproteobacteria and Bacilli with the largest variance between samples associated with the genera Psychrobacter and Lactobacillus. The variance in the material was mainly determined by the origin of the samples, i.e., the different producers. The microbiota from the individual producers was to a large extent reproducible from one year to the next and appeared to be determined by the relatively small differences in the salinity and the ripening temperature. This is the first study exploring the microbiota in rakfisk brine and it provides insights into environmental factors affecting the rakfisk ecosystems.

Bacterial growth, microbiota and sensory quality of cod loins (farmed Atlantic cod, Gadus morhua) was studied during 15 days of storage (2 °C), packaged in vacuum or in modified atmosphere package (MAP, 60% CO2/40% N2, g/p ratio 1.6/1), with a CO2 emitter pad or a liquid absorbent pad. The objective was to investigate how packaging with CO2 emitter can prolong shelf life in vacuum, and in low headspace MAP. Sensory analysis showed that the initial freshness was better preserved by adding a CO2 emitter in both vacuum and MAP. The MAP had equal antibacterial effect compared to vacuum added CO2 emitter. 16S rDNA sequencing showed that CO2 emitter did not alter the dominant bacterial composition and that Photobacterium sp. dominated all packaging methods. Vacuum packaged samples past shelf life after 7 days of storage, vacuum added a CO2 emitter and MAP after 9 days, and MAP added a CO2 emitter after 13 days.

This study investigated the bacterial dynamics along the beef chain for clean and dirty cattle in the slaughter and processing lines, using classic quantitative methods and molecular analyses. In addition, the Norwegian national guidelines for Good Hygiene Practices in Norway were evaluated. In these guidelines, cattle presented for slaughter are categorised according to hide cleanliness, resulting in separate processing lines for meat from very dirty animals and reduced prices to farmers. The study was conducted in two commercial abattoirs in Norway. Two groups were compared; 40 visually clean cattle and 40 visually dirty cattle presented for slaughter, with 20 from each group at each abattoir. The same animals were sampled at five sampling sites: hides, carcass surfaces after dehiding, just before chilling, after chilling, and meat trimmings. Meat trimmings were sampled in only one abattoir. Three hundred and sixty samples were collected by swabbing 100 cm(2) of the brisket area at the first four sampling sites, and sampling 200 g of meat trimmings at the fifth site. The results showed that the hides of dirty cattle had more Enterobacteriaceae and higher Aerobic Plate Counts (APC) than visually clean cattle (P

Listeria monocytogenes is a foodborne pathogen that causes the often-fatal disease listeriosis. We present here the complete genome sequences of six L. monocytogenes isolates of sequence type 9 (ST9) collected from two different meat processing facilities in Norway. The genomes were assembled using Illumina and Nanopore sequencing data.

Listeria monocytogenes is a foodborne pathogen that causes the often-fatal disease listeriosis. We present here the complete genome sequences of six L. monocytogenes isolates of sequence type 9 (ST9) collected from two different meat processing facilities in Norway. The genomes were assembled using Illumina and Nanopore sequencing data.

Background Although diet is known to have a major modulatory influence on gut microbiota, knowledge of the specific roles of particular vitamins, minerals, and other nutrients is limited. Modulation of the composition of the microbiota in pregnant women is especially important as maternal microbes are transferred during delivery and initiate the colonization process in the infant. We studied the associations between intake of specific dietary nutrients during pregnancy and gut microbiota composition. Methods Utilizing the Norwegian NoMIC cohort, we examined the relations between intakes of 28 dietary macro- and micronutrients during pregnancy, derived from food frequency questionnaires administered to 60 women in the second trimester, and observed taxonomic differences in their gut microbiota four days after delivery (assessed through Illumina 16S rRNA amplicon analysis). Results Higher dietary intakes of fat-soluble vitamins, especially vitamin D, were associated with reduced microbial alpha diversity (p value

The effect of extrusion of barley and oat on the fecal microbiota and the formation of SCFA was evaluated using growing pigs as model system. The pigs were fed a diet containing either whole grain barley (BU), oat groat (OU), or their respective extruded samples (BE and OE). 454 pyrosequencing showed that the fecal microbiota of growing pigs was affected by both extrusion and grain type. Extruded grain resulted in lower bacterial diversity and enrichment in operational taxonomic units (OTUs) affiliated with members of the Streptococcus, Blautia and Bulleidia genera, while untreated grain showed enrichment in OTUs affiliated with members of the Bifidobacterium and Lactobacillus genera, and the butyrate-producing bacteria Butyricicoccus, Roseburia, Coprococcus and Pseudobutyrivibrio. Untreated grain resulted in a significant increase of n-butyric, i-valeric and n-valeric acid, which correlated with an increase of Bifidobacterium and Lactobacillus. This is the first study showing that cereal extrusion affects the microbiota composition and diversity towards a state generally thought to be less beneficial for health, as well as less amounts of beneficial butyric acid.

Foods naturally high in dietary fiber are generally considered to protect against development of colorectal cancer (CRC). However, the intrinsic effect of dietary fiber on intestinal carcinogenesis is unclear. We used azoxymethane (AOM) treated A/J Min/+ mice, which developed a significantly higher tumor load in the colon than in the small intestine, to compare the effects of dietary inulin (IN), cellulose (CE) or brewers spent grain (BSG) on intestinal tumorigenesis and cecal microbiota. Each fiber was tested at two dose levels, 5% and 15% (w/w) content of the AIN-93M diet. The microbiota was investigated by next-generation sequencing of the 16S rRNA gene (V4). We found that mice fed IN had approximately 50% lower colonic tumor load than mice fed CE or BSG (p

Stainless steel coupons are frequently used in biofilm studies in the laboratory, as this material is commonly used in the food industry. The coupons are attached to different surfaces to create a "natural" biofilm to be studied further in laboratory trials. However, little has been done to investigate how well the microbiota on such coupons represents the surrounding environment. The microbiota on sink wall surfaces and on new stainless steel coupons attached to the sink wall for 3 months in 8 domestic kitchen sinks was investigated by next-generation sequencing (MiSeq) of the 16S rRNA gene derived from DNA and RNA (cDNA), and by plating and identification of colonies. The mean number of colony-forming units was about 10-fold higher for coupons than sink surfaces, and more variation in bacterial counts between kitchens was seen on sink surfaces than coupons. The microbiota in the majority of biofilms was dominated by Moraxellaceae (genus Moraxella/Enhydrobacter) and Micrococcaceae (genus Kocuria). The results demonstrated that the variation in the microbiota was mainly due to differences between kitchens (38.2%), followed by the different nucleic acid template (DNA vs RNA) (10.8%), and that only 5.1% of the variation was a result of differences between coupons and sink surfaces. The microbiota variation between sink surfaces and coupons was smaller for samples based on their RNA than on their DNA. Overall, our results suggest that new stainless steel coupons are suited to model the dominating part of the natural microbiota of the surrounding environment and, furthermore, are suitable for different downstream studies.

Aim We have tested the effect of feed structure and feeding regime to prevent the spread of the zoonotic pathogen Campylobacter jejuni in broiler chicken flocks. Methods and Results Birds were offered two types of feed, control diet and a diet supplemented with 15% oat/barley hulls for structure. In addition, the birds were either fed ad libitum or intermittent. One bird in each treatment group was infected with a three-strain-mix of Camp. jejuni, and the spread of Camp. jejuni within the group was investigated. Feed structure increased the gizzard weight, delayed the spread of Camp. jejuni within the group and reduced the relative amount of Camp. jejuni in the caecum compared with the control diet. Conclusions Our results show that stimulating the bird's natural barriers is a novel and promising intervention strategy to reduce the spread of Camp. jejuni in chicken flocks. Significance and impact of the Study Preventing Camp. jejuni in broiler chicken flocks is essential to ensure food safety because this bacterium is transferred to chicken carcasses during the slaughter process and readily survive in unprocessed poultry products. We have evaluated a novel approach for stimulation of the bird's natural barriers in the upper digestive tract with promising results.

Two experiments were conducted to test the following hypothesis: exposing broiler chickens to coarse insoluble fiber in the diet will result in enhanced gizzard function and performance, improved adaptability to an intermittent feeding program, and an increase in the occurrence of reverse peristalsis. In experiment 1, 102 Ross 308 broiler chickens were either intermittently or ad libitum fed a basal diet, the basal diet diluted with 15% coarse hulls (consisting of equal weights of hulls from oats and barley), or the basal diet diluted with 15% of the same hulls finely ground in a 2 × 3 factorial arrangement with 17 individually caged birds per treatment. Birds fed ad libitum had access to feed continuously for 18 h/d, whereas those on intermittent feeding had restricted access to feed from 7 d of age. From 18 d of age, the restrictive-feeding program consisted of four 1-h meals and one 2-h meal per day. In experiment 2, 156 broiler chickens in 12 pen cages with wood shaving-lined floors were exposed to 1 of 4 treatment groups with 3 pens/treatment: intermittent or ad libitum feeding of a basal diet and intermittent or ad libitum feeding of a coarse hull diet, as described above. At 31 and 32 d of age, birds in experiment 1 were inoculated with chromium EDTA via the cloaca. There was no interaction between diet and feeding regimen. The addition of hulls increased gizzard weight and content and lowered (P < 0.001) gizzard pH, but it had no effect on the ability of the birds to handle intermittent feeding. Despite the dilution with coarse hulls, weight gain and the gain:feed ratio were not affected, which could partly be explained by an increased (P < 0.001) starch digestibility. Dietary reflux was confirmed by the presence of chromium in all intestinal tract sections. Broilers exhibited reverse peristaltic contractions of sufficient magnitude to propel the marker from the cloaca to the gizzard.

Knowledge of the composition of a normal healthy gut microbiota during infancy is important for understanding the role of gut microbiota in disease. A limitation of previous studies is that they are based on infants who have been subject to factors, which can have a profound disruptive effect on the natural colonization process. We describe the colonization process, during the first 4 months after birth, in 85 infants who have experienced no major medical or dietary interventions. They were all vaginally delivered, healthy, term infants, who were not exposed to antibiotics, exclusively breastfed during their first month of life and at least partially breastfed up to 4 months. Selected microbial groups were identified by targeting small subunit microbial ribosomal RNA genes. In contrast to more recent studies, but in agreement with older studies, almost all our infants harbored γ-Proteobacteria and Bifidobacterium. Yet undefined non-cultivable species belonging to Bacteroides, as well as microbes identified as Lachnospiraceae 2, were common. Strong associations were observed between some specific constituents of microbiota at day 4 and the concentration of specific microbial groups at day 120, indicating that early gut microbiota may influence later microbiota. Novel information of the undisturbed composition of early gut microbiota in babies is presented.

Aim: We have tested the effect of various combinations of formic acid and sorbate on Campylobacter jejuni colonization in broiler chickens to reduce the colonization of this zoonotic pathogen in broiler chicken flocks. Methods and Results: Chickens were offered feed supplemented with different concentrations and combinations of formic acid and/or potassium sorbate. We found little or no effect on the Camp. jejuni colonization levels in chickens that were given feed supplemented with formic acid alone. A combination of 1·5% formic acid and 0·1% sorbate reduced the colonization of Camp. jejuni significantly, while a concentration of 2·0% formic acid in combination with 0·1% sorbate prevented Camp. jejuni colonization in chickens. This inhibition was replicated in two independent trials with a combination of three different Camp. jejuni strains. Conclusions: Our results show a novel and promising intervention strategy to reduce the incidence of Camp. jejuni in poultry products and to obtain safer food. Significance and Impact of the Study: To ensure food safety, a reduction of the carcass contamination with Camp. jejuni through reduced colonization of this pathogen in broiler chicken flocks is important. A range of organic acids as additives in feed and drinking water have already been evaluated for this purpose. However, no studies have yet shown a complete inhibition of Camp. jejuni colonization in broiler chickens.

Aims The microbiota surviving sanitation of salmon-processing conveyor belts was identified and its growth dynamics further investigated in a model mimicking processing surfaces in such plants. Methods and Results A diverse microbiota dominated by Gram-negative bacteria was isolated after regular sanitation in three salmon processing plants. A cocktail of 14 bacterial isolates representing all genera isolated from conveyor belts (Listeria, Pseudomonas, Stenotrophomonas, Brochothrix, Serratia, Acinetobacter, Rhodococcus and Chryseobacterium) formed stable biofilms on steel coupons (12°C, salmon broth) of about 109 CFU cm−2 after 2 days. High-throughput sequencing showed that Listeria monocytogenes represented 0·1–0·01% of the biofilm population and that Pseudomonas spp dominated. Interestingly, both Brochothrix sp. and a Pseudomonas sp. dominated in the surrounding suspension. Conclusions The microbiota surviving sanitation is dominated by Pseudomonas spp. The background microbiota in biofilms inhibit, but do not eliminate L. monocytogenes. Significance and Impact of the Study The results highlights that sanitation procedures have to been improved in the salmon-processing industry, as high numbers of a diverse microbiota survived practical sanitation. High-throughput sequencing enables strain level studies of population dynamics in biofilm.

Aims Few studies have compared the effectiveness of hygienic cleaning under simulated use conditions. This study compares commonly used and novel cleaning methods for food contact and hand contact surfaces in kitchens. Methods and Results We report results from two surveys on Norwegian consumers' cleaning procedures. Laboratory models involving cutting boards, tap handles and mobile phones contaminated with Escherichia coli and Staphylococcus aureus were used to compare the hygiene efficacy of commonly used cleaning methods together with new technologies (sprays, single-use wipes, and chlorine-based disinfectants). Commonly used cleaning methods produced a mean log10 reduction (LR) in contamination of 1·5–2·5. The efficacy could be improved by drying or including a disinfection step (mean LR 3·1–4·6). Cleaning of mobile phones was common and was improved by including humidity (1·5–1·9 mean LR). Conclusions In many situations, traditional methods used by consumers may be sufficient to hygienically clean surfaces. However, in some situations, such as where there are infected or immune-compromised individuals, or where high risk foods are being handled, hygiene practices resulting in higher LR should be recommended. Significance and Impact of the Study This study demonstrates that data from models simulating use conditions are required to estimate the effectiveness of detergent-based removal practices and how these can be enhanced by inactivation processes such as drying and disinfection to ensure that contamination from food-borne pathogens is reduced to acceptable levels to prevent infection transmission.

The aim of this study was to detect and interpret correlation patterns in several large data matrices from the same biological system using Partial Least Squares Regression (PLSR) in order to get information on the system under investigation. To do this, DNA microarray data and Fourier Transform Infrared (FT-IR) spectra from a designed study where Campylobacter jejuni was exposed to environmental stress conditions, were used. The experimental design included variation in atmospheric conditions, temperature and time. PLSR was first used to analyse each of the two data types separately in order to explore the effect of the experimental parameters on the data. The results showed that both the gene expression and FT-IR spectra were affected by the variations in atmosphere, temperature and time, but that the effect was different for the two types of data. When the DNA microarray data and FT-IR spectra were linked together by PLSR, covariation due to temperature was seen. Both specific genes and ranges in the FT-IR spectra that were connected to the variation in temperature were detected. Some of these are possibly connected to properties of the cell wall of the bacteria. The results in this study show the potential of PLSR for investigation of covariance structures in biological data. By doing this, valuable information about the biological system can be detected and interpreted. It was also shown that the use of FT-IR spectroscopy provided important information about the stress responses in the bacteria, information that was not detected from the DNA microarray data.

Explorative approaches such as DNA microarray experiments are becoming increasingly important in microbial research. Despite these major technical advancements, approaches to study multifactor experiments are still lacking. We have addressed this problem by using rotation testing and a novel multivariate analysis of variance (MANOVA) approach (50-50 MANOVA) to investigate interacting experimental factors in a complex experimental design. Furthermore, a new rotation testing based method was introduced to calculate false-discovery rates for each response. This novel analytical concept was used to investigate global survival mechanisms in the environment of the major food-borne pathogen C. jejuni. We simulated nongrowth environmental conditions by investigating combinations of the factors temperature (5 and 25°C) and oxygen tension (anaerobic, microaerobic, and aerobic). Data were generated with DNA microarrays for information about gene expression patterns and Fourier transform infrared (FT-IR) spectroscopy to study global macromolecular changes in the cell. Microarray analyses showed that most genes were either unchanged or down regulated compared to the reference (day 0) for the conditions tested and that the 25°C anaerobic condition gave the most distinct expression pattern with the fewest genes expressed. The few up-regulated genes were generally stress related and/or related to the cell envelope. We found, using FT-IR spectroscopy, that the amount of polysaccharides and oligosaccharides increased under the nongrowth survival conditions. Potential mechanisms for survival could be to down regulate most functions to save energy and to produce polysaccharides and oligosaccharides for protection against harsh environments. Basic knowledge about the survival mechanisms is of fundamental importance in preventing transmission of this bacterium through the food chain.

The distinction between viable and dead cells is a major issue in many aspects of biological research. The current technologies for determining viable versus dead cells cannot readily be used for quantitative differentiation of specific cells in mixed populations. This is a serious limitation. We have solved this problem by developing a new concept with the viable/dead stain ethidium monoazide (EMA) in combination with real-time PCR (EMA-PCR). A dynamic range of approximately 4 log 10 was obtained for the EMA-PCR viable/dead assay. Viable/dead differentiation is obtained by covalent binding of EMA to DNA in dead cells by photoactivation. EMA penetrates only dead cells with compromised membrane/cell wall systems. DNA covalently bound to EMA cannot be PCR amplified. Thus, only DNA from viable cells can be detected. We evaluated EMA-PCR with the major food-borne bacterium Campylobacter jejuni as an example. Traditional diagnosis of this bacterium is very difficult due to its specific growth requirements and because it may enter a state where it is viable but not cultivable. The conditions analyzed included detection in mixed and natural samples, survival in food, and survival after disinfection or antibiotic treatment. We obtained reliable viable/dead quantifications for all conditions tested. Comparison with standard fluorescence-based viable/dead techniques showed that the EMA-PCR has a broader dynamic range and enables quantification in mixed and complex samples. In conclusion, EMA-PCR offers a novel real-time PCR method for quantitative distinction between viable and dead cells with potentially very wide application.

Several challenges still persist in the analysis of microorganisms in foods, particularly in studies of complex communities. Nucleic acid-based methods are promising tools in addressing new questions concerning microbial communities. We have developed several new methods in the field of nucleic acid-based microbial community analyses. These methods cover both sample preparation and detection approaches. The sample preparation method involves simplified DNA purification using paramagnetic beads. As an extension of this method, the same paramagnetic beads are used for both cell separation and DNA purification. This enables full automation. The separate detection of viable and dead bacteria is a major issue in nucleic acid-based diagnostics. We have applied a living/dead dye that binds covalently to DNA and inhibits the PCR from dead cells. In addition, a DNA array-based detection assay has been developed. The assay combines the specificity obtained by enzymatic labeling of DNA probes with the possibility of detecting several targets simultaneously by DNA array hybridization. In combination with 16S rDNA amplification, this is a promising tool for community analyses. Also, we have developed a novel approach for multiplex quantitative PCR. The multiplex PCR has been combined with our DNA array-based detection method. Finally, we are now in the process of adapting a system for monitoring microbial growth and death in real-time through the tagging of bacteria with green fluorescent protein (GFP) combined with fluorescence detection using a high-resolution confocal laser scanner.

The autotrophic ammonia-oxidizing bacteria (AOB), which play an important role in the global nitrogen cycle, assimilate CO 2 by using ribulose 1,5-bisphosphate carboxylase/oxygenase (RubisCO). Here we describe the first detailed study of RubisCO ( cbb ) genes and proteins from the AOB. The cbbLS genes from Nitrosospira sp. isolate 40KI were cloned and sequenced. Partial sequences of the RubisCO large subunit (CbbL) from 13 other AOB belonging to the β and γ subgroups of the class Proteobacteria are also presented. All except one of the β-subgroup AOB possessed a red-like type I RubisCO with high sequence similarity to the Ralstonia eutropha enzyme. All of these new red-like RubisCOs had a unique six-amino-acid insert in CbbL. Two of the AOB, Nitrosococcus halophilus Nc4 and Nitrosomonas europaea Nm50, had a green-like RubisCO. With one exception, the phylogeny of the AOB CbbL was very similar to that of the 16S rRNA gene. The presence of a green-like RubisCO in N. europaea was surprising, as all of the other β-subgroup AOB had red-like RubisCOs. The green-like enzyme of N. europaea Nm50 was probably acquired by horizontal gene transfer. Functional expression of Nitrosospira sp. isolate 40KI RubisCO in the chemoautotrophic host R. eutropha was demonstrated. Use of an expression vector harboring the R. eutropha cbb control region allowed regulated expression of Nitrosospira sp. isolate 40KI RubisCO in an R. eutropha cbb deletion strain. The Nitrosospira RubisCO supported autotrophic growth of R. eutropha with a doubling time of 4.6 h. This expression system may allow further functional analysis of AOB cbb genes.

In this study, we investigated the effects of different feed structures and beddings on the spread of C. jejuni in broiler flocks, and the effect on the cecal microbiota. Broiler chickens raised in 24 eight-bird group cages on either rubber mat or wood shavings were fed either a wheat-based control diet (Control), a diet where 50% of the ground wheat was replaced by whole wheat prior to pelleting (Wheat), or a wheat-based diet, such as the control diet diluted with 12% oat hulls (Oat). Samples from the cloacal mucosa of all birds were taken daily for C. jejuni quantification and cecum samples were collected at the end of the experiment for C. jejuni quantification and microbiota analyses. We have shown a statistically significant effect of increased feed structure on the reduced spread of C. jejuni in chicken flocks, but no significant differences were detected between types of structure included in the feed. No significant changes in the dominating microbiota in the lower lower gastrointestinal (GI) tract were observed, which indicates that feed structure only has an effect on the upper GI tract. Delaying the spread of C. jejuni in broiler flocks could, at time of slaughter, result in fewer C. jejuni -positive broilers.

It is acknowledged that some obesity trajectories are set early in life, and that rapid weight gain in infancy is a risk factor for later development of obesity. Identifying modifiable factors associated with early rapid weight gain is a prerequisite for curtailing the growing worldwide obesity epidemic. Recently, much attention has been given to findings indicating that gut microbiota may play a role in obesity development. We aim at identifying how the development of early gut microbiota is associated with expected infant growth. We developed a novel procedure that allows for the identification of longitudinal gut microbiota patterns (corresponding to the gut ecosystem developing), which are associated with an outcome of interest, while appropriately controlling for the false discovery rate. Our method identified developmental pathways of Staphylococcus species and Escherichia coli that were associated with expected growth, and traditional methods indicated that the detection of Bacteroides species at day 30 was associated with growth. Our method should have wide future applicability for studying gut microbiota, and is particularly important for translational considerations, as it is critical to understand the timing of microbiome transitions prior to attempting to manipulate gut microbiota in early life., Author Summary Some obesity trajectories are set early in life, with rapid weight gain being a risk factor for later development of obesity. Recently, much attention has been given to findings indicating that gut microbiota may play a role in obesity development. The existence of time-dependent exposure windows, which rely on stimuli from the gut to initiate healthy development, gives the evolution of early life gut microbiota a critical role in human health. We identified children that followed their expected growth trajectories at six months of life, and those that had deviated. We then developed a novel statistical approach that allowed the identification of longitudinal gut microbiota patterns (e.g. a particular species was detected at days 4, 10, and 30 and not detected at day 120) that were associated with expected growth, while appropriately restricting the false discovery rate. We further identified when a deviation from the proposed longitudinal gut microbiota patterns would result in an abnormal growth outcome (either rapid or decreased growth at six months of life). We found developmental pathways of Staphylococcus species and Escherichia coli that were associated with expected growth, as well as indications that Bacteroides species at day 30 was associated with growth.

Exposure of Escherichia coli to a subminimal inhibitory concentration (25% below MIC) of benzalkonium chloride (BC), an antimicrobial membrane-active agent commonly used in medical and food-processing environments, resulted in cell death and changes in cell morphology (filamentation). A small subpopulation (1–5% of the initial population) survived and regained similar morphology and growth rate as non-exposed cells. This subpopulation maintained tolerance to BC after serial transfers in medium without BC. To withstand BC during regrowth the cells up regulated a drug efflux associated gene (the acrB gene, member of the AcrAB-TolC efflux system) and changed expression of outer membrane porin genes (ompFW) and several genes involved in protecting the cell from the osmotic- and oxidative stress. Cells pre-exposed to osmotic- and oxidative stress (sodium chloride, salicylic acid and methyl viologen) showed higher tolerance to BC. A control and two selected isolates showing increased BC-tolerance after regrowth in BC was genome sequenced. No common point mutations were found in the BC- isolates but one point mutation in gene rpsA (Ribosomal protein S1) was observed in one of the isolates. The observed tolerance can therefore not solely be explained by the observed point mutation. The results indicate that there are several different mechanisms responsible for the regrowth of a tolerant subpopulation in BC, both BC-specific and general stress responses, and that sub-MIC of BC may select for phenotypic variants in a sensitive E. coli culture.

Cattle represent a major source of fecal contamination worldwide. Understanding the natural variation of the bovine livestock fecal microbiota is therefore important. For this reason we addressed the yearly differences of the fecal microbiota for bovine livestock reared in the same geographical region from 1999 to 2007 - analyzing a total of 300 samples representing a range of experimental regimes. The aim of our work was to determine the effect of year visa experimental regime of the bovine livestock fecal microbiota. We used a newly developed high-throughput 165 rRNA sequencing approach (a single mixed Sanger sequence was generated per sample) in combination with deep pyrosequencing. We found that similar feeding and treatment regimes for different years showed major differences in the fecal microbiota, suggesting other factors important for shaping the fecal microbiota than those experimentally controlled. Ruminococcaeae, Peptostreptococcaceae, Acinetobacter, Escherichia/Shigella, Lachnospiraceae and Lactobacillales were the main taxa associated with the yearly fluctuations. Furthermore, we found that fecal samples with high levels of Lactobacillales, Ruminococcaea and Lachnospiraceae had the most even species distributions. The Peptostreptococcaceae and Acinetobacter dominated samples, on the other hand, showed a few highly dominant taxa. Testing of neutrality showed that the evenly distributed samples were explained by a neutral mode (that the assembly of the microbiota was random), while for the other samples there were overrepresentation of the dominant species (indicating bacterial-bacterial nice competition). We therefore propose that there are natural yearly fluctuations of the bovine livestock microbiota - both with respect to ecology and composition. This knowledge will have impact on our management of fecal bacteria in the environment, since it is very difficult to predict risk based on historical data.

A major challenge with single-nucleotide polymorphism (SNP) fingerprinting of bacteria and higher organisms is the combination of genome-wide screenings with the potential of multiplexing and accurate SNP detection. Single-nucleotide extension by the minisequencing principle represents a technology that both is highly accurate and enables multiplexing. A current bottleneck for direct genome analyses by minisequencing, however, is the sensitivity, since minisequencing relies on linear signal amplification. Here, we present SNPtrap, which is a novel approach that combines the specificity and possibility of multiplexing by minisequencing with the sensitivity obtained by logarithmic signal amplification by polymerase chain reaction (PCR). We show a SNPtrap proof of principle in a model system for two polymorphic SNP sites in the Salmonella tetrathionate reductase gene (ttrC).

The global gene expression and biomolecular composition in an Escherichia coli model strain exposed to 10 adverse conditions (sodium chloride, ethanol, glycerol, hydrochloric and acetic acid, sodium hydroxide, heat (46 °C), and cold (15 °C), as well as ethidium bromide and the disinfectant benzalkonium chloride) were determined using DNA microarrays and Fourier transform infrared (FT-IR) spectroscopy. In total, approximately 40% of all investigated genes (1682/4279 genes) significantly changed expression, compared with a nonstressed control. There were, however, only 3 genes (ygaW (unknown function), rmf (encoding a ribosomal modification factor), and ghrA (encoding a glyoxylate/hydroxypyruvate reductase)) that significantly changed expression under all conditions (not including benzalkonium chloride). The FT-IR analysis showed an increase in unsaturated fatty acids during ethanol and cold exposure, and a decrease during acid and heat exposure. Cold conditions induced changes in the carbohydrate composition of the cell, possibly related to the upregulation of outer membrane genes (glgAP and rcsA). Although some covariance was observed between the 2 data sets, principle component analysis and regression analyses revealed that the gene expression and the biomolecular responses are not well correlated in stressed populations of E. coli, underlining the importance of multiple strategies to begin to understand the effect on the whole cell.

Over the latest decades, there has been a rapid development of laboratory techniques used for analyzing the pathways from the genes to the final physiology of an organism. Insight into the biological mechanism controlling growth and development of an organism is thereby enabled at a level that was never before possible. This in turn can be utilized, for example, in health care for improved diagnostics and improved medical treatment of diseases, and in agricultural science for tailoring plants and animals with desirable properties. Both from a biological and from a data analytical point of view the scientists are facing major challenges in the research area of analyzing functional genomics data. This chapter aims at addressing some of these challenges, and discusses important questions that need to be considered as well as gives guidelines to some useful approaches for analyzing data from functional genomics. The focus of this chapter is on data modeling of empirical data from functional genomics.

International audience; Benzalkonium chloride (BC) is a commonly used disinfectant and preservative. This study describes changes in expression level at the transcriptomic and proteomic level for Escherichia coli K-12 gradually adapted to a tolerance level to BC of 7–8 times the initial MIC. Results from DNA arrays and two-dimensional gel electrophoresis for global gene and protein expression studies were confirmed by real-time quantitative PCR. Peptide mass fingerprinting by MALDI-TOF MS was used to identify differentially expressed proteins. Changes in expression level in adapted cells were shown for porins, drug transporters, glycolytic enzymes, ribosomal subunits and several genes and proteins involved in protection against oxidative stress and antibiotics. Adapted strains showed increased tolerance to several antibiotics. In conclusion, E. coli K-12 adapted to higher tolerance to BC acquired several general resistance mechanisms, including responses normally related to the multiple antibiotic resistance (Mar) regulon and protection against oxidative stress. The results revealed that BC treatment might result in superoxide stress in E. coli.

Colorectal cancer is among the most prevalent types of cancers and its development has been associated with components in the diet. Dietary fibers seem to display a protective role, and in the search for what mechanisms may be involved increased focus has been directed at the effect of fibers on the gut micro flora. This project has included studies aimed at generating new knowledge about how various types of dietary fibers influence the intestinal tumorigenesis, inflammatory status, composition of bacteria in various intestinal segments and the interrelationships between these factors. Three different fibers were tested; a finely ground cellulose preparate, inulin and mask (a dried and finely milled brewers spent grain). All fibers were present in the diet in a lower and a higher concentration (5 % and 15%). The test diets were given to multiple intestinal neoplasia (Min) and wild type (Wt) mice from weaning until euthanation at week 11. The Min mice carry the same mutation as seen in the human familial adenomatous polyposis (FAP) syndrome that results in spontaneous development of colorectal tumors. The majority of sporadic human colorectal tumors are also mutated in the same signaling pathway. Colon carcinogenesis was induced by two subcutaneous injections of azoxymethan (day 7 ±1 and14 ±1 after birth). At euthanation a blood sample from the hepatic portal vein was obtained and the small (Min mice) and large intestine (Min and Wt mice) were removed and fixed. The coecum and spleen were weighed and snap frozen. Individual weights and diet consumption were recorded. Scoring of tumors in the small and large intestine displayed a weak but significant difference in tumor development between the different diet groups. The inulin 5% diet resulted in reduced tumor number compared to the cellulose (small and large intestine) and the mask (large intestine) fibers. Analysis by Multiplex of a large range of inflammatory cytokines in plasma revealed the presence of individual animals with high levels of certain cytokines. This variation seems to be independent of fiber type in the diet. However, analysis of several data from the individual animals with respect to any correlation between cancer development and cytokine level is presently under evaluation. Furthermore, a pilot screening of the micro flora in feces from selected representative animals demonstrated interesting differences in species variation when related to the various fibers tested and the two different doses included. All animal recordings in this study are done on the individual level; correlations between tumor development, inflammatory status and intestinal micro flora will therefore be made at several levels. In summary, our results demonstrate that various types of fibers influence the development of tumors to various degrees. Further evaluation of the data is in progress and may reveal associations between tumor development, inflammatory status and the intestinal micro flora. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 5431. doi:1538-7445.AM2012-5431