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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.

New aquaculture production systems are evolving for prolong production of Atlantic salmon smolts or post-smolts before stocking in traditional net pens, such as semi-closed containment systems (S-CCS) in sea (Fig. 1) and recirculating aquaculture systems (RAS) on land. The microbiota in these systems can potentially have great impact on the robustness and health of the fish. These two types of aquaculture systems are likely to have different challenges regarding pathogenic invasion due to the different water management, e.g. different treatment of the intake water and different turnover of the water. In this study, we investigated the bacterial microbiota of both water and biofilms in a commercial RAS and in S-CCS in sea during a three months period of post-smolt production. Deep-sequencing of the bacterial 16S rRNA gene (V4) was used for the first time to obtain in depth compositional analysis of microbial communities in commercial scale facilities. Highly diverse communities were detected, with up to 2000 different Operational Taxonomic Units (OTUs) within samples. Both systems were dominated by Proteobacteria with Rhodobacteraceae as the dominating taxa, followed by Bacteroidetes that was dominated by Polaribacter among others. However, the microbiota composition was clearly different between the two aquaculture systems, and between water samples and biofilms. In RAS, it was also shown different microbiota composition with water salinity of 12 vs 22 parts per thousand (ppt). Higher abundance of e.g. Myxococcales and Nitrospiraceae was observed at 12 ppt, which coincided with lower total ammonia nitrogen (TAN) levels. Both taxa were also more abundant in the Moving Bed Bioreactor (MBBR)-biofilms than in water, as well as Planctomyces among others. In S-CCS, clear temporal changes of the microbiota was observed during the production, where potential pathogens like Tenacibaculum , Aliivibrio , Alteromonadaceae and Polaribacter were increasing in the spring time, as well as one unassigned taxa and chloroplast DNA likely from algae. The implication of these potential pathogens on fish health is unknown. A common observation for both RAS and S-CCS was higher abundance of the potential pathogens in the water compared to the biofilms. Further studies on the microbiota in closed-containment aquaculture systems are needed to obtain more knowledge about their impact on post-smolt production performance, welfare and health.

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.

BackgroundThis paper presents a strategy for statistical analysis and interpretation of longitudinal intervention effects on bacterial communities. Data from such experiments often suffers from small sample size, high degree of irrelevant variation, and missing data points. Our strategy is a combination of multi-way decomposition methods, multivariate ANOVA, multi-block regression, hierarchical clustering and phylogenetic network graphs. The aim is to provide answers to relevant research questions, which are both statistically valid and easy to interpret.ResultsThe strategy is illustrated by analysing an intervention design where two mice groups were subjected to a treatment that caused inflammation in the intestines. Total microbiota in fecal samples was analysed at five time points, and the clinical end point was the load of colon cancer lesions. By using different combinations of the aforementioned methods, we were able to show that:The treatment had a significant effect on the microbiota, and we have identified clusters of bacteria groups with different time trajectories.Individual differences in the initial microbiota had a large effect on the load of tumors, but not on the formation of early-stage lesions (flat ACFs).The treatment resulted in an increase in Bacteroidaceae, Prevotellaceae and Paraprevotellaceae, and this increase could be associated with the formation of cancer lesions.ConclusionThe results show that by applying several data analytical methods in combination, we are able to view the system from different angles and thereby answer different research questions. We believe that multiway methods and multivariate ANOVA should be used more frequently in the bioinformatics fields, due to their ability to extract meaningful components from data sets with many collinear variables, few samples and a high degree of noise or irrelevant variation.

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.

Summary The main objective of this study was to investigate the effect of excess cobalt (Co) on gene expression of stearoyl-CoA desaturase (SCD), fatty acid synthase (FASN), diacylglycerol acyltransferase 1 (DGAT1) and diacylglycerol acyltransferase 2 (DGAT2) of lactating dairy cows in relation to milk fatty acid profile. Seven multiparous cows of the Norwegian Red cattle breed (NRF) had their basal diet supplemented with 1.4 g Co as a 24 g/l solution of Co-acetate per os twice daily for 7 days followed by a 9-day depuration period. Udder biopsies were performed prior to the treatment period, after 1 week of treatment and immediately after the depuration period. Excess Co reduced the proportion of all cis-9 monounsaturated fatty acids and increased the proportion of 18:0 in milk. However, gene expression levels of SCD, DGAT1, DGAT2 and FASN were not significantly altered. Our results indicate that the effect of Co on milk fatty acid profile is mediated at the post-transcriptional level by reduced activity of SCD in the mammary gland. Potential mechanisms explaining how Co might reduce stearoyl-CoA desaturation are discussed.

Pigs are recognised as suitable biomedical models to study obesity and obesity-related diseases; however, little is known about adipose tissue development and adipogenesis in pigs. In this study, the temporal expression of key genes involved in lipid metabolism was investigated during porcine adipogenesis and the metabolic fate of exogenously administered palmitic acid (16:0) was examined in differentiating preadipocytes. The expression of genes encoding elongases and desaturases increased simultaneously with those involved in fatty acid and triacylglycerol synthesis during porcine adipogenesis, and a high biosynthesis of monounsaturated fatty acids was measured prior to storage in differentiating preadipocytes. Although the total fatty acid oxidation in differentiating preadipocytes was low, differentiating cells showed increased expression of hormone-sensitive lipase and mitochondrial and peroxisomal genes. These data provide new insight into the temporal expression of genes involved in lipid metabolism during porcine adipogenesis and suggest a possible role of elongation and desaturation events prior to lipid accumulation in porcine adipocytes.

The potential presence of widespread and stable bacterial core phylogroups in the human colon has promoted considerable attention. Despite major efforts, no such phylogroups have yet been identified. Therefore, using a novel phylogroup- and tree-independent approach, we present a reanalysis of 1 114 722 V2 region and 71 550 near full-length 16S rRNA sequences from a total of 210 human beings, with widespread geographic origin, ethnic background and diet, in addition to a wide range of other mammals. We found two highly prevalent core phylogroups (cores 1 and 2), belonging to the clostridial family Lachnospiraceae. These core phylogroups showed a log-normal distribution among human individuals, while non-core phylogroups showed more skewed distributions towards individuals with low levels compared with the log-normal distribution. Molecular clock analyses suggest that core 2 co-evolved with the radiation of vertebrates, while core 1 co-evolved with the mammals. Taken together, the stability, prevalence and potential functionality support the fact that the identified core phylogroups are pivotal in maintaining gut homeostasis and health.

Background Boar taint is an unpleasant odor and flavor of the meat and occurs in a high proportion of uncastrated male pigs. Androstenone, a steroid produced in testis and acting as a sex pheromone regulating reproductive function in female pigs, is one of the main compounds responsible for boar taint. The primary goal of the present investigation was to determine the differential gene expression of selected candidate genes related to levels of androstenone in pigs. Results Altogether 2560 boars from the Norwegian Landrace and Duroc populations were included in this study. Testicle samples from the 192 boars with most extreme high or low levels of androstenone in fat were used for RNA extraction, and 15 candidate genes were selected and analyzed by real-competitive PCR analysis. The genes Cytochrome P450 c17 (CYP17A1), Steroidogenic acute regulatory protein (STAR), Aldo-keto reductase family 1 member C4 (AKR1C4), Short-chain dehydrogenase/reductase family member 4 (DHRS4), Ferritin light polypeptide (FTL), Sulfotransferase family 2A, dehydroepiandrosterone-preferring member 1 (SULT2A1), Cytochrome P450 subfamily XIA polypeptide 1 (CYP11A1), Cytochrome b5 (CYB5A), and 17-beta-Hydroxysteroid dehydrogenase IV (HSD17B4) were all found to be significantly (P < 0.05) up-regulated in high androstenone boars in both Duroc and Landrace. Furthermore, Cytochrome P450 c19A2 (CYP19A2) was down-regulated and progesterone receptor membrane component 1 (PGRMC1) was up-regulated in high-androstenone Duroc boars only, while CYP21 was significantly down-regulated (2.5) in high-androstenone Landrace only. The genes Nuclear Receptor co-activator 4 (NCOA4), Sphingomyrlin phosphodiesterase 1 (SMPD1) and 3β-hydroxysteroid dehydrogenase (HSD3B) were not significantly differentially expressed in any breeds. Additionally, association studies were performed for the genes with one or more detected SNPs. Association between SNP and androstenone level was observed in CYB5A only, suggesting cis-regulation of the differential transcription in this gene. Conclusion A large pig material of highly extreme androstenone levels is investigated. The current study contributes to the knowledge about which genes that is differentially expressed regard to the levels of androstenone in pigs. Results in this paper suggest that several genes are important in the regulation of androstenone level in boars and warrant further evaluation of the above mentioned candidate genes, including analyses in different breeds, identification of causal mutations and possible gene interactions.

Microorganisms are constantly exposed to new and altered growth conditions, and respond by changing gene expression patterns. Several methods for studying gene expression exist. During the last decade, the analysis of microarrays has been one of the most common approaches applied for large scale gene expression studies. A relatively new method for gene expression analysis is MassARRAY, which combines real competitive-PCR and MALDI-TOF (matrix-assisted laser desorption/ionization time-of-flight) mass spectrometry. In contrast to microarray methods, MassARRAY technology is suitable for analysing a larger number of samples, though for a smaller set of genes. In this study we compare the results from MassARRAY with microarrays on gene expression responses of Staphylococcus aureus exposed to acid stress at pH 4.5. RNA isolated from the same stress experiments was analysed using both the MassARRAY and the microarray methods. The MassARRAY and microarray methods showed good correlation. Both MassARRAY and microarray estimated somewhat lower fold changes compared with quantitative real-time PCR (qRT-PCR). The results confirmed the up-regulation of the urease genes in acidic environments, and also indicated the importance of metal ion regulation. This study shows that the MassARRAY technology is suitable for gene expression analysis in prokaryotes, and has advantages when a set of genes is being analysed for an organism exposed to many different environmental conditions.

Four different subtypes of fatty acid binding proteins i.e. liver-type FABP1, heart/muscle-type FABP3, adipocyte-type FABP4 and epithelial/epidermal-type FABP5 are expressed in adipose tissue. However, only the regulatory role of FABP4 in adipogenesis has been thoroughly investigated. To increase the knowledge on possible roles of these FABP subtypes in preadipocyte differentiation, gene expression patterns were examined during adipogenesis in pig (Sus scrofa). FABP1 expression was induced in proliferating cells, whereas FABP3, FABP4 and FABP5 expression increased throughout preadipocyte differentiation. Interestingly, the FABP4 and FABP5 expression increased early in the differentiation, followed by FABP3 later in the differentiation process. This indicates a role of FABP4 and FABP5 in intracellular fatty acid transport during initiation of differentiation, whereas, FABP3 likely is involved in the transport of fatty acids during intermediate stages of adipogenesis. In this study we demonstrate that FABP3, FABP4 and FABP5 expression is correlated with that of the peroxisome proliferator-activated receptors alpha and gamma (PPARA and PPARG). Altogether, this suggests a role of FABP1 during cell proliferation, whereas a coordinated expression of FABP3, FABP4 and FABP5 together with that of PPARA, PPARG1 and PPARG2 might be critical for the metabolic regulation during porcine adipogenesis.

Recently a role of adipose tissue as an endocrine organ secreting factors involved in the regulation of whole-body energy homeostasis has emerged. Preadipocytes in different fat depots have distinct adipogenic potential and the metabolic activity differs between mature adipocytes of different depot origins. Here we describe the proliferation and differentiation of stromal-vascular cells derived from subcutaneous and visceral fat depots of adult pigs. We demonstrate that subcutaneous porcine preadipocytes proliferate more actively and that individual subcutaneous adipocytes have a more rapid accumulation of triacylglycerols than visceral cells. During differentiation, subcutaneous and visceral preadipocytes showed similar gene expression patterns with increased expression of adiponectin (APM1), adipocyte-specific fatty acid binding protein (FABP4), catalase (CAT), and peroxisome proliferator-activated receptor gamma 2 (PPARG2). Furthermore, initial data showing depot-originated effects on the expression of CAT, carnitine palmitoyl transferase 1B (CPT1B) and FABP4 suggest possible depot specific differences in the function and metabolism of mature porcine adipocytes.

Despite great conceptual promise, the use of microarrays in typing approaches has not yet gained wide acceptance. The establishment of proper criteria for determining discriminatory power as well as typability and the accuracy of microarray data remains to be solved. Purely experimental estimations of these parameters would far exceed what is experimentally practical. We therefore used simulations in combination with experimental results in parameter estimations. Our assay was based on 26 single nucleotide polymorphisms (SNPs) identified in the Campylobacter jejuni Multi Locus Sequence Typing (MLST) database (http://pubmlst.org/campylobacter/). The SNPs were detected using a single nucleotide extension (SNE) typing microarray. Unknown isolates were assigned to the known sequence type(s) by calculating weighted sum of matches minus a weighted sum of mismatches between predicted and candidate genotype. The weights were set according to the Bayesian posterior probability of the SNP classification. These studies showed that any typing or profiling method based on binary data requires an accuracy of

Toll-like receptors (TLRs) are a family of recognition receptors playing a crucial role in the innate immune system. Different combinations of TLRs are thought to be crucial for effective immune response, thus insight into the organization and expression of TLRs is important for understanding disease resistance. Mastitis is the most frequent and costly disease in dairy production, and the innate immune system is considered to be important in the first line defence against this disease. In the present paper we have characterized the genomic organization of TLR6-TLR1-TLR10 in a approximately 50 kb region of bovine chromosome 6, including 5'-untranslated exons not previously described. A method for gene expression analysis was developed and used for transcription profiling of the three paralogous genes in different bovine tissues. The expression analysis showed similar expression profiles for TLR1 and TLR6, which indicate a co-regulation of these two genes in cattle. TLR10 had a different expression profile, pointing toward a stronger functional diversification compared to TLR1 and TLR6. The differences in expression are in accordance with the evolutionary history of this gene cluster, where TLR10 diverged from the common ancestral gene before the duplication event that created TLR1 and TLR6.