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Nisin is a bacteriocin that is globally employed as a biopreservative in food systems to control gram-positive, and some gram-negative, bacteria. Here we tested the bioactivity of nisin A-producing Lactococcus lactis NZ9700 and producers of bioengineered variants thereof against representatives of the gram-negative genus Thermus, which has been associated with the pink discoloration defect in cheese. Starting with a total of 73 nisin variant-producing Lactococcus lactis, bioactivity against Thermus was assessed via agar diffusion assays, and 22 variants were found to have bioactivity greater than or equal to that of the nisin A-producing control. To determine to what extent this enhanced bioactivity was attributable to an increase in specific activity, minimum inhibitory concentrations were determined using the corresponding purified form of these 22 nisin A derivatives. From these experiments, nisin M17Q and M21F were identified as peptides with enhanced antimicrobial activity against the majority of Thermus target strains tested. In addition, several other peptide variants were found to exhibit enhanced specific activity against a subset of strains.

Kefir consumption has been demonstrated to improve lipid and cholesterol metabolism; however, our previous study identified that benefits vary between different commercial and traditional kefir. Here, we investigate the ability of pitched culture kefir, that is, kefir produced by a small number of specific strains, to recapitulate health benefits of a traditional kefir, in a diet-induced obesity mouse model, and examine how microbial composition of kefir impacts these benefits. Eight-week-old female C57BL/6 mice were fed a high-fat diet (40 % energy from fat) supplemented with one of five kefir varieties (traditional, pitched, pitched with no Lactobacillus, pitched with no yeast and commercial control) at 2 ml in 20 g of food for 8 weeks prior to analysis of plasma and liver lipid profiles, and liver gene expression profiles related to lipid metabolism. Both traditional and pitched kefir lowered plasma cholesterol by about 35 % (P = 0·0005) and liver TAG by about 55 % (P = 0·0001) when compared with commercial kefir despite no difference in body weight. Furthermore, pitched kefir produced without either yeast or Lactobacillus did not lower cholesterol. The traditional and pitched kefir with the full complement of microbes were able to impart corresponding decreases in the expression of the cholesterol and lipid metabolism genes encoding 3-hydroxy-3-methylglutaryl-coenzyme A reductase, PPARγ and CD36 in the liver. These results demonstrate that traditional kefir organisms can successfully be utilised in a commercial process, while highlighting the importance of microbial interactions during fermentation in the ability of fermented foods to benefit host health.

Cleaning of the production environment is vital to ensure the safety and quality of dairy products. Although cleaning with chlorine-based agents is widely adopted, it has been associated with detrimental effects on milk quality and safety, which has garnered increasing interest in chlorine-free cleaning. However, the influence of these methods on the milk microbiota is not well documented. This study investigated the factors that influence the raw milk microbiota, with a focus on the differences when chlorine-based and chlorine-free cleaning of milking equipment are used. Bulk tank raw milk was sampled during three sampling months (April, August, and November), from farms across Ireland selected to capture the use of different cleaning methods, i.e., exclusively chlorine-based (n = 51) and chlorine-free cleaning (n = 92) and farms that used chlorine-free agents for the bulk tank and chlorine-based cleaning agents for the rest of the equipment (n = 28). Shotgun metagenomic analysis revealed the significant influence of seasonal and geographic factors on the bulk tank milk microbiota, indicated by differences in diversity, taxonomic composition, and functional characteristics. Taxonomic and functional profiles of samples collected in November clustered separately from those of samples collected in other months. In contrast, cleaning methods only accounted for 1% of the variation in the bulk tank milk bacterial community, and samples collected from farms using chlorine-based versus chlorine-free cleaning did not differ significantly, suggesting that the chlorine-free approaches used did not negatively impact microbiological quality. This study shows the value of shotgun metagenomics in advancing our knowledge of the raw milk microbiota. IMPORTANCE The microbiota of raw milk is affected by many factors that can control or promote the introduction of undesirable microorganisms. Chlorine-based cleaning agents have been commonly used due to their effectiveness in controlling undesirable microorganisms, but they have been associated with the formation of chlorine residues that are detrimental to product quality and may impact consumer health. Chlorine-free alternatives have been recommended in some countries, but the influence of cleaning agents on the milk microbiota is unknown. Here, we investigated the influence of cleaning methods and other factors on bulk tank raw milk. Results showed that season and location had a greater influence on the milk microbiota than the cleaning agents used. Indeed, the similar microbiota compositions of raw milk from farms that used chlorine-based and those that used chlorine-free cleaning methods supports the further use of chlorine-free cleaning agents in dairy production.

The human intestinal commensal microbiota and associated metabolic products have long been regarded as contributors to host health. As the identity and activities of the various members of this community have become clearer, newly identified health-associated bacteria, such as Faecalibacterium prausnitzii, Akkermansia muciniphila, Ruminococcus bromii and Roseburia species, have emerged. Notably, the abundance of many of these bacteria is inversely correlated to several disease states. While technological and regulatory hurdles may limit the use of strains from these taxa as probiotics, it should be possible to utilize prebiotics and other dietary components to selectively enhance their growth in situ. Dietary components of potential relevance include well-established prebiotics, such as galacto-oligosaccharides, fructo-oligosaccharides and inulin, while other putative prebiotics, such as other oligosaccharides, polyphenols, resistant starch, algae and seaweed as well as host gut metabolites such as lactate and acetate, may also be applied with the aim of selectively and/or differentially affecting the beneficial bacterial community within the gastrointestinal environment. The present review provides an overview of the dietary components that could be applied in this manner.

The interactions between parasitic helminths and gut microbiota are considered to be an important, although as yet incompletely understood, factor in the regulation of immunity, inflammation and a range of diseases. Infection with intestinal helminths is ubiquitous in grazing horses, with cyathostomins (about 50 species of which are recorded) predominating. Consequences of infection include both chronic effects, and an acute inflammatory syndrome, acute larval cyathostominosis, which sometimes follows removal of adult helminths by administration of anthelmintic drugs. The presence of cyathostomins as a resident helminth population of the equine gut (the "helminthome") provides an opportunity to investigate the effect helminth infection, and its perturbation, has on both the immune system and bacterial microbiome of the gut, as well as to determine the specific mechanisms of pathophysiology involved in equine acute larval cyathostominosis. We studied changes in the faecal microbiota of two groups of horses following treatment with anthelmintics (fenbendazole or moxidectin). We found decreases in both alpha diversity and beta diversity of the faecal microbiota at Day 7 post-treatment, which were reversed by Day 14. These changes were accompanied by increases in inflammatory biomarkers. The general pattern of faecal microbiota detected was similar to that seen in the relatively few equine gut microbiome studies reported to date. We conclude that interplay between resident cyathostomin populations and the bacterial microbiota of the equine large intestine is important in maintaining homeostasis and that disturbance of this ecology can lead to gut dysbiosis and play a role in the aetiology of inflammatory conditions in the horse, including acute larval cyathostominosis.

Whey protein isolate (WPI) is considered a dietary solution to obesity. However, the exact mechanism of WPI action is still poorly understood but is probably connected to its beneficial effect on energy balance, adiposity, and metabolism. More recently its ability to modulate the gut microbiota has received increasing attention. Here, we used a microbiota depletion, by antibiotic cocktail (ABX) administration, to investigate if the gut microbiota mediates the physiological and metabolic changes observed during high‐fat diet (HFD)‐WPI consumption. C57BL/6J mice received a HFD containing WPI (HFD‐WPI) or the control non‐whey milk protein casein (HFD‐CAS) for 5 or 10 weeks. HFD‐fed mice supplemented with WPI showed reduced body weight gain, adiposity, Ob gene expression level in the epidydimal adipose tissue (eWAT) and plasma leptin relative to HFD‐CAS‐fed mice, after 5‐ or 10‐weeks intervention both with or without ABX treatment. Following 10‐weeks intervention, ABX and WPI had an additive effect in lowering adiposity and leptin availability. HFD‐WPI‐fed mice showed a decrease in the expression of genes encoding pro‐inflammatory markers (MCP‐1, TNFα and CD68) within the ileum and eWAT, compared to HFD‐CAS‐fed mice, without showing alterations following microbiota depletion. Additionally, WPI supplementation decreased HFD‐induced intestinal permeability disruption in the distal ileum; an effect that was reversed by chronic ABX treatment. In summary, WPI reverses the effects of HFD on metabolic and physiological functions through mainly microbiota‐independent mechanisms. Moreover, we demonstrate a protective effect of WPI on HFD‐induced inflammation and ileal permeability disruption, with the latter being reversed by gut microbiota depletion., In this study, we provided new insights on the anti‐obesity effect of whey protein consumption. In this instance, we investigated the role of the gut microbiota by using an antibiotic treatment‐based depletion approach. Moreover, we showed novel findings on the effect of a high‐fat diet containing whey protein on inflammation, intestinal permeability and metabolites profile.

Background/Aims Diarrhea-predominant irritable bowel syndrome (IBS-D) has been previously associated with evidence of immune activation and altered microbiota. Our aim is to assess the effect of the anti-inflammatory agent, mesalazine, on inflammatory gene expression and microbiota composition in IBS-D. Methods We studied a subset of patients (n = 43) from a previously published 12-week radomized placebo-controlled trial of mesalazine. Mucosal biopsies were assessed by immunohistochemistry and reverse transcription-polymerase chain reaction for a range of markers of inflammation, altered permeability, and sensory receptors including Toll-like receptors (TLRs) at randomization after treatment. All biopsy data were compared to 21 healthy controls. Patient's stool microbiota composition was analysed through 16S ribosomal RNA sequencing. Results We found no evidence of increased immune activation compared to healthy controls. However, we did find increased expression of receptors in both sensory pathways and innate immune response including TLR4. Higher TLR4 expression was associated with greater urgency. TLR4 expression correlated strongly with the expression of the receptors bradykinin receptor B2, chemerin chemokine-like receptor 1, and transient receptor potential cation channel, subfamily A, member 1 as well as TLR4's downstream adaptor myeloid differentiation factor 88. Mesalazine had minimal effect on either gene expression or microbiota composition. Conclusions Biopsies from a well-characterized IBS-D cohort showed no substantial inflammation. Mesalazine has little effect on gene expression and its previous reported effect on fecal microbiota associated with much greater inflammation found in inflammatory bowel diseases is likely secondary to reduced inflammation. Increased expression of TLR4 and correlated receptors in IBS may mediate a general increase in sensitivity to external stimuli, particularly those that signal via the TLR system.

The prebiotic activity of a commercially available oat product and a novel oat ingredient, at similar β-glucan loads, was tested using a validated in vitro gut model (M-SHIME®). The novel oat ingredient was tested further at lower β-glucan loads in vitro, while the commercially available oat product was assessed in a randomised, single-blind, placebo-controlled, and cross-over human study. Both approaches focused on healthy individuals with mild hypercholesterolemia. In vitro analysis revealed that both oat products strongly stimulated Lactobacillaceae and Bifidobacteriaceae in the intestinal lumen and the simulated mucus layer, and corresponded with enhanced levels of acetate and lactate with cross-feeding interactions leading to an associated increase in propionate and butyrate production. The in vitro prebiotic activity of the novel oat ingredient remained at lower β-glucan levels, indicating the prebiotic potential of the novel oat product. Finally, the stimulation of Lactobacillus spp. was confirmed during the in vivo trial, where lactobacilli abundance significantly increased in the overall population at the end of the intervention period with the commercially available oat product relative to the control product, indicating the power of in vitro gut models in predicting in vivo response of the microbial community to dietary modulation.

Made available in DSpace on 2022-04-28T17:22:08Z (GMT). No. of bitstreams: 0 Previous issue date: 2021-08-01 Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) Listeria monocytogenes is one of the foodborne pathogens of most concern for food safety. To limit its presence in foods, bacteriocins have been proposed as natural bio-preservatives. Herein, a bacteriocin was produced on hemicellulose hydrolysate of sugarcane bagasse by Pediococcus pentosaceous ET34, whose genome sequencing revealed an operon with 100% similarity to that of pediocin PA-1. ET34 grown on hydrolysate-containing medium led to an increase in the expression of PA-1 genes and a non-optimized purification step sequence resulted in a yield of 0.8 mg & sdot;L-1 of pure pediocin (purity > 95%). Culture conditions were optimized according to a central composite design using temperature and hydrolysate % as independent variables and validated in 3-L Erlenmeyers. Finally, a process for scaled-up implementation by sugar-ethanol industry was proposed, considering green chemistry and biorefinery concepts. This work stands up as an approach addressing a future proper sugarcane bagasse valorisation for pediocin production. Superscript/Subscript Available

We investigated how protein quantity (10%-30%) and quality (casein and whey) interact with dietary fat (20%-55%) to affect metabolic health in adult mice. Although dietary fat was the main driver of body weight gain and individual tissue weight, high (30%) casein intake accentuated and high whey intake reduced the negative metabolic aspects of high fat. Jejunum and liver transcriptomics revealed increased intestinal permeability, low-grade inflammation, altered lipid metabolism, and liver dysfunction in casein-fed but not whey-fed animals. These differential effects were accompanied by altered gut size and microbial functions related to amino acid degradation and lipid metabolism. Fecal microbiota transfer confirmed that the casein microbiota increases and the whey microbiota impedes weight gain. These data show that the effects of dietary fat on weight gain and tissue partitioning are further influenced by the quantity and quality of the associated protein, primarily via effects on the microbiota.

International audience; Kombucha is a fermented tea. Here we investigate the fermentation kinetics, metabolite production, microbiome and potential health promoting properties of three different kombucha consortia. Shotgun metagenomic sequencing revealed several dominant bacterial genera such as Komagataeibacter, Gluconacetobacter and Gluconobacter. Brettanomyces and Schizosaccharomyces were the most dominant yeasts identified. Species distribution reflected different patterns of sugar consumption, with S. pombe being present in samples with the highest sugar conversion. Liquid-liquid extractions were performed with organic solvents in order to obtain dried extracts, which were later characterized. HPLC-DAD and GC-MS analysis revealed differences in the production of organic acids, sugars, alcohols and phenolic compounds, where the presence of caffeine, propanoic acid and 2,3 butanediol differ greatly across the three kombuchas. Metabolomic analysis exhibited a link between the microbiota and the production of bioactive compounds in kombucha fermentation. In vitro assays were carried out in order to evaluate potential health-promoting features of the fermented teas, with notable outcomes including antioxidant ability against DPPH radical and against the 15-lipoxygenase enzyme, indicating a potential anti-inflammatory activity. These investigations considerably enhance our understanding of the relationship between the microbiota and metabolites as well as health promoting potential of kombucha and have the potential for the development of future generations of kombucha products in which these relationships are optimized.

ObjectiveThe composition and metabolic function of the gut microbiome in the elite athlete differs from that of non-athletes. However, short-term fitness improvement in the sedentary adult does not replicate the microbiome characteristics seen in the athlete. Whether sustained fitness improvement over a prolonged period can lead to pronounced and beneficial alteration in the gut microbiome is unknown. The objective was to explore this possibility.MethodsThis study used a repeated-measures, case-study approach to explore changes in the gut microbiome of two unfit volunteers undertaking progressive exercise training over a 6-month period. Training was to culminate in the completion of a marathon or Olympic-distance triathlon. The volunteers were sampled every two weeks for six months and microbiome, metabolome, diet, body composition, and cardiorespiratory fitness data were recorded.ResultsBoth participants completed their respective goals with improved body composition and fitness parameters over the training period. Increases in α-diversity of the gut microbiota occurred with sustained training and fluctuations occurred in response to training events (e.g., injury, illness and training peaks). Participants’ fat mass and BMI reduced during the study and was significantly associated with increased urinary measurements of N-methyl nicotinate (P value < 0.001) and hippurate (P value < 0.05), and decreased phenylacetylglutamine (P value < 0.05).ConclusionThese results suggest that sustained fitness improvements result in alterations to gut microbiota and physiologically-relevant metabolites. This study provides longitudinal analysis of the response of the gut microbiome to real-world events during progressive fitness training, including intercurrent illness and injury.

Food processing environments can harbor microorganisms responsible for food spoilage or foodborne disease. Efficient and accurate identification of microorganisms throughout the food chain can allow the identification of sources of contamination and the timely implementation of control measures. Currently, microbial monitoring of the food chain relies heavily on culture-based techniques. These assays are determined on the microbes expected to be present in the environment, and thus do not cater for unexpected contaminants. Many culture-based assays are also unable to distinguish between undesirable taxa and closely related harmless species. Furthermore, even when multiple culture-based approaches are used in parallel, it is still not possible to comprehensively characterize the entire microbiology of a food-chain sample.High throughput DNA sequencing represents a potential means through which microbial monitoring of the food chain can be enhanced. While sequencing platforms, such as the Illumina MiSeq, NextSeq and NovaSeq, are most typically found in research or commercial sequencing laboratories, newer portable platforms, such as the Oxford Nanopore Technologies (ONT) MinION, offer the potential for rapid analysis of food chain microbiomes. In this study, having initially assessed the ability of rapid MinION-based sequencing to discriminate between different microbes within a simple mock metagenomic mixture of related food spoilage, spore-forming microorganisms. Subsequently, we proceeded to compare the performance of both ONT and Illumina sequencing for environmental monitoring of an active food processing facility.Overall, ONT MinION sequencing provided accurate classification to species level, which was comparable to Illumina-derived outputs. However, while the MinION-based approach provided a means of easy library preparations and portability, the high concentrations of DNA needed to run the rapid sequencing protocols was a limiting factor, requiring the random amplification of template DNA in order to generate sufficient material for analysis.

In recent decades, bacteriocins have received substantial attention as antimicrobial compounds. Although bacteriocins have been predominantly exploited as food preservatives, they are now receiving increased attention as potential clinical antimicrobials and as possible immune-modulating agents. Infections caused by antibiotic-resistant bacteria have been declared as a global threat to public health. Bacteriocins represent a potential solution to this worldwide threat due to their broad- or narrow-spectrum activity against antibiotic-resistant bacteria. Notably, despite their role in food safety as natural alternatives to chemical preservatives, nisin remains the only bacteriocin legally approved by regulatory agencies as a food preservative. Moreover, insufficient data on the safety and toxicity of bacteriocins represent a barrier against the more widespread use of bacteriocins by the food and medical industry. Here, we focus on the most recent trends relating to the application of bacteriocins, their toxicity and impacts., Antimicrobial activity, gastrointestinal bihaviour and toxicity of bacteriocins.

Background Evidence is accruing to suggest that microbiota-gut-brain signalling plays a regulatory role in cardiorespiratory physiology. Chronic intermittent hypoxia (CIH), modelling human sleep apnoea, affects gut microbiota composition and elicits cardiorespiratory morbidity. We investigated if treatment with prebiotics ameliorates cardiorespiratory dysfunction in CIH-exposed rats. Methods Adult male rats were exposed to CIH (96 cycles/day, 6.0% O2 at nadir) for 14 consecutive days with and without prebiotic supplementation (fructo- and galacto-oligosaccharides) beginning two weeks prior to gas exposures. Findings CIH increased apnoea index and caused hypertension. CIH exposure had modest effects on the gut microbiota, decreasing the relative abundance of Lactobacilli species, but had no effect on microbial functional characteristics. Faecal short-chain fatty acid (SCFA) concentrations, plasma and brainstem pro-inflammatory cytokine concentrations and brainstem neurochemistry were unaffected by exposure to CIH. Prebiotic administration modulated gut microbiota composition and diversity, altering gut-metabolic (GMMs) and gut-brain (GBMs) modules and increased faecal acetic and propionic acid concentrations, but did not prevent adverse CIH-induced cardiorespiratory phenotypes. Interpretation CIH-induced cardiorespiratory dysfunction is not dependant upon changes in microbial functional characteristics and decreased faecal SCFA concentrations. Prebiotic-related modulation of microbial function and resultant increases in faecal SCFAs were not sufficient to prevent CIH-induced apnoea and hypertension in our model. Our results do not exclude the potential for microbiota-gut-brain axis involvement in OSA-related cardiorespiratory morbidity, but they demonstrate that in a relatively mild model of CIH, sufficient to evoke classic cardiorespiratory dysfunction, such changes are not obligatory for the development of morbidity, but may become relevant in the elaboration and maintenance of cardiorespiratory morbidity with progressive disease. Funding Department of Physiology and APC Microbiome Ireland, University College Cork, Ireland. APC Microbiome Ireland is funded by Science Foundation Ireland, through the Government's National Development Plan.

Recently there has been a considerable rise in the frequency of metabolic diseases, such as obesity, due to changes in lifestyle and resultant imbalances between energy intake and expenditure. Whey proteins are considered as potentially important components of a dietary solution to the obesity problem. However, the roles of individual whey proteins in energy balance remain poorly understood. This study investigated the effects of a high-fat diet (HFD) containing α-lactalbumin (LAB), a specific whey protein, or the non-whey protein casein (CAS), on energy balance, nutrient transporters expression and enteric microbial populations. C57BL/6J mice (n 8) were given an HFD containing either 20 % CAS or LAB as protein sources or a low-fat diet containing CAS for 10 weeks. HFD-LAB-fed mice showed a significant increase in cumulative energy intake (P=0·043), without differences in body weight, energy expenditure, locomotor activity, RER or subcutaneous and epididymal white adipose tissue weight. HFD-LAB intake led to a decrease in the expression of glut2 in the ileum (P=0·05) and in the fatty acid transporter cd36 (PLactobacillus, Parabacteroides and Bifidobacterium were present in significantly higher proportions in the HFD-LAB group. These data indicate a possible functional relationship between gut microbiota, intestinal nutrient transporters and energy balance, with no impact on weight gain.

Consuming fermented foods has been reported to result in improvements in a range of health parameters. These positive effects can be exerted by a combination of the live microorganisms that the fermented foods contain, as well as the bioactive components released into the foods as by-products of the fermentation process. In many instances, and particularly in dairy fermented foods, the microorganisms involved in the fermentation process belong to the lactic acid group of bacteria (LAB). An alternative approach to making some of the health benefits that have been attributed to fermented foods available is through the production of ‘fermentates’. The term ‘fermentate’ generally relates to a powdered preparation, derived from a fermented product and which can contain the fermenting microorganisms, components of these microorganisms, culture supernatants, fermented substrates, and a range of metabolites and bioactive components with potential health benefits. Here, we provide a brief overview of a selection of in vitro and in vivo studies and patents exclusively reporting the health benefits of LAB ‘fermentates’. Typically, in such studies, the potential health benefits have been attributed to the bioactive metabolites present in the crude fermentates and/or culture supernatants rather than the direct effects of the LAB strain(s) involved.

The COVID-19 pandemic has resulted in increased need for diagnostic testing using reverse transcriptase real-time PCR (RT-PCR). An exponential increase in demand has resulted in a shortage of numerous reagents in particular those associated with the lysis buffer required to extract the viral RNA. Herein, we describe a rapid collective effort by hospital laboratory scientists, academic researchers and the biopharma industry to generate a validated lysis buffer. We have formulated a 4M Guanidinium thiocyanate (GITC)/ Triton X-100 Lysis buffer which provides comparable results with the recommended reagents. This buffer will ease the burden on hospital labs in their heroic efforts to diagnose a large population of patients.

Introduction. Management of steroid-refractory ulcerative colitis has predominantly involved treatment with systemic cyclosporine A (CyA) and infliximab. Aim. The purpose of this study was to assess the effect of using a colon-targeted delivery system CyA formulation on the composition and functionality of the gut microbiota. Methodology. Ex vivo faecal fermentations from six healthy control subjects were treated with coated minispheres (SmPill) with (+) or without (−) CyA and compared with a non-treated control in a model colon system. In addition, the in vivo effect of the SmPill+CyA formulation was investigated by analysing the gut microbiota in faecal samples collected before the administration of SmPill+CyA and after 7 consecutive days of administration from eight healthy subjects who participated in a pilot study. Results. Analysis of faecal samples by 16S rRNA gene sequencing indicated little variation in the diversity or relative abundance of the microbiota composition before or after treatment with SmPill minispheres with or without CyA ex vivo or with CyA in vivo. Short-chain fatty acid profiles were evaluated using gas chromatography, showing an increase in the concentration of n-butyrate (P=0.02) and acetate (P=0.32) in the faecal fermented samples incubated in the presence of SmPill minispheres with or without CyA. This indicated that increased acetate and butyrate production was attributed to a component of the coated minispheres rather than an effect of CyA on the microbiota. Butyrate and acetate levels also increased significantly (P=0.05 for both) in the faecal samples of healthy individuals following 7 days’ treatment with SmPill+CyA in the pilot study. Conclusion. SmPill minispheres with or without CyA at the clinically relevant doses tested here have negligible direct effects on the gut microbiota composition. Butyrate and acetate production increased, however, in the presence of the beads in an ex vivo model system as well as in vivo in healthy subjects. Importantly, this study also demonstrates the relevance and value of using ex vivo colon models to predict the in vivo impact of colon-targeted drugs directly on the gut microbiota.

Nisin P is a natural nisin variant, the genetic determinants for which were previously identified in the genomes of two Streptococcus species, albeit with no confirmed evidence of production. Here we describe Streptococcus agalactiae DPC7040, a human faecal isolate, which exhibits antimicrobial activity against a panel of gut and food isolates by virtue of producing nisin P. Nisin P was purified, and its predicted structure was confirmed by nanoLC-MS/MS, with both the fully modified peptide and a variant without rings B and E being identified. Additionally, we compared its spectrum of inhibition and minimum inhibitory concentration (MIC) with that of nisin A and its antimicrobial effect in a faecal fermentation in comparison with nisin A and H. We found that its antimicrobial activity was less potent than nisin A and H, and we propose a link between this reduced activity and the peptide structure.

As previous studies have demonstrated a link between the porcine intestinal microbiome and feed efficiency (FE), microbiota manipulation may offer a means of improving FE in pigs. A fecal microbiota transplantation procedure (FMTp), using fecal extracts from highly feed-efficient pigs, was performed in pregnant sows (n = 11), with a control group (n = 11) receiving no FMTp. At weaning, offspring were allocated, within sow treatment, to (i) control (n = 67; no dietary supplement) or (ii) inulin (n = 65; 6-week dietary inulin supplementation) treatments. The sow FMTp, alone or in combination with inulin supplementation in offspring, reduced offspring body weight by 8.1 to 10.6 kg at ∼140 days of age, but there was no effect on feed intake. It resulted in better FE, greater bacterial diversity, and higher relative abundances of potentially beneficial bacterial taxa (Fibrobacter and Prevotella) in offspring. Due to the FMTp and/or inulin supplementation, relative abundances of potential pathogens (Chlamydia and Treponema) in the ileum and cecal concentrations of butyric acid were significantly lower. The maternal FMTp led to a greater number of jejunal goblet cells in offspring. Inulin supplementation alone did not affect growth or FE but upregulated duodenal genes linked to glucose and volatile fatty acid homeostasis and increased the mean platelet volume but reduced ileal propionic acid concentrations, granulocyte counts, and serum urea concentrations. Overall, the FMTp in pregnant sows, with or without dietary inulin supplementation in offspring, beneficially modulated offspring intestinal microbiota (albeit mostly low-relative-abundance taxa) and associated physiological parameters. Although FE was improved, the detrimental effect on growth limits the application of this FMTp-inulin strategy in commercial pig production. IMPORTANCE As previous research suggests a link between microbiota and FE, modulation of the intestinal microbiome may be effective in improving FE in pigs. The FMTp in gestating sows, alone or in combination with postweaning dietary inulin supplementation in offspring, achieved improvements in FE and resulted in a higher relative abundance of intestinal bacteria associated with fiber degradation and a lower relative abundance of potential pathogens. However, there was a detrimental effect on growth, although this may not be wholly attributable to microbiota transplantation, as antibiotic and other interventions were also part of the FMT regimen. Therefore, further work with additional control groups is needed to disentangle the effects of each component of the FMTp in order to develop a regimen with practical applications in pig production. Additional research based on findings from this study may also identify specific dietary supplements for the promotion/maintenance of the microbiota transferred via the maternal FMTp, thereby optimizing pig growth and FE.

Soil drainage and phosphorus (P) availability are considered indicators of management intensity in pasture-based agriculture supporting livestock. However, microbial adaptations to P-availability according to soil drainage class are rarely investigated. We hypothesized that well-drained grasslands with high P-availability will sustain a distinctive soil microbiota when compared to poorly-drained grasslands with low-P availability. The relationship between soil drainage, plant available P and grassland microbial communities was evaluated among well-drained sites with high- or low-P and poorly-drained sites with high- or low-P, using fingerprinting, next-generation sequencing and quantitative PCR. Bacterial community structures were primarily affected by drainage as well as significantly separated between a combination of drainage and P availability i.e. low-P and poorly drained versus high-P and well drained. Abundance of the bacterial phylum Actinobacteria was significantly higher in well-drained high-P soils while Firmicutes were more abundant in well-drained low-P soils. Soil fungal communities responded to both drainage status and plant available P. Fungal phyla such as Basidiomycota responded strongly towards availability of P, while Glomeromycota were most abundant in poorly drained low-P soils. The diversity of the alkaline phosphatase gene phoD responded more clearly to drainage than availability of P. Of the other environmental factors, soil pH significantly affected the bacterial and fungal communities structure analyzed. This study suggests that while bacteria and fungi are affected by a combination of soil drainage and P availability in Irish grassland soils, drainage has a more profound influence on bacterial communities than P availability.

Nonstarter lactic acid bacteria are commonly implicated in undesirable gas formation in several varieties, including Cheddar, Dutch-, and Swiss-type cheeses, primarily due to their ability to ferment a wide variety of substrates. This effect can be magnified due to factors that detrimentally affect the composition or activity of starter bacteria, resulting in the presence of greater than normal amounts of fermentable carbohydrates and citrate. The objective of this study was to determine the potential for a facultatively heterofermentative Lactobacillus (Lactobacillus casei DPC6987) isolated from a cheese plant environment to promote gas defects in the event of compromised starter activity. A Swiss-type cheese was manufactured, at pilot scale and in triplicate, containing a typical starter culture (Streptococcus thermophilus and Lactobacillus helveticus) together with propionic acid bacteria. Lactobacillus helveticus populations were omitted in certain vats to mimic starter failure. Lactobacillus casei DPC6987 was added to each experimental vat at 4 log cfu/g. Cheese compositional analysis and X-ray computed tomography revealed that the failure of starter bacteria, in this case L. helveticus, coupled with the presence of a faculatively heterofermentative Lactobacillus (L. casei) led to excessive eye formation during ripening. The availability of excess amounts of lactose, galactose, and citrate during the initial ripening stages likely provided the heterofermentative L. casei with sufficient substrates for gas formation. The accrual of these fermentable substrates was notable in cheeses lacking the L. helveticus starter population. The results of this study are commercially relevant, as they demonstrate the importance of viability of starter populations and the control of specific nonstarter lactic acid bacteria to ensure appropriate eye formation in Swiss-type cheese.

The objective of this study was to identify the carotenoids imparting the orange colour to the rind, and pale yellow color to the core, of selected smear-ripened cheeses. The cheeses investigated were Charloe, Ashbrook, Taleggio, and Limburger, and were sourced from artisanal markets. Samples of the rind and core were extracted using non-polar solvents, followed by saponification to hydrolyze triglycerides to remove fatty acids, and to release carotenoid esters. Extracts were tested using ultra-high pressure liquid chromatograph-diode array detector-high resolution mass spectrometry (UHPLC-DAD-MS and -MS/MS), and identities of α- and β-carotene, lycopene, and β-cryptoxanthin confirmed with authentic standards. β-Carotene was the predominant species in both the rind and core, absorbing ~70% of the signal at 450 nm in all cheese extracts tested, as well as minor quantities of β-cryptoxanthin and α-carotene. Carotenoids unique to the rind included lycopene as well as the rare bacterial carotenoids previously identified in bacterial isolates of cheeses (i.e. decaprenoxanthin, sarcinaxanthin, and echinenone). This is the first detailed characterisation of carotenoids extracted directly from smear-ripened cheeses, and reveals that smear-ripened cheese can contribute both provitamin A carotenoids as well as C50 carotenoids to the human diet.

Pediocin PA −1 is class IIa bacteriocin that displays efficient antimicrobial activity against pathogenic Listeriaspp. This bacteriocin is known to lose activity during long periods of storage especially at non optimal pH, thus reducing its usefulness for the pharmaceutical and food industries. Loss of activity has been attributed to oxidation of the methionine residue at position 31, however, replacement of this residue by leucine results in a peptide with activity equivalent to that of the native peptide. In this work, the heterologous expression of the structural (with Met31 to Leu substitution), accessory and transport genes from pediocin PA-1 operon was carried out in Escherichia coli TunerTM (DE3) cells. The sequences of all genes were redesigned using codon bias for the host and were cloned into an expression vector that allows control of plasmid copy number. The heterologous expression of pediocin Met31Leu was optimized for temperature, induction time, IPTG concentration and plasmid copy number and was evaluated via antimicrobial activity assays against Listeria innocua DPC3572. Maximum activity (2560 AU mL−1) was achieved using low plasmid copy number and 6 h of induction at 37 °C with 1 mM of IPTG. Recombinant pediocin PA-1M31L was successfully purified in 5 steps (>95 % purity) as confirmed by mass spectrometry (4606.27 Da) with a yield of 0.725 mg per liter of culture. This variant showed a similar spectrum of activity to the native pediocin PA-1 and is an interesting alternative for industrial applications due to its greater stability.

peer-reviewed The full text of this article will not be available in ULIR until the embargo expires on the 24/01/2020 The effect of fermentation on the antithrombotic properties of polar lipids in ovine milk has been assessed through the production of yoghurts. The total lipids (TL), total neutral lipids (TNL), and total polar lipids (TPL) were extracted. The fatty acid profiles of all yoghurt polar lipids were analysed by GC-MS. The levels of MUFA increased in the fatty acids of the polar lipids, but there was a reduction in PUFA as milk was fermented to yoghurt. The bioactivity of each lipid extract was assessed against platelet-activating factor (PAF) induced platelet aggregation. All yoghurt polar lipids exhibited potent antithrombotic activities with IC50 values ranging from 45 to 77 µg. Shotgun metagenomics determined the species-level microbial composition and functional potential of the yoghurts. Yoghurts containing L. acidophilus seem to correlate with greater bioactivity. Several phospholipid biosynthetic genes have been identified in the most antithrombotic yoghurts. This study has demonstrated that fermentation enhances the antithrombotic properties of yoghurt polar lipids against PAF.

Bacteria active against multi-drug resistant pathogens, isolated by direct selection of colonies from clover silage samples, produce zones of inhibition against two Gram-negative (Klebsiella pneumoniae Ni9 and Pseudomonas aeruginosa MMA83) and two Gram-positive (Staphylococcus aureus ATCC25923 and Listeria monocytogenes ATCC19111) pathogens. Isolates BGSP7, BGSP9, BGSP11 and BGSP12 produced the largest zones of inhibition against all four pathogens when grown in LB broth with aeration at 37 degrees C. Isolates BGSP7, BGSP9, BGSP11 and BGSP12 were identified as Brevibacillus laterosporus and pulsed field gel electrophoresis and extracellular protein profiles showed that three different strains (BGSP7, BGSP9 and BGSP11) were isolated. A semi-native SDS-PAGE (sodium dodecyl sulphate-polyacrylamide gel electrophoresis) gel overlay assay showed that BGSP7 and BGSP9 produce small antimicrobial molecules of about 1.5 kDa, while BGSP11 produces antimicrobial molecules of 1.5 and 6 kDa active against S. aureus ATCC25923. Amino acid analysis of two antimicrobial molecules (1583.73 Da; from BGSP7 and 1556.31 Da; from BGSP11) revealed that they have a similar composition and differ only by virtue of the presence of a methionine which is present only in BGSP11 molecule. Genome sequencing of the three isolates revealed the presence of gene clusters associated with the production of non-ribosomally synthesized peptides (brevibacillin, bogorol, gramicidin S, plipastatin and tyrocin) and bacteriocins (laterosporulin, a lactococcin 972-like bacteriocin, as well as putative linocin M18, sactipeptide, UviB and lantipeptide-like molecules). Ultimately, the purification of a number of antimicrobial molecules from each isolate suggests that they can be considered as potent biocontrol strains that produce an arsenal of antimicrobial molecules active against Gram-positive and Gram-negative multi-resistant pathogens, fungi and insects.

One hundred human-derived coagulase negative staphylococci (CoNS) were screened for antimicrobial activity using agar-based deferred antagonism assays with a range of indicator bacteria. Based on the findings of the screen and subsequent well assays with cell free supernatants and whole cell extracts, one strain, designated CIT060, was selected for further investigation. It was identified as Staphylococcus capitis and herein we describe the purification and characterisation of the novel bacteriocin that the strain produces. This bacteriocin which we have named capidermicin was extracted from the cell-free supernatant of S. capitis CIT060 and purified to homogeneity using reversed-phase high performance liquid chromatography (RP-HPLC). Matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometric (MS) analysis revealed that the capidermicin peptide has a mass of 5,464 Da. Minimal inhibitory concentration (MIC) experiments showed that capidermicin was active in the micro-molar range against all the Gram-positive bacteria that were tested. Antimicrobial activity was retained over a range of pHs (2–11) and temperatures (10–121°C x 15 mins). The draft genome sequence of S. capitis CIT060 was determined and the genes predicted to be involved in the biosynthesis of capidermicin were identified. These genes included the predicted capidermicin precursor gene, and genes that are predicted to encode a membrane transporter, an immunity protein and a transcriptional regulator. Homology searches suggest that capidermicin is a novel member of the family of class II leaderless bacteriocins.

The emergence and dissemination of antibiotic resistant bacteria is a major medical challenge. Lantibiotics are highly modified bacterially produced antimicrobial peptides that have attracted considerable interest as alternatives or adjuncts to existing antibiotics. Nisin, the most widely studied and commercially exploited lantibiotic, exhibits high efficacy against many pathogens. However, some clinically relevant bacteria express highly specific membrane‐associated nisin resistance proteins. One notable example is the nisin resistance protein (NSR) that acts by cleaving the peptide bond between ring E and the adjacent serine 29, resulting in a truncated peptide with significantly less activity. We utilised a complete bank of bioengineered nisin (nisin A) producers in which the serine 29 residue has been replaced with every alternative amino acid. The nisin A S29P derivative was found to be as active as nisin A against a variety of bacterial targets but, crucially, exhibited a 20‐fold increase in specific activity against a strain expressing the nisin resistance protein. Another derivative, nisin PV, exhibited similar properties but was much less prone to oxidation. This version of nisin with enhanced resistance to specific resistance mechanisms could prove useful in the fight against antibiotic resistant pathogens.

Methane is generated in the foregut of all ruminant animals by the microorganisms present. Dietary manipulation is regarded as the most effective and most convenient way to reduce methane emissions (and in turn energy loss in the animal) and increase nitrogen utilization efficiency. This review examines the impact of diet on bovine rumen function and outlines what is known about the rumen microbiome. Our understanding of this area has increased significantly in recent years due to the application of omics technologies to determine microbial composition and functionality patterns in the rumen. This information can be combined with data on nutrition, rumen physiology, nitrogen excretion and/or methane emission to provide comprehensive insights into the relationship between rumen microbial activity, nitrogen utilisation efficiency and methane emission, with an ultimate view to the development of new and improved intervention strategies.

Bactofencin A is a novel class IId bacteriocin, produced by the intestinal isolate Lactobacillus salivarius DPC6502, which has potent activity against medically significant pathogens including Staphylococcus aureus. This bacteriocin is unusual in that it has a highly cationic N terminus and a single disulfide bond between Cys7 and Cys22, resulting in a large C terminal loop. In this study, a library of synthetic bactofencin A variants were screened against the mastitis isolate, S. aureus DPC5246, to identify key residues responsible for activity. It was apparent that substituting either cysteine of the disulfide bond with either serine or alanine significantly reduced the activity of the bacteriocin, confirming the importance of the C terminal loop. Substituting N terminal amino acids with alanine had no effect on activity, whereas sequential removal of the N terminal positively charged residues resulted in an increasingly inactive peptide. A complete (synthetic) alanine scanning analysis revealed that the residues between Val9 and Gly17 were most affected by substitution suggesting that this area has a major influence on the potency of the bacteriocin. Substituting residues in the loop region between Cys7 and Cys22 for D-amino acid equivalents had a more detrimental effect on activity than L-alanine substitutions. Specifically Y10A, N11A, P15A and T16A are active at 4, 16, 1 and 16 μM respectively while their D equivalents were inactive at 1000 μM, the highest concentration tested. Ultimately, this study identifies the critical features in the primary structure of the bacteriocin which gives it such potent activity against pathogenic staphylococci.

Background Despite the relevance of viral populations, our knowledge of (bacterio) phage populations, i.e., the phageome, suffers from the absence of a “gold standard” protocol for viral DNA extraction with associated in silico sequence processing analyses. To overcome this apparent hiatus, we present here a comprehensive performance evaluation of various protocols and propose an optimized pipeline that covers DNA extraction, sequencing, and bioinformatic analysis of phageome data. Results Five widely used protocols for viral DNA extraction from fecal samples were tested for their performance in removal of non-viral DNA. Moreover, we developed a novel bioinformatic platform, METAnnotatorX, for metagenomic dataset analysis. This in silico tool facilitates a range of read- and assembly-based analyses, including taxonomic profiling using an iterative multi-database pipeline, classification of contigs at genus and species level, as well as functional characterizations of reads and assembled data. Performances of METAnnotatorX were assessed through investigation of seven mother-newborn pairs, leading to the identification of shared phage genotypes, of which two were genomically decoded and characterized. METAnnotatorX was furthermore employed to evaluate a protocol for the identification of contaminant non-viral DNA in sequenced datasets and was exploited to determine the amount of metagenomic data needed for robust evaluation of human adult-derived (fecal) phageomes. Conclusions Results obtained in this study demonstrate that a comprehensive pipeline for analysis of phageomes will be pivotal for future explorations of the ecology of phages in the gut environment as well as for understanding their impact on the physiology and bacterial community kinetics as players of dysbiosis and homeostasis in the gut microbiota. Electronic supplementary material The online version of this article (10.1186/s40168-018-0527-z) contains supplementary material, which is available to authorized users.

peer-reviewed There is a growing recognition of the role the gastrointestinal microbiota plays in health and disease. Ingested antimicrobial proteins and peptides have the potential to alter the gastrointestinal microbiota; particularly if protected from digestion. Nisin is an antimicrobial peptide that is used as a food preservative. This study examined the ability of nisin to affect the murine microbiota when fed to mice in two different starch based matrices; a starch dough comprising raw starch granules and a starch gel comprising starch that was gelatinized and retrograded. The effects of the two starch matrices by themselves on the microbiota were also examined. Following 16S rRNA compositional sequencing, beta diversity analysis highlighted a significant difference (p = 0.001, n = 10) in the murine microbiota between the four diet groups. The differences between the two nisin containing diets were mainly attributable to differences in the nisin release from the starch matrices while the differences between the carriers were mainly attributable to the type of resistant starch they possessed. Indeed, the differences in the relative abundance of several genera in the mice consuming the starch dough and starch gel diets, in particular Akkermansia, the relative abundance of which was 0.5 and 11.9%, respectively (p = 0.0002, n = 10), points to the potential value of resistance starch as a modulator of beneficial gut microbes. Intact nisin and nisin digestion products (in particular nisin fragment 22–31) were detected in the feces and the nisin was biologically active. However, despite a three-fold greater consumption of nisin in the group fed the nisin in starch dough diet, twice as much nisin was detected in the feces of the group which consumed the nisin in starch gel diet. In addition, the relative abundance of three times as many genera from the lower gastrointestinal tract (GIT) were significantly different (p < 0.001, n = 10) to the control for the group fed the nisin in starch gel diet, implying that the starch gel afforded a degree of protection from digestion to the nisin entrapped within it. This work was funded by the Irish Department of Agriculture, Food and the Marine under the FIRM program (grant reference 10/RD/TMFRC/701). RG was funded under the Teagasc Walsh Fellowship scheme (grant reference 2012221). RC, PO, FC, AB, SM, CH, RR, PC, KN and MR were funded by Teagasc, Ireland. RC, PO, FC, CH, RR, and MR were supported in part by a research grant from Science Foundation Ireland (grant reference SFI/12/RC2273). KN was also funded by Science Foundation Ireland under the grant reference SFI/16/BBSRC/3389 and the UK Biotechnology and Biological Sciences Research Council under grant reference BB/P009875/1.

Biofilms are sessile communities of bacteria typically embedded in an extracellular polymeric matrix. Bacterial cells embedded in biofilms are inherently recalcitrant to antimicrobials, compared to cells existing in a planktonic state, and are notoriously difficult to eradicate once formed. Avenues to tackle biofilms thus far have largely focussed on attempting to disrupt the initial stages of biofilm formation, including adhesion and maturation of the biofilm. Such an approach is advantageous as the concentrations required to inhibit formation of biofilms are generally much lower than removing a fully established biofilm. The crisis of antibiotic resistance in clinical settings worldwide has been further exacerbated by the ability of certain pathogenic bacteria to form biofilms. Perhaps the most notorious biofilm formers described from a clinical viewpoint have been methicillin-resistant Staphylococcus aureus (MRSA), Staphylococcus epidermidis, Pseudomonas aeruginosa, Gardnerella vaginalis and Streptococcus mutans, the latter of which is found in oral biofilms. Due to the dearth of novel antibiotics in recent decades, compounded by the increasing rate of emergence of resistance amongst pathogens with a propensity for biofilm formation, solutions are urgently required to mitigate these crises. Bacteriocins are a class of antimicrobial peptides, which are ribosomally synthesised and often are more potent than their antibiotic counterparts. Here, we review a selection of studies conducted with bacteriocins with the ultimate objective of inhibiting biofilms. Overall, a deeper understanding of the precise means by which a biofilm forms on a substrate as well as insights into the mechanisms by which bacteriocins inhibit biofilms is warranted.

The generation of the antimicrobial peptide caseicin A (IKHQGLPQE) from a casein substrate using proteolytic enzymes was assessed in silico. This bioinformatic analysis predicted that thermolysin (EC 3.4.24.27) and thermolysin-like enzymes (EC 3.4.24.28) are likely candidates to liberate the bioactive peptide from αS1-casein. Commercially available sources of the thermolysin enzyme from industrial suppliers were subsequently shown to liberate the 1049 Da caseicin A peptide, under various conditions of hydrolysis, at both lab and pilot scale. The antimicrobial ability of the hydrolysates to reduce pathogen numbers spiked in infant formula trials was subsequently confirmed. For example, numbers of Cronobacter sakazakii were reduced 1000 fold after 3 h of incubation at 37 °C. In conclusion, this study demonstrates the potential to improve the safety of infant milk formula using milk-derived bioactive peptides.

Accumulating evidence suggests that bacteriocin production represents a probiotic trait for intestinal strains to promote dominance, fight infection, and even signal the immune system. In this respect, in a previous study, we isolated from the porcine intestine a strain of Streptococcus hyointestinalis DPC6484 that displays antimicrobial activity against a wide range of Gram-positive bacteria and produces a bacteriocin with a mass of 3,453 Da. Interestingly, the strain was also found to be immune to a nisin-producing strain. Genome sequencing revealed the genetic determinants responsible for a novel version of nisin, designated nisin H, consisting of the nshABTCPRKGEF genes, with transposases encoded between nshP and nshR and between nshK and nshG . A similar gene cluster is also found in S. hyointestinalis LMG14581. Notably, the cluster lacks an equivalent of the nisin immunity gene, nisI . Nisin H is proposed to have the same structure as the prototypical nisin A but differs at 5 amino acid positions—Ile1Phe (i.e., at position 1, nisin A has Ile while nisin H has Phe), Leu6Met, Gly18Dhb (threonine dehydrated to dehydrobutyrine), Met21Tyr, and His31Lys—-and appears to represent an intermediate between the lactococcal nisin A and the streptococcal nisin U variant of nisin. Purified nisin H inhibits a wide range of Gram-positive bacteria, including staphylococci, streptococci, Listeria spp., bacilli, and enterococci. It represents the first example of a natural nisin variant produced by an intestinal isolate of streptococcal origin.

Fermented foods, in particular, surface-ripened cheese, represent a model to explain the metabolic interactions which regulate microbial succession in complex environments. This study explains the role of individual species in a heterogeneous microbial environment, i.e., the exterior of surface-ripened cheese. Through whole-metagenome shotgun sequencing, it was possible to investigate the metabolic potential of the resident microorganisms and show how variations in the microbial populations influence important aspects of cheese ripening, especially flavor development. Overall, in addition to providing fundamental insights, this research has considerable industrial relevance relating to the production of fermented food with specific qualities., In this study, a young Cheddar curd was used to produce two types of surface-ripened cheese, using two commercial smear-culture mixes of yeasts and bacteria. Whole-metagenome shotgun sequencing was used to screen the microbial population within the smear-culture mixes and on the cheese surface, with comparisons of microorganisms at both the species and the strain level. The use of two smear mixes resulted in the development of distinct microbiotas on the surfaces of the two test cheeses. In one case, most of the species inoculated on the cheese established themselves successfully on the surface during ripening, while in the other, some of the species inoculated were not detected during ripening and the most dominant bacterial species, Glutamicibacter arilaitensis, was not a constituent of the culture mix. Generally, yeast species, such as Debaryomyces hansenii and Geotrichum candidum, were dominant during the first stage of ripening but were overtaken by bacterial species, such as Brevibacterium linens and G. arilaitensis, in the later stages. Using correlation analysis, it was possible to associate individual microorganisms with volatile compounds detected by gas chromatography-mass spectrometry in the cheese surface. Specifically, D. hansenii correlated with the production of alcohols and carboxylic acids, G. arilaitensis with alcohols, carboxylic acids and ketones, and B. linens and G. candidum with sulfur compounds. In addition, metagenomic sequencing was used to analyze the metabolic potential of the microbial populations on the surfaces of the test cheeses, revealing a high relative abundance of metagenomic clusters associated with the modification of color, variation of pH, and flavor development. IMPORTANCE Fermented foods, in particular, surface-ripened cheese, represent a model to explain the metabolic interactions which regulate microbial succession in complex environments. This study explains the role of individual species in a heterogeneous microbial environment, i.e., the exterior of surface-ripened cheese. Through whole-metagenome shotgun sequencing, it was possible to investigate the metabolic potential of the resident microorganisms and show how variations in the microbial populations influence important aspects of cheese ripening, especially flavor development. Overall, in addition to providing fundamental insights, this research has considerable industrial relevance relating to the production of fermented food with specific qualities.

Objective It is evident that the gut microbiota and factors that influence its composition and activity effect human metabolic, immunological and developmental processes. We previously reported that extreme physical activity with associated dietary adaptations, such as that pursued by professional athletes, is associated with changes in faecal microbial diversity and composition relative to that of individuals with a more sedentary lifestyle. Here we address the impact of these factors on the functionality/metabolic activity of the microbiota which reveals even greater separation between exercise and a more sedentary state. Design Metabolic phenotyping and functional metagenomic analysis of the gut microbiome of professional international rugby union players (n=40) and controls (n=46) was carried out and results were correlated with lifestyle parameters and clinical measurements (eg, dietary habit and serum creatine kinase, respectively). Results Athletes had relative increases in pathways (eg, amino acid and antibiotic biosynthesis and carbohydrate metabolism) and faecal metabolites (eg, microbial produced short-chain fatty acids (SCFAs) acetate, propionate and butyrate) associated with enhanced muscle turnover (fitness) and overall health when compared with control groups. Conclusions Differences in faecal microbiota between athletes and sedentary controls show even greater separation at the metagenomic and metabolomic than at compositional levels and provide added insight into the diet–exercise–gut microbiota paradigm.

Bacteriocins from lactic acid bacteria (LAB) are of increasing interest in recent years due to their potential as natural preservatives against food and beverage spoilage microorganisms. In a screening study for LAB, we isolated from olives a strain, Lactobacillus plantarum NI326, with activity against the beverage-spoilage bacterium Alicyclobacillus acidoterrestris . Genome sequencing of NI326 enabled the identification of a gene cluster (designated plc ) encoding a putative circular bacteriocin and proteins involved in its modification, transport, and immunity. This novel bacteriocin, named plantaricyclin A (PlcA), was grouped into the circular bacteriocin subgroup II due to its high degree of similarity with other gassericin A-like bacteriocins. Purification of PlcA from the supernatant of Lb. plantarum NI326 resulted in an active peptide with a molecular mass of 5,570 Da, corresponding to that predicted from the (processed) PlcA amino acid sequence. The plc gene cluster was cloned and expressed in Lactococcus lactis NZ9000, resulting in the production of an active 5,570-Da bacteriocin in the supernatant. PlcA is believed to be produced as a 91-amino-acid precursor with a 33-amino-acid leader peptide, which is predicted to be removed, followed by joining of the N and C termini via a covalent linkage to form the mature 58-amino-acid circular bacteriocin PlcA. We report the characterization of a circular bacteriocin produced by Lb. plantarum . The inhibition displayed against A. acidoterrestris highlights its potential use as a preservative in food and beverages. IMPORTANCE In this work, we describe the purification and characterization of an antimicrobial peptide, termed plantaricyclin A (PlcA), produced by a Lactobacillus plantarum strain isolated from olives. This peptide has a circular structure, and all genes involved in its production, circularization, and secretion were identified. PlcA shows antimicrobial activity against different strains, including Alicyclobacillus acidoterrestris , a common spoilage bacterium, which causes substantial economic losses in the beverage industry every year. In this study, we describe a circular antimicrobial peptide, PlcA, for a Lactobacillus plantarum strain.

The main aim of the present study was to investigate the effects of dietarytrans-10,cis-12-conjugated linoleic acid (t10c12-CLA) on intestinal microbiota composition and SCFA production. C57BL/6 mice (n8 per group) were fed a standard diet either supplemented witht10c12-CLA (0·5 %, w/w) (intervention) or with no supplementation (control), daily for 8 weeks. Metabolic markers (serum glucose, leptin, insulin and TAG, and liver TAG) were assessed by ELISA commercial kits, tissue long-chain fatty acids and caecal SCFA by GC, and microbial composition by 16S rRNA pyrosequencing. Dietaryt10c12-CLA significantly decreased visceral fat mass (Pt10c12-CLA intake. Caecal acetate, propionate and isobutyrate concentrations were higher (Pt10c12-CLA-supplemented group than in the control group. The analysis of the microbiota composition following 8 weeks oft10c12-CLA supplementation revealed lower proportions of Firmicutes (P= 0·003) and higher proportions of Bacteroidetes (P= 0·027) compared with no supplementation. Furthermore,t10c12-CLA supplementation for 8 weeks significantly altered the gut microbiota composition, harbouring higher proportions of Bacteroidetes, including Porphyromonadaceae bacteria previously linked with negative effects on lipid metabolism and induction of hepatic steatosis. These results indicate that the mechanism of dietaryt10c12-CLA on lipid metabolism in mice may be, at least, partially mediated by alterations in gut microbiota composition and functionality.

peer-reviewed Centrifugation is a common milk pretreatment method for removal of Clostridium spores which, on germination, can produce high levels of butyric acid and gas, resulting in rancid, gassy cheese. The aim of this study was to determine the effect of centrifugation of milk, as well as incorporation of high heat-treated centrifugate into cheese milk, on the microbial and volatile profile of Maasdam cheese. To facilitate this, 16S rRNA amplicon sequencing in combination with a selective media-based approach were used to study the microbial composition of cheese during maturation, and volatile organic compounds within the cheese matrix were analyzed by HPLC and solid-phase microextraction coupled with gas chromatography–mass spectrometry. Both culture-based and molecular approaches revealed major differences in microbial populations within the cheese matrix before and after warm room ripening. During warm room ripening, an increase in counts of propionic acid bacteria (by ∼101.5 cfu) and nonstarter lactic acid bacteria (by ∼108 cfu) and a decrease in the counts of Lactobacillus helveticus (by ∼102.5 cfu) were observed. Lactococcus species dominated the curd population throughout ripening, followed by Lactobacillus, Propionibacterium, and Leuconostoc, and the relative abundance of these accounted for more than 99% of the total genera, as revealed by high-throughput sequencing. Among subdominant microflora, the overall relative abundance of Clostridium sensu stricto was lower in cheeses made from centrifuged milk than control cheeses, which coincided with lower levels of butyric acid. Centrifugation as well as incorporation of high heat-treated centrifugate into cheese milk seemed to have little effect on the volatile profile of Maasdam cheese, except for butyric acid levels. Overall, this study suggests that centrifugation of milk before cheesemaking is a suitable method for controlling undesirable butyric acid fermentation without significantly altering the levels of other volatile organic compounds of Maasdam cheese.

Probiotic bacteria have been associated with a reduction in cardiovascular disease risk, a leading cause of death and disability.The aim of this study was to assess the impact of dietary administration of exopolysaccharide-producing probiotic Lactobacillus cultures on lipid metabolism and gut microbiota in apolipoprotein E (apoE)-deficient mice.First, we examined lipid metabolism in response to dietary supplementation with recombinant β-glucan-producing Lactobacillus paracasei National Food Biotechnology Centre (NFBC) 338 expressing the glycosyltransferase (Gtf) gene from Pediococcus parvulus 2.6 (GTF), and naturally exopolysaccharide-producing Lactobacillus mucosae Dairy Product Culture Collection (DPC) 6426 (DPC 6426) compared with the non-β-glucan-producing isogenic control strain Lactobacillus paracasei NFBC 338 (PNZ) and placebo (15% wt:vol trehalose). Second, we examined the effects on the gut microbiota of dietary administration of DPC 6426 compared with placebo. Probiotic Lactobacillus strains at 1 × 10(9) colony-forming units/d per animal were administered to apoE(-/-) mice fed a high-fat (60% fat)/high-cholesterol (2% wt:wt) diet for 12 wk. At the end of the study, aortic plaque development and serum, liver, and fecal variables involved in lipid metabolism were analyzed, and culture-independent microbial analyses of cecal content were performed.Total cholesterol was reduced in serum (P0.001; ∼33-50%) and liver (P0.05; ∼30%) and serum triglyceride concentrations were reduced (P0.05; ∼15-25%) in mice supplemented with GTF or DPC 6426 compared with the PNZ or placebo group, respectively. In addition, dietary intervention with GTF led to increased amounts of fecal cholesterol excretion (P0.05) compared with all other groups. Compositional sequencing of the gut microbiota revealed a greater prevalence of Porphyromonadaceae (P = 0.001) and Prevotellaceae (P = 0.001) in the DPC 6426 group and lower proportions of Clostridiaceae (P0.05), Peptococcaceae (P0.001), and Staphylococcaceae (P0.01) compared with the placebo group.Ingestion of exopolysaccharide-producing lactobacilli resulted in seemingly favorable improvements in lipid metabolism, which were associated with changes in the gut microbiota of mice.

Antibiotic resistance among pathogenic microorganisms is becoming ever more common. Unfortunately, the development of new antibiotics which may combat resistance has decreased. Recently, however the oceans and the marine animals that reside there have received increased attention as a potential source for natural product discovery. Many marine eukaryotes interact and form close associations with microorganisms that inhabit their surfaces, many of which can inhibit the attachment, growth or survival of competitor species. It is the bioactive compounds responsible for the inhibition that is of interest to researchers on the hunt for novel bioactives. The genus Pseudovibrio has been repeatedly identified from the bacterial communities isolated from marine surfaces. In addition, antimicrobial activity assays have demonstrated significant antimicrobial producing capabilities throughout the genus. This review will describe the potency, spectrum and possible novelty of the compounds produced by these bacteria, while highlighting the capacity for this genus to produce natural antimicrobial compounds which could be employed to control undesirable bacteria in the healthcare and food production sectors.

OBJECTIVES: Irritable Bowel Syndrome (IBS) is a functional gastrointestinal (GI) disorder featuring chronic or recurrent abdominal discomfort, usually with changes in GI habit. To improve our understanding of links between the microbiome and IBS, and how these links can be manipulated through diet, we undertook shotgun metagenomic sequencing of fecal samples from a case-control study. METHODS: Fecal samples from 172 individuals were analyzed by shotgun sequencing using the Illumina NextSeq platform. Of these, 77 were classified as controls, 16 were constipation-predominant IBS (IBS-C), 39 were diarrhea-predominant IBS (IBS-D), 29 were diagnosed with functional constipation (FC), and 11 had functional diarrhea (FD). Taxonomic classifications were determined using Metaxa2 and the SILVA 128 database. Gene functions were assigned by alignment of sequences against a protein reference database using DIAMOND. Mean relative abundance of bacterial taxa and functional genes were compared using permutation ANOVA. Ethical approval was obtained from the University of Otago Human Ethics Committee (Health) (Reference H16/094). RESULTS: Bacterial genera that discriminated case-controls (P

Different dietary fat and energy subtypes have an impact on both the metabolic health and the intestinal microbiota population of the host. The present study assessed the impact of dietary fat quality, with a focus on dietary fatty acid compositions of varying saturation, on the metabolic health status and the intestinal microbiota composition of the host. C57BL/6J mice (n9–10 mice per group) were fed high-fat (HF) diets containing either (1) palm oil, (2) olive oil, (3) safflower oil or (4) flaxseed/fish oil for 16 weeks and compared with mice fed low-fat (LF) diets supplemented with either high maize starch or high sucrose. Tissue fatty acid compositions were assessed by GLC, and the impact of the diet on host intestinal microbiota populations was investigated using high-throughput 16S rRNA sequencing. Compositional sequencing analysis revealed that dietary palm oil supplementation resulted in significantly lower populations of Bacteroidetes at the phylum level compared with dietary olive oil supplementation (PBacteroidaceaecompared with dietary supplementation of palm oil, flaxseed/fish oil and high sucrose (PPBifidobacteriumat the genus level compared with the LF-high-maize starch diet (P

peer-reviewed Lactic acid bacteria (LAB) produce a wide variety of antimicrobial peptides (bacteriocins) which contribute to the safety and preservation of fermented foods. This review discusses strategies that have been or could be employed to further enhance the commercial application of bacteriocins and/or bacteriocin-producing LAB for for food use. Food Institutional Research Measure of the Department of Agriculture, Fisheries and Food (grant no. 04R and DC); Science Foundation of Ireland (grant no. 07/CE/B1368).

Cronobacter sakazakii and Escherichia coli O157:H7 are well known food-borne pathogens that can cause severe disease. The identification of new alternatives to heating to control these pathogens in foods, while reducing the impact on organoleptic properties and nutritional value, is highly desirable. In this study, nisin and its bioengineered variants, nisin V and nisin S29A, are used alone, or in combination with plant essential oils (thymol, carvacrol and trans-cinnamaldehyde) or citric acid, with a view to controlling C. sakazakii and E. coli O157:H7 in laboratory-based assays and model food systems. The use of nisin variants (30 μM) with low concentrations of thymol (0.015%), carvacrol (0.03%) and trans-cinnamaldehyde (0.035%) resulted in extended lag phases of growth compared to those for corresponding nisin A-essential oil combinations. Furthermore, nisin variants (60 μM) used in combination with carvacrol (0.03%) significantly reduced viable counts of E. coli O157:H7 (3-log) and C. sakazakii (4-log) compared to nisin A-carvacrol treatment. Importantly, this increased effectiveness translated into food. More specifically, sub-inhibitory concentrations of nisin variants and carvacrol caused complete inactivation of E. coli O157:H7 in apple juice within 3 h at room temperature compared to that of the equivalent nisin A combination. Furthermore, combinations of commercial Nisaplin and the food additive citric acid reduced C. sakazakii numbers markedly in infant formula within the same 3 h period. These results highlight the potential benefits of combining nisin and variants thereof with carvacrol and/or citric acid for the inhibition of Gram negative food-borne pathogens.