RSS 1.0Food Biosciences
The core objective of the Food Biosciences Department is to engage in advanced research and technology development in support of the Irish Agri-Food industry sector. Activities fall into three research areas: Food for Health; Cheese Microbiology and Biochemistry and Milk and Product Quality.
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Food Biosciences
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Diverse Bacteriocins Produced by Strains From the Human Milk MicrobiotaMicrobial colonization of the infant gut is a convoluted process dependent on numerous contributing factors, including age, mode of delivery and diet among others that has lifelong implication for human health. Breast milk also contains a microbiome which acts as a source of colonizing bacteria for the infant. Here, we demonstrate that human milk harbors a wide diversity of bacteriocin-producing strains with the potential to compete among the developing gut microbiota of the infant. We screened 37 human milk samples and found isolates with antimicrobial activity and distinct cross-immunity profiles. From these isolates, we detected 73 putative gene clusters for bacteriocins of all known sub-classes, including 16 novel prepeptides. More specifically, we detected two novel lantibiotics, four sactibiotics and three class IIa bacteriocins with an unusual modification of the pediocin box that is composed of YDNGI instead of the highly conserved motif YGNGV. Moreover, we identified a novel class IIb bacteriocin, four novel class IIc and two class IId bacteriocins. In conclusion, human milk contains a variety of bacteriocin-producing strains which may provide them a competitive advantage in the colonization of the infant gut and suggests that the milk microbiota is a source of antimicrobial potential.
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Dairy Products and Dairy-Processing Environments as a Reservoir of Antibiotic Resistance and Quorum-Quenching Determinants as Revealed through Functional MetagenomicsHere, the role of the dairy-processing chain as a reservoir of antimicrobial resistance (AR) determinants and a source of novel biocontrol quorum-sensing inhibitors is assessed through a functional metagenomics approach. A metagenomic library comprising ∼22,000 recombinant clones was built from DNA isolated from raw milk, raw milk cheeses, and cheese-processing environment swab samples. The high-throughput sequencing of 9,216 recombinant clones showed that lactic acid bacteria (LAB) dominated the microbial communities of raw milk cheese, while Gram-negative microorganisms of animal or soil origin dominated the microbiota of raw milk and cheese-processing environments. Although functional screening of the metagenomic library did not recover potential quorum-sensing inhibitors, in silico analysis using an in-house database built specifically for this study identified homologues to several genes encoding proteins with predicted quorum-quenching activity, among which, the QsdH hydrolase was the most abundant. In silico screening of the library identified LAB, and especially Lactococcus lactis, as a relevant reservoir of AR determinants in cheese. Functional screening of the library allowed the isolation of 13 recombinant clones showing an increased resistance toward ampicillin, which in all cases was accompanied by a reduced susceptibility to a wide range of β-lactam antibiotics. This study shows that the dairy-processing environment is a rich reservoir of AR determinants, which vary by sample source, and suggests that combining next-generation sequencing with functional metagenomics can be of use in overcoming the limitations of both approaches. IMPORTANCE The study shows the potential of functional metagenomics analyses to uncover the diversity of functions in microbial communities prevailing in dairy products and their processing environments, evidencing that lactic acid bacteria (LAB) dominate the cheese microbiota, whereas Gram-negative microorganisms of animal or soil origin dominate the microbiota of milk and cheese-processing environments. The functional and in silico screening of the library allowed the identification of LAB, and especially Lactococcus lactis, as a relevant reservoir of antimicrobial resistance (AR) determinants in cheese. Quorum-quenching (QQ) determinants were not recovered through the execution of wet-lab function-based screenings but were detected through in silico sequencing-based analyses.
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Bifidobacterium and Lactobacillus Composition at Species Level and Gut Microbiota Diversity in Infants before 6 WeeksOur objective was to investigate the effects of different delivery and feeding modes on the gut microbiota composition of early infants with special emphasis on Bifidobacterium and Lactobacillus profiles at species level. 16S rRNA V3-V4 regions, bifidobacterial, and lactobacilli groEL genes from infant feces were sequenced by Illumina MiSeq. Gut microbiota abundance was significantly different, where standard vaginally delivered (SVD) and breast-fed (BF) groups were higher in comparison with caesarean section (CS), milk-powder-fed (MPF), and mixed-fed (MF) groups. The genus unclassified Enterobacteriaceae was dominant, followed by Bifidobacterium, which was highly abundant in SVD and BF groups. The dominant Bifidobacterium species in all groups were B. longum subsp. longum, B. longum subsp. infantis and B. animalis subsp. lactis. B. dentium and the diversity of Bifidobacterium in SVD and BF groups were significantly higher. For Lactobacillus profiles, L. rhamnosus and L. gasseri were dominant among all the groups, while Lactobacillus species in CS and MPF groups were more diverse. Functional predictions showed significant differences between delivery mode and feeding groups, such as phosphotransferase system as well as taurine and hypotaurine metabolism. In early infants with different delivery and feeding methods, gut microbiota—particularly bifidobacteria and lactobacilli communities—showed significant differences, with strong implications for physiological functions.
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Effects of a polysaccharide-rich extract derived from Irish-sourced Laminaria digitata on the composition and metabolic activity of the human gut microbiota using an in vitro colonic modelBackground Brown seaweeds are known to be a rich source of fiber with the presence of several non-digestible polysaccharides including laminarin, fucoidan and alginate. These individual polysaccharides have previously been shown to favorably alter the gut microbiota composition and activity albeit the effect of the collective brown seaweed fiber component on the microbiota remains to be determined. Methods This study investigated the effect of a crude polysaccharide-rich extract obtained from Laminaria digitata (CE) and a depolymerized CE extract (DE) on the gut microbiota composition and metabolism using an in vitro fecal batch culture model though metagenomic compositional analysis using 16S rRNA FLX amplicon pyrosequencing and short-chain fatty acid (SCFA) analysis using GC-FID. Results Selective culture analysis showed no significant changes in cultured lactobacilli or bifidobacteria between the CE or DE and the cellulose-negative control at any time point measured (0, 5, 10, 24, 36, 48 h). Following metagenomic analysis, the CE and DE significantly altered the relative abundance of several families including Lachnospiraceae and genera including Streptococcus, Ruminococcus and Parabacteroides of human fecal bacterial populations in comparison to cellulose after 24 h. The concentrations of acetic acid, propionic acid, butyric acid and total SCFA were significantly higher for both the CE and DE compared to cellulose after 10, 24, 36 and 48 h fermentation (p < 0.05). Furthermore, the acetate:propionate ratio was significantly reduced (p < 0.05) for both CD and DE following 24, 36 and 48 h fermentation. Conclusion The microbiota-associated metabolic and compositional changes noted provide initial indication of putative beneficial health benefits of L. digitata in vitro; however, research is needed to clarify if L. digitata-derived fiber can favorably alter the gut microbiota and confer health benefits in vivo.
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Isolation and Characterization of Listeria monocytogenes Phage vB_LmoH_P61, a Phage With Biocontrol Potential on Different Food MatricesThe high mortality rate associated with Listeria monocytogenes as well as its ability to adapt to the harsh conditions employed in food processing have ensured that this pathogen has become a significant concern in the ready-to-eat food industry. Lytic bacteriophages are viruses that hijack the metabolic mechanisms of their bacterial host as a means to grow and replicate, subsequently leading to host cell death due to lysis. This study reports the biological and genomic characterization of L. monocytogenes phage vB_LmoH_P61 (P61) and its potential application for the reduction of L. monocytogenes in artificially contaminated foods. Phage P61 is a virulent bacteriophage belonging to the family Herelleviridae and has a genome size of 136,485bp. The lytic spectrum of phage P61 was investigated and it was shown to infect serotypes 1/2a, 1/2b, 1/2c, 4b, 4e, and 6a. Treatment of artificially contaminated milk stored at 8 and 12°C with phage P61 resulted in a significant reduction in L. monocytogenes numbers over the product shelf life. Similarly, phage P61 reduced L. monocytogenes numbers on artificially contaminated baby spinach stored at 8, 12, and 25°C. The research findings indicate that biocontrol of L. monocytogenes with phage P61 may offer a safe and environmentally friendly approach for the reduction of L. monocytogenes numbers in certain ready-to-eat foods.
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Future of Probiotics and Prebiotics and the Implications for Early Career ResearchersThe opportunities in the fields of probiotics and prebiotics to a great degree stem from what we can learn about how they influence the microbiota and interact with the host. We discuss recent insights, cutting-edge technologies and controversial results from the perspective of early career researchers innovating in these areas. This perspective emerged from the 2019 meeting of the International Scientific Association for Probiotics and Prebiotics - Student and Fellows Association (ISAPP-SFA). Probiotic and prebiotic research is being driven by genetic characterization and modification of strains, state-of-the-art in vitro, in vivo, and in silico techniques designed to uncover the effects of probiotics and prebiotics on their targets, and metabolomic tools to identify key molecules that mediate benefits on the host. These research tools offer unprecedented insights into the functionality of probiotics and prebiotics in the host ecosystem. Young scientists need to acquire these diverse toolsets, or form inter-connected teams to perform comprehensive experiments and systematic analysis of data. This will be critical to identify microbial structure and co-dependencies at body sites and determine how administered probiotic strains and prebiotic substances influence the host. This and other strategies proposed in this review will pave the way for translating the health benefits observed during research into real-life outcomes. Probiotic strains and prebiotic products can contribute greatly to the amelioration of global issues threatening society. The intent of this article is to provide an early career researcher’s perspective on where the biggest opportunities lie to advance science and impact human health.
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Complete Genome Sequence of the Gamma-Aminobutyric Acid-Producing Strain Streptococcus thermophilus APC151Here is presented the whole-genome sequence of Streptococcus thermophilus APC151, isolated from a marine fish. This bacterium produces gamma-aminobutyric acid (GABA) in high yields and is biotechnologically suitable to produce naturally GABAenriched biofunctional yogurt. Its complete genome comprises 2,097 genes and 1,839,134 nucleotides, with an average GC content of 39.1%.
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Separation of the effects of denaturation and aggregation on whey-casein protein interactions during the manufacture of a model infant formulaDenaturation and aggregation of whey protein have been extensively studied but there is limited knowledge of their effects on processing properties of infant milk formulae (IMF) systems. In this study, the separate effects of denaturation and aggregation of whey protein on the physicochemical characteristics during processing of a model IMF were examined. Whey protein solutions (8%, w/w, protein) were pre-heated for 2 min at 72 or 85 °C, followed by addition of 2.2 mM calcium (Ca) (H-BCa), or at 85 °C after addition of the same level of Ca (H-ACa), to give pre-treated whey protein for inclusion in three model IMF systems, encoded as H-72-BCa, H-85-BCa and H-85-ACa, respectively. Unheated control samples without (UH-C) and with (UH-C-Ca) added Ca were also prepared. Model IMF systems (5.2%, w/w, protein, 60:40 whey protein:casein ratio, pH 6.8) were then prepared incorporating these pre-treated whey protein ingredients and subjected to lab-scale high-temperature short-time (HTST) heating at 85 °C for 2 min; whey protein denaturation was >81.2% in all samples after HTST. Aggregation of whey protein resulted in a significantly (P < 0.05) higher viscosity in sample H-85-ACa (8.3 mPa.s) compared to UH-C (4.0 mPa.s), and measurement of Ca ion concentration on heating showed that Ca ions enhanced whey protein aggregation, resulting in larger mean protein particle size. The results also suggest that pre-heating of whey protein had a preventative effect on aggregation of protein during HTST of IMF. This study clearly showed that aggregation is more influential than denaturation in determining viscosity development during HTST treatment of IMF, and that such viscosity development can be controlled by altering protein-protein interactions using, for example, pre-heat treatment of whey protein ingredients.
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Forgotten fungi—the gut mycobiome in human health and diseaseThe human body is home to a complex and diverse microbial ecosystem that plays a central role in host health. This includes a diversity of fungal species that is collectively referred to as our ‘mycobiome’. Although research into the mycobiome is still in its infancy, its potential role in human disease is increasingly recognised. Here we review the existing literature available on the human mycobiota with an emphasis on the gut mycobiome, including how fungi interact with the human host and other microbes. In doing so, we provide a comprehensive critique of the methodologies available to research the human mycobiota as well as highlighting the latest research findings from mycological surveys of different groups of interest including infants, obese and inflammatory bowel disease cohorts. This in turn provides new insights and directions for future studies in this burgeoning research area.
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Glycomacropeptide Reduces Intestinal Epithelial Cell Barrier Dysfunction and Adhesion of Entero-Hemorrhagic and Entero-Pathogenic Escherichia coli in VitroIn recent years, the potential of glycosylated food components to positively influence health has received considerable attention. Milk is a rich source of biologically active glycoconjugates which are associated with antimicrobial, immunomodulatory, anti-adhesion, anti-inflammatory and prebiotic properties. Glycomacropeptide (GMP) is the C-terminal portion of kappa-casein that is released from whey during cheese-making by the action of chymosin. Many of the biological properties associated with GMP, such as anti-adhesion, have been linked with the carbohydrate portion of the protein. In this study, we investigated the ability of GMP to inhibit the adhesion of a variety of pathogenic Escherichia coli strains to HT-29 and Caco-2 intestinal cell lines, given the importance of E. coli in causing bacterial gastroenteritis. GMP significantly reduced pathogen adhesion, albeit with a high degree of species specificity toward enteropathogenic E. coli (EPEC) strains O125:H32 and O111:H2 and enterohemorrhagic E. coli (EHEC) strain 12900 O157:H7. The anti-adhesive effect resulted from the interaction of GMP with the E. coli cells and was also dependent on GMP concentration. Pre-incubation of intestinal Caco-2 cells with GMP reduced pathogen translocation as represented by a decrease in transepithelial electrical resistance (TEER). Thus, GMP is an effective in-vitro inhibitor of adhesion and epithelial injury caused by E. coli and may have potential as a biofunctional ingredient in foods to improve gastrointestinal health.
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Boarfish (Capros aper): review of a new capture fishery and its valorization potentialThe world’s fish stocks, although renewable, are a finite resource. European capture fisheries have remained stagnant in terms of volume for many years. To remain profitable, fishers are looking for new opportunities to diversify, reduce costs, and maximize profits. The targeted fishing of boarfish (Capros aper) in Europe is an excellent example of such adaptation. Using this fishery as a case study, we highlight how established fisheries are adapting to changes faced by the industry. We begin by compiling the knowledge to date on the taxonomy, biology, and ecology of the understudied boarfish and go on to provide a comprehensive overview of its expansion as a targeted fishery in Europe, examining the range of valorization options currently being investigated.
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Metagenome-based surveillance and diagnostic approaches to studying the microbial ecology of food production and processing environmentsMetagenomic-based analyses have the potential to revolutionize our understanding of the microbiology of food production and processing environments. By adopting such approaches, it will be possible to more accurately determine sources of microbial contamination, identify critical control points for such contaminants, and select practices that optimize quality and safety. This mini-review will discuss the merits of adopting metagenostic-based approaches, highlight novel insights that they have provided to date and consider how they could be further implemented.
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Impacts of Seasonal Housing and Teat Preparation on Raw Milk Microbiota: a High-Throughput Sequencing StudyIn pasture-based systems, changes in dairy herd habitat due to seasonality results in the exposure of animals to different environmental niches. These niches contain distinct microbial communities that may be transferred to raw milk, with potentially important food quality and safety implications for milk producers. It is postulated that the extent to which these microorganisms are transferred could be limited by the inclusion of a teat preparation step prior to milking. High-throughput sequencing on a variety of microbial niches on farms was used to study the patterns of microbial movement through the dairy production chain and, in the process, to investigate the impact of seasonal housing and the inclusion/exclusion of a teat preparation regime on the raw milk microbiota from the same herd over two sampling periods, i.e., indoor and outdoor. Beta diversity and network analyses showed that environmental and milk microbiotas separated depending on whether they were sourced from an indoor or outdoor environment. Within these respective habitats, similarities between the milk microbiota and that of teat swab samples and, to a lesser extent, fecal samples were apparent. Indeed, SourceTracker identified the teat surface as the most significant source of contamination, with herd feces being the next most prevalent source of contamination. In milk from cows grazing outdoors, teat prep significantly increased the numbers of total bacteria present. In summary, sequence-based microbiota analysis identified possible sources of raw milk contamination and highlighted the influence of environment and farm management practices on the raw milk microbiota.
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Bacteriophages and Bacterial Plant DiseasesLosses in crop yields due to disease need to be reduced in order to meet increasing global food demands associated with growth in the human population. There is a well-recognized need to develop new environmentally friendly control strategies to combat bacterial crop disease. Current control measures involving the use of traditional chemicals or antibiotics are losing their efficacy due to the natural development of bacterial resistance to these agents. In addition, there is an increasing awareness that their use is environmentally unfriendly. Bacteriophages, the viruses of bacteria, have received increased research interest in recent years as a realistic environmentally friendly means of controlling bacterial diseases. Their use presents a viable control measure for a number of destructive bacterial crop diseases, with some phage-based products already becoming available on the market. Phage biocontrol possesses advantages over chemical controls in that tailor-made phage cocktails can be adapted to target specific disease-causing bacteria. Unlike chemical control measures, phage mixtures can be easily adapted for bacterial resistance which may develop over time. In this review, we will examine the progress and challenges for phage-based disease biocontrol in food crops.
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Things Are Getting Hairy: Enterobacteria Bacteriophage vB_PcaM_CBBEnterobacteria phage vB_PcaM_CBB is a “jumbo” phage belonging to the family Myoviridae. It possesses highly atypical whisker-like structures along the length of its contractile tail. It has a broad host range with the capability of infecting species of the genera Erwinia, Pectobacterium, and Cronobacter. With a genome of 355,922 bp, excluding a predicted terminal repeat of 22,456 bp, phage CBB is the third largest phage sequenced to date. Its genome was predicted to encode 554 ORFs with 33 tRNAs. Based on prediction and proteome analysis of the virions, 29% of its predicted ORFs could be functionally assigned. Protein comparison shows that CBB shares between 33–38% of its proteins with Cronobacter phage GAP32, coliphages PBECO4 and 121Q as well as Klebsiella phage vB_KleM_Rak2. This work presents a detailed and comparative analysis of vB_PcaM_CBB of a highly atypical jumbo myoviridae phage, contributing to a better understanding of phage diversity and biology.
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Predicted Release and Analysis of Novel ACE-I, Renin, and DPP-IV Inhibitory Peptides from Common Oat (Avena sativa) Protein Hydrolysates Using in Silico AnalysisThe renin-angiotensin-aldosterone system (RAAS) plays an important role in regulating hypertension by controlling vasoconstriction and intravascular fluid volume. RAAS itself is largely regulated by the actions of renin (EC 3.4.23.15) and the angiotensin-I-converting enzyme (ACE-I; EC 3.4.15.1). The enzyme dipeptidyl peptidase-IV (DPP-IV; EC 3.4.14.5) also plays a role in the development of type-2 diabetes. The inhibition of the renin, ACE-I, and DPP-IV enzymes has therefore become a key therapeutic target for the treatment of hypertension and diabetes. The aim of this study was to assess the bioactivity of different oat (Avena sativa) protein isolates and their ability to inhibit the renin, ACE-I, and DPP-IV enzymes. In silico analysis was carried out to predictthe likelihood of bioactive inhibitory peptides occurring from oat protein hydrolysates following in silico hydrolysis with the proteases papain and ficin. Nine peptides, including FFG, IFFFL, PFL, WWK, WCY, FPIL, CPA, FLLA, and FEPL were subsequently chemically synthesised, and their bioactivities were confirmed using in vitro bioassays. The isolated oat proteins derived from seven different oat varieties were found to inhibit the ACE-I enzyme by between 86.5 ± 10.7% and 96.5 ± 25.8%, renin by between 40.5 ± 21.5% and 70.9 ± 7.6%, and DPP-IV by between 3.7 ± 3.9% and 46.2 ± 28.8%. The activity of the synthesised peptides was also determined.
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A novel method of microsatellite genotyping-by-sequencing using individual combinatorial barcodingThis study examines the potential of next-generation sequencing based ‘genotyping-by-sequencing’ (GBS) of microsatellite loci for rapid and cost-effective genotyping in large-scale population genetic studies. The recovery of individual genotypes from large sequence pools was achieved by PCR-incorporated combinatorial barcoding using universal primers. Three experimental conditions were employed to explore the possibility of using this approach with existing and novel multiplex marker panels and weighted amplicon mixture. The GBS approach was validated against microsatellite data generated by capillary electrophoresis. GBS allows access to the underlying nucleotide sequences that can reveal homoplasy, even in large datasets and facilitates cross laboratory transfer. GBS of microsatellites, using individual combinatorial barcoding, is potentially faster and cheaper than current microsatellite approaches and offers better and more data.
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Draft Genome Sequence of Lactobacillus casei DPC6800, an Isolate with the Potential to Diversify Flavor in CheeseLactobacillus casei is a nonstarter lactic acid bacterium commonly present in various types of cheeses. It is believed that strains of this species have a significant impact on the development of cheese flavor. The draft genome sequence of L. casei DPC6800, isolated from a semi-hard Dutch cheese, is reported.
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Thermus and the Pink Discoloration Defect in CheeseA DNA sequencing-based strategy was applied to study the microbiology of Continental-type cheeses with a pink discoloration defect. The basis for this phenomenon has remained elusive, despite decades of research. The bacterial composition of cheese containing the defect was compared to that of control cheese using 16S rRNA gene and shotgun metagenomic sequencing as well as quantitative PCR (qPCR). Throughout, it was apparent that Thermus, a carotenoid-producing genus, was present at higher levels in defect-associated cheeses than in control cheeses. Prompted by this finding and data confirming the pink discoloration to be associated with the presence of a carotenoid, a culture-based approach was employed, and Thermus thermophilus was successfully cultured from defect-containing cheeses. The link between Thermus and the pinking phenomenon was then established through the cheese defect equivalent of Koch’s postulates when the defect was recreated by the reintroduction of a T. thermophilus isolate to a test cheese during the manufacturing process. IMPORTANCE Pink discoloration in cheese is a defect affecting many cheeses throughout the world, leading to significant financial loss for the dairy industry. Despite decades of research, the cause of this defect has remained elusive. The advent of high-throughput, next-generation sequencing has revolutionized the field of food microbiology and, with respect to this study, provided a means of testing a possible microbial basis for this defect. In this study, a combined 16S rRNA, whole-genome sequencing, and quantitative PCR approach was taken. This resulted in the identification of Thermus, a carotenoid-producing thermophile, in defect-associated cheeses and the recreation of the problem in cheeses to which Thermus was added. This finding has the potential to lead to new strategies to eliminate this defect, and our method represents an approach that can be employed to investigate the role of microbes in other food defects of unknown origin.
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The Omega-3 Polyunsaturated Fatty Acid Docosahexaenoic Acid (DHA) Reverses Corticosterone-Induced Changes in Cortical NeuronsBackground: Chronic exposure to the glucocorticoid hormone corticosterone exerts cellular stress-induced toxic effects that have been associated with neurodegenerative and psychiatric disorders. Docosahexaenoic acid is a polyunsaturated fatty acid that has been shown to be of benefit in stress-related disorders, putatively through protective action in neurons. Methods: We investigated the protective effect of docosahexaenoic acid against glucocorticoid hormone corticosterone-induced cellular changes in cortical cell cultures containing both astrocytes and neurons. Results: We found that glucocorticoid hormone corticosterone (100, 150, 200 μM) at different time points (48 and 72 hours) induced a dose- and time-dependent reduction in cellular viability as assessed by methyl thiazolyl tetrazolium. Moreover, glucocorticoid hormone corticosterone (200 μM, 72 hours) decreased the percentage composition of neurons while increasing the percentage of astrocytes as assessed by βIII-tubulin and glial fibrillary acidic protein immunostaining, respectively. In contrast, docosahexaenoic acid treatment (6 μM) increased docosahexaenoic acid content and attenuated glucocorticoid hormone corticosterone (200 μM)-induced cell death (72 hours) in cortical cultures. This translates into a capacity for docosahexaenoic acid to prevent neuronal death as well as astrocyte overgrowth following chronic exposure to glucocorticoid hormone corticosterone. Furthermore, docosahexaenoic acid (6 μM) reversed glucocorticoid hormone corticosterone-induced neuronal apoptosis as assessed by terminal deoxynucleotidyl transferase-mediated nick-end labeling and attenuated glucocorticoid hormone corticosterone-induced reductions in brain derived neurotrophic factor mRNA expression in these cultures. Finally, docosahexaenoic acid inhibited glucocorticoid hormone corticosterone-induced downregulation of glucocorticoid receptor expression on βIII- tubulin-positive neurons. Conclusions: This work supports the view that docosahexaenoic acid may be beneficial in ameliorating stress-related cellular changes in the brain and may be of value in psychiatric disorders.
