• Identification of existing and emerging chemical residue contamination concerns in milk

      Danaher, Martin; Jordan, Kieran; Department of Agriculture, Food and the Marine, Ireland (Teagasc (Agriculture and Food Development Authority), Ireland, 2013)
      In order to maintain the quality of Irish milk and meet increasingly demanding specifications, it is necessary to focus on chemical residues in milk, in addition to other quality issues. The objective of the work was to assess the current status of chemical contaminant analysis and to identify technological and knowledge needs. This was achieved through a review of literature with respect to chemical contaminants. Quaternary ammonium compounds (QACs) have been identified as an area of concern for the dairy industry because of the recent reports of QAC residues in dairy products internationally. Analytical support to analyse QAC residues in milk and dairy products on an ongoing basis is required. Furthermore, the source of QAC residues along the milk production chain needs to be identified. Similarly, analytical support and research is needed in the area of phthalates, to support the development of intervention strategies to reduce contamination, if present. Cephalosporin antibiotics have been a concern for the dairy industry because of the lack of suitable chemical tests to measure these substances.
    • Identification of short peptide sequences in the nanofiltration permeate of a bioactive whey protein hydrolysate

      Le Maux, Solene; Nongoniermaa, Alice B.; Murray, Brian; Kelly, Philip M.; Fitzgerald, Richard J.; Enterprise Ireland; TC2013-0001 (Elsevier, 16/09/2015)
      Short peptides in food protein hydrolysates are of significant interest as they may be highly bioactive whilst also being bioavailable. A dipeptidyl peptidase IV (DPP-IV) inhibitory whey protein hydrolysate (WPH) was fractionated using nanofiltration (NF) with a 200 Da MWCO membrane. The DPP-IV half maximal inhibitory concentration of the NF permeate (IC50 = 0.66 ± 0.08 mg protein equivalent mL− 1) was significantly more potent (P > 0.05) than that of the starting WPH (IC50 = 0.94 ± 0.24 mg protein equivalent mL− 1) and associated retentate (IC50 = 0.82 ± 0.13 mg protein equivalent mL− 1). This confirmed the contribution of short peptides within the NF permeate to the overall DPP-IV inhibitory activity. An hydrophilic interaction liquid chromatography (HILIC-) and reverse-phase (RP-) liquid chromatography tandem mass spectrometry (LC–MS/MS) strategy, based on two retention time models, allowed detection of eight free amino acids and eight di- to tetrapeptides in the NF permeate. The potential sequences of the peptides within the NF permeate were then ranked on the basis of their highest probability of occurrence. A confirmatory study with synthetic peptides showed that valine–alanine (VA), valine–leucine (VL), tryptophan–leucine (WL) and tryptophan–isoleucine (WI) displayed DPP-IV IC50 values < 170 μM. The NF and LC–MS strategies employed herein represent a new approach for the targeted identification of short peptides within bioactive food protein hydrolysates.
    • Image Processing of Outer-Product matrices - a new way to classify samples: Examples using visible/NIR/MIR spectral data.

      Jaillais, B.; Morrin, V.; Downey, Gerard (Elsevier, 2007)
      A chemometric analysis has been developed to emphasise the discrimination power of spectroscopic techniques such as near infrared, mid-infrared and visible spectroscopy. The combination of two spectral domains using outer product analysis (OPA) leads to the calculation of an outer product (OP) matrix. The representation of this matrix is called the "analytical fingerprint" of the samples and their classification is performed in the following steps. First, two different techniques are tested by subtracting the images one-by-one and the sum of all the elements of the resulting difference matrix gives a scalar, characteristic of the distance between the two images. Combining chemical analysis with image processing techniques provides an original approach to study butters and margarines in relation to their fat content. Best results were obtained with the OP matrix built from NIR and visible signals following the use of city block distance and average linkage. Samples were arranged in four groups: 100 %, 82-75 %, 70-59 % and 38-25 % w/w fat. The cophenetic correlation coefficient (validity of the cluster information generated by the linkage function) associated with these spectral data has a value of 0.973. Similar results were obtained using Ward's algorithm which generated four groups and a cophenetic correlation coefficient equal to 0.959.
    • Impact of Environmental Factors on Bacteriocin Promoter Activity in Gut-Derived Lactobacillus salivarius

      Guinane, Caitriona M.; Piper, Clare; Draper, Lorraine A.; O'Connor, Paula M.; Hill, Colin; Ross, R Paul; Cotter, Paul D.; Science Foundation Ireland; SFI/11/PI/1137 (American Society for Microbiology, 04/09/2015)
      Bacteriocin production is regarded as a desirable probiotic trait that aids in colonization and persistence in the gastrointestinal tract (GIT). Strains of Lactobacillus salivarius, a species associated with the GIT, are regarded as promising probiotic candidates and have a number of associated bacteriocins documented to date. These include multiple class IIb bacteriocins (salivaricin T, salivaricin P, and ABP-118) and the class IId bacteriocin bactofencin A, which show activity against medically important pathogens. However, the production of a bacteriocin in laboratory media does not ensure production under stressful environmental conditions, such as those encountered within the GIT. To allow this issue to be addressed, the promoter regions located upstream of the structural genes encoding the L. salivarius bacteriocins mentioned above were fused to a number of reporter proteins (green fluorescent protein [GFP], red fluorescent protein [RFP], and luciferase [Lux]). Of these, only transcriptional fusions to GFP generated signals of sufficient strength to enable the study of promoter activity in L. salivarius. While analysis of the class IIb bacteriocin promoter regions indicated relatively weak GFP expression, assessment of the promoter of the antistaphylococcal bacteriocin bactofencin A revealed a strong promoter that is most active in the absence of the antimicrobial peptide and is positively induced in the presence of mild environmental stresses, including simulated gastric fluid. Taken together, these data provide information on factors that influence bacteriocin production, which will assist in the development of strategies to optimize in vivo and in vitro production of these antimicrobials.
    • Improved emulsion stability and modified nutrient release by structuring O/W emulsions using konjac glucomannan

      Lu, Wei; Zheng, Baodong; Miao, Song; National Natural Science Foundation of China; China Scholarship Council; 31628016; 201508300001 (Elsevier, 2018-02-22)
      Functional konjac glucomannan (KGM) was used to structure the water phase of O/W emulsions containing a lipophilic bioactive compound (β-carotene). KGM greatly increased the viscosity of the water phase and thus the viscosity of final emulsions. Results of Fourier-transform infrared spectroscopy (FT-IR) showed that there is no significant non-covalent interaction between KGM and whey proteins in the water phase. KGM significantly improved the creaming and pH stability of whey-protein-stabilized emulsions (p < 0.05), and significantly decreased the oiling-off of emulsions during freeze-thaw test. Emulsions with or without KGM all had good thermal stability at 80 °C. Microscopy observations indicated obvious aggregation of free proteins and oil droplets in gastric phase and an enzymatic-induced break-down of droplets, mainly in the intestinal phase of the simulated gastrointestinal tract (GIT) digestion. Emulsions with KGM-structured water phase showed a lower final release rate of encapsulated β-carotene than emulsion without KGM (p < 0.05), and the release rate decreased with the increasing KGM content. The findings of this study contribute to a better understanding of the influence of the water phase on the release of encapsulated compounds from emulsions, and make it possible to achieve controlled release of encapsulated compounds, and/or to deliver multiple health-beneficial nutrients at once by structuring emulsion-based carriers with functional natural biopolymers.
    • Improving the quality of gluten-free products

      Gallagher, Eimear; McCarthy, Denise; Gormley, Ronan T.; Arendt, Elke (Teagasc, 2004-03)
      The incidence of coeliac disease or other allergic reactions/intolerances to gluten is increasing, largely due to improved diagnostic procedures and changes in eating habits. The worldwide number of sufferers of coeliac disease has been predicted to increase by a factor of ten over the next number of years, resulting in a growing market for gluten-free cereal-based products. Market research has shown that many of the products currently on sale are of inferior quality. The replacement of gluten presents a major technological challenge, as it is an essential structure-building protein which is necessary for formulating high quality cereal-based goods. Therefore, the production of high quality gluten-free bread is difficult.
    • In silico analysis highlights the frequency and diversity of type 1 lantibiotic gene clusters in genome sequenced bacteria

      Marsh, Alan J.; O'Sullivan, Orla; Ross, R Paul; Cotter, Paul D.; Hill, Colin; Science Foundation Ireland (Biomed Central, 30/11/2010)
      Background: Lantibiotics are lanthionine-containing, post-translationally modified antimicrobial peptides. These peptides have significant, but largely untapped, potential as preservatives and chemotherapeutic agents. Type 1 lantibiotics are those in which lanthionine residues are introduced into the structural peptide (LanA) through the activity of separate lanthionine dehydratase (LanB) and lanthionine synthetase (LanC) enzymes. Here we take advantage of the conserved nature of LanC enzymes to devise an in silico approach to identify potential lantibiotic-encoding gene clusters in genome sequenced bacteria. Results: In total 49 novel type 1 lantibiotic clusters were identified which unexpectedly were associated with species, genera and even phyla of bacteria which have not previously been associated with lantibiotic production. Conclusions: Multiple type 1 lantibiotic gene clusters were identified at a frequency that suggests that these antimicrobials are much more widespread than previously thought. These clusters represent a rich repository which can yield a large number of valuable novel antimicrobials and biosynthetic enzymes.
    • In Silico Assigned Resistance Genes Confer Bifidobacterium with Partial Resistance to Aminoglycosides but Not to Β-Lactams

      Fouhy, Fiona; O’Connell Motherway, Mary; Fitzgerald, Gerald F.; Ross, R Paul; Stanton, Catherine; van Sinderen, Douwe; Cotter, Paul D.; Irish Research Council for Science, Engineering and Technology; Science Foundation Ireland; Health Research Board; Teagasc Walsh Fellowship Programme; SFI/11/PI/1137; SFI/02/CE/B124; SFI/07/CE/B1368; PDTM/20011/9) (PLoS, 06/12/2013)
      Bifidobacteria have received significant attention due to their contribution to human gut health and the use of specific strains as probiotics. It is thus not surprising that there has also been significant interest with respect to their antibiotic resistance profile. Numerous culture-based studies have demonstrated that bifidobacteria are resistant to the majority of aminoglycosides, but are sensitive to β-lactams. However, limited research exists with respect to the genetic basis for the resistance of bifidobacteria to aminoglycosides. Here we performed an in-depth in silico analysis of putative Bifidobacterium-encoded aminoglycoside resistance proteins and β-lactamases and assess the contribution of these proteins to antibiotic resistance. The in silico-based screen detected putative aminoglycoside and β-lactam resistance proteins across the Bifidobacterium genus. Laboratory-based investigations of a number of representative bifidobacteria strains confirmed that despite containing putative β-lactamases, these strains were sensitive to β-lactams. In contrast, all strains were resistant to the aminoglycosides tested. To assess the contribution of genes encoding putative aminoglycoside resistance proteins in Bifidobacterium sp. two genes, namely Bbr_0651 and Bbr_1586, were targeted for insertional inactivation in B. breve UCC2003. As compared to the wild-type, the UCC2003 insertion mutant strains exhibited decreased resistance to gentamycin, kanamycin and streptomycin. This study highlights the associated risks of relying on the in silico assignment of gene function. Although several putative β-lactam resistance proteins are located in bifidobacteria, their presence does not coincide with resistance to these antibiotics. In contrast however, this approach has resulted in the identification of two loci that contribute to the aminoglycoside resistance of B. breve UCC2003 and, potentially, many other bifidobacteria.
    • In silico identification of bacteriocin gene clusters in the gastrointestinal tract, based on the Human Microbiome Project’s reference genome database

      Walsh, Calum J.; Guinane, Caitriona M.; Hill, Colin; Ross, R Paul; O’Toole, Paul W.; Cotter, Paul D.; Science Foundation Ireland; SFI/11/PI/1137 (Biomed Central, 16/09/2015)
      Background The human gut microbiota comprises approximately 100 trillion microbial cells which significantly impact many aspects of human physiology - including metabolism, nutrient absorption and immune function. Disturbances in this population have been implicated in many conditions and diseases, including obesity, type-2 diabetes and inflammatory bowel disease. This suggests that targeted manipulation or shaping of the gut microbiota, by bacteriocins and other antimicrobials, has potential as a therapeutic tool for the prevention or treatment of these conditions. With this in mind, several studies have used traditional culture-dependent approaches to successfully identify bacteriocin-producers from the mammalian gut. In silico-based approaches to identify novel gene clusters are now also being utilised to take advantage of the vast amount of data currently being generated by next generation sequencing technologies. In this study, we employed an in silico screening approach to mine potential bacteriocin clusters in genome-sequenced isolates from the gastrointestinal tract (GIT). More specifically, the bacteriocin genome-mining tool BAGEL3 was used to identify potential bacteriocin producers in the genomes of the GIT subset of the Human Microbiome Project’s reference genome database. Each of the identified gene clusters were manually annotated and potential bacteriocin-associated genes were evaluated. Results We identified 74 clusters of note from 59 unique members of the Firmicutes, Bacteroidetes, Actinobacteria, Fusobacteria and Synergistetes. The most commonly identified class of bacteriocin was the >10 kDa class, formerly known as bacteriolysins, followed by lantibiotics and sactipeptides. Conclusions Multiple bacteriocin gene clusters were identified in a dataset representative of the human gut microbiota. Interestingly, many of these were associated with species and genera which are not typically associated with bacteriocin production.
    • In Vitro Activities of Nisin and Nisin Derivatives Alone and In Combination with Antibiotics against Staphylococcus Biofilms

      Field, Des; O'Connor, Rory; Cotter, Paul D; Ross, R Paul; Hill, Colin; Science Foundation Ireland; TIDA14/TIDA/2286); 10/IN.1/B3027; 11/PI/1137; SFI/12/RC/2273 (Frontiers Media S. A., 18/04/2016)
      The development and spread of pathogenic bacteria that are resistant to the existing catalog of antibiotics is a major public health threat. Biofilms are complex, sessile communities of bacteria embedded in an organic polymer matrix which serve to further enhance antimicrobial resistance. Consequently, novel compounds and innovative methods are urgently required to arrest the proliferation of drug-resistant infections in both nosocomial and community environments. Accordingly, it has been suggested that antimicrobial peptides could be used as novel natural inhibitors that can be used in formulations with synergistically acting antibiotics. Nisin is a member of the lantibiotic family of antimicrobial peptides that exhibit potent antibacterial activity against many Gram-positive bacteria. Recently we have used bioengineering strategies to enhance the activity of nisin against several high profile targets, including multi-drug resistant clinical pathogens such as methicillin-resistant Staphylococcus aureus, vancomycinresistant enterococci, staphylococci, and streptococci associated with bovine mastitis. We have also identified nisin derivatives with an enhanced ability to impair biofilm formation and to reduce the density of established biofilms of methicillin resistant S. pseudintermedius. The present study was aimed at evaluating the potential of nisin and nisin derivatives to increase the efficacy of conventional antibiotics and to assess the possibility of killing and/or eradicating biofilm-associated cells of a variety of staphylococcal targets. Growth curve-based comparisons established that combinations of derivatives nisin V C penicillin or nisin I4V C chloramphenicol had an enhanced inhibitory effect against S. aureus SA113 and S. pseudintermedius DSM21284, respectively, compared to the equivalent nisin A C antibiotic combinations or when each antimicrobial was administered alone. Furthermore, the metabolic activity of established biofilms treated with nisin V C chloramphenicol and nisin I4V C chloramphenicol combinations revealed a significant decrease in S. aureus SA113 and S. pseudintermedius DSM21284 biofilm viability, respectively, compared to the nisin A C antibiotic combinations as determined by the rapid colorimetric XTT assay. The results indicate that the activities of the nisin derivative and antibiotic combinations represent a significant improvement over that of the wild-type nisin and antibiotic combination and merit further investigation with a view to their use as anti-biofilm agents.
    • In vivo activity of Nisin A and Nisin V against Listeria monocytogenes in mice

      Campion, Alicia; Casey, Patrick G.; Field, Des; Cotter, Paul D.; Hill, Colin; Ross, R Paul; Programme for Research in Third-Level Institutions; Irish Research Council for Science, Engineering and Technology; Enterprise Ireland; Science Foundation Ireland (Biomed Central, 01/02/2013)
      Background: Lantibiotics are post-translationally modified antimicrobial peptides, of which nisin A is the most extensively studied example. Bioengineering of nisin A has resulted in the generation of derivatives with increased in vitro potency against Gram-positive bacteria. Of these, nisin V (containing a Met21Val change) is noteworthy by virtue of exhibiting enhanced antimicrobial efficacy against a wide range of clinical and food-borne pathogens, including Listeria monocytogenes. However, this increased potency has not been tested in vivo. Results: Here we address this issue by assessing the ability of nisin A and nisin V to control a bioluminescent strain of Listeria monocytogenes EGDe in a murine infection model. More specifically, Balb/c mice were infected via the intraperitoneal route at a dose of 1 × 105 cfu/animal and subsequently treated intraperitoneally with either nisin V, nisin A or a PBS control. Bioimaging of the mice was carried out on day 3 of the trial. Animals were then sacrificed and levels of infection were quantified in the liver and spleen. Conclusion: This analysis revealed that nisin V was more effective than Nisin A with respect to controlling infection and therefore merits further investigation with a view to potential chemotherapeutic applications.
    • Indicator organisms to determine the use of chilling as a critical point in beef slaughter HACCP

      Prendergast, Deirdre M.; Sheridan, James J.; Downey, Gerard (Teagasc, 01/11/2008)
      During chilling, temperatures of carcass surfaces at different sites change over time as do other parameters such as water activity (aw), the structure of the muscle and other tissues, as the carcass enters rigor mortis. Many of these factors are known to have a major effect on cell survival and growth and must be considered in determining the influence of chilling on bacterial survival on carcass surfaces. This study aimed to determine if chilling could be used as a critical control point (CCP) in beef slaughter in relation to pathogens such as E. coli O157:H7 and L. monocytogenes, using E. coli and Listeria innocua as pathogen indicators. The present study was designed to determine the influence of (a) chilling at 10oC for 72 h on the survival of E. coli and (b) chilling at 4oC for 72 h on the survival of L. innocua inoculated at different sites on beef carcasses. Three sites (neck, outside round and brisket) were inoculated (1) immediately after dressing while hot (E. coli and L. innocua) and (2) when cold after chilling (L. innocua). The influence of changes in surface aw was also considered and their relationship to the survival of E. coli and L. innocua over time was assessed. The data are discussed in relation to the use of chilling as a CCP in beef hazard analysis (HACCP) and the monitoring of neck temperature as the most suitable CCP.
    • Indicator organisms to determine the use of chilling as a critical point in beef slaughter HACCP

      Prendergast, Deirdre M.; Sheridan, James J.; Downey, Gerard (Teagasc, 2008-11)
      During chilling, temperatures of carcass surfaces at different sites change over time as do other parameters such as water activity (aw), the structure of the muscle and other tissues, as the carcass enters rigor mortis. Many of these factors are known to have a major effect on cell survival and growth and must be considered in determining the influence of chilling on bacterial survival on carcass surfaces. This study aimed to determine if chilling could be used as a critical control point (CCP) in beef slaughter in relation to pathogens such as E. coli O157:H7 and L. monocytogenes, using E. coli and Listeria innocua as pathogen indicators. The present study was designed to determine the influence of (a) chilling at 10oC for 72 h on the survival of E. coli and (b) chilling at 4oC for 72 h on the survival of L. innocua inoculated at different sites on beef carcasses. Three sites (neck, outside round and brisket) were inoculated (1) immediately after dressing while hot (E. coli and L. innocua) and (2) when cold after chilling (L. innocua). The influence of changes in surface aw was also considered and their relationship to the survival of E. coli and L. innocua over time was assessed. The data are discussed in relation to the use of chilling as a CCP in beef hazard analysis (HACCP) and the monitoring of neck temperature as the most suitable CCP.
    • The influence of bovine serum albumin on β-lactoglobulin denaturation, aggregation and gelation

      Kehoe, Joseph James; Morris, Edwin R; Brodkorb, Andre; Department of Agriculture, Food and the Marine; Teagasc Walsh Fellowship Programme (Elsevier, 22/11/2006)
      The effect of bovine serum albumin (BSA) on the heat-induced denaturation, aggregation and subsequent acid-induced gelation of β-lactoglobulin (β-lg) was investigated in this work. Changes in the denaturation kinetics of β-lg during heating at 78 °C were determined by monitoring the disappearance of the native protein by reverse-phase chromatography. Replacing β-lg with increasing amounts of BSA, while keeping the total protein concentration constant at 5% (w/w), significantly increased the denaturation rate of β-lg from 2.57±0.30×10−3(g L−1)(1−n)s−1 to 5.07±0.72×10−3(g L−1)(1−n)s−1 (β-lg: BSA ratio of 3:1 w/w). The reaction order for β-lg was 1.40±0.09. Partial replacement of β-lg with BSA (β-lg: BSA ratio of 3:1 w/w) significantly increased the reaction order to 1.67±0.13. Heat-induced aggregates between β-lg and BSA were studied by dynamic light scattering, two-dimensional electrophoresis and size exclusion chromatography. The partial replacement of β-lg with BSA significantly changed the gelling properties of the acid-induced gels. A rapid rate of acidification resulted in a significant decrease, while a slow acidification rate resulted in a significant increase in gel strength. Size exclusion chromatography demonstrated that intermolecular disulphide bond formation occurred during both heat-induced denaturation/aggregation and subsequent acid-induced gelation. Results clearly indicate that BSA contributed to the formation of these disulphide bonds.
    • Influence of feeding systems on the eating quality of beef

      Troy, Declan J.; Murray, Brendan; O'Sullivan, Aidan; Mooney, Teresa; Moloney, Aidan P; Kerry, Joseph (Teagasc, 2002-10)
      The objective was to determine pre-slaughter factors which may enhance the eating quality of beef and to assist the Irish beef production chain to exploit these factors to produce beef of higher quality and increased consumer acceptability. The effects of pre-slaughter growth rate, high energy diets, feed type and age at slaughter on beef quality were examined.
    • Influence of GABA and GABA-producing Lactobacillus brevis DPC 6108 on the development of diabetes in a streptozotocin rat model

      Marques, T. M.; Patterson, E.; Wall, R.; O'Sullivan, Orla; Fitzgerald, G. F.; Cotter, Paul D.; Dinan, T. G.; Cryan, J. F.; Ross, R Paul; Stanton, Catherine; Science Foundation Ireland; SFI/12/RC/2273 (Wageningen Academic Publishers, 26/05/2016)
      The aim of this study was to investigate if dietary administration of γ-aminobutyric acid (GABA)-producing Lactobacillus brevis DPC 6108 and pure GABA exert protective effects against the development of diabetes in streptozotocin (STZ)-induced diabetic Sprague Dawley rats. In a first experiment, healthy rats were divided in 3 groups (n=10/group) receiving placebo, 2.6 mg/kg body weight (bw) pure GABA or L. brevis DPC 6108 (~109microorganisms). In a second experiment, rats (n=15/group) were randomised to five groups and four of these received an injection of STZ to induce type 1 diabetes. Diabetic and non-diabetic controls received placebo [4% (w/v) yeast extract in dH2O], while the other three diabetic groups received one of the following dietary supplements: 2.6 mg/kg bw GABA (low GABA), 200 mg/kg bw GABA (high GABA) or ~109 L. brevis DPC 6108. L. brevis DPC 6108 supplementation was associated with increased serum insulin levels (P<0.05), but did not alter other metabolic markers in healthy rats. Diabetes induced by STZ injection decreased body weight (P<0.05), increased intestinal length (P<0.05) and stimulated water and food intake. Insulin was decreased (P<0.05), whereas glucose was increased (P<0.001) in all diabetic groups, compared with non-diabetic controls. A decrease (P<0.01) in glucose levels was observed in diabetic rats receiving L. brevis DPC 6108, compared with diabetic-controls. Both the composition and diversity of the intestinal microbiota were affected by diabetes. Microbial diversity in diabetic rats supplemented with low GABA was not reduced (P>0.05), compared with non-diabetic controls while all other diabetic groups displayed reduced diversity (P<0.05). L. brevis DPC 6108 attenuated hyperglycaemia induced by diabetes but additional studies are needed to understand the mechanisms involved in this reduction.
    • INFOGEST static in vitro simulation of gastrointestinal food digestion

      Brodkorb, Andre; Egger, Lotti; Alminger, Marie; et al; French National Institute for Agricultural Research; European Cooperation in Science and Technology (COST); FA1005 (Nature Publishing Group, 2019-03-18)
      Developing a mechanistic understanding of the impact of food structure and composition on human health has increasingly involved simulating digestion in the upper gastrointestinal tract. These simulations have used a wide range of different conditions that often have very little physiological relevance, and this impedes the meaningful comparison of results. The standardized protocol presented here is based on an international consensus developed by the COST INFOGEST network. The method is designed to be used with standard laboratory equipment and requires limited experience to encourage a wide range of researchers to adopt it. It is a static digestion method that uses constant ratios of meal to digestive fluids and a constant pH for each step of digestion. This makes the method simple to use but not suitable for simulating digestion kinetics. Using this method, food samples are subjected to sequential oral, gastric and intestinal digestion while parameters such as electrolytes, enzymes, bile, dilution, pH and time of digestion are based on available physiological data. This amended and improved digestion method (INFOGEST 2.0) avoids challenges associated with the original method, such as the inclusion of the oral phase and the use of gastric lipase. The method can be used to assess the endpoints resulting from digestion of foods by analyzing the digestion products (e.g., peptides/amino acids, fatty acids, simple sugars) and evaluating the release of micronutrients from the food matrix. The whole protocol can be completed in ~7 d, including ~5 d required for the determination of enzyme activities.
    • Inhibitory activity of Lactobacillus plantarum LMG P-26358 against Listeria innocua when used as an adjunct starter in the manufacture of cheese

      Mills, Susan; Serrano, L Mariela; Griffin, Carmel; O'Connor, Paula M.; Schaad, Gwenda; Bruining, Chris; Hill, Colin; Ross, R Paul; Meijer, Wilco C (Biomed Central, 30/08/2011)
      Lactobacillus plantarum LMG P-26358 isolated from a soft French artisanal cheese produces a potent class IIa bacteriocin with 100% homology to plantaricin 423 and bacteriocidal activity against Listeria innocua and Listeria monocytogenes. The bacteriocin was found to be highly stable at temperatures as high as 100°C and pH ranges from 1-10. While this relatively narrow spectrum bacteriocin also exhibited antimicrobial activity against species of enterococci, it did not inhibit dairy starters including lactococci and lactobacilli when tested by well diffusion assay (WDA). In order to test the suitability of Lb. plantarum LMG P-26358 as an anti-listerial adjunct with nisin-producing lactococci, laboratory-scale cheeses were manufactured. Results indicated that combining Lb. plantarum LMG P-26358 (at 108 colony forming units (cfu)/ml) with a nisin producer is an effective strategy to eliminate the biological indicator strain, L. innocua. Moreover, industrial-scale cheeses also demonstrated that Lb. plantarum LMG P-26358 was much more effective than the nisin producer alone for protection against the indicator. MALDI-TOF mass spectrometry confirmed the presence of plantaricin 423 and nisin in the appropriate cheeses over an 18 week ripening period. A spray-dried fermentate of Lb. plantarum LMG P-26358 also demonstrated potent anti-listerial activity in vitro using L. innocua. Overall, the results suggest that Lb. plantarum LMG P-26358 is a suitable adjunct for use with nisin-producing cultures to improve the safety and quality of dairy products.
    • Insights into the Mode of Action of the Sactibiotic Thuricin CD

      Mathur, Harsh; Fallico, Vicenzo; O'Connor, Paula M.; Rea, Mary C.; Cotter, Paul D.; Hill, Colin; Ross, R Paul; Science Foundation Ireland; SFI/12/RC/2273 (Frontiers, 20/04/2017)
      Thuricin CD is a two-component bacteriocin, consisting of the peptides Trnα and Trnβ, and belongs to the newly designated sactibiotic subclass of bacteriocins. While it is clear from studies conducted thus far that it is a narrow-spectrum bacteriocin, requiring the synergistic activity of the two peptides, the precise mechanism of action of thuricin CD has not been elucidated. This study used a combination of flow cytometry and traditional culture-dependent assays to ascertain the effects of the thuricin CD peptides on the morphology, physiology and viability of sensitive Bacillus firmus DPC6349 cells. We show that both Trnα and Trnβ are membrane-acting and cause a collapse of the membrane potential, which could not be reversed even under membrane-repolarizing conditions. Furthermore, the depolarizing action of thuricin CD is accompanied by reductions in cell size and granularity, producing a pattern of physiological alterations in DPC6349 cells similar to those triggered by the pore-forming single-component bacteriocin Nisin A, and two-component lacticin 3147. Taken together, these results lead us to postulate that the lytic activity of thuricin CD involves the insertion of thuricin CD peptides into the membrane of target cells leading to permeabilization due to pore formation and consequent flux of ions across the membrane, resulting in membrane depolarization and eventual cell death.
    • Intake, growth and carcass traits in male progeny of sires differing in genetic merit for beef production

      Clarke, A. M.; Drennan, Michael J; McGee, Mark; Kenny, David A.; Evans, Ross D; Berry, Donagh P. (Cambridge University Press, 2009-06)
      Validation of economic indexes under a controlled experimental environment, can aid in their acceptance and use as breeding tools to increase herd profitability. The objective of this study was to compare intake, growth and carcass traits in bull and steer progeny of high and low ranking sires, for genetic merit in an economic index. The Beef Carcass Index (BCI; expressed in euro (€) and based on weaning weight, feed intake, carcass weight, carcass conformation and fat scores) was generated by the Irish Cattle Breeding Federation as a tool to compare animals on genetic merit for the expected profitability of their progeny at slaughter. A total of 107 male suckler herd progeny, from 22 late-maturing ‘continental’ beef sires of high (n = 11) or low (n = 11) BCI were compared under either a bull or steer production system, and slaughtered at approximately 16 and 24 months of age, respectively. All progeny were purchased after weaning at approximately 6 to 8 months of age. Dry matter (DM) intake and live-weight gain in steer progeny offered grazed grass or grass silage alone, did not differ between the two genetic groups. Similarly, DM intake and feed efficiency did not differ between genetic groups during an ad libitum concentrate-finishing period on either production system. Carcasses of progeny of high BCI sires were 14 kg heavier (P < 0.05) than those of low BCI sires. In a series of regression analyses, increasing sire BCI resulted in increases in carcass weight (P < 0.01) and carcass conformation (P = 0.051) scores, and decreases in carcass fat (P < 0.001) scores, but had no effect on weaning weight or DM intake of the progeny. Each unit increase in sire expected progeny difference led to an increase in progeny weaning weight, DM intake, carcass weight, carcass conformation score and carcass fat score of 1.0 (s.e. = 0.53) kg, 1.1 (s.e. = 0.32) kg, 1.3 (s.e. = 0.31) kg, 0.9 (s.e. = 0.32; scale 1 to 15) and 1.0 (s.e. = 0.25; scale 1 to 15), respectively, none of which differed from the theoretical expectation of unity. The expected difference in profitability at slaughter between progeny of the high and low BCI sires was €42, whereas the observed phenotypic profit differential of the progeny was €53 in favour of the high BCI sires. Results from this study indicate that the BCI is a useful tool in the selection of genetically superior sires, and that actual progeny performance under the conditions of this study is within expectations for both bull and steer beef production systems.