• Alternative uses for co-products: Harnessing the potential of valuable compounds from meat processing chains

      Mullen, Anne Maria; Álvarez García, Carlos; Zeugolis, Dimitrios; Henchion, Maeve; O'Neill, Eileen; Drummond, Liana; Department of Agriculture, Food and the Marine, Ireland; 11/F/043 (Elsevier, 03/05/2017)
      Opportunities for exploiting the inherent value of protein-rich meat processing co-products, in the context of increased global demand for protein and for sustainable processing systems, are discussed. While direct consumption maybe the most profitable route for some, this approach is influenced greatly by local and cultural traditions. A more profitable and sustainable approach may be found in recognizing this readily available and under-utilised resource can provide high value components, such as proteins, with targeted high value functionality of relevance to a variety of sectors. Applications in food & beverages, petfood biomedical and nutrition arenas are discussed. Utilization of the raw material in its entirety is a necessary underlying principle in this approach to help maintain minimum waste generation. Understanding consumer attitudes to these products, in particular when used in food or beverage systems, is critical in optimizing commercialization strategies.
    • A case of bovine raw milk contamination with Listeria monocytogenes

      Hunt, Karen; Drummond, Niall; Murphy, Mary; Butler, Francis; Buckley, James F.; Jordan, Kieran; Teagasc Walsh Fellowship Programme; Department of Agriculture, Food and the Marine, Ireland; European Union (Biomed Central, 06/07/2012)
      During routine sampling of bulk raw milk on a dairy farm, the pathogenic bacteria Listeria monocytogenes was found to be a contaminant, at numbers < 100 cfu/ml. A strain with an indistinguishable pulsed-field gel electrophoresis pattern was isolated from the bulk milk two months later. Environmental swabs taken at the dairy environment were negative for the presence of L. monocytogenes, indicating a possible case of excretion of the L. monocytogenes directly into the milk. Milk samples were collected from the individual cows and analysed, resulting in the identification of L. monocytogenes excretion (at 280 cfu/ml) from one of the 4 mammary quarters of one dairy cow out of 180. When the infected cow was isolated from the herd, no L. monocytogenes was detected from the remaining herd. The pulsed-field gel electrophoresis pattern of the strain from the individual cow was indistinguishable from that originally isolated from the bulk milk. The infected cow did not show any clinical signs of disease, nor did the appearance of the milk have any physical abnormalities. Antibiotic treatment of the infected mammary quarter was found to be ineffective. This study shows that there can be risks associated with direct contamination of raw milk with L. monocytogenes.
    • Characterisation and application of fruit by-products as novel ingredients in gluten-free products

      O'Shea, Norah; Department of Agriculture, Food and the Marine, Ireland (2014-01)
      Literature has revealed that “waste” left from the processing of fruit can still contain a substantial quantity of macro and minor nutrients. The aim of this thesis was to ascertain the nutritional and structural properties and potential uses of two fruit by-products [apple pomace (Malus domestica Cv. “Karmijn de Sonnaville”) and orange pomace (Citrus sinensis L. Cv. “Valencia”)] in glutenfree bread and extruded snack formulations. The physicochemical and nutritional properties of the fruit by-products were initially studied. Apple pomace contained a high level of fibre and pectin. The isolated pectin was demonstrated to have a high level of methylation which developed viscous pastes. Orange pomace also had high levels of fibre and pectin, and it was an abundant source of minerals such as potassium and magnesium. Orange pomace had a poor gelling ability. The flour obtained after milling dried orange pomace was used in the formulation of gluten-free bread with the aid of a response surface design. Due to the fibrous properties of orange pomace flour, proofing and water addition were also studied. When added at levels greater than 6%, the loaf volume decreased. The number of cells per slice also decreased with increasing orange pomace addition. Inclusion of orange pomace at levels of up to 4% increased crumb softness. An optimised formulation and proofing time was derived using the optimisation tool; these consisted of 5.5% orange pomace, 94.6% water inclusion and with 49 minutes proofing. These optimised parameters doubled the total dietary fibre content of the bread compared to the original control. The pasting properties, rheology, microstructure and sensory characteristics of the optimised formulation (batter and bread) were investigated. Pasting results showed how orange pomace inclusions reduced the final viscosity of the batter, hence reducing the occurrence of starch gelatinisation. Rheological properties such as the storage modulus (G') and complex modulus (G*) increased in the orange pomace batter compared to the control batter. This demonstrates how the orange pomace as an ingredient improved the robustness of the formulation. Sensory panellists scored the orange pomace bread comparably to the control bread. Milled apple pomace was studied as a potential novel ingredient in an extruded snack. As extrusion requires the trialling of a number of extruder parameters, a response surface design was again used to develop an optimised snack. The parameters studied were apple pomace addition, die head temperature and screw speed. Screw speed had the most significant impact on extrudate characteristics. As screw speed increased the favourable extrudate characteristics such as radical expansion ratio, porosity and specific volume decreased. The inclusion of apple pomace had a negative effect on extrudate characteristics at levels greater than 8% addition. Including apple pomace reduced the hardness and increased the crispiness of the snack. Using the optimisation tool, the optimised and validated formulation and extrusion process contained the following parameters: 7.7% apple pomace, 150oC die head temperature and a screw speed of 69 rpm.
    • Comparative and functional genomics of the Lactococcus lactis taxon; insights into evolution and niche adaptation

      Kelleher, Philip; Bottacini, Francesca; Mahony, Jennifer; Kilcawley, Kieran; van Sinderen, Douwe; Department of Agriculture, Food and the Marine, Ireland; Science Foundation Ireland; 10/RD/TMFRC/704; 13/IA/1953; 14/TIDA/2287; et al. (Biomed Central, 29/03/2017)
      Background Lactococcus lactis is among the most widely studied lactic acid bacterial species due to its long history of safe use and economic importance to the dairy industry, where it is exploited as a starter culture in cheese production. Results In the current study, we report on the complete sequencing of 16 L. lactis subsp. lactis and L. lactis subsp. cremoris genomes. The chromosomal features of these 16 L. lactis strains in conjunction with 14 completely sequenced, publicly available lactococcal chromosomes were assessed with particular emphasis on discerning the L. lactis subspecies division, evolution and niche adaptation. The deduced pan-genome of L. lactis was found to be closed, indicating that the representative data sets employed for this analysis are sufficient to fully describe the genetic diversity of the taxon. Conclusions Niche adaptation appears to play a significant role in governing the genetic content of each L. lactis subspecies, while (differential) genome decay and redundancy in the dairy niche is also highlighted.
    • Comparative Proteomic Profiling of Divergent Phenotypes for Water Holding Capacity across the Post Mortem Ageing Period in Porcine Muscle Exudate

      Di Luca, Alessio; Hamill, Ruth; Mullen, Anne Maria; Slavov, Nikolai; Giuliano, Elia; Department of Agriculture, Food and the Marine, Ireland; 06RDNUIG470 (PLOS, 07/03/2016)
      Two dimensional Difference Gel Electrophoresis (2-D DIGE) and mass spectrometry were applied to investigate the changes in metabolic proteins that occur over a seven day (day 1, 3 and 7) post mortem ageing period in porcine centrifugal exudate from divergent meat quality phenotypes. The objectives of the research were to enhance our understanding of the phenotype (water holding capacity) and search for biomarkers of this economically significant pork quality attribute. Major changes in protein abundance across nine phenotype-by-time conditions were observed. Proteomic patterns were dominated by post mortem ageing timepoint. Using a machine learning algorithm (l1-regularized logistic regression), a model was derived with the ability to discriminate between high drip and low drip phenotypes using a subset of 25 proteins with an accuracy of 63%. Models discriminating between divergent phenotypes with accuracy of 72% and 73% were also derived comparing respectively, high drip plus intermediate phenotype (considered as one phenotype) versus low drip and comparing low drip plus intermediate phenotype (considered as one phenotype) versus high drip. In all comparisons, the general classes of discriminatory proteins identified include metabolic enzymes, stress response, transport and structural proteins. In this research we have enhanced our understanding of the protein related processes underpinning this phenotype and provided strong data to work toward development of protein biomarkers for water holding capacity.
    • Encapsulation of a Lactic Acid Bacteria Cell-Free Extract in Liposomes and Use in Cheddar Cheese Ripening

      Nongonierma, Alice B.; Abrlova, Magdalena; Kilcawley, Kieran; Department of Agriculture, Food and the Marine, Ireland (MDPI AG., Basel, Switzerland, 13/03/2013)
      A concentrated form of cell free extract (CFE) derived from attenuated Lactococcus lactis supsb. lactis 303 CFE was encapsulated in liposomes prepared from two different proliposome preparations (Prolipo Duo and Prolipo S) using microfluidization. Entrapment efficiencies of 19.7 % (Prolipo S) and 14.0 % (Prolipo Duo) were achieved and the preparations mixed in the ratio 4 (Prolipo Duo):1 (Prolipo S). Cheddar cheese trials were undertaken evaluating the performance of CFE entrapped in liposomes, empty liposomes and free CFE in comparison to a control cheese without any CFE or liposomes. Identical volumes of liposome and amounts of CFE were used in triplicate trials. The inclusion of liposomes did not adversely impact on cheese composition water activity, or microbiology. Entrapment of CFE in liposomes reduced loss of CFE to the whey. No significant differences were evident in proteolysis or expressed PepX activity during ripening in comparison to the cheeses containing free CFE, empty liposomes or the control, as the liposomes did not degrade during ripening. This result highlights the potential of liposomes to minimize losses of encapsulated enzymes into the whey during cheese production but also highlights the need to optimize the hydrophobicity, zeta potential, size and composition of the liposomes to maximize their use as vectors for enzyme addition in cheese to augment ripening.
    • Enzyme Modified Cheese Flavour Ingredients

      Wilkinson, M.G.; Kilcawley, Kieran; Mulholland, E.; Department of Agriculture, Food and the Marine, Ireland (Teagasc, 01/09/2000)
      Enzyme-modified cheeses (EMCs) are defined as concentrated cheese flavours produced enzymatically from cheeses of various ages and are principally used as an ingredient in processed foods, where they provide a cost-effective alternative to natural cheese. They can be used as the sole source of cheese flavour to intensify an existing cheese taste, or to impart a specific cheese character to a more bland product. Their main applications are in processed cheese, analogue cheese, cheese spreads, snack foods, soups, sauces, biscuits, dips and pet foods. Their main advantages over other cheese flavour ingredients are: low production costs, consistency, high flavour intensity, diverse flavour range, extended shelf- life, low storage costs and increased functionality. EMCs are generated utilising the same flavour pathways that occur in natural cheese ripening i.e. proteolysis, lipolysis and glycolysis. They are not as easy to differentiate as natural cheeses, as they are characterised by flavour and aroma alone as texture is not a factor in EMC production. The relationship of the flavour of EMCs to the flavour of the corresponding natural cheese remains unclear. This is especially true for Cheddar EMC which is commercially available in a range of Cheddar flavours . Despite the fact that a wide range of commercial EMCs are available, there is very little detailed information available regarding their properties or the specific production processes used. The main objective of this research was to build a knowledge base on EMC products and to utilise this to develop a biotechnological process for the production of improved enzyme modified cheeses for use as flavour ingredients. The strategy was to establish quantitative relationships between the compositional, proteolytic and lipolytic parameters and the sensory characteristics of EMCs. This data would then be used to develop a predictive model for flavour development in EMC production and the subsequent generation of an optimised EMC process enabling the generation of a range of cheese flavours from single or multiple substrates.
    • Evaluation of beef eating quality by Irish consumers

      McCarthy, Sinead N.; Henchion, Maeve; White, A.; Brandon, K.; Allen, Paul; Department of Agriculture, Food and the Marine, Ireland; 04/R&D/TN/256 (Elsevier, 08/05/2017)
      A consumer's decision to purchase beef is strongly linked to its sensory properties and consistent eating quality is one of the most important attributes. Consumer taste panels were held according to the Meat Standards Australia guidelines and consumers scored beef according to its palatability attributes and completed a socio-demographic questionnaire. Consumers were able to distinguish between beef quality on a scale from unsatisfactory to premium with high accuracy. Premium cuts of beef scored significantly higher on all of the scales compared to poorer quality cuts. Men rated grilled beef higher on juiciness and flavour scales compared to women. Being the main purchaser of beef had no impact on rating scores. Overall the results show that consumers can judge eating quality with high accuracy. Further research is needed to determine how best to communicate inherent benefits that are not visible into extrinsic eating quality indicators, to provide the consumer with consistent indications of quality at the point of purchase.
    • Future Protein Supply and Demand: Strategies and Factors Influencing a Sustainable Equilibrium

      Henchion, Maeve; Hayes, Maria; Mullen, Anne Maria; Fenelon, Mark; Tiwari, Brijesh K; Department of Agriculture, Food and the Marine, Ireland; 11/F/043 (MDPI, 20/07/2017)
      A growing global population, combined with factors such as changing socio-demographics, will place increased pressure on the world’s resources to provide not only more but also different types of food. Increased demand for animal-based protein in particular is expected to have a negative environmental impact, generating greenhouse gas emissions, requiring more water and more land. Addressing this “perfect storm” will necessitate more sustainable production of existing sources of protein as well as alternative sources for direct human consumption. This paper outlines some potential demand scenarios and provides an overview of selected existing and novel protein sources in terms of their potential to sustainably deliver protein for the future, considering drivers and challenges relating to nutritional, environmental, and technological and market/consumer domains. It concludes that different factors influence the potential of existing and novel sources. Existing protein sources are primarily hindered by their negative environmental impacts with some concerns around health. However, they offer social and economic benefits, and have a high level of consumer acceptance. Furthermore, recent research emphasizes the role of livestock as part of the solution to greenhouse gas emissions, and indicates that animal-based protein has an important role as part of a sustainable diet and as a contributor to food security. Novel proteins require the development of new value chains, and attention to issues such as production costs, food safety, scalability and consumer acceptance. Furthermore, positive environmental impacts cannot be assumed with novel protein sources and care must be taken to ensure that comparisons between novel and existing protein sources are valid. Greater alignment of political forces, and the involvement of wider stakeholders in a governance role, as well as development/commercialization role, is required to address both sources of protein and ensure food security.
    • Future Protein Supply and Demand: Strategies and Factors Influencing a Sustainable Equilibrium

      Henchion, Maeve; Hayes, Maria; Mullen, Anne Maria; Fenelon, Mark; Tiwari, Brijesh K; Department of Agriculture, Food and the Marine, Ireland (MDPI, 20/07/2017)
      A growing global population, combined with factors such as changing socio-demographics, will place increased pressure on the world’s resources to provide not only more but also different types of food. Increased demand for animal-based protein in particular is expected to have a negative environmental impact, generating greenhouse gas emissions, requiring more water and more land. Addressing this “perfect storm” will necessitate more sustainable production of existing sources of protein as well as alternative sources for direct human consumption. This paper outlines some potential demand scenarios and provides an overview of selected existing and novel protein sources in terms of their potential to sustainably deliver protein for the future, considering drivers and challenges relating to nutritional, environmental, and technological and market/consumer domains. It concludes that different factors influence the potential of existing and novel sources. Existing protein sources are primarily hindered by their negative environmental impacts with some concerns around health. However, they offer social and economic benefits, and have a high level of consumer acceptance. Furthermore, recent research emphasizes the role of livestock as part of the solution to greenhouse gas emissions, and indicates that animal-based protein has an important role as part of a sustainable diet and as a contributor to food security. Novel proteins require the development of new value chains, and attention to issues such as production costs, food safety, scalability and consumer acceptance. Furthermore, positive environmental impacts cannot be assumed with novel protein sources and care must be taken to ensure that comparisons between novel and existing protein sources are valid. Greater alignment of political forces, and the involvement of wider stakeholders in a governance role, as well as development/commercialization role, is required to address both sources of protein and ensure food security
    • Model System for the Production of Enzyme Modified Cheese (EMC) Flavours.

      Kilcawley, Kieran; Beresford, Tom; Lee, B.; Wilkinson, M.G.; Department of Agriculture, Food and the Marine, Ireland; Irish Dairy Levy Research Trust (Teagasc, 01/04/2002)
      Natural cheese flavour ingredients, in the form of enzyme modified cheeses (EMCs), are widely used in the convenience food industry and can provide high volume added opportunities for the cheese industry. Many EMCs are produced using commercial enzyme preparations and previous studies have indicated that they contain side activities in addition to their stated main activity (see DPRC Report No.10). Therefore, it is critical that the exact enzyme complement of these preparations are known before they can be used to produce EMC of specific requirements on a consistent basis. The scientific basis of rapid enzyme mediated flavour formation in the production of EMCs is not fully understood. Consequently this knowledge gap is a major obstacle in the development of high value cheese flavour ingredients. Hence, a major objective of this project was to deepen the scientific understanding of flavour formation with a view to the production of natural enzyme-mediated dairy flavour ingredients with commercial potential. The ultimate aim was to develop the technology to produce customised high value dairy flavour ingredients in an optimised process.
    • Monitoring post mortem changes in porcine muscle through 2-D DIGE proteome analysis of Longissimus muscle exudate

      Di Luca, Alessio; Elia, Giuliano; Mullen, Anne Maria; Hamill, Ruth M; Department of Agriculture, Food and the Marine, Ireland; 06RDNUIG470 (Biomed Central, 20/03/2013)
      Background: Meat quality is a complex trait influenced by a range of factors with post mortem biochemical processes highly influential in defining ultimate quality. High resolution two-dimensional DIfference Gel Electrophoresis (2-D DIGE) and Western blot were applied to study the influence of post mortem meat ageing on the proteome of pork muscle. Exudate collected from the muscle following centrifugation was analysed at three timepoints representing a seven day meat ageing period. Results: The intensity of 136 spots varied significantly (p < 0.05) across this post mortem period and 40 spots were identified using mass spectrometry. The main functional categories represented were metabolic proteins, stress-related proteins, transport and structural proteins. Metabolic and structural proteins were generally observed to increase in abundance post mortem and many likely represent the accumulation of the degradation products of proteolytic enzyme activity. In contrast, stress-related proteins broadly decreased in abundance across the ageing period. Stress response proteins have protective roles in maintaining cellular integrity and a decline in their abundance over time may correlate with a reduction in cellular integrity and the onset of meat ageing. Since cellular conditions alter with muscle ageing, changes in solubility may also contribute to observed abundance profiles. Conclusions: Muscle exudate provided valuable information about the pathways and processes underlying the post mortem ageing period, highlighting the importance of post mortem modification of proteins and their interaction for the development of meat quality traits.
    • Optimisation of plant protein and transglutaminase content in novel beef restructured steaks for older adults by central composite design

      Baugreet, Sephora; Kerry, Joseph P.; Brodkorb, Andre; Gomez, Carolina; Auty, Mark; Allen, Paul; Hamill, Ruth; Department of Agriculture, Food and the Marine, Ireland; Teagasc Walsh Fellowship Programme; 11/F/045 (Elsevier, 2018-03-29)
      With the goal of optimising a protein-enriched restructured beef steak targeted at the nutritional and chemosensory requirements of older adults, technological performance of thirty formulations, containing plant-based ingredients, pea protein isolate (PPI), rice protein (RP) and lentil flour (LF) with transglutaminase (TG) to enhance binding of meat pieces, were analysed. Maximal protein content of 28% in cooked product was achieved with PPI, RP and LF. Binding strength was primarily affected by TG, while textural parameters were improved with LF inclusion. Optimal formulation (F) to obtain a protein-enriched steak with lowest hardness values was achieved with TG (2%), PPI (8%), RP (9.35%) and LF (4%). F, F1S (optimal formulation 1 with added seasoning) and control restructured products (not containing plant proteins or seasonings) were scored by 120 consumers' aged over-65 years. Controls were most preferred (P < .05), while F1S were least liked by the older consumers. Consumer testing suggests further refinement and optimisation of restructured products with plant proteins should be undertaken.
    • Optimization of protein recovery from bovine lung by pH shift process using response surface methodology

      Lynch, Sarah A.; Álvarez García, Carlos; O'Neill, Eileen; Keenan, Derek F.; Mullen, Anne Maria; Department of Agriculture, Food and the Marine, Ireland; Teagasc Walsh Fellowship Programme; 11/F/043 (Wiley, 2017-09)
      BACKGROUND Response surface methodology (RSM) was used in a sequential manner to optimize solubilization and precipitation conditions in the recovery of protein from bovine lung using pH shift. RESULTS Separate D‐optimal designs were employed for protein solubilization and precipitation. Independent variables investigated for protein solubilization were time (10–120 min), temperature (4–20 °C), pH (8.0–11.0) and solvent/sample ratio (2.5–10). Variables for protein precipitation were time (0–60 min) and pH (4.25–6.00). Soluble protein yields ranged from 323 to 649 g kg−1 and the quadratic model for protein solubilization revealed a coefficient of determination R2 of 0.9958. Optimal conditions for maximum protein solubility were extraction time 140 min, temperature 19 °C, pH 10.8 and solvent/sample ratio 13.02. Protein precipitation yields varied from 407 to 667 g kg−1, giving a coefficient of determination R2 of 0.9335. Optimal conditions for maximum protein precipitation were pH 5.03 and 60 min. Based on the RSM model, solubilization conditions were manipulated to maximize protein solubilization under reduced water and alkaline usage. These conditions were also validated. CONCLUSION Models for solubilization and precipitation using bovine and porcine lung were validated; predicted and actual yields were in good agreement, showing cross‐species applicability of the results. © 2017 Society of Chemical Industry
    • Partitioning of starter bacteria and added exogenous enzyme activities between curd and whey during Cheddar cheese manufacture

      Doolan, I. A.; Nongonierma, Alice B.; Kilcawley, Kieran; Wilkinson, M.G.; Department of Agriculture, Food and the Marine, Ireland; 04/R&D/C/238 (Elsevier, 26/07/2013)
      Partitioning of starter bacteria and enzyme activities was investigated at different stages of Cheddar cheese manufacture using three exogenous commercial enzyme preparations added to milk or at salting. The enzyme preparations used were: Accelase AM317, Accelase AHC50, Accelerzyme CPG. Flow cytometric analysis indicated that AHC50 or AM317 consisted of permeabilised or dead cells and contained a range of enzyme activities. The CPG preparation contained only carboxypeptidase activity. Approximately 90% of starter bacteria cells partitioned with the curd at whey drainage. However, key enzyme activities partitioned with the bulk whey in the range of 22%–90%. An increased level of enzyme partitioning with the curd was observed for AHC50 which was added at salting, indicating that the mode of addition influenced partitioning. These findings suggest that further scope exists to optimise both bacterial and exogenous enzyme incorporation into cheese curd to accelerate ripening.
    • Regulatory polymorphisms in the bovine Ankyrin 1 gene promoter are associated with tenderness and intra-muscular fat content

      Aslan, Ozlem; Sweeney, Torres; Mullen, Anne Maria; Hamill, Ruth M; Department of Agriculture, Food and the Marine, Ireland (Biomed Central, 15/12/2010)
      Recent QTL and gene expression studies have highlighted ankyrins as positional and functional candidate genes for meat quality. Our objective was to characterise the promoter region of the bovine ankyrin 1 gene and to test polymorphisms for association with sensory and technological meat quality measures. Results Seven novel promoter SNPs were identified in a 1.11 kb region of the ankyrin 1 promoter in Angus, Charolais and Limousin bulls (n = 15 per breed) as well as 141 crossbred beef animals for which meat quality data was available. Eighteen haplotypes were inferred with significant breed variation in haplotype frequencies. The five most frequent SNPs and the four most frequent haplotypes were subsequently tested for association with sensory and technological measures of meat quality in the crossbred population. SNP1, SNP3 and SNP4 (which were subsequently designated regulatory SNPs) and SNP5 were associated with traits that contribute to sensorial and technological measurements of tenderness and texture; Haplotype 1 and haplotype 4 were oppositely correlated with traits contributing to tenderness (P < 0.05). While no single SNP was associated with intramuscular fat (IMF), a clear association with increased IMF and juiciness was observed for haplotype 2. Conclusion The conclusion from this study is that alleles defining haplotypes 2 and 4 could usefully contribute to marker SNP panels used to select individuals with improved IMF/juiciness or tenderness in a genome-assisted selection framework.
    • SNP variation in the promoter of the PRKAG3 gene and association with meat quality traits in pig

      Ryan, Marion T; Hamill, Ruth M; O'Halloran, Aisling M; Davey, Grace C; McBryan, Jean; Mullen, Anne Maria; McGee, Chris; Gispert, Marina; Southwood, Olwen I; Sweeney, Torres; et al. (Biomed Central, 25/07/2012)
      Background: The PRKAG3 gene encodes the γ3 subunit of adenosine monophosphate activated protein kinase (AMPK), a protein that plays a key role in energy metabolism in skeletal muscle. Non-synonymous single nucleotide polymorphisms (SNPs) in this gene such as I199V are associated with important pork quality traits. The objective of this study was to investigate the relationship between gene expression of the PRKAG3 gene, SNP variation in the PRKAG3 promoter and meat quality phenotypes in pork. Results: PRKAG3 gene expression was found to correlate with a number of traits relating to glycolytic potential (GP) and intramuscular fat (IMF) in three phenotypically diverse F1 crosses comprising of 31 Large White, 23 Duroc and 32 Pietrain sire breeds. The majority of associations were observed in the Large White cross. There was a significant association between genotype at the g.-311A>G locus and PRKAG3 gene expression in the Large White cross. In the same population, ten novel SNPs were identified within a 1.3 kb region spanning the promoter and from this three major haplotypes were inferred. Two tagging SNPs (g.-995A>G and g.-311A>G) characterised the haplotypes within the promoter region being studied. These two SNPs were subsequently genotyped in larger populations consisting of Large White (n = 98), Duroc (n = 99) and Pietrain (n = 98) purebreds. Four major haplotypes including promoter SNP’s g.-995A>G and g.-311A>G and I199V were inferred. In the Large White breed, HAP1 was associated with IMF% in the M. longissmus thoracis et lumborum (LTL) and driploss%. HAP2 was associated with IMFL% GP-influenced traits pH at 24 hr in LTL (pHULT), pH at 45 min in LTL (pH45LT) and pH at 45 min in the M. semimembranosus muscle (pH45SM). HAP3 was associated with driploss%, pHULT pH45LT and b* Minolta. In the Duroc breed, associations were observed between HAP1 and driploss% and pHUSM. No associations were observed with the remaining haplotypes (HAP2, HAP3 and HAP4) in the Duroc breed. The Pietrain breed was monomorphic in the promoter region. The I199V locus was associated with several GP-influenced traits across all three breeds and IMF% in the Large White and Pietrain breed. No significant difference in promoter function was observed for the three main promoter haplotypes when tested in vitro. Conclusion: Gene expression levels of the porcine PRKAG3 are associated with meat quality phenotypes relating to glycolytic potential and IMF% in the Large White breed, while SNP variation in the promoter region of the gene is associated with PRKAG3 gene expression and meat quality phenotypes.
    • Transcriptome analysis of porcine M. semimembranosus divergent in intramuscular fat as a consequence of dietary protein restriction

      Hamill, Ruth M; Aslan, Ozlem; Mullen, Anne Maria; O'Doherty, John V.; McBryan, Jean; Morris, Dermot G.; Sweeney, Torres; Department of Agriculture, Food and the Marine, Ireland (Biomed Central, 06/07/2013)
      Background: Intramuscular fat (IMF) content is positively correlated with aspects of pork palatability, including flavour, juiciness and overall acceptability. The ratio of energy to protein in the finishing diet of growing pigs can impact on IMF content with consequences for pork quality. The objective of this study was to compare gene expression profiles of Musculus semimembranosus (SM) of animals divergent for IMF as a consequence of protein dietary restriction in an isocaloric diet. The animal model was derived through the imposition of low or high protein diets during the finisher stage in Duroc gilts. RNA was extracted from post mortem SM tissue, processed and hybridised to Affymetrix porcine GeneChip® arrays. Results: IMF content of SM muscle was increased on the low protein diet (3.60 ± 0.38% versus 1.92 ± 0.35%). Backfat depth was also greater in animals on the low protein diet, and average daily gain and feed conversion ratio were lower, but muscle depth, protein content and moisture content were not affected. A total of 542 annotated genes were differentially expressed (DE) between animals on low and high protein diets, with 351 down-regulated and 191 up-regulated on the low protein diet. Transcript differences were validated for a subset of DE genes by qPCR. Alterations in functions related to cell cycle, muscle growth, extracellular matrix organisation, collagen development, lipogenesis and lipolysis, were observed. Expression of adipokines including LEP, TNFα and HIF1α were increased and the hypoxic stress response was induced. Many of the identified transcriptomic responses have also been observed in genetic and fetal programming models of differential IMF accumulation, indicating they may be robust biological indicators of IMF content. Conclusion: An extensive perturbation of overall energy metabolism in muscle occurs in response to protein restriction. A low protein diet can modulate IMF content of the SM by altering gene pathways involved in lipid biosynthesis and degradation; however this nutritional challenge negatively impacts protein synthesis pathways, with potential consequences for growth.