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  • Culicoides species composition and abundance on Irish cattle farms: implications for arboviral disease transmission

    Collins, Áine B; Mee, John F; Doherty, Michael L; Barrett, Damien J; England, Marion E; Teagasc Walsh Fellowship Programme (Biomed Central, 2018-08-17)
    Background Following the emergence of Schmallenberg virus (SBV) in Ireland in 2012, a sentinel herd surveillance program was established in the south of Ireland with the primary aim of investigating the species composition and abundance of Culicoides on livestock farms in the region. Methods Ultraviolet-light trapping for Culicoides was carried out on 10 sentinel farms. Each site was sampled fortnightly over 16 weeks (21st July to 5th November 2014). One Onderstepoort Veterinary Institute UV light trap was run overnight at each site and catches were transferred immediately into 70% ethanol. Culicoides were morphologically identified to species level. Collection site habitats were characterised using the Phase 1 habitat survey technique (Joint Nature Conservation Committee). Results A total of 23,929 individual Culicoides from 20 species was identified, including one species identified in Ireland for the first time, Culicoides cameroni. The most abundant species identified were Culicoides obsoletus/Culicoides scoticus (38%), Culicoides dewulfi (36%), Culicoides pulicaris (9%), Culicoides chiopterus (5%) and Culicoides punctatus (5%), comprising 93% of all Culicoides specimens identified. Collection site habitats were dominated by improved grassland and a combination of broadleaf woodland and native woodland species. Conclusions The most abundant species of Culicoides identified were the putative vectors of bluetongue virus (BTV) and SBV in northern Europe. Their presence and abundance demonstrates the potential for future transmission of arboviruses among livestock in this region.
  • Gene-trait matching across the Bifidobacterium longum pan-genome reveals considerable diversity in carbohydrate catabolism among human infant strains

    Arboleya, Silvia; Bottacini, Francesca; O’Connell-Motherway, Mary; Ryan, C. A; Ross, R. P; van Sinderen, Douwe; Stanton, Catherine; Science Foundation Ireland; Department of Agriculture, Food and the Marine, Ireland; SFI/12/RC/2273; 10FDairy (Biomed Central, 08/01/2018)
    Background Bifidobacterium longum is a common member of the human gut microbiota and is frequently present at high numbers in the gut microbiota of humans throughout life, thus indicative of a close symbiotic host-microbe relationship. Different mechanisms may be responsible for the high competitiveness of this taxon in its human host to allow stable establishment in the complex and dynamic intestinal microbiota environment. The objective of this study was to assess the genetic and metabolic diversity in a set of 20 B. longum strains, most of which had previously been isolated from infants, by performing whole genome sequencing and comparative analysis, and to analyse their carbohydrate utilization abilities using a gene-trait matching approach. Results We analysed their pan-genome and their phylogenetic relatedness. All strains clustered in the B. longum ssp. longum phylogenetic subgroup, except for one individual strain which was found to cluster in the B. longum ssp. suis phylogenetic group. The examined strains exhibit genomic diversity, while they also varied in their sugar utilization profiles. This allowed us to perform a gene-trait matching exercise enabling the identification of five gene clusters involved in the utilization of xylo-oligosaccharides, arabinan, arabinoxylan, galactan and fucosyllactose, the latter of which is an abundant human milk oligosaccharide (HMO). Conclusions The results showed high diversity in terms of genes and predicted glycosyl-hydrolases, as well as the ability to metabolize a large range of sugars. Moreover, we corroborate the capability of B. longum ssp. longum to metabolise HMOs. Ultimately, their intraspecific genomic diversity and the ability to consume a wide assortment of carbohydrates, ranging from plant-derived carbohydrates to HMOs, may provide an explanation for the competitive advantage and persistence of B. longum in the human gut microbiome.
  • Effect of early calf-hood nutrition on the transcriptomic profile of subcutaneous adipose tissue in Holstein-Friesian bulls

    English, Anne-Marie; Waters, Sineád M; Cormican, Paul; Byrne, Colin J; Fair, Seán; Kenny, David A; Department of Agriculture, Food and the Marine, Ireland; Irish Research Council; 11/S/116; GOIPG/2013/1391 (Biomed Central, 2018-04-24)
    Background Adipose tissue is a major endocrine organ and is thought to play a central role in the metabolic control of reproductive function in cattle. Plane of nutrition during early life has been shown to influence the timing of puberty in both male and female cattle, though the exact biological mechanisms involved are currently unknown. The aim of this study was to investigate the effect of early calf-hood nutrition on the transcriptomic profile of subcutaneous adipose tissue in Holstein-Friesian bulls to identify possible downstream effects on reproductive physiology. Results Holstein-Friesian bull calves with a mean (±S.D.) age and bodyweight of 19 (±8.2) days and 47.5 (±5.3) kg, respectively, were assigned to either a high (n = 10) or low (n = 10) plane of nutrition. Calves were fed in order to achieve an overall growth rate of 1.08 and 0.57 kg/day for the high and low plane of nutrition treatments, respectively. At 126 days of age, the bulls were euthanized, subcutaneous adipose tissue samples were harvested and RNAseq analysis was performed. There were 674 genes differentially expressed in adipose tissue of calves on the low compared with the high plane of nutrition (P < 0.05; FDR < 0.05; fold change > 2.0). High plane of nutrition positively altered the expression of genes across an array of putative biological processes but the most dominant cellular processes affected were cellular energy production and branched chain amino acid degradation. A high plane of nutrition caused upregulation of genes such as leptin (LEP) and adiponectin (ADIPOQ), which are known to directly affect reproductive function. Conclusions These results provide an insight into the effect of augmenting the plane of nutrition of Holstein-Friesian bull calves in the prepubertal period on the transcriptome of adipose tissue.
  • Genomic prediction of crown rust resistance in Lolium perenne

    Arojju, Sai Krishna; Conaghan, Patrick; Barth, Susanne; Milbourne, Dan; Casler, Michael D.; Hodkinson, Trevor R.; Michel, Thibauld; Byrne, Stephen L.; Department of Agriculture, Food and the Marine, Ireland; Marie Sklodowska-Curie; Teagasc Walsh Fellowship Programme Ireland; RSF 11/S/109; 658031; 658031 (Biomed Central, 29/05/2018)
    Background Genomic selection (GS) can accelerate genetic gains in breeding programmes by reducing the time it takes to complete a cycle of selection. Puccinia coronata f. sp lolli (crown rust) is one of the most widespread diseases of perennial ryegrass and can lead to reductions in yield, persistency and nutritional value. Here, we used a large perennial ryegrass population to assess the accuracy of using genome wide markers to predict crown rust resistance and to investigate the factors affecting predictive ability. Results Using these data, predictive ability for crown rust resistance in the complete population reached a maximum of 0.52. Much of the predictive ability resulted from the ability of markers to capture genetic relationships among families within the training set, and reducing the marker density had little impact on predictive ability. Using permutation based variable importance measure and genome wide association studies (GWAS) to identify and rank markers enabled the identification of a small subset of SNPs that could achieve predictive abilities close to those achieved using the complete marker set. Conclusion Using a GWAS to identify and rank markers enabled a small panel of markers to be identified that could achieve higher predictive ability than the same number of randomly selected markers, and predictive abilities close to those achieved with the entire marker set. This was particularly evident in a sub-population characterised by having on-average higher genome-wide linkage disequilibirum (LD). Higher predictive abilities with selected markers over random markers suggests they are in LD with QTL. Accuracy due to genetic relationships will decay rapidly over generations whereas accuracy due to LD will persist, which is advantageous for practical breeding applications.
  • Prevalence of welfare outcomes in the weaner and finisher stages of the production cycle on 31 Irish pig farms

    van Staaveren, Nienke; Calderón Díaz, Julia A; Garcia Manzanilla, Edgar; Hanlon, Alison; Boyle, Laura A; Department of Agriculture, Food and the Marine, Ireland; 11/S/107 (Biomed Central, 2018-03-27)
    Background Knowledge on the most prevalent welfare problems for pigs in different production stages is required to improve herd management plans. Thirty-one farrow-to-finish pig farms were visited between July and November 2015 to assess the welfare of pigs using the multicriteria approach of the Welfare Quality® protocol. On each farm, 6 pens were selected using proportionate stratified sampling in the first weaner (S1, 4 to 8 wks), second weaner (S2, 8 to 13 wks) and finisher stage (S3, 13 to 23 wks), excluding hospital pens. Each pen was observed for 10 min and the number of pigs affected by different welfare outcomes was recorded. The percentage of pigs affected was calculated and ranked to identify the most prevalent outcomes within each production stage. Differences between production stages were analysed using generalised linear mixed models for binomial data with pen within stage and farm as a random effect. Results Tail and ear lesions showed the highest prevalence; however, large variation was observed between farms. In S1 the most prevalent welfare outcomes (presented as median prevalence) were poor body condition (4.4%), lethargic pigs (1.5%), scouring (20.3% of pens) and huddling (3.7%). In S2 and S3 outcomes related to injurious behaviour (tail lesions: 5.9% [S2] and 10.5% [S3], ear lesions: 9.1% [S2] and 3.3% [S3], and flank lesions: 0.4% [S2] and 1.3% [S3]), lameness (0.8% [S2] and 1.1% [S3]), bursitis (3.9% [S2] and 7.5% [S3]) and hernias (1.6% [S2] and 1.8% [S3]) were more prevalent. Conclusions A large variation was observed for the recorded welfare outcomes corresponding to the different challenges pigs experience during the different stages of production on commercial pig farms. The prevalence of pigs affected by lesions caused by injurious behavior is a cause for concern and requires a collaborative approach to identify appropriate intervention strategies. This information could be used to further investigate appropriate benchmark values for different welfare outcomes that would assist the pig industry to develop appropriate health and welfare management plans to minimise welfare problems. At herd level such plans should include information on aspects of intervention, treatment, and the management of hospital pens as well as euthanasia.
  • Species classifier choice is a key consideration when analysing low-complexity food microbiome data

    Walsh, Aaron M; Crispie, Fiona; O’Sullivan, Orla; Finnegan, Laura; Claesson, Marcus J; Cotter, Paul D; Science Foundation Ireland; SFI/12/RC/2273; 11/PI/1137; 13/SIRG/2160 (Biomed Central, 2018-03-20)
    Background The use of shotgun metagenomics to analyse low-complexity microbial communities in foods has the potential to be of considerable fundamental and applied value. However, there is currently no consensus with respect to choice of species classification tool, platform, or sequencing depth. Here, we benchmarked the performances of three high-throughput short-read sequencing platforms, the Illumina MiSeq, NextSeq 500, and Ion Proton, for shotgun metagenomics of food microbiota. Briefly, we sequenced six kefir DNA samples and a mock community DNA sample, the latter constructed by evenly mixing genomic DNA from 13 food-related bacterial species. A variety of bioinformatic tools were used to analyse the data generated, and the effects of sequencing depth on these analyses were tested by randomly subsampling reads. Results Compositional analysis results were consistent between the platforms at divergent sequencing depths. However, we observed pronounced differences in the predictions from species classification tools. Indeed, PERMANOVA indicated that there was no significant differences between the compositional results generated by the different sequencers (p = 0.693, R2 = 0.011), but there was a significant difference between the results predicted by the species classifiers (p = 0.01, R2 = 0.127). The relative abundances predicted by the classifiers, apart from MetaPhlAn2, were apparently biased by reference genome sizes. Additionally, we observed varying false-positive rates among the classifiers. MetaPhlAn2 had the lowest false-positive rate, whereas SLIMM had the greatest false-positive rate. Strain-level analysis results were also similar across platforms. Each platform correctly identified the strains present in the mock community, but accuracy was improved slightly with greater sequencing depth. Notably, PanPhlAn detected the dominant strains in each kefir sample above 500,000 reads per sample. Again, the outputs from functional profiling analysis using SUPER-FOCUS were generally accordant between the platforms at different sequencing depths. Finally, and expectedly, metagenome assembly completeness was significantly lower on the MiSeq than either on the NextSeq (p = 0.03) or the Proton (p = 0.011), and it improved with increased sequencing depth. Conclusions Our results demonstrate a remarkable similarity in the results generated by the three sequencing platforms at different sequencing depths, and, in fact, the choice of bioinformatics methodology had a more evident impact on results than the choice of sequencer did.
  • Reproducible protocols for metagenomic analysis of human faecal phageomes

    Shkoporov, Andrey N.; Ryan, Feargal J.; Draper, Lorraine A.; Forde, Amanda; Stockdale, Stephen R.; Daly, Karen M.; McDonnell, Siobhan A.; Nolan, James A.; Sutton, Thomas D.S.; Dalmasso, Marion; McCann, Angela; Ross, R. Paul; Hill, Colin; Science Foundation Ireland; SFI/12/RC/2273; SFI/14/SP APC/B3032 (Biomed Central, 2018-04-10)
    Background Recent studies have demonstrated that the human gut is populated by complex, highly individual and stable communities of viruses, the majority of which are bacteriophages. While disease-specific alterations in the gut phageome have been observed in IBD, AIDS and acute malnutrition, the human gut phageome remains poorly characterised. One important obstacle in metagenomic studies of the human gut phageome is a high level of discrepancy between results obtained by different research groups. This is often due to the use of different protocols for enriching virus-like particles, nucleic acid purification and sequencing. The goal of the present study is to develop a relatively simple, reproducible and cost-efficient protocol for the extraction of viral nucleic acids from human faecal samples, suitable for high-throughput studies. We also analyse the effect of certain potential confounding factors, such as storage conditions, repeated freeze-thaw cycles, and operator bias on the resultant phageome profile. Additionally, spiking of faecal samples with an exogenous phage standard was employed to quantitatively analyse phageomes following metagenomic sequencing. Comparative analysis of phageome profiles to bacteriome profiles was also performed following 16S rRNA amplicon sequencing. Results Faecal phageome profiles exhibit an overall greater individual specificity when compared to bacteriome profiles. The phageome and bacteriome both exhibited moderate change when stored at + 4 °C or room temperature. Phageome profiles were less impacted by multiple freeze-thaw cycles than bacteriome profiles, but there was a greater chance for operator effect in phageome processing. The successful spiking of faecal samples with exogenous bacteriophage demonstrated large variations in the total viral load between individual samples. Conclusions The faecal phageome sequencing protocol developed in this study provides a valuable additional view of the human gut microbiota that is complementary to 16S amplicon sequencing and/or metagenomic sequencing of total faecal DNA. The protocol was optimised for several confounding factors that are encountered while processing faecal samples, to reduce discrepancies observed within and between research groups studying the human gut phageome. Rapid storage, limited freeze-thaw cycling and spiking of faecal samples with an exogenous phage standard are recommended for optimum results.
  • Effect of pre-milking teat disinfection on new mastitis infection rates of dairy cows

    Gleeson, David; Flynn, Jimmy; O'Brien, Bernadette (Biomed Central, 2018-04-18)
    Background The practise of teat disinfection prior to cluster attachment for milking is being adopted by farmers in Ireland, particularly where there are herd issues with new infection rates. Pre-milking teat disinfection has been shown to reduce bacterial numbers on teat skin and to be most effective against environmental bacteria such as Escherichia coli and Streptococcus uberis. A split udder design experiment was undertaken on two research herds (A = 96 cows: B = 168 cows) to test the benefit of pre-milking teat disinfection on new mastitis infection levels. The disinfectant was applied to the left front and right hind teats of all cows in each herd and the right front and left hind teats received no disinfectant treatment prior to milking over a complete lactation. Individual quarter foremilk samples were taken on 5 occasions during the lactation and all clinical cases were recorded. The presence and number of staphylococcus and streptococcus bacteria on teat skin of a random sample of experimental cows (n = 20) was measured on 3 occasions during lactation (April, June, and October). Results Pre-milking teat disinfection had no significant impact on quarter SCC and new infection rates (P > 0.05). The median SCC was 169 (95% CI = 144–198) × 103 cells/mL and 170 (95% CI = 145–199) × 103 cells/mL for disinfected teats and non-disinfected teats, respectively. There were no differences in SCC observed between herds (A = 161 (95% CI = 127–205) × 103 cells/mL; B = 169 (95% CI = 144–198) × 103 cells/mL) over the complete lactation. Bacterial levels on teat skin were reduced significantly with pre-milking teat disinfection compared to teats receiving no disinfectant (P < 0.001). Total infections (clinical and sub-clinical) were similar for disinfected teats (n = 36) and not disinfected teats (n = 40), respectively. Staphylococcus aureus (n = 47) and Strep. uberis (n = 9) were identified as the predominant bacteria in quarter foremilk samples with both clinical and sub-clinical infections. Conclusion SCC and new infection rates were similar in non-disinfected teats and disinfected (pre-milking) teats. The routine application of pre-milking teat disinfectant in pasture-grazed herds is unlikely to be of benefit where herd SCC is below 200 × 103 cells/mL.
  • Gene-trait matching across the Bifidobacterium longum pan-genome reveals considerable diversity in carbohydrate catabolism among human infant strains

    Arboleya, Silvia; Bottacini, Francesca; O’Connell-Motherway, Mary; Ryan, C. A; Ross, R. P; van Sinderen, Douwe; Stanton, Catherine; Science Foundation Ireland; Department of Agriculture, Food and the Marine, Ireland; SFI/12/RC/2273; 10FDairy (Biomed Central, 08/01/2018)
    Background Bifidobacterium longum is a common member of the human gut microbiota and is frequently present at high numbers in the gut microbiota of humans throughout life, thus indicative of a close symbiotic host-microbe relationship. Different mechanisms may be responsible for the high competitiveness of this taxon in its human host to allow stable establishment in the complex and dynamic intestinal microbiota environment. The objective of this study was to assess the genetic and metabolic diversity in a set of 20 B. longum strains, most of which had previously been isolated from infants, by performing whole genome sequencing and comparative analysis, and to analyse their carbohydrate utilization abilities using a gene-trait matching approach. Results We analysed their pan-genome and their phylogenetic relatedness. All strains clustered in the B. longum ssp. longum phylogenetic subgroup, except for one individual strain which was found to cluster in the B. longum ssp. suis phylogenetic group. The examined strains exhibit genomic diversity, while they also varied in their sugar utilization profiles. This allowed us to perform a gene-trait matching exercise enabling the identification of five gene clusters involved in the utilization of xylo-oligosaccharides, arabinan, arabinoxylan, galactan and fucosyllactose, the latter of which is an abundant human milk oligosaccharide (HMO). Conclusions The results showed high diversity in terms of genes and predicted glycosyl-hydrolases, as well as the ability to metabolize a large range of sugars. Moreover, we corroborate the capability of B. longum ssp. longum to metabolise HMOs. Ultimately, their intraspecific genomic diversity and the ability to consume a wide assortment of carbohydrates, ranging from plant-derived carbohydrates to HMOs, may provide an explanation for the competitive advantage and persistence of B. longum in the human gut microbiome.
  • Risk factors associated with exposure to bovine respiratory disease pathogens during the peri-weaning period in dairy bull calves

    Murray, Gerard M; More, Simon J; Clegg, Tracy A; Earley, Bernadette; O’Neill, Rónan G; Johnston, Dayle; Gilmore, John; Nosov, Mikhail; McElroy, Máire C; Inzana, Thomas J; Cassidy, Joseph P (Biomed Central, 2018-02-27)
    Background Bovine respiratory disease (BRD) remains among the leading causes of death of cattle internationally. The objective of this study was to identify risk factors associated with exposure to BRD pathogens during the peri-weaning period (day (d)-14 to d 14 relative to weaning at 0) in dairy bull calves using serological responses to these pathogens as surrogate markers of exposure. Clinically normal Holstein-Friesian and Jersey breed bull calves (n = 72) were group housed in 4 pens using a factorial design with calves of different breeds and planes of nutrition in each pen. Intrinsic, management and clinical data were collected during the pre-weaning (d − 56 to d − 14) period. Calves were gradually weaned over 14 days (d − 14 to d 0). Serological analysis for antibodies against key BRD pathogens (BRSV, BPI3V, BHV-1, BHV-4, BCoV, BVDV and H. somni) was undertaken at d − 14 and d 14. Linear regression models (for BVDV, BPI3V, BHV-1, BHV-4, BCoV and H. somni) and a single mixed effect random variable model (for BRSV) were used to identify risk factors for changes in antibody levels to these pathogens. Results BRSV was the only pathogen which demonstrated clustering by pen. Jersey calves experienced significantly lower changes in BVDV S/P than Holstein-Friesian calves. Animals with a high maximum respiratory score (≥8) recorded significant increases in H. somni S/P during the peri-weaning period when compared to those with respiratory scores of ≤3. Haptoglobin levels of between 1.32 and 1.60 mg/ml at d − 14 were significantly associated with decreases in BHV-1 S/N during the peri-weaning period. Higher BVDV S/P ratios at d − 14 were significantly correlated with increased changes in serological responses to BHV-4 over the peri-weaning period. Conclusions Haptoglobin may have potential as a predictor of exposure to BHV-1. BRSV would appear to play a more significant role at the ‘group’ rather than ‘individual animal’ level. The significant associations between the pre-weaning levels of antibodies to certain BRD pathogens and changes in the levels of antibodies to the various pathogens during the peri-weaning period may reflect a cohort of possibly genetically linked ‘better responders’ among the study population.
  • Prevalence of Bovine Viral Diarrhoea Virus (BVDV), Bovine Herpes Virus 1 (BHV 1), Leptospirosis and Neosporosis, and associated risk factors in 161 Irish beef herds

    Barrett, Damien; Parr, Mervyn; Fagan, John; Johnson, Alan; Tratalos, Jamie; Lively, Francis; Diskin, Michael; Kenny, David; Department of Agriculture, Food and the Marine, Ireland (Biomed Central, 2018-01-06)
    Background There are limited data available, in Ireland or elsewhere, to determine the extent of exposure to various endemic diseases among beef cows and factors associated with exposure to causative pathogens. The objectives of this study were to determine the herd and within herd prevalence of Bovine Viral Diarrhoea Virus (BVDV), Bovine Herpes Virus 1 (BHV-1), Leptospirosis and Neosporosis in a large scale study of commercial beef herds on the island of Ireland, and to examine herd level factors associated with exposure to these pathogens in these herds. Results The average number of cows tested per herd was 35.5 (median 30). Herd level seroprevalence to Bovine Herpesvirus-1(BHV-1), Bovine Viral-Diarrhoea Virus (BVDV), Leptospirosis and Neosporosis was 90%, 100%, 91% and 67%, respectively, while the mean within herd prevalence for the these pathogens was 40%, 77.7%, 65.7% and 5.7%, respectively. The study confirms that the level of seroconversion for the four pathogens of interest increases with herd size. There was also evidence that exposure to one pathogen may increase the risk of exposure to another pathogen. Conclusions Herd level seroprevalences were in excess of 90% for BVDV, BHV-1 and Leptosporosis. Larger herds were subject to increased exposure to disease pathogens. This study suggests that exposure to several pathogens may be associated with the further exposure to other pathogens.
  • Comparative genomic and metabolic analysis of three Lactobacillus paracasei cheese isolates reveals considerable genomic differences in strains from the same niche

    Stefanovic, Ewelina; McAuliffe, Olivia; Teagasc Walsh Fellowship Programme; 2012040 (Biomed Central, 2018-03-20)
    Background Strains of Lactobacillus paracasei are present in many diverse environments, including dairy and plant materials and the intestinal tracts of humans and animals. Their adaptation to various niches is correlated to intra-species diversity at the genomic and metabolic level. In this study, we compared the genome sequences of three L. paracasei strains isolated from mature Cheddar cheeses, two of which (DPC4206 and DPC4536) shared the same genomic fingerprint by PFGE, but demonstrated varying metabolic capabilities. Results Genome sizes varied from 2.9 Mbp for DPC2071, to 3.09 Mbp for DPC4206 and 3.08 Mpb for DPC4536. The presence of plasmids was a distinguishing feature between the strains with strain DPC2071 possessing an unusually high number of plasmids (up to 11), while DPC4206 had one plasmid and DPC4536 harboured no plasmids. Each of the strains possessed specific genes not present in the other two analysed strains. The three strains differed in their abundance of sugar-converting genes, and in the types of sugars that could be used as energy sources. Genes involved in the metabolism of sugars not usually connected with the dairy niche, such as myo-inositol and pullulan were also detected, but strains did not utilise these sugars. The genetic content of the three strains differed in regard to specific genes for arginine and sulfur-containing amino acid metabolism and genes contributing to resistance to heavy metal ions. In addition, variability in the presence of phage remnants and phage protection systems was evident. Conclusions The findings presented in this study confirm a considerable level of heterogeneity of Lactobacillus paracasei strains, even between strains isolated from the same niche.
  • Gene-trait matching across the Bifidobacterium longum pan-genome reveals considerable diversity in carbohydrate catabolism among human infant strains

    Arboleya, Silvia; Bottacini, Francesca; O’Connell-Motherway, Mary; Ryan, C. A; Ross, R. P; van Sinderen, Douwe; Stanton, Catherine; Science Foundation Ireland; Department of Agriculture, Food and the Marine, Ireland; SFI/12/RC/2273; 10FDairy (Biomed Central, 08/01/2018)
    Background Bifidobacterium longum is a common member of the human gut microbiota and is frequently present at high numbers in the gut microbiota of humans throughout life, thus indicative of a close symbiotic host-microbe relationship. Different mechanisms may be responsible for the high competitiveness of this taxon in its human host to allow stable establishment in the complex and dynamic intestinal microbiota environment. The objective of this study was to assess the genetic and metabolic diversity in a set of 20 B. longum strains, most of which had previously been isolated from infants, by performing whole genome sequencing and comparative analysis, and to analyse their carbohydrate utilization abilities using a gene-trait matching approach. Results We analysed their pan-genome and their phylogenetic relatedness. All strains clustered in the B. longum ssp. longum phylogenetic subgroup, except for one individual strain which was found to cluster in the B. longum ssp. suis phylogenetic group. The examined strains exhibit genomic diversity, while they also varied in their sugar utilization profiles. This allowed us to perform a gene-trait matching exercise enabling the identification of five gene clusters involved in the utilization of xylo-oligosaccharides, arabinan, arabinoxylan, galactan and fucosyllactose, the latter of which is an abundant human milk oligosaccharide (HMO). Conclusions The results showed high diversity in terms of genes and predicted glycosyl-hydrolases, as well as the ability to metabolize a large range of sugars. Moreover, we corroborate the capability of B. longum ssp. longum to metabolise HMOs. Ultimately, their intraspecific genomic diversity and the ability to consume a wide assortment of carbohydrates, ranging from plant-derived carbohydrates to HMOs, may provide an explanation for the competitive advantage and persistence of B. longum in the human gut microbiome.
  • Bile acids at the cross-roads of gut microbiome–host cardiometabolic interactions

    Ryan, Paul M; Stanton, Catherine; Caplice, Noel M; Science Foundation Ireland; Enterprise Ireland Commercialization Fund; SFI/12/RC/2273; CF/2013/3030A/B (Biomed Central, 28/12/2017)
    While basic and clinical research over the last several decades has recognized a number of modifiable risk factors associated with cardiometabolic disease progression, additional and alternative biological perspectives may offer novel targets for prevention and treatment of this disease set. There is mounting preclinical and emerging clinical evidence indicating that the mass of metabolically diverse microorganisms which inhabit the human gastrointestinal tract may be implicated in initiation and modulation of cardiovascular and metabolic disease outcomes. The following review will discuss this gut microbiome–host metabolism axis and address newly proposed bile-mediated signaling pathways through which dysregulation of this homeostatic axis may influence host cardiovascular risk. With a central focus on the major nuclear and membrane-bound bile acid receptor ligands, we aim to review the putative impact of microbial bile acid modification on several major phenotypes of metabolic syndrome, from obesity to heart failure. Finally, attempting to synthesize several separate but complementary hypotheses, we will review current directions in preclinical and clinical investigation in this evolving field.
  • Birth delivery method affects expression of immune genes in lung and jejunum tissue of neonatal beef calves

    Surlis, Carla; McNamara, Keelan; O’Hara, Eoin; Waters, Sinead; Beltman, Marijke; Cassidy, Joseph; Kenny, David (Biomed Central, 2017-12-14)
    Background Caesarean section is a routine veterinary obstetrical procedure employed to alleviate dystocia in cattle. However, CS, particularly before the onset of labour, is known to negatively affect neonatal respiration and metabolic adaptation in humans, though there is little published information for cattle. The aim of this study was to investigate the effect of elective caesarean section (ECS) or normal trans-vaginal (TV) delivery, on lung and jejunal gene expression profiles of neonatal calves. Results Paternal half-sib Angus calves (gestation length 278 + 1.8 d) were delivered either transvaginally (TV; n = 8) or by elective caesarean section (ECS; n = 9) and immediately euthanized. Lung and jejunum epithelial tissue was isolated and snap frozen. Total RNA was extracted using Trizol reagent and reverse transcribed to generate cDNA. For lung tissue, primers were designed to target genes involved in immunity, surfactant production, cellular detoxification, membrane transport and mucin production. Primers for jejunum tissue were chosen to target mucin production, immunoglobulin uptake, cortisol reaction and membrane trafficking. Quantitative real-time PCR reactions were performed and data were statistically analysed using mixed models ANOVA. In lung tissue the expression of five genes were affected (p < 0.05) by delivery method. Four of these genes were present at lower (LAP, CYP1A1, SCN11α and SCN11β) and one (MUC5AC) at higher abundance in ECS compared with TV calves. In jejunal tissue, expression of TNFα, Il-1β and 1 l-6 was higher in ECS compared with TV calves. Conclusions This novel study shows that ECS delivery affects the expression of key genes involved in the efficiency of the pulmonary liquid to air transition at birth, and may lead to an increased inflammatory response in jejunal tissue, which could compromise colostral immunoglobulin absorption. These findings are important to our understanding of the viability and management of neonatal calves born through ECS.
  • Effect of short term diet restriction on gene expression in the bovine hypothalamus using next generation RNA sequencing technology

    Matthews, Daragh; Diskin, Michael G; Kenny, David A; Creevey, Christopher J; Keogh, Kate; Waters, Sinead M (Biomed Central, 2017-11-09)
    Background Negative energy balance (NEB) is an imbalance between energy intake and energy requirements for lactation and body maintenance affecting high-yielding dairy cows and is of considerable economic importance due to its negative impact on fertility and health in dairy herds. It is anticipated that the cow hypothalamus experiences extensive biochemical changes during the early post partum period in an effort to re-establish metabolic homeostasis. However, there is variation in the tolerance to NEB between individual cows. In order to understand the genomic regulation of ovulation in hypothalamic tissue during NEB, mRNA transcriptional patterns between tolerant and sensitive animals were examined. A short term dietary restriction heifer model was developed which induced abrupt onset of anoestrus in some animals (Restricted Anovulatory; RA) while others maintained oestrous cyclicity (Restricted Ovulatory; RO). A third control group (C) received a higher level of normal feeding. Results A total of 15,295 genes were expressed in hypothalamic tissue. Between RA and C groups 137 genes were differentially expressed, whereas between RO and C, 32 genes were differentially expressed. Differentially expressed genes were involved in the immune response and cellular motility in RA and RO groups, respectively, compared to C group. The largest difference between groups was observed in the comparison between RA and RO heifers, with 1094 genes shown to be significantly differentially expressed (SDE). Pathway analysis showed that these SDE genes were associated with 6 canonical pathways (P < 0.01), of which neuroactive ligand-receptor interaction was the most significant. Within the comparisons the main over-represented pathway functions were immune response including neuroprotection (CXCL10, Q1KLR3, IFIH1, IL1 and IL8; RA v C and RA v RO); energy homeostasis (AgRP and NPY; RA v RO); cell motility (CADH1, DSP and TSP4; RO v C) and prevention of GnRH release (NTSR1 IL1α, IL1β, NPY and PACA; RA v RO). Conclusions This information will assist in understanding the genomic factors regulating the influence of diet restriction on fertility and may assist in optimising nutritional and management systems for the improvement in reproductive performance.
  • Quantitative trait loci associated with different polar metabolites in perennial ryegrass - providing scope for breeding towards increasing certain polar metabolites

    Foito, Alexandre; Hackett, Christine A; Stewart, Derek; Velmurugan, Janaki; Milbourne, Dan; Byrne, Stephen L; Barth, Susanne; Department of Agriculture, Food and the Marine, Ireland; RSF 06–346 (Biomed Central, 10/10/2017)
    Background Recent advances in the mapping of biochemical traits have been reported in Lolium perenne. Although the mapped traits, including individual sugars and fatty acids, contribute greatly towards ruminant productivity, organic acids and amino acids have been largely understudied despite their influence on the ruminal microbiome. Results In this study, we used a targeted gas-chromatography mass spectrometry (GC-MS) approach to profile the levels of 25 polar metabolites from different classes (sugars, amino acids, phenolic acids, organic acids and other nitrogen-containing compounds) present in a L. perenne F2 population consisting of 325 individuals. A quantitative trait (QTL) mapping approach was applied and successfully identified QTLs regulating seven of those polar metabolites (L-serine, L-leucine, glucose, fructose, myo-inositol, citric acid and 2, 3-hydroxypropanoic acid).Two QTL mapping approaches were carried out using SNP markers on about half of the population only and an imputation approach using SNP and DArT markers on the entire population. The imputation approach confirmed the four QTLs found in the SNP-only analysis and identified a further seven QTLs. Conclusions These results highlight the potential of utilising molecular assisted breeding in perennial ryegrass to modulate a range of biochemical quality traits with downstream effects in livestock productivity and ruminal digestion.
  • Stress and immunological response of heifers divergently ranked for residual feed intake following an adrenocorticotropic hormone challenge

    Kelly, A. K; Lawrence, P.; Earley, Bernadette; Kenny, David A.; McGee, Mark (Biomed Central, 2017-08-08)
    Background When an animal is exposed to a stressor, metabolic rate, energy consumption and utilisation increase primarily through activation of the hypothalamic-pituitary-adrenal (HPA) axis. Changes to partitioning of energy by an animal are likely to influence the efficiency with which it is utilised. Therefore, this study aimed to determine the physiological stress response to an exogenous adrenocorticotropic hormone (ACTH) challenge in beef heifers divergently ranked on phenotypic residual feed intake (RFI). Results Data were collected on 34 Simmental weaning beef heifers the progeny of a well characterized and divergently bred RFI suckler beef herd. Residual feed intake was determined on each animal during the post-weaning stage over a 91-day feed intake measurement period during which they were individually offered adlibitum grass silage and 2 kg of concentrate per head once daily. The 12 highest [0.34 kg DM/d] and 12 lowest [−0.48 kg DM/d] ranking animals on RFI were selected for use in this study. For the physiological stress challenge heifers (mean age 605 ± 13 d; mean BW 518 ± 31.4 kg) were fitted aseptically with indwelling jugular catheters to facilitate intensive blood collection. The response of the adrenal cortex to a standardised dose of ACTH (1.98 IU/kg metabolic BW0.75) was examined. Serial blood samples were analysed for plasma cortisol, ACTH and haematology variables. Heifers differing in RFI did not differ (P = 0.59) in ACTH concentrations. Concentration of ACTH peaked (P < 0.001) in both RFI groups at 20 min post-ACTH administration, following which concentration declined to baseline levels by 150 min. Similarly, cortisol systemic profile peaked at 60 min and concentrations remained continuously elevated for 150 min. A RFI × time interaction was detected for cortisol concentrations (P = 0.06) with high RFI heifers had a greater cortisol response than Low RFI from 40 min to 150 min relative to ACTH administration. Cortisol response was positively associated with RFI status (r = 0.32; P < 0.01). No effect of RFI was evident for neutrophil, lymphocytes, monocyte, eosinophils and basophil count. Plasma red blood cell number (6.07 vs. 6.23; P = 0.02) and hematocrit percentage (23.2 vs. 24.5; P = 0.02) were greater for low than high RFI animals. Conclusions Evidence is provided that feed efficiency is associated with HPA axis function and susceptibility to stress, and responsiveness of the HPA axis is likely to contribute to appreciable variation in the efficiency feed utilisation of cattle.
  • Illumina MiSeq 16S amplicon sequence analysis of bovine respiratory disease associated bacteria in lung and mediastinal lymph node tissue

    Johnston, Dayle; Earley, Bernadette; Cormican, Paul; Murray, Gerard; Kenny, David A; Waters, Sinead M; McGee, Mark; Kelly, Alan K; McCabe, Matthew S (Biomed Central, 2017-05-02)
    Background Bovine respiratory disease (BRD) is caused by growth of single or multiple species of pathogenic bacteria in lung tissue following stress and/or viral infection. Next generation sequencing of 16S ribosomal RNA gene PCR amplicons (NGS 16S amplicon analysis) is a powerful culture-independent open reference method that has recently been used to increase understanding of BRD-associated bacteria in the upper respiratory tract of BRD cattle. However, it has not yet been used to examine the microbiome of the bovine lower respiratory tract. The objective of this study was to use NGS 16S amplicon analysis to identify bacteria in post-mortem lung and lymph node tissue samples harvested from fatal BRD cases and clinically healthy animals. Cranial lobe and corresponding mediastinal lymph node post-mortem tissue samples were collected from calves diagnosed as BRD cases by veterinary laboratory pathologists and from clinically healthy calves. NGS 16S amplicon libraries, targeting the V3-V4 region of the bacterial 16S rRNA gene were prepared and sequenced on an Illumina MiSeq. Quantitative insights into microbial ecology (QIIME) was used to determine operational taxonomic units (OTUs) which corresponded to the 16S rRNA gene sequences. Results Leptotrichiaceae, Mycoplasma, Pasteurellaceae, and Fusobacterium were the most abundant OTUs identified in the lungs and lymph nodes of the calves which died from BRD. Leptotrichiaceae, Fusobacterium, Mycoplasma, Trueperella and Bacteroides had greater relative abundances in post-mortem lung samples collected from fatal cases of BRD in dairy calves, compared with clinically healthy calves without lung lesions. Leptotrichiaceae, Mycoplasma and Pasteurellaceae showed higher relative abundances in post-mortem lymph node samples collected from fatal cases of BRD in dairy calves, compared with clinically healthy calves without lung lesions. Two Leptotrichiaceae sequence contigs were subsequently assembled from bacterial DNA-enriched shotgun sequences. Conclusions The microbiomes of the cranial lung lobe and mediastinal lymph node from calves which died from BRD and from clinically healthy H-F calves have been characterised. Contigs corresponding to the abundant Leptotrichiaceae OTU were sequenced and found not to be identical to any known bacterial genus. This suggests that we have identified a novel bacterial species associated with BRD.
  • The altered gut microbiota in adults with cystic fibrosis

    Burke, D.G.; Fouhy, Fiona; Harrison, M. J; Rea, Mary C; Cotter, Paul D; O’Sullivan, Orla; Stanton, Catherine; Hill, C.; Shanahan, F.; Plant, B. J; Ross, R. Paul (Biomed Central, 09/03/2017)
    Background Cystic Fibrosis (CF) is an autosomal recessive disease that affects the function of a number of organs, principally the lungs, but also the gastrointestinal tract. The manifestations of cystic fibrosis transmembrane conductance regulator (CFTR) dysfunction in the gastrointestinal tract, as well as frequent antibiotic exposure, undoubtedly disrupts the gut microbiota. To analyse the effects of CF and its management on the microbiome, we compared the gut microbiota of 43 individuals with CF during a period of stability, to that of 69 non-CF controls using 454-pyrosequencing of the 16S rRNA gene. The impact of clinical parameters, including antibiotic therapy, on the results was also assessed. Results The CF-associated microbiome had reduced microbial diversity, an increase in Firmicutes and a reduction in Bacteroidetes compared to the non-CF controls. While the greatest number of differences in taxonomic abundances of the intestinal microbiota was observed between individuals with CF and the healthy controls, gut microbiota differences were also reported between people with CF when grouped by clinical parameters including % predicted FEV1 (measure of lung dysfunction) and the number of intravenous (IV) antibiotic courses in the previous 12 months. Notably, CF individuals presenting with severe lung dysfunction (% predicted FEV1 ≤ 40%) had significantly (p < 0.05) reduced gut microbiota diversity relative to those presenting with mild or moderate dysfunction. A significant negative correlation (−0.383, Simpson’s Diversity Index) was also observed between the number of IV antibiotic courses and gut microbiota diversity. Conclusions This is one of the largest single-centre studies on gut microbiota in stable adults with CF and demonstrates the significantly altered gut microbiota, including reduced microbial diversity seen in CF patients compared to healthy controls. The data show the impact that CF and it's management have on gut microbiota, presenting the opportunity to develop CF specific probiotics to minimise microbiota alterations.

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