ORCID Profile
0000-0002-8527-1445
Current Organisations
National University of Ireland
,
University College Cork
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Publisher: PeerJ
Date: 07-05-2018
DOI: 10.7717/PEERJ.4694
Abstract: Establishing a erse gut microbiota after birth is being increasingly recognised as important for preventing illnesses later in life. It is well established that bacterial ersity rapidly increases post-partum however, few studies have examined the infant gut virome hageome during this developmental period. We performed a metagenomic analysis of 20 infant faecal viromes at one year of age to determine whether spontaneous vaginal delivery (SVD) or caesarean section (CS) influenced viral composition. We find that birth mode results in distinctly different viral communities, with SVD infants having greater viral and bacteriophage ersity. We demonstrate that CrAssphage is acquired early in life, both in this cohort and two others, although no difference in birth mode is detected. A previous study has shown that bacterial OTU’s (operational taxonomic units) identified in the same infants could not discriminate between birth mode at 12 months of age. Therefore, our results indicate that vertical transmission of viral communities from mother to child may play a role in shaping the early life microbiome, and that birth mode should be considered when studying the early life gut virome.
Publisher: Elsevier BV
Date: 12-2014
Abstract: Probiotic bacteria have been associated with a reduction in cardiovascular disease risk, a leading cause of death and disability. The aim of this study was to assess the impact of dietary administration of exopolysaccharide-producing probiotic Lactobacillus cultures on lipid metabolism and gut microbiota in apolipoprotein E (apoE)-deficient mice. First, we examined lipid metabolism in response to dietary supplementation with recombinant β-glucan-producing Lactobacillus paracasei National Food Biotechnology Centre (NFBC) 338 expressing the glycosyltransferase (Gtf) gene from Pediococcus parvulus 2.6 (GTF), and naturally exopolysaccharide-producing Lactobacillus mucosae Dairy Product Culture Collection (DPC) 6426 (DPC 6426) compared with the non-β-glucan-producing isogenic control strain Lactobacillus paracasei NFBC 338 (PNZ) and placebo (15% wt:vol trehalose). Second, we examined the effects on the gut microbiota of dietary administration of DPC 6426 compared with placebo. Probiotic Lactobacillus strains at 1 × 10(9) colony-forming units/d per animal were administered to apoE(-/-) mice fed a high-fat (60% fat)/high-cholesterol (2% wt:wt) diet for 12 wk. At the end of the study, aortic plaque development and serum, liver, and fecal variables involved in lipid metabolism were analyzed, and culture-independent microbial analyses of cecal content were performed. Total cholesterol was reduced in serum (P < 0.001 ∼33-50%) and liver (P < 0.05 ∼30%) and serum triglyceride concentrations were reduced (P < 0.05 ∼15-25%) in mice supplemented with GTF or DPC 6426 compared with the PNZ or placebo group, respectively. In addition, dietary intervention with GTF led to increased amounts of fecal cholesterol excretion (P < 0.05) compared with all other groups. Compositional sequencing of the gut microbiota revealed a greater prevalence of Porphyromonadaceae (P = 0.001) and Prevotellaceae (P = 0.001) in the DPC 6426 group and lower proportions of Clostridiaceae (P < 0.05), Peptococcaceae (P < 0.001), and Staphylococcaceae (P < 0.01) compared with the placebo group. Ingestion of exopolysaccharide-producing lactobacilli resulted in seemingly favorable improvements in lipid metabolism, which were associated with changes in the gut microbiota of mice.
Publisher: Elsevier BV
Date: 10-2019
DOI: 10.1016/J.CHOM.2019.09.009
Abstract: The human gut contains a vast array of viruses, mostly bacteriophages. The majority remain uncharacterized, and their roles in shaping the gut microbiome and in impacting on human health remain poorly understood. We performed longitudinal metagenomic analysis of fecal viruses in healthy adults that reveal high temporal stability, in idual specificity, and correlation with the bacterial microbiome. Using a database-independent approach that uses most of the sequencing data, we uncovered the existence of a stable, numerically predominant in idual-specific persistent personal virome. Clustering of viral genomes and de novo taxonomic annotation identified several groups of crAss-like and Microviridae bacteriophages as the most stable colonizers of the human gut. CRISPR-based host prediction highlighted connections between these stable viral communities and highly predominant gut bacterial taxa such as Bacteroides, Prevotella, and Faecalibacterium. This study provides insights into the structure of the human gut virome and serves as an important baseline for hypothesis-driven research.
Publisher: Springer Science and Business Media LLC
Date: 28-01-2019
Publisher: Springer Science and Business Media LLC
Date: 13-06-2008
Abstract: The foodborne, gram-positive pathogen, Listeria monocytogenes , is capable of causing lethal infections in compromised in iduals. In the post genomic era of L. monocytogenes research, techniques are required to identify and validate genes involved in the pathogenicity and environmental biology of the organism. The aim here was to develop a widely applicable method to tag L. monocytogenes strains, with a particular emphasis on the development of multiple strain competitive index assays. We have constructed a new site-specific integrative vector, pIMC, based on pPL2, for the selection of L. monocytogenes from complex s les. The pIMC vector was further modified through the incorporation of IPTG inducible markers (antibiotic and phenotypic) to produce a suite of four vectors which allowed the discrimination of multiple strains from a single s le. We were able to perform murine infection studies with up to four EGDe isolates within a single mouse and showed that the tags did not impact upon growth rate or virulence. The system also allowed the identification of subtle differences in virulence between strains of L. monocytogenes commonly used in laboratory studies. This study has developed a competitive index assay that can be broadly applied to all L. monocytogenes strains. Improved statistical robustness of the data was observed, resulting in fewer mice being required for virulence assays. The competitive index assays provide a powerful method to analyse the virulence or fitness of L. monocytogenes in complex biological s les.
Publisher: Springer Science and Business Media LLC
Date: 19-08-2020
Publisher: American Society for Microbiology
Date: 04-2006
DOI: 10.1128/AEM.72.4.2876-2884.2006
Abstract: In this paper we describe construction of a luciferase-based vector, pPL2 lux , and use of this vector to study gene expression in Listeria monocytogenes . pPL2 lux is a derivative of the listerial integration vector pPL2 and harbors a synthetic luxABCDE operon encoding a fatty acid reductase complex (LuxCDE) involved in synthesis of the fatty aldehyde substrate for the bioluminescence reaction catalyzed by the LuxAB luciferase. We constructed pPL2 lux derivatives in which the secA and hlyA promoters were translationally fused to luxABCDE and integrated as a single copy into the chromosome of L. monocytogenes EGD-e. Growth experiments revealed that hlyA was expressed predominantly in the stationary phase in LB medium buffered at pH 7.4, whereas secA expression could be detected in the exponential growth phase. Moreover, the correlation between luciferase activity and transcription levels, as determined by reverse transcriptase PCR, was confirmed using conditions known to lead to repression and activation of hemolysin expression (addition of cellobiose and activated charcoal, respectively). Furthermore, hemolysin expression could be monitored in real time during invasion of an intact monolayer of C2Bbe1 (Caco-2-derived) cells. Finally, hemolysin expression could be detected in the livers, spleens, and kidneys of mice 3 days postinfection. These experiments clearly established the effectiveness of pPL2 lux as a quantitative reporter system for real-time, noninvasive evaluation of gene expression in L. monocytogenes .
Publisher: Springer Science and Business Media LLC
Date: 15-02-2021
DOI: 10.1038/S41538-021-00087-2
Abstract: Efficient and accurate identification of microorganisms throughout the food chain can potentially allow the identification of sources of contamination and the timely implementation of control measures. High throughput DNA sequencing represents a potential means through which microbial monitoring can be enhanced. While Illumina sequencing platforms are most typically used, newer portable platforms, such as the Oxford Nanopore Technologies (ONT) MinION, offer the potential for rapid analysis of food chain microbiomes. Initial assessment of the ability of rapid MinION-based sequencing to identify microbes within a simple mock metagenomic mixture is performed. Subsequently, we compare the performance of both ONT and Illumina sequencing for environmental monitoring of an active food processing facility. Overall, ONT MinION sequencing provides accurate classification to species level, comparable to Illumina-derived outputs. However, while the MinION-based approach provides a means of easy library preparations and portability, the high concentrations of DNA needed is a limiting factor.
Publisher: Springer Science and Business Media LLC
Date: 10-04-2018
Publisher: Wiley
Date: 18-02-2009
DOI: 10.1111/J.1365-2958.2008.06589.X
Abstract: The Listeria monocytogenes Agr peptide-sensing system has been analysed by creating a deletion mutant in agrD, the structural gene for the putative quorum-sensing peptide. The DeltaagrD mutant displayed significantly reduced biofilm formation, a defect which could be restored by genetic or physical complementation. A reduced invasion of Caco-2 intestinal epithelial cells was observed for the DeltaagrD mutant while phagocytosis by THP-1 macrophages was unaffected. Additionally, the level of internalin A (InlA) in the cell wall was decreased in the DeltaagrD mutant. Expression profiling of virulence genes (hlyA, actA, plcA, prfA and inlA) identified a finely tuned regulation which resulted in an impaired virulence response in the DeltaagrD mutant. The mutant is also significantly attenuated for virulence in mice, as revealed by bioluminescent in vivo imaging. On day 3 post infection, systemic dissemination to livers and spleens had occurred for the wild type, whereas the DeltaagrD mutant remained localized to the liver. Microarray analysis identified 126 and 670 genes as significantly regulated in exponential and stationary phase respectively. The results presented here suggest that peptide sensing plays an important role in the biology of L. monocytogenes, with relevant phenotypes in both the saprophytic and parasitic lifecycles.
Publisher: Cold Spring Harbor Laboratory
Date: 03-06-2019
DOI: 10.1101/657528
Abstract: The human gut contains a vast array of viruses, mostly bacteriophages. The majority remain uncharacterised and their roles in shaping the gut microbiome and in impacting on human health remain poorly understood. Here we performed a longitudinal focused metagenomic study of faecal bacteriophage populations in healthy adults. Our results reveal high temporal stability and in idual specificity of bacteriophage consortia which correlates with the bacterial microbiome. We report the existence of a stable, numerically predominant in idual-specific persistent personal virome. Clustering of bacteriophage genomes and de novo taxonomic annotation identified several groups of crAss-like and Microviridae bacteriophages as the most stable colonizers of the human gut. CRISPR-based host prediction highlighted connections between these stable viral communities and highly predominant gut bacterial taxa such as Bacteroides , Prevotella and Faecalibacterium . This study provides insights into the structure of the human gut virome and serves as an important baseline for hypothesis-driven research.
Publisher: Springer Science and Business Media LLC
Date: 15-04-2008
Publisher: American Society for Microbiology
Date: 26-03-2021
DOI: 10.1128/AEM.02459-20
Abstract: In general, disruptions to the gut microbiota are associated with multiple disorders in humans. The presence of high levels of Bifidobacterium spp. in the human gut is commonly considered beneficial.
Publisher: Royal Society of Chemistry (RSC)
Date: 2013
DOI: 10.1039/C3NP90040A
Publisher: American Society for Microbiology
Date: 07-2008
DOI: 10.1128/AEM.00314-08
Abstract: We describe the development of genetic tools for regulated gene expression, the introduction of chromosomal mutations, and improved plasmid transfer by electroporation in the food-borne pathogen Listeria monocytogenes . pIMK, a kanamycin-resistant, site-specific, integrative listeriophage vector was constructed and then modified for overexpression (pIMK2) or for isopropyl-β- d -thiogalactopyranoside (IPTG)-regulated expression (pIMK3 and pIMK4). The dynamic range of promoters was assessed by determining luciferase activity, P60 secretion, and internalin A-mediated invasion. These analyses demonstrated that pIMK4 and pIMK3 have a stringently controlled dynamic range of 540-fold. Stable gene overexpression was achieved with pIMK2, giving a range of expression for the three vectors of 1,350-fold. The lactococcal pORI280 system was optimized for the generation of chromosomal mutations and used to create five new prfA star mutants. The combination of pIMK4 and pORI280 allowed streamlined creation of “IPTG-dependent” mutants. This was exemplified by creation of a clean deletion mutant with deletion of the universally essential secA gene, and this mutant exhibited a rapid loss of viability upon withdrawal of IPTG. We also improved plasmid transfer by electroporation into three commonly used laboratory strains of L. monocytogenes . A 125-fold increase in transformation efficiency for EGDe compared with the widely used protocol of Park and Stewart (S. F. Park and G. S. Stewart, Gene 94:129-132, 1990) was observed. Maximal transformation efficiencies of 5.7 × 10 6 and 6.7 × 10 6 CFU per μg were achieved for EGDe and 10403S, respectively, with a replicating plasmid. An efficiency of 2 × 10 7 CFU per μg is the highest efficiency reported thus far for L. monocytogenes F2365.
Publisher: Springer Science and Business Media LLC
Date: 18-01-2019
Publisher: Elsevier BV
Date: 12-2019
DOI: 10.1016/J.CHOM.2019.10.009
Abstract: The human gut virome is thought to significantly impact the microbiome and human health. However, most virome analyses have been performed on a limited fraction of known viruses. Using whole-virome analysis on a published keystone inflammatory bowel disease (IBD) cohort and an in-house ulcerative colitis dataset, we shed light on the composition of the human gut virome in IBD beyond this identifiable minority. We observe IBD-specific changes to the virome and increased numbers of temperate phage sequences in in iduals with Crohn's disease. Unlike prior database-dependent methods, no changes in viral richness were observed. Among IBD subjects, the changes in virome composition reflected alterations in bacterial composition. Furthermore, incorporating both bacteriome and virome composition offered greater classification power between health and disease. This approach to analyzing whole virome across cohorts highlights significant IBD signals, which may be crucial for developing future biomarkers and therapeutics.
Publisher: American Society for Microbiology
Date: 11-2007
DOI: 10.1128/AEM.01394-07
Abstract: A novel vector has been constructed for the constitutive luminescent tagging of gram-negative bacteria by site-specific integration into the 16S locus of the bacterial chromosome. A number of gram-negative pathogens were successfully tagged using this vector, and the system was validated during murine infections of living animals.
Publisher: Elsevier BV
Date: 11-2018
DOI: 10.1016/J.CHOM.2018.10.002
Abstract: CrAssphages represent the most abundant virus in the human gut microbiota, but the lack of available genome sequences for comparison has kept them enigmatic. Recently, sequence-based classification of distantly related crAss-like phages from multiple environments was reported, leading to a proposed familial-level taxonomic group. Here, we assembled the metagenomic sequencing reads from 702 human fecal virome hageome s les and analyzed 99 complete circular crAss-like phage genomes and 150 contigs ≥70 kb. In silico comparative genomics and taxonomic analysis enabled a classification scheme of crAss-like phages from human fecal microbiomes into four candidate subfamilies composed of ten candidate genera. Laboratory analysis was performed on fecal s les from an in idual harboring seven distinct crAss-like phages. We achieved crAss-like phage propagation in ex vivo human fecal fermentations and visualized short-tailed podoviruses by electron microscopy. Mass spectrometry of a crAss-like phage capsid protein could be linked to metagenomic sequencing data, confirming crAss-like phage structural annotations.
Publisher: Microbiology Society
Date: 07-2012
Abstract: Isoprenoids may be synthesized via one of two pathways, the classical mevalonate pathway or the alternative 2-C-methyl-D-erythritol 4-phosphate (MEP) pathway. While the majority of bacteria utilize a single pathway for isoprenoid biosynthesis, Listeria monocytogenes is unusual in possessing the complete set of genes for both pathways. Here, we utilized new molecular tools to create precise gene deletions in selected genes encoding enzymes of both pathways, gcpE, lytB (encoding proteins in the MEP pathway) and hmgR (encoding a protein in the mevalonate pathway). We demonstrate that the hmgR gene can only be deleted when the growth medium is supplemented with exogenous mevalonate. Furthermore, full growth of the mutant in the absence of mevalonate was only possible when the intact hmgR gene was supplied in trans using an IPTG-inducible expression system. Murine competitive index assays performed via the oral and intraperitoneal routes of infection revealed that the mevalonate hmgR mutant could not be recovered from livers and spleens 3 days post-infection. We propose that HmgR in L. monocytogenes EGDe is involved in essential metabolic functions and that an intact MEP pathway is not capable of sustaining growth.
Publisher: Public Library of Science (PLoS)
Date: 12-05-2011
Publisher: American Physical Society (APS)
Date: 06-02-2015
Publisher: Springer Science and Business Media LLC
Date: 12-2010
Abstract: Internalin A (InlA) is a critical virulence factor which mediates the initiation of Listeria monocytogenes infection by the oral route in permissive hosts. The interaction of InlA with the host cell ligand E-cadherin efficiently stimulates L. monocytogenes entry into human enterocytes, but has only a limited interaction with murine cells. We have created a surface display library of randomly mutated InlA in a non-invasive heterologous host Lactococcus lactis in order to create and screen novel variants of this invasion factor. After sequential passage through a murine cell line (CT-26), multiple clones with enhanced invasion characteristics were identified. Competitive index experiments were conducted in mice using selected mutations introduced into L. monocytogenes EGD-e background. A novel single amino acid change was identified which enhanced virulence by the oral route in the murine model and will form the basis of further engineering approaches. As a control a previously described EGD-InlA m murinized strain was also re-created as part of this study with minor modifications and designated EGD-e InlA m * . The strain was created using a procedure that minimizes the likelihood of secondary mutations and incorporates Listeria -optimized codons encoding the altered amino acids. L. monocytogenes EGD-e InlA m * yielded consistently higher level murine infections by the oral route when compared to EGD-e, but did not display the two-fold increased invasion into a human cell line that was previously described for the EGD-InlA m strain. We have used both site-directed mutagenesis and directed evolution to create variants of InlA which may inform future structure-function analyses of this protein. During the course of the study we engineered a murinized strain of L. monocytogenes EGD-e which shows reproducibly higher infectivity in the intragastric murine infection model than the wild type, but does not display enhanced entry into human cells as previously observed. This murinized L. monocytogenes strain will provide a useful tool for the analysis of the gastrointestinal phase of listeriosis.
Publisher: Informa UK Limited
Date: 2021
Publisher: Elsevier BV
Date: 11-2014
DOI: 10.1016/J.JHEP.2014.06.025
Abstract: Despite the mortality associated with liver disease observed in patients with short bowel syndrome (SBS), mechanisms underlying the development of SBS-associated liver disease (SBS-ALD) are poorly understood. This study examines the impact of bacterially-mediated bile acid (BA) dysmetabolism on farnesoid X receptor (FXR) signalling pathways and clinical outcome in a piglet model of SBS-ALD. 4-week old piglets underwent 75% small bowel resection (SBR) or sham operation. Liver histology and hepatic inflammatory gene expression were examined. Abundance of BA biotransforming bacteria was determined and metabolomic studies detailed the alterations in BA composition of stool, portal serum and bile s les. Gene expression of intestinal and hepatic FXR target genes and small heterodimer partner (SHP) transrepression targets were assessed. Histological evidence of SBS-ALD included liver bile duct proliferation, hepatocyte ballooning and fibrosis. Inflammatory gene expression was increased. Microbiota changes included a 10-fold decrease in Clostridium and a two-fold decrease in Bacteroides in SBS-ALD piglets. BA composition was altered and reflected a primary BA dominant composition. Intestinal and hepatic regulation of BA synthesis was characterised by a blunted intestinal FXR activation response and a failure of SHP to repress key hepatic targets. We propose a pathological scenario in which microbial dysbiosis following SBR results in significant BA dysmetabolism and consequent outcomes including steatorrhoea, persistent diarrhoea and liver damage. Furthermore alterations in BA composition may have contributed to the observed disturbance in FXR-mediated signalling pathways. These findings provide an insight into the complex mechanisms mediating the development of liver disease in patients with SBS.
Publisher: American Society for Microbiology
Date: 05-2007
DOI: 10.1128/AEM.02940-06
Abstract: An improved system for luciferase tagging Listeria monocytogenes was developed by constructing a highly active, constitutive promoter. This construct gave 100-fold-higher activity in broth than any native promoter tested and allowed for imaging of lux -tagged L. monocytogenes in food products, during murine infections, and in tumor targeting studies.
Publisher: Cold Spring Harbor Laboratory
Date: 26-01-2019
DOI: 10.1101/527796
Abstract: Microbiomes are vast communities of microbes and viruses that populate all natural ecosystems. Viruses have been considered the most variable component of microbiomes, as supported by virome surveys and ex les of high genomic mosaicism. However, recent evidence suggests that the human gut virome is remarkably stable compared to other environments. Here we investigate the origin, evolution, and epidemiology of crAssphage, a widespread human gut virus. Through a global collaboratory, we obtained DNA sequences of crAssphage from over one-third of the world's countries, and showed that its phylogeography is locally clustered within countries, cities, and in iduals. We also found colinear crAssphage-like genomes in both Old-World and New-World primates, challenging genomic mosaicism and suggesting that the association of crAssphage with primates may be millions of years old. We conclude that crAssphage is a benign globetrotter virus that may have co-evolved with the human lineage and an integral part of the normal human gut virome.
Publisher: American Society for Microbiology
Date: 15-01-2020
DOI: 10.1128/JB.00639-19
Abstract: This study describes the characterization of nisin J, the first ex le of a natural nisin variant, produced by a human skin isolate of staphylococcal origin. Nisin J displays inhibitory activity against a wide range of bacterial targets, including MRSA. This work demonstrates the potential of human commensals as a source for novel antimicrobials that could form part of the solution to antibiotic resistance across a broad range of bacterial pathogens.
Publisher: American Society for Microbiology
Date: 02-2008
DOI: 10.1128/IAI.01132-07
Abstract: The ability of Pseudomonas aeruginosa to cause a broad range of infections in humans is due, at least in part, to its adaptability and its capacity to regulate the expression of key virulence genes in response to specific environmental conditions. Multiple two-component response regulators have been shown to facilitate rapid responses to these environmental conditions, including the coordinated expression of specific virulence determinants. RsmA is a posttranscriptional regulatory protein which controls the expression of a number of virulence-related genes with relevance for acute and chronic infections. Many membrane-bound sensors, including RetS, LadS, and GacS, are responsible for the reciprocal regulation of genes associated with acute infection and chronic persistence. In P. aeruginosa this is due to sensors influencing the expression of the regulatory RNA RsmZ, with subsequent effects on the level of free RsmA. While interactions between an rsmA mutant and human airway epithelial cells have been examined in vitro, the role of RsmA during infection in vivo has not been determined yet. Here the function of RsmA in both acute and chronic models of infection was examined. The results demonstrate that RsmA is involved in initial colonization and dissemination in a mouse model of acute pneumonia. Furthermore, while loss of RsmA results in reduced colonization during the initial stages of acute infection, the data show that mutation of rsmA ultimately favors chronic persistence and results in increased inflammation in the lungs of infected mice.
No related grants have been discovered for Colin Hill.