Publications by authors named "Martin Woodward"

131 Publications

Assessment of the anti-pathogenic effects of condensed tannin extracts using scanning electron microscopy.

Arch Microbiol 2021 Jan 5. Epub 2021 Jan 5.

School of Agriculture, Policy and Development, University of Reading, Reading, RG6 6AT, UK.

Two different types of condensed tannins (CTs), which were extracted and purified from tilia (Tilia L.) and black locust (Robinia pseudoacacia), were studied and tested against two kinds of bacteria, including Gram-negative and Gram-positive, avian pathogenic E. coli (APEC) and Staphylococcus epidermidis (S. epidermidis) respectively, by minimal bactericidal concentrations (MBCs) and scanning electron microscopy (SEM). Both CT extracts were significantly effective (p ≤ 0.05) at MBCs of 5-10 mg CT/ml against APEC (Gram-negative), and at 1.25-5 mg CT/ml on S. epidermidis (Gram-positive). This indicated that the CTs were more potent against the Gram-positive than the Gram-negative bacteria. Further, SEM revealed that CTs caused mainly morphological deformations of the bacterial cells and some conjoined cell growth.
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http://dx.doi.org/10.1007/s00203-020-02147-9DOI Listing
January 2021

Antimicrobial Activities of Ellagitannins against , , and .

Molecules 2020 Aug 14;25(16). Epub 2020 Aug 14.

Natural Chemistry Research Group, Department of Chemistry, University of Turku, FI-20014 Turku, Finland.

In this study, we tested the growth inhibition effect of 22 individual ellagitannins and of pentagalloylglucose on four bacterial species, i.e., and . All tested compounds showed antimicrobial effects against , and almost all against and . For , no or very weak growth inhibition was detected. The level of inhibition was the greatest for and the weakest for . For , the molecular size or flexibility of ellagitannins did not show a clear relationship with their antimicrobial activity, even though rugosins E and D and pentagalloylglucose with four or five free galloyl groups had a stronger growth inhibition effect than the other ellagitannins with glucopyranose cores but with less free galloyl groups. Additionally, our results with showed that the oligomeric linkage of ellagitannin might have an effect on its antimicrobial activity. For , the molecular size, but not the molecular flexibility, of ellagitannins seemed to be an important factor. For , both the molecular size and the flexibility of ellagitannin were important factors. In previous studies, corilagin was used as a model for ellagitannins, but our results showed that other ellagitannins are much more efficacious; therefore, the antimicrobial effects of ellagitannins could be more significant than previously thought.
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http://dx.doi.org/10.3390/molecules25163714DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7465317PMC
August 2020

Sub-lethal Concentrations of Phytochemicals (Carvacrol and Oregano) Select for Reduced Susceptibility Mutants of O23:H52.

Pol J Microbiol 2020 ;69:1-5

University of Reading , Reading, Berkshire , United Kingdom.

studies aimed at studying the mechanism of action of carvacrol and oregano as natural anti-bacterial agents to control multiple antibiotic-resistant avian pathogenic (APEC) strain O23:H52 isolated from chicken were performed. Derivatives with increased minimum inhibitory concentrations (MIC) to the phytochemicals were selected after growing () strain O23:H52 at sub-lethal concentrations of carvacrol and oregano for a period of 60 days. Whole-genome sequencing (WGS) of two derivatives revealed a missense mutation in and : the genes responsible for survival mechanisms and antibiotic resistance by efflux, respectively.

studies aimed at studying the mechanism of action of carvacrol and oregano as natural anti-bacterial agents to control multiple antibiotic-resistant avian pathogenic (APEC) strain O23:H52 isolated from chicken were performed. Derivatives with increased minimum inhibitory concentrations (MIC) to the phytochemicals were selected after growing () strain O23:H52 at sub-lethal concentrations of carvacrol and oregano for a period of 60 days. Whole-genome sequencing (WGS) of two derivatives revealed a missense mutation in and : the genes responsible for survival mechanisms and antibiotic resistance by efflux, respectively.
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http://dx.doi.org/10.33073/pjm-2020-003DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7256741PMC
December 2020

Thymol tolerance in Escherichia coli induces morphological, metabolic and genetic changes.

BMC Microbiol 2019 12 16;19(1):294. Epub 2019 Dec 16.

Department of Food and Nutrition Science, School of Chemistry, University of Reading, Reading, RG6 6AP, UK.

Background: Thymol is a phenolic compound used for its wide spectrum antimicrobial activity. There is a limited understanding of the antimicrobial mechanisms underlying thymol activity. To investigate this, E. coli strain JM109 was exposed to thymol at sub-lethal concentrations and after 16 rounds of exposure, isolates with a 2-fold increased minimal inhibitory concentration (MIC) were recovered (JM109-Thy). The phenotype was stable after multiple sub-cultures without thymol.

Results: Cell morphology studies by scanning electron microscopy (SEM) suggest that thymol renders bacterial cell membranes permeable and disrupts cellular integrity. H Nuclear magnetic resonance (NMR) data showed an increase in lactate and the lactic acid family amino acids in the wild type and JM109-Thy in the presence of thymol, indicating a shift from aerobic respiration to fermentation. Sequencing of JM109-Thy defined multiple mutations including a stop mutation in the acrR gene resulting in a truncation of the repressor of the AcrAB efflux pump. AcrAB is a multiprotein complex traversing the cytoplasmic and outer membrane, and is involved in antibiotic clearance.

Conclusions: Our data suggests that thymol tolerance in E. coli induces morphological, metabolic and genetic changes to adapt to thymol antimicrobial activity.
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http://dx.doi.org/10.1186/s12866-019-1663-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6915861PMC
December 2019

Antibiotic treatment triggers gut dysbiosis and modulates metabolism in a chicken model of gastro-intestinal infection.

BMC Vet Res 2019 Jan 25;15(1):37. Epub 2019 Jan 25.

Department of Food and Nutritional Sciences, University of Reading, Whiteknights, Reading, RG6 6AP, UK.

Background: Infection of the digestive track by gastro-intestinal pathogens results in the development of symptoms ranging from mild diarrhea to more severe clinical signs such as dysentery, severe dehydration and potentially death. Although, antibiotics are efficient to tackle infections, they also trigger dysbiosis that has been suggested to result in variation in weight gain in animal production systems.

Results: Here is the first study demonstrating the metabolic impact of infection by a gastro-intestinal pathogen (Brachyspira pilosicoli) and its resolution by antibiotic treatment (tiamulin) on the host (chicken) systemic metabolism and gut microbiota composition using high-resolution H nuclear magnetic resonance (NMR) spectroscopy and 16S rDNA next generation sequencing (NGS). Clear systemic metabolic markers of infections such as glycerol and betaine were identified. Weight loss in untreated animals was in part explained by the observation of a modification of systemic host energy metabolism characterized by the utilization of glycerol as a glucose precursor. However, antibiotic treatment triggered an increased VLDL/HDL ratio in plasma that may contribute to reducing weight loss observed in treated birds. All metabolic responses co-occurred with significant shift of the microbiota upon infection or antibiotic treatment.

Conclusion: This study indicates that infection and antibiotic treatment trigger dysbiosis that may impact host systemic energy metabolism and cause phenotypic and health modifications.
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http://dx.doi.org/10.1186/s12917-018-1761-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6347850PMC
January 2019

Characterization of a colistin-resistant Avian Pathogenic Escherichia coli ST69 isolate recovered from a broiler chicken in Germany.

J Med Microbiol 2019 Jan 26;68(1):111-114. Epub 2018 Nov 26.

1​Department of Pathology and Infectious Diseases, School of Veterinary Medicine, Faculty of Health & Medical Sciences, University of Surrey, Guildford, Surrey, GU2 7AL, UK.

In recent years, several plasmids harbouring genes encoding phosphoethanolamine transferases conferring colistin resistance have been described in multiple Enterobacteriaceae species. Avian Pathogenic E. coli (APEC) causes colibacillosis and is responsible for a considerable proportion of the disease burden in commercial poultry flocks, and may be linked to zoonotic infections in humans. Here, we describe the genotypic and phenotypic characteristics of a multidrug-resistant APEC ST69 isolate (APECA2), recovered in 2016 from a diseased broiler at post-mortem examination in Germany. The isolate was resistant to several antibiotics of human and veterinary importance, including colistin. The mcr-1 gene was detected on a mobile genetic element located on an IncHI2/ST4 plasmid, which was characterized using long-read Nanopore and short-read Illumina sequencing of purified plasmid. Isolate APECA2 displayed resistance to chicken serum and harbours numerous virulence genes. This study highlights the public health importance of enhanced antimicrobial resistance surveillance and strict antimicrobial stewardship in human and veterinary healthcare.
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http://dx.doi.org/10.1099/jmm.0.000882DOI Listing
January 2019

Heteromeric interactions regulate butyrophilin (BTN) and BTN-like molecules governing γδ T cell biology.

Proc Natl Acad Sci U S A 2018 01 16;115(5):1039-1044. Epub 2018 Jan 16.

Peter Gorer Department of Immunobiology, King's College London, London SE1 9RT, United Kingdom;

The long-held view that gamma delta (γδ) T cells in mice and humans are fundamentally dissimilar, as are γδ cells in blood and peripheral tissues, has been challenged by emerging evidence of the cells' regulation by butyrophilin (BTN) and butyrophilin-like (BTNL) molecules. Thus, murine and the related gene, , mediate T cell receptor (TCR)-dependent selection of murine intraepithelial γδ T cell repertoires in gut and skin, respectively; and are TCR-dependent regulators of human gut γδ cells; and is essential for TCR-dependent activation of human peripheral blood Vγ9Vδ2 T cells. However, some observations concerning BTN/Btnl molecules continue to question the extent of mechanistic conservation. In particular, murine and human gut γδ cell regulation depends on pairings of Btnl1 and Btnl6 and BTNL3 and BTNL8, respectively, whereas blood γδ cells are reported to be regulated by BTN3A1 independent of other BTNs. Addressing this paradox, we show that BTN3A2 regulates the subcellular localization of BTN3A1, including functionally important associations with the endoplasmic reticulum (ER), and is specifically required for optimal BTN3A1-mediated activation of Vγ9Vδ2 T cells. Evidence that BTNL3/BTNL8 and Btnl1/Btnl6 likewise associate with the ER reinforces the prospect of broadly conserved mechanisms underpinning the selection and activation of γδ cells in mice and humans, and in blood and extralymphoid sites.
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http://dx.doi.org/10.1073/pnas.1701237115DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5798315PMC
January 2018

Identification of temperature regulated factors of and their potential roles in virulence.

AIMS Microbiol 2017 7;3(4):885-898. Epub 2017 Nov 7.

Animal and Plant Health Agency, Woodham Lane, New Haw, Addlestone, Surrey KT15 3NB, UK.

is the major cause of bacterial gastroenteritis in man, while it is generally regarded as a commensal of the avian gut. Consumption and handling of contaminated poultry meat products are major risk factors for human infection. The body temperature in man (37 °C) and chickens (42 °C) differ markedly, and differential gene regulation and protein expression at different temperatures may in part explain the behaviour in the two hosts. We performed proteomics analyses with cells grown at 37 °C and 42 °C. Time-of-flight mass spectrometry (Q-Tof) analysis was carried out after samples were digested with the Filter-Aided Sample Preparation (FASP) method and peptides were fractionated by strong anion exchanges. Differentially regulated proteins were identified by Mascot and Scaffold analyses. Triple quadrupole (QQQ) mass spectrometer analysis confirmed that a total of 33 proteins were differentially regulated between 37 °C and 42 °C. Several upregulated proteins were selected for their corresponding gene knock-out mutants to be tested for their virulence in the model. To correlate with other tissue/animal models, the GADH mutant was selected for its reduced ability to colonize chickens. At 37 °C, the mutants of outer membrane protein Omp50 and Chaperone GroEL significantly increased virulence; while at 42 °C, the mutants of YceI, Omp50, and GADH reduced virulence against compared with the wild type strains. The results of current and previous studies indicate that GADH is a virulent factor in and a colonization factor in chickens. The workflow of this study may prove a new way to identify stress related virulent factors. The implications of these findings are discussed for pathogenesis in the model and other hosts.
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http://dx.doi.org/10.3934/microbiol.2017.4.885DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6604965PMC
November 2017

An Chicken Gut Model Demonstrates Transfer of a Multidrug Resistance Plasmid from to Commensal .

mBio 2017 07 18;8(4). Epub 2017 Jul 18.

Department of Bacteriology, Animal and Plant Health Agency, New Haw, Addlestone, Surrey, United Kingdom

The chicken gastrointestinal tract is richly populated by commensal bacteria that fulfill various beneficial roles for the host, including helping to resist colonization by pathogens. It can also facilitate the conjugative transfer of multidrug resistance (MDR) plasmids between commensal and pathogenic bacteria which is a significant public and animal health concern as it may affect our ability to treat bacterial infections. We used an chemostat system to approximate the chicken cecal microbiota, simulate colonization by an MDR pathogen, and examine the dynamics of transfer of its MDR plasmid harboring several genes, including the extended-spectrum beta-lactamase We also evaluated the impact of cefotaxime administration on plasmid transfer and microbial diversity. Bacterial community profiles obtained by culture-independent methods showed that inoculation resulted in no significant changes to bacterial community alpha diversity and beta diversity, whereas administration of cefotaxime caused significant alterations to both measures of diversity, which largely recovered. MDR plasmid transfer from to commensal was demonstrated by PCR and whole-genome sequencing of isolates purified from agar plates containing cefotaxime. Transfer occurred to seven sequence types at high rates, even in the absence of cefotaxime, with resistant strains isolated within 3 days. Our chemostat system provides a good representation of bacterial interactions, including antibiotic resistance transfer It can be used as an ethical and relatively inexpensive approach to model dissemination of antibiotic resistance within the gut of any animal or human and refine interventions that mitigate its spread before employing studies. The spread of antimicrobial resistance presents a grave threat to public health and animal health and is affecting our ability to respond to bacterial infections. Transfer of antimicrobial resistance via plasmid exchange is of particular concern as it enables unrelated bacteria to acquire resistance. The gastrointestinal tract is replete with bacteria and provides an environment for plasmid transfer between commensals and pathogens. Here we use the chicken gut microbiota as an exemplar to model the effects of bacterial infection, antibiotic administration, and plasmid transfer. We show that transfer of a multidrug-resistant plasmid from the zoonotic pathogen to commensal occurs at a high rate, even in the absence of antibiotic administration. Our work demonstrates that the gut model provides a powerful screening tool that can be used to assess and refine interventions that mitigate the spread of antibiotic resistance in the gut before undertaking animal studies.
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http://dx.doi.org/10.1128/mBio.00777-17DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5516254PMC
July 2017

Metabonomics-based analysis of Brachyspira pilosicoli's response to tiamulin reveals metabolic activity despite significant growth inhibition.

Anaerobe 2017 Jun 31;45:71-77. Epub 2017 Mar 31.

Department of Food and Nutritional Sciences, University of Reading, Whiteknights, Reading, UK6 6AP, UK. Electronic address:

Pathogenic anaerobes Brachyspira spp. are responsible for an increasing number of Intestinal Spirochaetosis (IS) cases in livestock against which few approved treatments are available. Tiamulin is used to treat swine dysentery caused by Brachyspira spp. and recently has been used to handle avian intestinal spirochaetosis (AIS). The therapeutic dose used in chickens requires further evaluation since cases of bacterial resistance to tiamulin have been reported. In this study, we evaluated the impact of tiamulin at varying concentrations on the metabolism of B. pilosicoli using a H-NMR-based metabonomics approach allowing the capture of the overall bacterial metabolic response to antibiotic treatment. Based on growth curve studies, tiamulin impacted bacterial growth even at very low concentration (0.008 μg/mL) although its metabolic activity was barely affected 72 h post exposure to antibiotic treatment. Only the highest dose of tiamulin tested (0.250 μg/mL) caused a major metabolic shift. Results showed that below this concentration, bacteria could maintain a normal metabolic trajectory despite significant growth inhibition by the antibiotic, which may contribute to disease reemergence post antibiotic treatment. Indeed, we confirmed that B. pilosicoli remained viable even after exposition to the highest antibiotic dose. This paper stresses the need to ensure new evaluation of bacterial viability post bacteriostatic exposure such as tiamulin to guarantee treatment efficacy and decrease antibiotic resistance development.
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http://dx.doi.org/10.1016/j.anaerobe.2017.03.018DOI Listing
June 2017

Colonisation of poultry by Salmonella Enteritidis S1400 is reduced by combined administration of Lactobacillus salivarius 59 and Enterococcus faecium PXN-33.

Vet Microbiol 2017 Feb 26;199:100-107. Epub 2016 Dec 26.

Department of Food and Nutrition, The University of Reading, Whiteknights Park, Reading, RG6 6AP, UK.

Salmonella Enteritidis remains a significant issue within the poultry industry and one potential solution is to use probiotic bacteria to prevent Salmonella colonisation through competitive exclusion (CE). We demonstrate that combined administration of Lactobacillus salivarius 59 and Enterococcus faecium PXN33 were effective competitive excluders of Salmonella Enteritidis S1400 in poultry. Two models were developed to evaluate the efficacy of probiotic where birds received Salmonella Enteritidis S1400 by a) oral gavage and b) sentinel bird to bird transmission. A statistically significant (p<0.001) 2 log reduction of Salmonella Enteritidis S1400 colonisation was observed in the ileum, caecum and colon at day 43 using combined administration of the two probiotic bacteria. However, no Salmonella Enteritidis S1400 colonisation reduction was observed when either probiotic was administered individually. In the sentinel bird model the combined probiotic administered at days 12 and 20 was more effective than one-off or double administrations at age 1 and 12days. In vitro cell free culture supernatant studies suggest the mechanism of Salmonella Enteritidis S1400 inhibition was due to a reduction in pH by the probiotic bacteria. Our current study provides further evidence that probiotics can significantly reduce pathogenic bacterial colonisation in poultry and that mixed preparation of probiotics provide superior performance when compared to individual bacterial preparations.
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http://dx.doi.org/10.1016/j.vetmic.2016.12.029DOI Listing
February 2017

Comparative genomics of European avian pathogenic E. Coli (APEC).

BMC Genomics 2016 11 22;17(1):960. Epub 2016 Nov 22.

Department of Pathology and Infectious Diseases, School of Veterinary Medicine, Faculty of Health and Medical Sciences, University of Surrey, Guildford, GU2 7AL, UK.

Background: Avian pathogenic Escherichia coli (APEC) causes colibacillosis, which results in significant economic losses to the poultry industry worldwide. However, the diversity between isolates remains poorly understood. Here, a total of 272 APEC isolates collected from the United Kingdom (UK), Italy and Germany were characterised using multiplex polymerase chain reactions (PCRs) targeting 22 equally weighted factors covering virulence genes, R-type and phylogroup. Following these analysis, 95 of the selected strains were further analysed using Whole Genome Sequencing (WGS).

Results: The most prevalent phylogroups were B2 (47%) and A1 (22%), although there were national differences with Germany presenting group B2 (35.3%), Italy presenting group A1 (53.3%) and UK presenting group B2 (56.1%) as the most prevalent. R-type R1 was the most frequent type (55%) among APEC, but multiple R-types were also frequent (26.8%). Following compilation of all the PCR data which covered a total of 15 virulence genes, it was possible to build a similarity tree using each PCR result unweighted to produce 9 distinct groups. The average number of virulence genes was 6-8 per isolate, but no positive association was found between phylogroup and number or type of virulence genes. A total of 95 isolates representing each of these 9 groupings were genome sequenced and analysed for in silico serotype, Multilocus Sequence Typing (MLST), and antimicrobial resistance (AMR). The UK isolates showed the greatest variability in terms of serotype and MLST compared with German and Italian isolates, whereas the lowest prevalence of AMR was found for German isolates. Similarity trees were compiled using sequencing data and notably single nucleotide polymorphism data generated ten distinct geno-groups. The frequency of geno-groups across Europe comprised 26.3% belonging to Group 8 representing serogroups O2, O4, O18 and MLST types ST95, ST140, ST141, ST428, ST1618 and others, 18.9% belonging to Group 1 (serogroups O78 and MLST types ST23, ST2230), 15.8% belonging to Group 10 (serogroups O8, O45, O91, O125ab and variable MLST types), 14.7% belonging to Group 7 (serogroups O4, O24, O35, O53, O161 and MLST type ST117) and 13.7% belonging to Group 9 (serogroups O1, O16, O181 and others and MLST types ST10, ST48 and others). The other groups (2, 3, 4, 5 and 6) each contained relatively few strains. However, for some of the genogroups (e.g. groups 6 and 7) partial overlap with SNPs grouping and PCR grouping (matching PCR groups 8 (13 isolates on 22) and 1 (14 isolates on 16) were observable). However, it was not possible to obtain a clear correlation between genogroups and unweighted PCR groupings. This may be due to the genome plasticity of E. coli that enables strains to carry the same virulence factors even if the overall genotype is substantially different.

Conclusions: The conclusion to be drawn from the lack of correlations is that firstly, APEC are very diverse and secondly, it is not possible to rely on any one or more basic molecular or phenotypic tests to define APEC with clarity, reaffirming the need for whole genome analysis approaches which we describe here. This study highlights the presence of previously unreported serotypes and MLSTs for APEC in Europe. Moreover, it is a first step on a cautious reconsideration of the merits of classical identification criteria such as R typing, phylogrouping and serotyping.
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http://dx.doi.org/10.1186/s12864-016-3289-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5120500PMC
November 2016

NMR-based metabolic characterization of chicken tissues and biofluids: a model for avian research.

Metabolomics 2016 15;12(10):157. Epub 2016 Sep 15.

Department of Food and Nutritional Sciences, University of Reading, Whiteknights, Reading, RG6 6AP UK.

Introduction: Poultry is one of the most consumed meat in the world and its related industry is always looking for ways to improve animal welfare and productivity. It is therefore essential to understand the metabolic response of the chicken to new feed formulas, various supplements, infections and treatments.

Objectives: As a basis for future research investigating the impact of diet and infections on chicken's metabolism, we established a high-resolution proton nuclear magnetic resonance (NMR)-based metabolic atlas of the healthy chicken ().

Methods: Metabolic extractions were performed prior to H-NMR and 2D NMR spectra acquisition on twelve biological matrices: liver, kidney, spleen, plasma, egg yolk and white, colon, caecum, faecal water, ileum, pectoral muscle and brain of 6 chickens. Metabolic profiles were then exhaustively characterized.

Results: Nearly 80 metabolites were identified. A cross-comparison of these matrices was performed to determine metabolic variations between and within each section and highlighted that only eight core metabolites were systematically found in every matrice.

Conclusion: This work constitutes a database for future NMR-based metabolomic investigations in relation to avian production and health.
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http://dx.doi.org/10.1007/s11306-016-1105-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5025519PMC
September 2016

AIRE-Deficient Patients Harbor Unique High-Affinity Disease-Ameliorating Autoantibodies.

Cell 2016 Jul 14;166(3):582-595. Epub 2016 Jul 14.

Peter Gorer Department of Immunobiology, King's College, London SE19RT, UK. Electronic address:

APS1/APECED patients are defined by defects in the autoimmune regulator (AIRE) that mediates central T cell tolerance to many self-antigens. AIRE deficiency also affects B cell tolerance, but this is incompletely understood. Here we show that most APS1/APECED patients displayed B cell autoreactivity toward unique sets of approximately 100 self-proteins. Thereby, autoantibodies from 81 patients collectively detected many thousands of human proteins. The loss of B cell tolerance seemingly occurred during antibody affinity maturation, an obligatorily T cell-dependent step. Consistent with this, many APS1/APECED patients harbored extremely high-affinity, neutralizing autoantibodies, particularly against specific cytokines. Such antibodies were biologically active in vitro and in vivo, and those neutralizing type I interferons (IFNs) showed a striking inverse correlation with type I diabetes, not shown by other anti-cytokine antibodies. Thus, naturally occurring human autoantibodies may actively limit disease and be of therapeutic utility.
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http://dx.doi.org/10.1016/j.cell.2016.06.024DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4967814PMC
July 2016

Curing vector for IncI1 plasmids and its use to provide evidence for a metabolic burden of IncI1 CTX-M-1 plasmid pIFM3791 on Klebsiella pneumoniae.

J Med Microbiol 2016 Jul 9;65(7):611-618. Epub 2016 May 9.

Department of Food and Nutritional Sciences, University of Reading, Whiteknights Park, Reading RG6 5AP, UK.

Using a sequence-based approach we previously identified an IncI1 CTX-M-1 plasmid, pIFM3791, on a single pig farm in the UK that was harboured by Klebsiella pneumoniae, Escherichia coli and Salmonella enterica serotype 4,5,12:i:-. To test the hypothesis that the plasmid had spread rapidly into these differing host bacteria we wished to assess whether the plasmid conferred a fitness advantage. To do this an IncI1 curing vector was constructed and used to displace the IncI1 CTX-M-1 plasmids from K. pneumoniae strain B3791 and several other unrelated IncI1-harbouring strains indicating the potential wider application of the curing vector. The IncI1 CTX-M-1 plasmid was reintroduced by conjugation into the cured K. pneumoniae strain and also a naturally IncI1 plasmid free S. enterica serotype 4,5,12:i:-, S348/11. Original, cured and complemented strains were tested for metabolic competence using Biolog technology and in competitive growth, association to mammalian cells and biofilm formation experiments. The plasmid-cured K. pneumoniae strain grew more rapidly than either the original plasmid-carrying strain or plasmid-complemented strains in competition experiments. Additionally, the plasmid-cured strain was significantly better at respiring with l-sorbose as a carbon source and putrescine, γ-amino-n-butyric acid, l-alanine and l-proline as nitrogen sources. By contrast, no differences in phenotype were found when comparing plasmid-harbouring and plasmid-free S. enterica S348/11. In conclusion, the IncI1 curing vector successfully displaced multiple IncI plasmids. The IncI1 CTX-M1 plasmid conferred a growth disadvantage upon K. pneumoniae, possibly by imposing a metabolic burden, the mechanism of which remains to be determined.
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http://dx.doi.org/10.1099/jmm.0.000271DOI Listing
July 2016

Novel European free-living, non-diazotrophic Bradyrhizobium isolates from contrasting soils that lack nodulation and nitrogen fixation genes - a genome comparison.

Sci Rep 2016 05 10;6:25858. Epub 2016 May 10.

Department of AgroEcology, Rothamsted Research, Harpenden, AL5 2JQ, UK.

The slow-growing genus Bradyrhizobium is biologically important in soils, with different representatives found to perform a range of biochemical functions including photosynthesis, induction of root nodules and symbiotic nitrogen fixation and denitrification. Consequently, the role of the genus in soil ecology and biogeochemical transformations is of agricultural and environmental significance. Some isolates of Bradyrhizobium have been shown to be non-symbiotic and do not possess the ability to form nodules. Here we present the genome and gene annotations of two such free-living Bradyrhizobium isolates, named G22 and BF49, from soils with differing long-term management regimes (grassland and bare fallow respectively) in addition to carbon metabolism analysis. These Bradyrhizobium isolates are the first to be isolated and sequenced from European soil and are the first free-living Bradyrhizobium isolates, lacking both nodulation and nitrogen fixation genes, to have their genomes sequenced and assembled from cultured samples. The G22 and BF49 genomes are distinctly different with respect to size and number of genes; the grassland isolate also contains a plasmid. There are also a number of functional differences between these isolates and other published genomes, suggesting that this ubiquitous genus is extremely heterogeneous and has roles within the community not including symbiotic nitrogen fixation.
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http://dx.doi.org/10.1038/srep25858DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4861915PMC
May 2016

Characterization of a Putative Receptor Binding Surface on Skint-1, a Critical Determinant of Dendritic Epidermal T Cell Selection.

J Biol Chem 2016 Apr 25;291(17):9310-21. Epub 2016 Feb 25.

From the Cancer Immunology and Immunotherapy Centre, Institute of Immunology and Immunotherapy, University of Birmingham, Edgbaston, Birmingham B15 2TT,

Dendritic epidermal T cells (DETC) form a skin-resident γδ T cell population that makes key contributions to cutaneous immune stress surveillance, including non-redundant contributions to protection from cutaneous carcinogens. How DETC become uniquely associated with the epidermis was in large part solved by the identification of Skint-1, the prototypic member of a novel B7-related multigene family. Expressed only by thymic epithelial cells and epidermal keratinocytes, Skint-1 drives specifically the development of DETC progenitors, making it the first clear candidate for a selecting ligand for non-MHC/CD1-restricted T cells. However, the molecular mechanisms underpinning Skint-1 activity are unresolved. Here, we provide evidence that DETC selection requires Skint-1 expression on the surface of thymic epithelial cells, and depends upon specific residues on the CDR3-like loop within the membrane-distal variable domain of Skint-1 (Skint-1 DV). Nuclear magnetic resonance of Skint-1 DV revealed a core tertiary structure conserved across the Skint family, but a highly distinct surface charge distribution, possibly explaining its unique function. Crucially, the CDR3-like loop formed an electrostatically distinct surface, featuring key charged and hydrophobic solvent-exposed residues, at the membrane-distal tip of DV. These results provide the first structural insights into the Skint family, identifying a putative receptor binding surface that directly implicates Skint-1 in receptor-ligand interactions crucial for DETC selection.
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http://dx.doi.org/10.1074/jbc.M116.722066DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4861494PMC
April 2016

Population structure and associated phenotypes of Salmonella enterica serovars Derby and Mbandaka overlap with host range.

BMC Microbiol 2016 Feb 4;16:15. Epub 2016 Feb 4.

Department of Food and Nutritional Sciences, Reading University, Whiteknights, Reading, RG6 6AP, UK.

Background: The Salmonella enterica serovar Derby is frequently isolated from pigs and turkeys whereas serovar Mbandaka is frequently isolated from cattle, chickens and animal feed in the UK. Through comparative genomics, phenomics and mutant construction we previously suggested possible mechanistic reasons why these serovars demonstrate apparently distinct host ranges. Here, we investigate the genetic and phenotypic diversity of these two serovars in the UK. We produce a phylogenetic reconstruction and perform several biochemical assays on isolates of S. Derby and S. Mbandaka acquired from sites across the UK between the years 2000 and 2010.

Results: We show that UK isolates of S. Mbandaka comprise of one clonal lineage which is adapted to proficient utilisation of metabolites found in soya beans under ambient conditions. We also show that this clonal lineage forms a biofilm at 25 °C, suggesting that this serovar maybe well adapted to survival ex vivo, growing in animal feed. Conversely, we show that S. Derby is made of two distinct lineages, L1 and L2. These lineages differ genotypically and phenotypically, being divided by the presence and absence of SPI-23 and the ability to more proficiently invade porcine jejunum derived cell line IPEC-J2.

Conclusion: The results of this study lend support to the hypothesis that the differences in host ranges of S. Derby and S. Mbandaka are adaptations to pathogenesis, environmental persistence, as well as utilisation of metabolites abundant in their respective host environments.
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http://dx.doi.org/10.1186/s12866-016-0628-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4743429PMC
February 2016

Diversity of STs, plasmids and ESBL genes among Escherichia coli from humans, animals and food in Germany, the Netherlands and the UK.

J Antimicrob Chemother 2016 May 23;71(5):1178-82. Epub 2016 Jan 23.

Public Health England, London, UK.

Objectives: This study aimed to compare ESBL-producing Escherichia coli causing infections in humans with infecting or commensal isolates from animals and isolates from food of animal origin in terms of the strain types, the ESBL gene present and the plasmids that carry the respective ESBL genes.

Methods: A collection of 353 ESBL-positive E. coli isolates from the UK, the Netherlands and Germany were studied by MLST and ESBL genes were identified. Characterization of ESBL gene-carrying plasmids was performed using PCR-based replicon typing. Moreover, IncI1-Iγ and IncN plasmids were characterized by plasmid MLST.

Results: The ESBL-producing E. coli represented 158 different STs with ST131, ST10 and ST88 being the most common. Overall, blaCTX-M-1 was the most frequently detected ESBL gene, followed by blaCTX-M-15, which was the most common ESBL gene in the human isolates. The most common plasmid replicon type overall was IncI1-Iγ followed by multiple IncF replicons.

Conclusions: ESBL genes were present in a wide variety of E. coli STs. IncI1-Iγ plasmids that carried the blaCTX-M-1 gene were widely disseminated amongst STs in isolates from animals and humans, whereas other plasmids and STs appeared to be more restricted to isolates from specific hosts.
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http://dx.doi.org/10.1093/jac/dkv485DOI Listing
May 2016

Drinking water application of Denagard® Tiamulin for control of Brachyspira pilosicoli infection of laying poultry.

Res Vet Sci 2015 Dec 25;103:87-95. Epub 2015 Sep 25.

Animal Health Agency, Woodham Lane, New Haw, Addlestone, Surrey KT15 3NB, UK; School of Veterinary Medicine, Faculty of Health and Medical Sciences, University of Surrey, Guildford, Surrey GU2 7XH, UK.

Avian intestinal spirochaetosis (AIS) caused by Brachyspira spp., and notably Brachyspira pilosicoli, is common in layer flocks and reportedly of increasing incidence in broilers and broiler breeders. Disease manifests as diarrhoea, increased feed consumption, reduced growth rates and occasional mortality in broilers and these signs are shown in layers also associated with a delayed onset of lay, reduced egg weights, faecal staining of eggshells and non-productive ovaries. Treatment with Denagard® Tiamulin has been used to protect against B. pilosicoli colonisation, persistence and clinical presentation of AIS in commercial layers, but to date there has been no definitive study validating efficacy. Here, we used a poultry model of B. pilosicoli infection of layers to compare the impact of three doses of Denagard® Tiamulin. Four groups of thirty 17 week old commercial pre-lay birds were all challenged with B. pilosicoli strain B2904 with three oral doses two days apart. All birds were colonised within 2 days after the final oral challenge and mild onset of clinical signs were observed thereafter. A fifth group that was unchallenged and untreated was also included for comparison as healthy birds. Five days after the final oral Brachypira challenge three groups were given Denagard® Tiamulin in drinking water made up following the manufacturer's recommendations with doses verified as 58.7 ppm, 113 ppm and 225 ppm. Weight gain body condition and the level of diarrhoea of birds infected with B. pilosicoli were improved and shedding of the organism reduced significantly (p=0.001) following treatment with Denagard® Tiamulin irrespective of dose given. The level and duration of colonisation of organs of birds infected with B. pilosicoli was also reduced. Confirming previous findings we showed that the ileum, caeca, colon, and both liver and spleen were colonised and here we demonstrated that treatment with Denagard® Tiamulin resulted in significant reduction in the numbers of Brachyspira found in each of these sites and dramatic reduction in faecal shedding (p<0.001) to approaching zero as assessed by culture of cloacal swabs. Although the number of eggs produced per bird and the level of eggshell staining appeared unaffected, egg weights of treated birds were greater than those of untreated birds for a period of approximately two weeks following treatment. These data conclusively demonstrate the effectiveness of Denagard® Tiamulin in reducing B. pilosicoli infection in laying hens.
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http://dx.doi.org/10.1016/j.rvsc.2015.09.015DOI Listing
December 2015

Brachyspira pilosicoli-induced avian intestinal spirochaetosis.

Microb Ecol Health Dis 2015 15;26:28853. Epub 2015 Dec 15.

Department of Food and Nutritional Sciences, University of Reading, Reading, UK;

Avian intestinal spirochaetosis (AIS) is a common disease occurring in poultry that can be caused by Brachyspira pilosicoli, a Gram-negative bacterium of the order Spirochaetes. During AIS, this opportunistic pathogen colonises the lower gastrointestinal (GI) tract of poultry (principally, the ileum, caeca, and colon), which can cause symptoms such as diarrhoea, reduced growth rate, and reduced egg production and quality. Due to the large increase of bacterial resistance to antibiotic treatment, the European Union banned in 2006 the prophylactic use of antibiotics as growth promoters in livestock. Consequently, the number of outbreaks of AIS has dramatically increased in the UK resulting in significant economic losses. This review summarises the current knowledge about AIS infection caused by B. pilosicoli and discusses various treatments and prevention strategies to control AIS.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4683989PMC
http://dx.doi.org/10.3402/mehd.v26.28853DOI Listing
December 2015

Temperature and oxygen dependent metabolite utilization by Salmonella enterica serovars Derby and Mbandaka.

PLoS One 2015 23;10(3):e0120450. Epub 2015 Mar 23.

School of Biological Sciences, Royal Holloway University of London, Egham, Surrey, TW20 0EX, United Kingdom.

Salmonella enterica is a zoonotic pathogen of clinical and veterinary significance, with over 2500 serovars. In previous work we compared two serovars displaying host associations inferred from isolation statistics. Here, to validate genome sequence data and to expand on the role of environmental metabolite constitution in host range determination we use a phenotypic microarray approach to assess the ability of these serovars to metabolise ~500 substrates at 25°C with oxygen (aerobic conditions) to represent the ex vivo environment and at 37°C with and without oxygen (aerobic/anaerobic conditions) to represent the in vivo environment. A total of 26 substrates elicited a significant difference in the rate of metabolism of which only one, D-galactonic acid-g-lactone, could be explained by the presence (S. Mbandaka) or the absence (S. Derby) of metabolic genes. We find that S. Mbandaka respires more efficiently at ambient temperatures and under aerobic conditions on 18 substrates including: glucosominic acid, saccharic acid, trehalose, fumaric acid, maltotriose, N-acetyl-D-glucosamine, N-acetyl-beta-D-mannosamine, fucose, L-serine and dihydroxy-acetone; whereas S. Derby is more metabolically competent anaerobically at 37°C for dipeptides, glutamine-glutamine, alanine-lysine, asparagine-glutamine and nitrogen sources glycine and nitrite. We conclude that the specific phenotype cannot be reliably predicted from the presence of metabolic genes directly relating to the metabolic pathways under study.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0120450PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4370486PMC
December 2015

SPI-23 of S. Derby: role in adherence and invasion of porcine tissues.

PLoS One 2014 19;9(9):e107857. Epub 2014 Sep 19.

Department of Food and Nutritional Sciences, University of Reading, Reading, United Kingdom.

Salmonella enterica serovars Derby and Mbandaka are isolated from different groups of livestock species in the UK. S. Derby is predominantly isolated from pigs and turkeys and S. Mbandaka is predominantly isolated from cattle and chickens. Alignment of the genome sequences of two isolates of each serovar led to the discovery of a new putative Salmonella pathogenicity island, SPI-23, in the chromosome sequence of S. Derby isolates. SPI-23 is 37 kb in length and contains 42 ORFs, ten of which are putative type III effector proteins. In this study we use porcine jejunum derived cell line IPEC-J2 and in vitro organ culture of porcine jejunum and colon, to characterise the association and invasion rates of S. Derby and S. Mbandaka, and tissue tropism of S. Derby respectively. We show that S. Derby invades and associates to an IPEC-J2 monolayer in significantly greater numbers than S. Mbandaka, and that S. Derby preferentially attaches to porcine jejunum over colon explants. We also show that nine genes across SPI-23 are up-regulated to a greater degree in the jejunum compared to the colon explants. Furthermore, we constructed a mutant of the highly up-regulated, pilV-like gene, potR, and find that it produces an excess of surface pili compared to the parent strain which form a strong agglutinating phenotype interfering with association and invasion of IPEC-J2 monolayers. We suggest that potR may play a role in tissue tropism.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0107857PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4169617PMC
November 2015

Identification of potential drug targets in Salmonella enterica sv. Typhimurium using metabolic modelling and experimental validation.

Microbiology (Reading) 2014 Jun 28;160(Pt 6):1252-1266. Epub 2014 Apr 28.

Department of Medical and Biological Sciences, Oxford Brookes University, Gipsy Lane, Headington, Oxford OX3 OBP, UK.

Salmonella enterica sv. Typhimurium is an established model organism for Gram-negative, intracellular pathogens. Owing to the rapid spread of resistance to antibiotics among this group of pathogens, new approaches to identify suitable target proteins are required. Based on the genome sequence of S. Typhimurium and associated databases, a genome-scale metabolic model was constructed. Output was based on an experimental determination of the biomass of Salmonella when growing in glucose minimal medium. Linear programming was used to simulate variations in the energy demand while growing in glucose minimal medium. By grouping reactions with similar flux responses, a subnetwork of 34 reactions responding to this variation was identified (the catabolic core). This network was used to identify sets of one and two reactions that when removed from the genome-scale model interfered with energy and biomass generation. Eleven such sets were found to be essential for the production of biomass precursors. Experimental investigation of seven of these showed that knockouts of the associated genes resulted in attenuated growth for four pairs of reactions, whilst three single reactions were shown to be essential for growth.
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http://dx.doi.org/10.1099/mic.0.076091-0DOI Listing
June 2014

Sequence analysis of a CTX-M-1 IncI1 plasmid found in Salmonella 4,5,12:i:-, Escherichia coli and Klebsiella pneumoniae on a UK pig farm.

J Antimicrob Chemother 2014 Aug 11;69(8):2098-101. Epub 2014 Apr 11.

Department of Food and Nutritional Sciences, The University of Reading, Whiteknights Park, Reading RG6 5AP, UK.

Objectives: In 2009, CTX-M Enterobacteriaceae and Salmonella isolates were recovered from a UK pig farm, prompting studies into the dissemination of the resistance and to establish any relationships between the isolates.

Methods: PFGE was used to elucidate clonal relationships between isolates whilst plasmid profiling, restriction analysis, sequencing and PCR were used to characterize the CTX-M-harbouring plasmids.

Results: Escherichia coli, Klebsiella pneumoniae and Salmonella 4,5,12:i:- and Bovismorbificans resistant to cefotaxime (n = 65) were recovered and 63 were shown by PCR to harbour a group 1 CTX-M gene. The harbouring hosts were diverse, but the group 1 CTX-M plasmids were common. Three sequenced CTX-M plasmids from E. coli, K. pneumoniae and Salmonella enterica serotype 4,5,12:i:- were identical except for seven mutations and highly similar to IncI1 plasmid ColIb-P9. Two antimicrobial resistance regions were identified: one inserted upstream of yacABC harbouring ISCR2 transposases, sul2 and floR; and the other inserted within shfB of the pilV shufflon harbouring the ISEcp1 transposase followed by blaCTX-M-1.

Conclusions: These data suggest that an ST108 IncI1 plasmid encoding a blaCTX-M-1 gene had disseminated across multiple genera on this farm, an example of horizontal gene transfer of the blaCTX-M-1 gene.
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http://dx.doi.org/10.1093/jac/dku098DOI Listing
August 2014

Array based detection of antibiotic resistance genes in Gram negative bacteria isolated from retail poultry meat in the UK and Ireland.

Int J Food Microbiol 2014 Jun 27;179:24-32. Epub 2014 Mar 27.

Northern Ireland Centre for Food and Health, School of Biomedical Sciences, University of Ulster, Cromore Road, Coleraine, Co. Londonderry BT52 1SA, United Kingdom. Electronic address:

The use of antibiotics in birds and animals intended for human consumption within the European Union (EU) and elsewhere has been subject to regulation prohibiting the use of antimicrobials as growth promoters and the use of last resort antibiotics in an attempt to reduce the spread of multi-resistant Gram negative bacteria. Given the inexorable spread of antibiotic resistance there is an increasing need for improved monitoring of our food. Using selective media, Gram negative bacteria were isolated from retail chicken of UK-Intensively reared (n=27), Irish-Intensively reared (n=19) and UK-Free range (n=30) origin and subjected to an oligonucleotide based array system for the detection of 47 clinically relevant antibiotic resistance genes (ARGs) and two integrase genes. High incidences of β-lactamase genes were noted in all sample types, acc (67%), cmy (80%), fox (55%) and tem (40%) while chloramphenicol resistant determinants were detected in bacteria from the UK poultry portions and were absent in bacteria from the Irish samples. Denaturing Gradient Gel Electrophoresis (DGGE) was used to qualitatively analyse the Gram negative population in the samples and showed the expected diversity based on band stabbing and DNA sequencing. The array system proved to be a quick method for the detection of antibiotic resistance gene (ARG) burden within a mixed Gram negative bacterial population.
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http://dx.doi.org/10.1016/j.ijfoodmicro.2014.03.019DOI Listing
June 2014

Brachyspira and its role in avian intestinal spirochaetosis.

Vet Microbiol 2014 Jan 28;168(2-4):245-60. Epub 2013 Nov 28.

Department of Food and Nutritional Sciences, University of Reading, Reading, Berkshire RG6 6AP, UK.

The fastidious, anaerobic spirochaete Brachyspira is capable of causing enteric disease in avian, porcine and human hosts, amongst others, with a potential for zoonotic transmission. Avian intestinal spirochaetosis (AIS), the resulting disease from colonisation of the caeca and colon of poultry by Brachyspira leads to production losses, with an estimated annual cost of circa £ 18 million to the commercial layer industry in the United Kingdom. Of seven known and several proposed species of Brachyspira, three are currently considered pathogenic to poultry; B. alvinipulli, B. intermedia and B. pilosicoli. Currently, AIS is primarily prevented by strict biosecurity controls and is treated using antimicrobials, including tiamulin. Other treatment strategies have been explored, including vaccination and probiotics, but such developments have been hindered by a limited understanding of the pathobiology of Brachyspira. A lack of knowledge of the metabolic capabilities and little genomic information for Brachyspira has resulted in a limited understanding of the pathobiology. In addition to an emergence of antibiotic resistance amongst Brachyspira, bans on the prophylactic use of antimicrobials in livestock are driving an urgent requirement for alternative treatment strategies for Brachyspira-related diseases, such as AIS. Advances in the molecular biology and genomics of Brachyspira heralds the potential for the development of tools for genetic manipulation to gain an improved understanding of the pathogenesis of Brachyspira.
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http://dx.doi.org/10.1016/j.vetmic.2013.11.019DOI Listing
January 2014

Metaproteomics analysis reveals the adaptation process for the chicken gut microbiota.

Appl Environ Microbiol 2014 Jan 8;80(2):478-85. Epub 2013 Nov 8.

Veterinary Laboratories Agency, Addlestone, Surrey, United Kingdom.

The animal gastrointestinal tract houses a large microbial community, the gut microbiota, that confers many benefits to its host, such as protection from pathogens and provision of essential metabolites. Metagenomic approaches have defined the chicken fecal microbiota in other studies, but here, we wished to assess the correlation between the metagenome and the bacterial proteome in order to better understand the healthy chicken gut microbiota. Here, we performed high-throughput sequencing of 16S rRNA gene amplicons and metaproteomics analysis of fecal samples to determine microbial gut composition and protein expression. 16 rRNA gene sequencing analysis identified Clostridiales, Bacteroidaceae, and Lactobacillaceae species as the most abundant species in the gut. For metaproteomics analysis, peptides were generated by using the Fasp method and subsequently fractionated by strong anion exchanges. Metaproteomics analysis identified 3,673 proteins. Among the most frequently identified proteins, 380 proteins belonged to Lactobacillus spp., 155 belonged to Clostridium spp., and 66 belonged to Streptococcus spp. The most frequently identified proteins were heat shock chaperones, including 349 GroEL proteins, from many bacterial species, whereas the most abundant enzymes were pyruvate kinases, as judged by the number of peptides identified per protein (spectral counting). Gene ontology and KEGG pathway analyses revealed the functions and locations of the identified proteins. The findings of both metaproteomics and 16S rRNA sequencing analyses are discussed.
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http://dx.doi.org/10.1128/AEM.02472-13DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3911106PMC
January 2014

Comparative analysis of ESBL-positive Escherichia coli isolates from animals and humans from the UK, The Netherlands and Germany.

PLoS One 2013 26;8(9):e75392. Epub 2013 Sep 26.

Animal Health and Veterinary Laboratories Agency (AHVLA, Weybridge), Addlestone, United Kingdom.

The putative virulence and antimicrobial resistance gene contents of extended spectrum β-lactamase (ESBL)-positive E. coli (n=629) isolated between 2005 and 2009 from humans, animals and animal food products in Germany, The Netherlands and the UK were compared using a microarray approach to test the suitability of this approach with regard to determining their similarities. A selection of isolates (n=313) were also analysed by multilocus sequence typing (MLST). Isolates harbouring bla(CTX-M-group-1) dominated (66%, n=418) and originated from both animals and cases of human infections in all three countries; 23% (n=144) of all isolates contained both bla(CTX-M-group-1) and bla(OXA-1-like) genes, predominantly from humans (n=127) and UK cattle (n=15). The antimicrobial resistance and virulence gene profiles of this collection of isolates were highly diverse. A substantial number of human isolates (32%, n=87) did not share more than 40% similarity (based on the Jaccard coefficient) with animal isolates. A further 43% of human isolates from the three countries (n=117) were at least 40% similar to each other and to five isolates from UK cattle and one each from Dutch chicken meat and a German dog; the members of this group usually harboured genes such as mph(A), mrx, aac(6')-Ib, catB3, bla(OXA-1-like) and bla(CTX-M-group-1). forty-four per cent of the MLST-typed isolates in this group belonged to ST131 (n=18) and 22% to ST405 (n=9), all from humans. Among animal isolates subjected to MLST (n=258), only 1.2% (n=3) were more than 70% similar to human isolates in gene profiles and shared the same MLST clonal complex with the corresponding human isolates. The results suggest that minimising human-to-human transmission is essential to control the spread of ESBL-positive E. coli in humans.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0075392PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3784421PMC
July 2014