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Here we investigate the faecal microbiome of wild European badgers Meles meles using samples collected at post-mortem as part of the All Wales Badger Found Dead study. This is the first published characterisation of the badger microbiome. We initially undertook a sex-matched age comparison between the adult and cub microbiomes, based on sequencing the V3–V4 region of the 16S rRNA gene. Analysis used the QIIME 2 pipeline utilising DADA2 and the Silva database for taxonomy assignment. Fusobacteria appeared to be more abundant in the microbiomes of the cubs than the adults although no significant difference was seen in alpha or beta diversity between the adult and cub badger microbiomes. Comparisons were also made against other wild, omnivorous, mammals’ faecal microbiomes using publicly available data. Significant differences were seen in both alpha and beta diversity between the microbiomes from different species. As a wildlife species of interest to the disease bovine tuberculosis, knowledge of the faecal microbiome could assist in identification of infected badgers. Our work here suggests that, if comparisons were made between the faeces of bTB infected and non-infected badgers, age may not have a significant impact on the microbiome.

Introduction The European badger (Meles meles) is a known wildlife reservoir for bovine tuberculosis (bTB) and a better understanding of the epidemiology of bTB in this wildlife species is required for disease control in both wild and farmed animals. Flow infusion electrospray—high-resolution mass spectrometry (FIE-HRMS) may potentially identify novel metabolite biomarkers based on which new, rapid, and sensitive point of care tests for bTB infection could be developed. Objectives In this foundational study, we engaged on assessing the baseline metabolomic variation in the non-bTB infected badger population (“metabotyping”) across Wales. Methods FIE-HRMS was applied on thoracic fluid samples obtained by post-mortem of bTB negative badgers (n = 285) which were part of the Welsh Government ‘All Wales Badger Found Dead’ study. Results Using principal component analysis and partial least squares—discriminant analyses, the major sources of variation were linked to sex, and to a much lesser extent age, as indicated by tooth wear. Within the female population, variation was seen between lactating and non-lactating individuals. No significant variation linked to the presence of bite wounds, obvious lymphatic lesions or geographical region of origin was observed. Conclusion Future metabolomic work when making comparisons between bTB infected and non-infected badger samples will only need be sex-matched and could focus on males only, to avoid lactation bias.

Introduction Paratuberculosis, commonly known as Johne’s disease, is a chronic granulomatous infection of ruminants caused by Mycobacterium avium subspecies paratuberculosis (MAP). Clinical signs, including reduced milk yields, weight loss and diarrhoea, are typically absent until 2 to 6 years post exposure. Objectives To identify metabolomic changes profiles of MAP challenged Holstein–Friesian (HF) cattle and correlate identified metabolites to haematological and immunological parameters. Methods At approximately 6 weeks of age, calves (n = 9) were challenged with 3.8 × 109 cells of MAP (clinical isolate CIT003) on 2 consecutive days. Additional unchallenged calves (n = 9) formed the control group. The study used biobanked serum from cattle sampled periodically from 3- to 33-months post challenge. The assessment of sera using flow infusion electrospray high resolution mass spectrometry (FIE-HRMS) for high throughput, sensitive, non-targeted metabolite fingerprinting highlighted differences in metabolite levels between the two groups. Results In total, 25 metabolites which were differentially accumulated in MAP challenged cattle were identified, including 20 which displayed correlation to haematology parameters, particularly monocyte levels. Conclusion The targeted metabolites suggest shifts in amino acid metabolism that could reflect immune system activation linked to MAP and as well as differences in phosphocholine levels which could reflect activation of the Th1 (tending towards pro-inflammatory) immune response. If verified by future work, selected metabolites could be used as biomarkers to diagnose and manage MAP infected cattle.

Here we investigate the faecal microbiome of wild European badgers Meles meles using samples collected at post-mortem as part of the All Wales Badger Found Dead study based on sequencing the V3-V4 region of the 16S rRNA gene. This is the first published characterisation of the badger microbiome. We initially undertook a sex-matched age comparison between the adult and cub microbiomes. Analysis used the QIIME 2 pipeline utilising DADA2 and the Silva database for taxonomy assignment. Fusobacteria appeared to be more abundant in the microbiomes of the cubs than the adults although no significant difference was seen in alpha or beta diversity between the adult and cub badger microbiomes. Comparisons were also made against other wild, omnivorous, mammals’ faecal microbiomes using publicly available data. Significant differences were seen in both alpha and beta diversity between the microbiomes from different species. As a wildlife species of interest to the disease bovine tuberculosis, knowledge of the faecal microbiome could assist in identification of infected badgers. Our work here suggests that if comparisons were made between the faeces of bTB infected and non-infected badgers, its possible age may not have a significant impact on the microbiome.

Bacteriophages (phages) are viruses that target bacteria, with the ability to lyse and kill host bacterial cells. Due to this, they have been of some interest as a therapeutic since their discovery in the early 1900s, but with the recent increase in antibiotic resistance, phages have seen a resurgence in attention. Current methods of isolation and purification of phages can be long and tedious, with caesium chloride concentration gradients the gold standard for purifying a phage fraction. Isolation of novel phages requires centrifugation and ultrafiltration of mixed samples, such as water sources, effluent or faecal samples etc, to prepare phage filtrates for further testing. We propose countercurrent chromatography as a novel and alternative approach to use when studying phages, as a scalable and high-yield method for obtaining phage fractions. However, the full extent of the usefulness and resolution of separation with this technique has not been researched; it requires optimization and ample testing before this can be revealed. Here we present an initial study to determine survivability of two phages, T4 and ϕX174, using only water as a mobile phase in a Spectrum Series 20 HPCCC. Both phages were found to remain active once eluted from the column. Phages do not fully elute from the column and sodium hydroxide is necessary to flush the column between runs to deactivate remaining phages.

Introduction Mycobacterium bovis, the causative agent of bovine tuberculosis (bTB) in cattle, represents a major disease burden to UK cattle farming, with considerable costs associated with its control. The European badger (Meles meles) is a known wildlife reservoir for bTB and better knowledge of the epidemiology of bTB through testing wildlife is required for disease control. Current tests available for the diagnosis of bTB in badgers are limited by cost, processing time or sensitivities. Materials and Methods We assessed the ability of flow infusion electrospray—high-resolution mass spectrometry (FIE-HRMS) to determine potential differences between infected and non-infected badgers based on thoracic blood samples obtained from badgers found dead in Wales. Thoracic blood samples were autoclaved for handling in a containment level 2 (CL2) hazard laboratory. Results Here we show the major differences associated with with M. bovis infection were changes to folate, pyrimidine, histidine, glycerophospholipid and phosphonate metabolism. Conclusions Our studies have indicated differences in the metabolomic signature of badgers found dead in relation to their infection status, suggesting metabolomics could hold potential for developing novel diagnostics for bTB in badgers. As well as highlighting a potential way to handle samples containing a highly pathogenic agent at CL2 for metabolomics studies.

1.AbstractBacteriophages (phages) are viruses that target bacteria, with the ability to lyse and kill host bacterial cells. Due to this, they have been of some interest as a therapeutic since their discovery in the early 1900s, but with the recent increase in antibiotic resistance, phages have seen a resurgence in attention. Current methods of isolation and purification of phages can be long and tedious, with caesium chloride concentration gradients the gold standard for purifying a phage fraction. Isolation of novel phages requires centrifugation and ultrafiltration of mixed samples, such as water sources, effluent or faecal samples etc, to prepare phage filtrates for further testing. We propose countercurrent chromatography as a novel and alternative approach to use when studying phages, as a scalable and high-yield method for obtaining phage fractions. However, the full extent of the usefulness and resolution of separation with this technique has not been researched; it requires optimisation and ample testing before this can be revealed. Here we present an initial study to determine survivability of two phages, T4 and ϕX174, using only water as a mobile phase in a Spectrum Series 20 HPCCC. Both phages were found to remain active once eluted from the column. Phages do not fully elute from the column and sodium hydroxide is necessary to flush the column between runs to deactivate remaining phages.

COPD is one of the top 10 causes of death worldwide. Omic technologies represent a “big-data” approach to more accurate describe key features of COPD and inform treatment strategies. Herein we explore metabolomic approaches to study biochemical changes in the sputum of COPD patients and volunteers (n=14) not diagnosed with a respiratory disease (CON). Sputum samples were assessed by high resolution - Direct infusion mass spectroscopy (DI-MS) based in the Q Exactive hybrid quadrupole-Orbitrap was used to assess the sample’s metabolomes. COPD samples were shown to be distinct using PLS-DA from CON and the sources of variation were identified. Five key metabolites allowed the discrimination between COPD and CON samples with an AUC value of 99% (CI 100%- 0.943 %). Three of the key metabolites were monoacylglycerides also stearic acid suggesting considerable changes in fatty processing in COPD. Other metabolites were involved in pathways involved in inflammatory mediation, bronchoconstriction and mucus secretion, known features of COPD. Ongoing working is assessing metabolomic changes at different GOLD stages, examining the impact of comorbidities and smoking status. We expect new insights that could improve the monitoring of COPD and its treatment.

Although the prokaryotic communities of the rumen microbiome are being uncovered through genome sequencing, little is known about the resident viral populations. Whilst temperate phages can be predicted as integrated prophages when analyzing bacterial and archaeal genomes, the genetics underpinning lytic phages remain poorly characterized. To the five genomes of bacteriophages isolated from rumen-associated samples sequenced and analyzed previously, this study adds a further five novel genomes and predictions gleaned from them to further the understanding of the rumen phage population. Lytic bacteriophages isolated from fresh ovine and bovine fecal and rumen fluid samples were active against the predominant fibrolytic ruminal bacterium Butyrivibrio fibrisolvens. The double stranded DNA genomes were sequenced and reconstructed into single circular complete contigs. Based on sequence similarity and genome distances, the five phages represent four species from three separate genera, consisting of: (1) Butyrivibrio phages Arian and Bo-Finn; (2) Butyrivibrio phages Idris and Arawn; and (3) Butyrivibrio phage Ceridwen. They were predicted to all belong to the Siphoviridae family, based on evidence in the genomes such as size, the presence of the tail morphogenesis module, genes that share similarity to those in other siphovirus isolates and phylogenetic analysis using phage proteomes. Yet, phylogenomic analysis and sequence similarity of the entire phage genomes revealed that these five phages are unique and novel. These phages have only been observed undergoing the lytic lifecycle, but there is evidence in the genomes of phages Arawn and Idris for the potential to be temperate. However, there is no evidence in the genome of the bacterial host Butyrivibrio fibrisolvens of prophage genes or genes that share similarity with the phage genomes.

Background/aimThis paper summarises and extends information presented at the Lilly Lecture 2017. It is based on the author’s experience and research in working to develop senior leaders in many sectors over the last 30 years. The aim is to provide practical, usable insights into an aspect of leadership capability that is often overlooked in order to enable greater effectiveness in leaders.MethodsThe paper draws on the author’s own experience, of the research of Harthill and other organisations working with the concepts that have been labelled constructivist development or more lately ‘vertical’ development.ResultsResearch, both qualitative and quantitative, points to the need for leaders to develop the meaning-making capacities to deal with increased levels of complexity and ambiguity. Fewer than 5% of leaders across multisectors are seen to develop these capacities.ConclusionLeaders within the National Health Service are particularly challenged by complex and ambiguous contexts. They need to extend their leadership capacities to be able to lead effectively in such contexts. Leaders today are not being supported to develop these capacities and more needs to be done. There are actions that individuals as well as organisations can take to support this development.