How does reviewing the evidence change veterinary surgeons' beliefs regarding the treatment of ovine footrot? A quantitative and qualitative study

Footrot is a widespread, infectious cause of lameness in sheep, with major economic and welfare costs. The aims of this research were: (i) to quantify how veterinary surgeons’ beliefs regarding the efficacy of two treatments for footrot changed following a review of the evidence (ii) to obtain a consensus opinion following group discussions (iii) to capture complementary qualitative data to place their beliefs within a broader clinical context. Grounded in a Bayesian statistical framework, probabilistic elicitation (roulette method) was used to quantify the beliefs of eleven veterinary surgeons during two one-day workshops. There was considerable heterogeneity in veterinary surgeons’ beliefs before they listened to a review of the evidence. After hearing the evidence, seven participants quantifiably changed their beliefs. In particular, two participants who initially believed that foot trimming with topical oxytetracycline was the better treatment, changed to entirely favour systemic and topical oxytetracycline instead. The results suggest that a substantial amount of the variation in beliefs related to differences in veterinary surgeons’ knowledge of the evidence. Although considerable differences in opinion still remained after the evidence review, with several participants having non-overlapping 95% credible intervals, both groups did achieve a consensus opinion. Two key findings from the qualitative data were: (i) veterinary surgeons believed that farmers are unlikely to actively seek advice on lameness, suggesting a proactive veterinary approach is required (ii) more attention could be given to improving the way in which veterinary advice is delivered to farmers. In summary this study has: (i) demonstrated a practical method for probabilistically quantifying how veterinary surgeons’ beliefs change (ii) revealed that the evidence that currently exists is capable of changing veterinary opinion (iii) suggested that improved transfer of research knowledge into veterinary practice is needed (iv) identified some potential obstacles to the implementation of veterinary advice by farmers

Ovine pedomics : the first study of the ovine foot 16S rRNA-based microbiome

We report the first study of the bacterial microbiome of ovine interdigital skin based on 16S rRNA by pyrosequencing and conventional cloning with Sanger-sequencing. Three flocks were selected, one a flock with no signs of footrot or interdigital dermatitis, a second flock with interdigital dermatitis alone and a third flock with both interdigital dermatitis and footrot. The sheep were classified as having either healthy interdigital skin (H), interdigital dermatitis (ID) or virulent footrot (VFR). The ovine interdigital skin bacterial community varied significantly by flock and clinical condition. The diversity and richness of operational taxonomic units was greater in tissue from sheep with ID than H or VFR affected sheep. Actinobacteria, Bacteriodetes, Firmicutes and Proteobacteria were the most abundant phyla comprising 25 genera. Peptostreptococcus, Corynebacterium and Staphylococcus were associated with H, ID and VFR respectively. Sequences of Dichelobacter nodosus, the causal agent of ovine footrot, were not amplified due to mismatches in the 16S rRNA universal forward primer (27F). A specific real time PCR assay was used to demonstrate the presence of D. nodosus which was detected in all samples including the flock with no signs of ID or VFR. Sheep with ID had significantly higher numbers of D. nodosus (104-109 cells/g tissue) than those with H or VFR feet

A Spaetzle-like role for Nerve Growth Factor β in vertebrate immunity to Staphylococcus aureus

Many key components of innate immunity to infection are shared between Drosophila and humans. However, the fly Toll ligand Spaetzle is not thought to have a vertebrate equivalent. We have found that the structurally related cystine-knot protein, nerve growth factor β (NGFβ), plays an unexpected Spaetzle-like role in immunity to Staphylococcus aureus infection in chordates. Deleterious mutations of either human NGFβ or its high-affinity receptor tropomyosin-related kinase receptor A (TRKA) were associated with severe S. aureus infections. NGFβ was released by macrophages in response to S. aureus exoproteins through activation of the NOD-like receptors NLRP3 and NLRC4 and enhanced phagocytosis and superoxide-dependent killing, stimulated proinflammatory cytokine production, and promoted calcium-dependent neutrophil recruitment. TrkA knockdown in zebrafish increased susceptibility to S. aureus infection, confirming an evolutionarily conserved role for NGFβ-TRKA signaling in pathogen-specific host immunity

A Spaetzle-like role for nerve growth factor beta in vertebrate immunity to Staphylococcus aureus

Many key components of innate immunity to infection are shared between Drosophila and humans. However, the fly Toll ligand Spaetzle is not thought to have a vertebrate equivalent. We have found that the structurally related cystine-knot protein, nerve growth factor β (NGFβ), plays an unexpected Spaetzle-like role in immunity to Staphylococcus aureus infection in chordates. Deleterious mutations of either human NGFβ or its high-affinity receptor tropomyosin-related kinase receptor A (TRKA) were associated with severe S. aureus infections. NGFβ was released by macrophages in response to S. aureus exoproteins through activation of the NOD-like receptors NLRP3 and NLRC4 and enhanced phagocytosis and superoxide-dependent killing, stimulated proinflammatory cytokine production, and promoted calcium-dependent neutrophil recruitment. TrkA knockdown in zebrafish increased susceptibility to S. aureus infection, confirming an evolutionarily conserved role for NGFβ-TRKA signaling in pathogen-specific host immunity

Biogenesis and functions of bacterial S-layers.

The outer surface of many archaea and bacteria is coated with a proteinaceous surface layer (known as an S-layer), which is formed by the self-assembly of monomeric proteins into a regularly spaced, two-dimensional array. Bacteria possess dedicated pathways for the secretion and anchoring of the S-layer to the cell wall, and some Gram-positive species have large S-layer-associated gene families. S-layers have important roles in growth and survival, and their many functions include the maintenance of cell integrity, enzyme display and, in pathogens and commensals, interaction with the host and its immune system. In this Review, we discuss our current knowledge of S-layer and related proteins, including their structures, mechanisms of secretion and anchoring and their diverse functions

Characterisation of dichelobacter nodosus on misshapen and damaged ovine feet: a longitudinal study of four UK sheep flocks

Dichelobacter nodosus is the causal agent of ovine footrot, a contagious disease of welfare and economic concern worldwide. Damaged feet may be subclinical carriers of D. nodosus and covertly spread infection. Accordingly, we evaluated the risk of misshapen and damaged feet on D. nodosus presence and load in four commercial UK sheep flocks. Foot-level observations and swabs (n = 972) were collected from ewes (n = 85) over 12 months. On average, ewes were sampled three times. Feet were inspected for disease and scored (good/poor) for three hoof conformation traits (sole and heel, wall, and wall overgrowth). Swabs were analysed for presence and load of D. nodosus, and mixed models were constructed. Poor hoof conformation traits were present in 92.5% of foot-level observations. Feet with poor sole and heel conformation were more likely to have higher D. nodosus loads (β = 0.19, 95% CI: 0.04–0.35) than those with good conformation. Furthermore, on feet positive for D. nodosus, wall overgrowth was associated with higher D. nodosus loads (β = 0.27, 95% CI: 0.01–0.52). Feet with aspects of poor conformation covertly harbour D. nodosus and are a source of infection. Flock management should be guided by hoof conformation to reduce disease challenge

Within-flock population dynamics of Dichelobacter nodosus

Footrot causes 70 – 90 % of lameness in sheep in Great Britain. With approximately 5 % of 18 million adult sheep lame at any one time, it costs the UK sheep industry £24 - 84 million per year. The Gram-negative anaerobe Dichelobacter nodosus is the causative agent, with disease severity influenced by bacterial load, virulence and climate. The aim of the current study was to characterise strains of D. nodosus isolated by culture from swabs from healthy and diseased feet of 99 ewes kept as a closed flock over a 10-month period; and investigate persistence and transmission of strains within feet, sheep and the flock. Overall 268 isolates were characterised into strains by serogroup, proline-glycine repeat (pgr) status and multilocus variable number tandem repeat analysis (MLVA). The culture collection contained 87 unique MLVA profiles and two major MLVA complexes that persisted over time. A subset of 189 isolates tested for the virulence marker aprV2 were all positive. The two MLVA complexes (76 and 114) comprised 62 and 22 MLVA types and 237 and 28 isolates respectively. Serogroups B and I, and pgrB were associated with MLVA complex 76, whereas serogroups D and H were associated with MLVA complex 114. We conclude that within-flock D. nodosus evolution appeared to be driven by clonal diversification. There was no association (P > 0.05) between serogroup, pgr or MLVA type and disease state of feet. Strains of D. nodosus clustered within sheep and were transmitted between ewes over time. D. nodosus was isolated at more than one timepoint from 21 feet, including five feet where the same strain was isolated on two occasions at an interval of 1 - 33 weeks. Collectively our results indicate that D. nodosus strains persisted in the flock, spread between sheep and possibly persisted on feet over time

Specialist intervention is associated with improved patient outcomes in patients with decompensated heart failure: evaluation of the impact of a multidisciplinary inpatient heart failure team

Objective. The study aimed to evaluate the impact of a multidisciplinary inpatient heart failure team (HFT) on treatment, hospital readmissions and mortality of patients with decompensated heart failure (HF). Methods A retrospective service evaluation was undertaken in a UK tertiary centre university hospital comparing 196 patients admitted with HF in the 6 months prior to the introduction of the HFT (pre-HFT) with all 211 patients seen by the HFT (post-HFT) during its first operational year. Results. There were no significant differences in patient baseline characteristics between the groups. Inpatient mortality (22% pre-HFT vs 6% post-HFT; p<0.0001) and 1-year mortality (43% pre-HFT vs 27% post-HFT; p=0.001) were significantly lower in the post-HFT cohort. Post-HFT patients were significantly more likely to be discharged on loop diuretics (84% vs 98%; p=<0.0001), ACE inhibitors (65% vs 76%; p=0.02), ACE inhibitors and/or angiotensin receptor blockers (83% vs 91%; p=0.02), and mineralocorticoid receptor antagonists (44% vs 68%; p<0.0001) pre-HFT versus post-HFT, respectively. There was no difference in discharge prescription rates of betablockers (59% pre-HFT vs 63% post-HFT; p=0.45). The mean length of stay (17±19 days pre-HFT vs 19±18 days post-HFT; p=0.06), 1-year all-cause readmission rates (46% pre-HFT vs 47% post-HFT; p=0.82) and HF readmission rates (28% pre-HFT vs 20% post-HFT; p=0.09) were not different between the groups. Conclusions. The introduction of a specialist inpatient HFT was associated with improved patient outcome. Inpatient and 1-year mortality were significantly reduced. Improved use of evidence-based drug therapies, more intensive diuretic use and multidisciplinary care may contribute to these differences in outcome

Stepwise pathogenic evolution of Mycobacterium abscessus.

Although almost all mycobacterial species are saprophytic environmental organisms, a few, such as Mycobacterium tuberculosis, have evolved to cause transmissible human infection. By analyzing the recent emergence and spread of the environmental organism M. abscessus through the global cystic fibrosis population, we have defined key, generalizable steps involved in the pathogenic evolution of mycobacteria. We show that epigenetic modifiers, acquired through horizontal gene transfer, cause saltational increases in the pathogenic potential of specific environmental clones. Allopatric parallel evolution during chronic lung infection then promotes rapid increases in virulence through mutations in a discrete gene network; these mutations enhance growth within macrophages but impair fomite survival. As a consequence, we observe constrained pathogenic evolution while person-to-person transmission remains indirect, but postulate accelerated pathogenic adaptation once direct transmission is possible, as observed for M. tuberculosis Our findings indicate how key interventions, such as early treatment and cross-infection control, might restrict the spread of existing mycobacterial pathogens and prevent new, emergent ones

A Role for TLR4 in Clostridium difficile Infection and the Recognition of Surface Layer Proteins

Clostridium difficile is the etiological agent of antibiotic-associated diarrhoea (AAD) and pseudomembranous colitis in humans. The role of the surface layer proteins (SLPs) in this disease has not yet been fully explored. The aim of this study was to investigate a role for SLPs in the recognition of C. difficile and the subsequent activation of the immune system. Bone marrow derived dendritic cells (DCs) exposed to SLPs were assessed for production of inflammatory cytokines, expression of cell surface markers and their ability to generate T helper (Th) cell responses. DCs isolated from C3H/HeN and C3H/HeJ mice were used in order to examine whether SLPs are recognised by TLR4. The role of TLR4 in infection was examined in TLR4-deficient mice. SLPs induced maturation of DCs characterised by production of IL-12, TNFα and IL-10 and expression of MHC class II, CD40, CD80 and CD86. Furthermore, SLP-activated DCs generated Th cells producing IFNγ and IL-17. SLPs were unable to activate DCs isolated from TLR4-mutant C3H/HeJ mice and failed to induce a subsequent Th cell response. TLR4−/− and Myd88−/−, but not TRIF−/− mice were more susceptible than wild-type mice to C. difficile infection. Furthermore, SLPs activated NFκB, but not IRF3, downstream of TLR4. Our results indicate that SLPs isolated from C. difficile can activate innate and adaptive immunity and that these effects are mediated by TLR4, with TLR4 having a functional role in experimental C. difficile infection. This suggests an important role for SLPs in the recognition of C. difficile by the immune system