A rhodanine agent active against non-replicating intracellular Mycobacterium avium subspecies paratuberculosis.

BACKGROUND: Antibiotic therapy targeting chronic mycobacterial disease is often ineffective due to problems with the emergence of drug resistance and non-replicating persistent intracellular antibiotic resistant phenotypes. Strategies which include agents able to enhance host cell killing mechanisms could represent an alternative to conventional methods with the potential for host clearance if active against dormant phenotypes. Investigations of agents with potential activity against non-replicating mycobacteria however are restricted due to a need for assays that can assess bacterial viability without having to culture. RESULTS: This study describes the development and use of a pre16S ribosomal gene RNA/DNA ratio viability assay which is independent of the need for culture, supported by a novel thin layer accelerated mycobacterial colony forming method for determining viability and culturability of MAP in intracellular environments. We describe the use of these tools to demonstrate intracellular killing activity of a novel rhodanine agent (D157070) against the intracellular pathogen Mycobacterium avium subspecies paratuberculosis (MAP) and show that the culturability of MAP decreases relative to its viability on intracellular entry suggesting the induction of a non-culturable phenotype. We further demonstrate that D157070, although having no direct activity against the culturability of extracellular MAP, can bind to cultured MAP cells and has significant influence on the MAP transcriptome, particularly with respect of delta(L )associated genes. D157070 is shown to be taken up by bovine and human cells and able to enhance host cell killing, as measured by significant decreases in both culturability and viability of intracellular MAP. CONCLUSIONS: This work suggests that pre16srRNA gene ratios represent a viable method for studying MAP viability. In addition, the rhodanine agent D157070 tested is non-toxic and enhances cell killing activity against both growing and latent MAP phenotypes

Genomic variations associated with attenuation in Mycobacterium avium subsp paratuberculosis vaccine strains

BACKGROUND: Mycobacterium avium subspecies paratuberculosis (MAP) whole cell vaccines have been widely used tools in the control of Johne's disease in animals despite being unable to provide complete protection. Current vaccine strains derive from stocks created many decades ago; however their genotypes, underlying mechanisms and relative degree of their attenuation are largely unknown. RESULTS: Using mouse virulence studies we confirm that MAP vaccine strains 316 F, II and 2e have diverse but clearly attenuated survival and persistence characteristics compared with wild type strains. Using a pan genomic microarray we characterise the genomic variations in a panel of vaccine strains sourced from stocks spanning over 40 years of maintenance. We describe multiple genomic variations specific for individual vaccine stocks in both deletion (26-32 Kbp) and tandem duplicated (11-40 Kbp) large variable genomic islands and insertion sequence copy numbers. We show individual differences suitable for diagnostic differentiation between vaccine and wild type genotypes and provide evidence for functionality of some of the deleted MAP-specific genes and their possible relation to attenuation. CONCLUSIONS: This study shows how culture environments have influenced MAP genome diversity resulting in large tandem genomic duplications, deletions and transposable element activity. In combination with classical selective systematic subculture this has led to fixation of specific MAP genomic alterations in some vaccine strain lineages which link the resulting attenuated phenotypes with deficiencies in high reactive oxygen species handling

The potential of integrated nurse-led models to improve care for people with functional gastrointestinal disorders: a systematic review

Functional gastrointestinal disorders (FGID) such as irritable bowel syndrome (IBS) and functional dyspepsia (FD) are extremely common, debilitating and costly. Although diagnostic guidelines and effective management options exist, management is sub-optimal, with long waiting lists, delayed diagnosis and poor patient outcomes. The aim of this systematic review was to explore and evaluate evidence for existing models of care for functional gastrointestinal disorders. 38 studies pertaining to the diagnosis or management of FGIDs were found, however only 6 investigated a full model of care. Five studies assessed a nurse-led model and one a structured gastroenterologist consultation. Nurse-led models were cheaper to current treatments, and resulted in symptomatic improvement, high patient satisfaction, reduced healthcare usage, and improved psychosocial functioning and quality of life, whilst standard gastroenterological care did not improve pain or quality of life. There is minimal research trialling integrated models of care for the diagnosis and management of functional gastrointestinal disorders. This represents a lost opportunity for timely, effective, healthcare provision to a large patient group. Although low in quality, preliminary data suggest that integrated nurse-led models of care are economically viable and may facilitate timely diagnosis and management, and improve patient outcomes. Further, studies to robustly evaluate the efficacy, safety and acceptability of such models are needed.Ecushla C. Linedale, Antonina Mikocka-Walus, Peter R. Gibson, Jane M. Andrew

Serum antibodies against Toxoplasma gondii and Neospora caninum in southeast Queensland dugongs

The dugong (Dugong dugon) is an herbivorous marine mammal that inhabits tropical inshore waters and thus may be vulnerable to pollutants and terrestrial pathogens as a result of coastal runoff. In this study, serum samples collected from live, wild dugongs (n = 114) in an embayment located on the urbanized southeast Queensland coast of Australia during 2008–2014, were measured for IgG antibody levels specific to Toxoplasma gondii and Neospora caninum. An ELISA used to measure T. gondii tachyzoite antibodies indicated a non-Gaussian distribution of antibody level, with five dugongs identified as high outliers. Mean levels of antibodies specific for T. gondii in dugongs sampled in 2014 were significantly higher than in 2010 (p =.006) and 2011 (p =.009) with an elevation in mean antibody levels after a major 2011 flood event relative to antibody levels prior to the flood (

Nutrient intake in patients with inflammatory bowel disease: Are they meeting their dietary requirements?

First published: 25 September 2015A Pilichiewicz, C Goess, E Linedale, S Ooi, R Bryant, R Grafton, J Hughes, J Andrew

Transgenic expression of a viral oncoprotein exclusively in K14(+) epithelial cells results in peripheral but not central CD8 T cell tolerance

The role of thymic versus peripheral epithelial cells in the negative selection of the peptide-specific CD8 T cell repertoire is still largely unresolved. We have generated TCRb chain transgenic mice in which 20–35% of peripheral CD8 T cells recognize an epitope from a viral, nuclear oncoprotein (human papillomavirus type 16 E7) in the context ofMHC class I, H-2Db. When T cells from these transgenic mice develop through the thymus of a second transgenic mouse expressing E7 from a keratin 14 promoter, no major perturbation to thymic T cell development is observed over a 7 month period. In contrast, peripheral CD8 T cell responses in these same mice (E7TCRxK14E7 double transgenic) become reduced over time. This data suggests that peripheral tolerance mechanisms predominate over thymic negative selection in controlling CD8 T cell responses to this epithelial, nuclear oncoprotein