Prophage induction reduces Shiga toxin producing \u3ci\u3eEscherichia coli\u3c/i\u3e (STEC) and Salmonella enterica on tomatoes and spinach: A model study

Fresh produce is increasingly implicated in foodborne outbreaks and most fresh produce is consumed raw, emphasizing the need to develop non-thermal methods to control foodborne pathogens. This study investigates bacterial cell lysis through induction of prophages as a novel approach to control foodborne bacterial pathogens on fresh produce. Shiga toxin producing Escherichia coli (STEC) and Salmonella enterica isolates were exposed to different prophage inducers (i.e. mitomycin C or streptonigrin) and growth of the cells was monitored by measuring the optical density (OD600) during incubation at 37C. Beginning at three hours after addition of the inducer, all concentrations (0.5, 1, 2 mg/mL) of mitomycin C, or 2 mg/mL streptonigrin significantly reduced the OD600 in broth cultures, in a concentration dependent manner, relative to cultures where no inducer was added. PCR confirmed bacterial release of induced bacteriophages and demonstrated that a single compound could successfully induce multiple types of prophages. The ability of mitomycin C to induce prophages in STEC O157:H7 and in S. enterica (serovars Typhimurium and Newport) on fresh produce was evaluated by inoculating red greenhouse tomatoes or spinach leaves with 5 x 107 and 5 x 108 colony forming units, respectively. After allowing time for the inoculum to dry on the fresh produce samples, 6 mg/mL mitomycin C was sprayed onto each sample, while control samples were sprayed with water. Following overnight incubation at 4C, the bacterial cells were recovered and plate counts were performed. A 3 log reduction in STEC O157:H7 cells was observed on tomatoes sprayed with mitomycin C compared to those sprayed with water, while a 1 log reduction was obtained on spinach. Similarly, spraying mitomycin C on tomatoes and spinach inoculated with S. enterica isolates resulted in a 1-1.5 log and 2 log reduction, respectively. These findings serve as a proof of concept that prophage induction can effectively control bacterial foodborne pathogens on fresh produce

MICL controls inflammation in rheumatoid arthritis

Acknowledgments We thank G Milne, D MacCallum, S Hardison, G Wilson, C Wallace, S Hadebe and A Richmond for assistance; H. El-Gabalawy for tissues and the animal facility staff for the care of our animals. Flow cytometry was undertaken in the Iain Fraser Cytometry Centre, University of Aberdeen. Funding: GDB was funded by the Wellcome Trust and MRC (UK). AA and CDB are supported by the Arthritis Research UK Tissue Engineering Centre (grant 19429). This study makes use of data generated by the Wellcome Trust Case Control Consortium. A full list of the investigators who contributed to the generation of the data is available from http://www.wtccc.org.uk, and was funded by the Wellcome Trust (076113). MJGF was funded by The Arthritis Society and the Canadian Arthritis Network and J-ML by a scholarship from the Canadian Arthritis Network.Peer reviewedPublisher PD

Comparative genomics of isolates of a pseudomonas aeruginosa epidemic strain associated with chronic lung infections of cystic fibrosis patients

Pseudomonas aeruginosa is the main cause of fatal chronic lung infections among individuals suffering from cystic fibrosis (CF). During the past 15 years, particularly aggressive strains transmitted among CF patients have been identified, initially in Europe and more recently in Canada. The aim of this study was to generate high-quality genome sequences for 7 isolates of the Liverpool epidemic strain (LES) from the United Kingdom and Canada representing different virulence characteristics in order to: (1) associate comparative genomics results with virulence factor variability and (2) identify genomic and/or phenotypic divergence between the two geographical locations. We performed phenotypic characterization of pyoverdine, pyocyanin, motility, biofilm formation, and proteolytic activity. We also assessed the degree of virulence using the Dictyostelium discoideum amoeba model. Comparative genomics analysis revealed at least one large deletion (40-50 kb) in 6 out of the 7 isolates compared to the reference genome of LESB58. These deletions correspond to prophages, which are known to increase the competitiveness of LESB58 in chronic lung infection. We also identified 308 non-synonymous polymorphisms, of which 28 were associated with virulence determinants and 52 with regulatory proteins. At the phenotypic level, isolates showed extensive variability in production of pyocyanin, pyoverdine, proteases and biofilm as well as in swimming motility, while being predominantly avirulent in the amoeba model. Isolates from the two continents were phylogenetically and phenotypically undistinguishable. Most regulatory mutations were isolate-specific and 29% of them were predicted to have high functional impact. Therefore, polymorphism in regulatory genes is likely to be an important basis for phenotypic diversity among LES isolates, which in turn might contribute to this strain's adaptability to varying conditions in the CF lung

Longitudinal In Vivo Imaging of Retinal Ganglion Cells and Retinal Thickness Changes Following Optic Nerve Injury in Mice

Retinal ganglion cells (RGCs) die in sight-threatening eye diseases. Imaging RGCs in humans is not currently possible and proof of principle in experimental models is fundamental for future development. Our objective was to quantify RGC density and retinal thickness following optic nerve transection in transgenic mice expressing cyan fluorescent protein (CFP) under control of the Thy1 promoter, expressed by RGCs and other neurons.A modified confocal scanning laser ophthalmoscopy (CSLO)/spectral-domain optical coherence tomography (SD-OCT) camera was used to image and quantify CFP+ cells in mice from the B6.Cg-Tg(Thy1-CFP)23Jrs/J line. SD-OCT circle (1 B-scan), raster (37 B-scans) and radial (24 B-scans) scans of the retina were also obtained. CSLO was performed at baseline (n = 11) and 3 (n = 11), 5 (n = 4), 7 (n = 10), 10 (n = 6), 14 (n = 7) and 21 (n = 5) days post-transection, while SD-OCT was performed at baseline and 7, 14 and 35 days (n = 9) post-transection. Longitudinal change in CFP+ cell density and retinal thickness were computed. Compared to baseline, the mean (SD) percentage CFP+ cells remaining at 3, 5, 7, 10, 14 and 21 days post-transection was 86 (9)%, 63 (11)%, 45 (11)%, 31 (9)%, 20 (9)% and 8 (4)%, respectively. Compared to baseline, the mean (SD) retinal thickness at 7 days post-transection was 97 (3)%, 98 (2)% and 97 (4)% for the circle, raster and radial scans, respectively. The corresponding figures at 14 and 35 days post-transection were 96 (3)%, 97 (2)% and 95 (3)%; and 93 (3)%, 94 (3)% and 92 (3)%.Longitudinal imaging showed an exponential decline in CFP+ cell density and a small (≤8%) reduction in SD-OCT measured retinal thickness post-transection. SD-OCT is a promising tool for detecting structural changes in experimental optic neuropathy. These results represent an important step towards translation for clinical use

Re-evaluation of blood mercury, lead and cadmium concentrations in the Inuit population of Nunavik (Québec): a cross-sectional study

<p>Abstract</p> <p>Background</p> <p>Arctic populations are exposed to mercury, lead and cadmium through their traditional diet. Studies have however shown that cadmium exposure is most often attributable to tobacco smoking. The aim of this study is to examine the trends in mercury, lead and cadmium exposure between 1992 and 2004 in the Inuit population of Nunavik (Northern Québec, Canada) using the data obtained from two broad scale health surveys, and to identify sources of exposure in 2004.</p> <p>Methods</p> <p>In 2004, 917 adults aged between 18 and 74 were recruited in the 14 communities of Nunavik to participate to a broad scale health survey. Blood samples were collected and analysed for metals by inductively coupled plasma mass spectrometry, and dietary and life-style characteristics were documented by questionnaires. Results were compared with data obtained in 1992, where 492 people were recruited for a similar survey in the same population.</p> <p>Results</p> <p>Mean blood concentration of mercury was 51.2 nmol/L, which represent a 32% decrease (p < 0.001) between 1992 and 2004. Mercury blood concentrations were mainly explained by age (partial r²= 0.20; p < 0.0001), and the most important source of exposure to mercury was marine mammal meat consumption (partial r²= 0.04; p < 0.0001). In 2004, mean blood concentration of lead was 0.19 μmol/L and showed a 55% decrease since 1992. No strong associations were observed with any dietary source, and lead concentrations were mainly explained by age (partial r²= 0.20.; p < 0.001). Blood cadmium concentrations showed a 22% decrease (p < 0.001) between 1992 and 2004. Once stratified according to tobacco use, means varied between 5.3 nmol/L in never-smokers and 40.4 nmol/L in smokers. Blood cadmium concentrations were mainly associated with tobacco smoking (partial r²= 0.56; p < 0.0001), while consumption of caribou liver and kidney remain a minor source of cadmium exposure among never-smokers.</p> <p>Conclusion</p> <p>Important decreases in mercury, lead and cadmium exposure were observed. Mercury decrease could be explained by dietary changes and the ban of lead cartridges use likely contributed to the decrease in lead exposure. Blood cadmium concentrations remain high and, underscoring the need for intensive tobacco smoking prevention campaigns in the Nunavik population.</p

RA-MAP, molecular immunological landscapes in early rheumatoid arthritis and healthy vaccine recipients

Rheumatoid arthritis (RA) is a chronic inflammatory disorder with poorly defined aetiology characterised by synovial inflammation with variable disease severity and drug responsiveness. To investigate the peripheral blood immune cell landscape of early, drug naive RA, we performed comprehensive clinical and molecular profiling of 267 RA patients and 52 healthy vaccine recipients for up to 18 months to establish a high quality sample biobank including plasma, serum, peripheral blood cells, urine, genomic DNA, RNA from whole blood, lymphocyte and monocyte subsets. We have performed extensive multi-omic immune phenotyping, including genomic, metabolomic, proteomic, transcriptomic and autoantibody profiling. We anticipate that these detailed clinical and molecular data will serve as a fundamental resource offering insights into immune-mediated disease pathogenesis, progression and therapeutic response, ultimately contributing to the development and application of targeted therapies for RA.</p

Intracellular Localization in Zebrafish Muscle and Conserved Sequence Features Suggest Roles for Gelatinase A Moonlighting in Sarcomere Maintenance

Gelatinase A (Mmp2 in zebrafish) is a well-characterized effector of extracellular matrix remodeling, extracellular signaling, and along with other matrix metalloproteinases (MMPs) and extracellular proteases, it plays important roles in the establishment and maintenance of tissue architecture. Gelatinase A is also found moonlighting inside mammalian striated muscle cells, where it has been implicated in the pathology of ischemia-reperfusion injury. Gelatinase A has no known physiological function in muscle cells, and its localization within mammalian cells appears to be due to inefficient recognition of its N-terminal secretory signal. Here we show that Mmp2 is abundant within the skeletal muscle cells of zebrafish, where it localizes to the M-line of sarcomeres and degrades muscle myosin. The N-terminal secretory signal of zebrafish Mmp2 is also challenging to identify, and this is a conserved characteristic of gelatinase A orthologues, suggesting a selective pressure acting to prevent the efficient secretion of this protease. Furthermore, there are several strongly conserved phosphorylation sites within the catalytic domain of gelatinase A orthologues, some of which are phosphorylated in vivo, and which are known to regulate the activity of this protease. We conclude that gelatinase A likely participates in uncharacterized physiological functions within the striated muscle, possibly in the maintenance of sarcomere proteostasis, that are likely regulated by kinases and phosphatases present in the sarcomere.Dentistry, Faculty ofNon UBCOral Biological and Medical Sciences (OBMS), Department ofReviewedFacult

Refining the tethering of American oysters (Crassostrea virginica) to measure the effects of two environmental stressors

Little is known about the microbiota shift induced by exacerbation in chronic obstructive pulmonary disease (COPD) patients. The sputa microbiota of COPD patients was evaluated when clinically stable and during acute exacerbations of the disease. Sputa microbiota was analyzed using 16S ribosomal RNA gene pyrosequencing and quantitative polymerase chain reaction-based pathogen detection. Nine COPD patients were enrolled. Pyrosequencing of 16S rRNA genes identified 2,267 unique bacterial operational taxonomic units. Principal microbiota shifts during exacerbation were in either Proteobacteria, Firmicutes or Bacteroidetes. Streptococcus and Moraxella levels were detected during exacerbation in severe (Global Initiative for Chronic Obstructive Lung Disease 3) COPD patients. Most of the clinically-important genera found in the sputum with the pyrosequencing of 16S rRNA gene correlated with specific quantitative polymerase chain reactions for bacteria while respiratory viruses were nearly absent. Sputum microbiotas of exacerbated COPD patients are complex. This pilot study shows a clear shift in the microbiota of patients during exacerbation. The nature of this shift varies from patient to patient in such a way that the treatment should be patient-specific. Further studies are needed to establish the impact of microbial exacerbations on the pulmonary microbiota

Prophage induction reduces Shiga toxin producing \u3ci\u3eEscherichia coli\u3c/i\u3e (STEC) and Salmonella enterica on tomatoes and spinach: A model study

Fresh produce is increasingly implicated in foodborne outbreaks and most fresh produce is consumed raw, emphasizing the need to develop non-thermal methods to control foodborne pathogens. This study investigates bacterial cell lysis through induction of prophages as a novel approach to control foodborne bacterial pathogens on fresh produce. Shiga toxin producing Escherichia coli (STEC) and Salmonella enterica isolates were exposed to different prophage inducers (i.e. mitomycin C or streptonigrin) and growth of the cells was monitored by measuring the optical density (OD600) during incubation at 37C. Beginning at three hours after addition of the inducer, all concentrations (0.5, 1, 2 mg/mL) of mitomycin C, or 2 mg/mL streptonigrin significantly reduced the OD600 in broth cultures, in a concentration dependent manner, relative to cultures where no inducer was added. PCR confirmed bacterial release of induced bacteriophages and demonstrated that a single compound could successfully induce multiple types of prophages. The ability of mitomycin C to induce prophages in STEC O157:H7 and in S. enterica (serovars Typhimurium and Newport) on fresh produce was evaluated by inoculating red greenhouse tomatoes or spinach leaves with 5 x 107 and 5 x 108 colony forming units, respectively. After allowing time for the inoculum to dry on the fresh produce samples, 6 mg/mL mitomycin C was sprayed onto each sample, while control samples were sprayed with water. Following overnight incubation at 4C, the bacterial cells were recovered and plate counts were performed. A 3 log reduction in STEC O157:H7 cells was observed on tomatoes sprayed with mitomycin C compared to those sprayed with water, while a 1 log reduction was obtained on spinach. Similarly, spraying mitomycin C on tomatoes and spinach inoculated with S. enterica isolates resulted in a 1-1.5 log and 2 log reduction, respectively. These findings serve as a proof of concept that prophage induction can effectively control bacterial foodborne pathogens on fresh produce