RocA truncation underpins hyper-encapsulation, carriage longevity and transmissibility of serotype M18 group A streptococci

Group A streptococcal isolates of serotype M18 are historically associated with epidemic waves of pharyngitis and the non-suppurative immune sequela rheumatic fever. The serotype is defined by a unique, highly encapsulated phenotype, yet the molecular basis for this unusual colony morphology is unknown. Here we identify a truncation in the regulatory protein RocA, unique to and conserved within our serotype M18 GAS collection, and demonstrate that it underlies the characteristic M18 capsule phenotype. Reciprocal allelic exchange mutagenesis of rocA between M18 GAS and M89 GAS demonstrated that truncation of RocA was both necessary and sufficient for hyper-encapsulation via up-regulation of both precursors required for hyaluronic acid synthesis. Although RocA was shown to positively enhance covR transcription, quantitative proteomics revealed RocA to be a metabolic regulator with activity beyond the CovR/S regulon. M18 GAS demonstrated a uniquely protuberant chain formation following culture on agar that was dependent on excess capsule and the RocA mutation. Correction of the M18 rocA mutation reduced GAS survival in human blood, and in vivo naso-pharyngeal carriage longevity in a murine model, with an associated drop in bacterial airborne transmission during infection. In summary, a naturally occurring truncation in a regulator explains the encapsulation phenotype, carriage longevity and transmissibility of M18 GAS, highlighting the close interrelation of metabolism, capsule and virulence

The changing nature of risk and risk management: the challenge of borders, uncertainty and resilience

No abstract available

Quality Assurance and its impact on ovarian visualisation rates in the multicentre United Kingdom Collaborative Trial of Ovarian Cancer Screening (UKCTOCS)

OBJECTIVE: To describe the quality assurance (QA) processes and their impact on visualisation of postmenopausal ovaries in the ultrasound arm of a multicentre ovarian cancer screening trial. METHODS: In UKCTOCS, 50,640 women aged 50-74 at recruitment were randomised to the ultrasound arm and underwent annual transvaginal scans. QA processes were developed during the course of the trial and included regular monitoring of Visualisation Rate (VR) of right ovary. Non-subjective factors previously identified as impacting on VR of right ovary were included in a generalised estimating equation(GEE) model for binary outcomes to enable comparison of observed versus adjusted VR between individual sonographers who had undertaken >1000scans on trial /centres. Analysis of annual VR of sonographers / centres was undertaken. RESULTS: Between June 2001 and December 2010, across 13 centres, 48230 (of 50639) women attended for 270035 annual transvaginal scans. One or both ovaries were seen in 84.5% (228145/270035) of scans. The observed VR of the right ovary was 72.7% (196426/270035). For the 78 sonographers included in the model, the median difference between observed and adjusted VR was 2% (range 0-8%) and median change in rank was 3 (range 0-18). For the 13 centres, the median difference between observed versus adjusted VR was 0% (range 0-2%) with no change in ranking. The median adjusted VR for sonographers was 73% (IQR 65-82%) and for centres was 74.7% (IQR 67.1-79.0%). Despite increasing age of the cohort, there was a steady decrease in the number of sonographers with VR80% (14.3% in 2002 to 40.8 % in 2010). Median centre VR increased from 65.5% (range 55.7-81.0%) in 2001 to 80.3% (range74.5%-90.9%) in 2010. CONCLUSIONS: A robust QA programme can improve visualisation of postmenopausal ovaries and is an essential component of ultrasound-based ovarian cancer screening trials. While VR should be adjusted for non-subjective factors that impact on ovarian visualisation, subjective factors are likely to be the largest contributors to VR differences

The Thermal Design, Characterization, and Performance of the SPIDER Long-Duration Balloon Cryostat

We describe the SPIDER flight cryostat, which is designed to cool six millimeter-wavelength telescopes during an Antarctic long-duration balloon flight. The cryostat, one of the largest to have flown on a stratospheric payload, uses liquid helium-4 to deliver cooling power to stages at 4.2 and 1.6 K. Stainless steel capillaries facilitate a high flow impedance connection between the main liquid helium tank and a smaller superfluid tank, allowing the latter to operate at 1.6 K as long as there is liquid in the 4.2 K main tank. Each telescope houses a closed cycle helium-3 adsorption refrigerator that further cools the focal planes down to 300 mK. Liquid helium vapor from the main tank is routed through heat exchangers that cool radiation shields, providing negative thermal feedback. The system performed successfully during a 17 day flight in the 2014-2015 Antarctic summer. The cryostat had a total hold time of 16.8 days, with 15.9 days occurring during flight.Comment: 15 pgs, 17 fig

Genetic dissection of photoperiod response based on GWAS of pre-anthesis phase duration in spring barley

Heading time is a complex trait, and natural variation in photoperiod responses is a major factor controlling time to heading, adaptation and grain yield. In barley, previous heading time studies have been mainly conducted under field conditions to measure total days to heading. We followed a novel approach and studied the natural variation of time to heading in a world-wide spring barley collection (218 accessions), comprising of 95 photoperiod-sensitive (Ppd-H1) and 123 accessions with reduced photoperiod sensitivity (ppd-H1) to long-day (LD) through dissecting pre-anthesis development into four major stages and sub-phases. The study was conducted under greenhouse (GH) conditions (LD; 16/8 h; ∼20/∼16°C day/night). Genotyping was performed using a genome-wide high density 9K single nucleotide polymorphisms (SNPs) chip which assayed 7842 SNPs. We used the barley physical map to identify candidate genes underlying genome-wide association scans (GWAS). GWAS for pre-anthesis stages/sub-phases in each photoperiod group provided great power for partitioning genetic effects on floral initiation and heading time. In addition to major genes known to regulate heading time under field conditions, several novel QTL with medium to high effects, including new QTL having major effects on developmental stages/sub-phases were found to be associated in this study. For example, highly associated SNPs tagged the physical regions around HvCO1 (barley CONSTANS1) and BFL (BARLEY FLORICAULA/LEAFY) genes. Based upon our GWAS analysis, we propose a new genetic network model for each photoperiod group, which includes several newly identified genes, such as several HvCO-like genes, belonging to different heading time pathways in barley

Probiotic-Derived Polyphosphate Enhances the Epithelial Barrier Function and Maintains Intestinal Homeostasis through Integrin–p38 MAPK Pathway

Probiotics exhibit beneficial effects on human health, particularly in the maintenance of intestinal homeostasis in a complex manner notwithstanding the diversity of an intestinal flora between individuals. Thus, it is highly probable that some common molecules secreted by probiotic and/or commensal bacteria contribute to the maintenance of intestinal homeostasis and protect the intestinal epithelium from injurious stimuli. To address this question, we aimed to isolate the cytoprotective compound from a lactobacillus strain, Lactobacillus brevis SBC8803 which possess the ability to induce cytoprotective heat shock proteins in mouse small intestine. L. brevis was incubated in MRS broth and the supernatant was passed through with a 0.2-µm filter. Caco2/bbe cells were treated with the culture supernatant, and HSP27 expression was evaluated by Western blotting. HSP27-inducible components were separated by ammonium sulfate precipitation, DEAE anion exchange chromatography, gel filtration, and HPLC. Finally, we identified that the HSP27-inducible fraction was polyphosphate (poly P), a simple repeated structure of phosphates, which is a common product of lactobacilli and other bacteria associated with intestinal microflora without any definitive physiological functions. Then, poly P was synthesized by poly P-synthesizing enzyme polyphosphate kinase. The synthesized poly P significantly induced HSP27 from Caco2/BBE cells. In addition, Poly P suppressed the oxidant-induced intestinal permeability in the mouse small intestine and pharmacological inhibitors of p38 MAPK and integrins counteract its protective effect. Daily intrarectal administration of poly P (10 µg) improved the inflammation grade and survival rate in 4% sodium dextran sulfate-administered mice. This study, for the first time, demonstrated that poly P is the molecule responsible for maintaining intestinal barrier actions which are mediated through the intestinal integrin β1-p38 MAPK

Steady-state modulation of voltage-gated K+ channels in rat arterial smooth muscle by cyclic AMP-dependent protein kinase and protein phosphatase 2B

Voltage-gated potassium channels (Kv) are important regulators of membrane potential in vascular smooth muscle cells, which is integral to controlling intracellular Ca2+ concentration and regulating vascular tone. Previous work indicates that Kv channels can be modulated by receptor-driven alterations of cyclic AMP-dependent protein kinase (PKA) activity. Here, we demonstrate that Kv channel activity is maintained by tonic activity of PKA. Whole-cell recording was used to assess the effect of manipulating PKA signalling on Kv and ATP-dependent K+ channels of rat mesenteric artery smooth muscle cells. Application of PKA inhibitors, KT5720 or H89, caused a significant inhibition of Kv currents. Tonic PKA-mediated activation of Kv appears maximal as application of isoprenaline (a β-adrenoceptor agonist) or dibutyryl-cAMP failed to enhance Kv currents. We also show that this modulation of Kv by PKA can be reversed by protein phosphatase 2B/calcineurin (PP2B). PKA-dependent inhibition of Kv by KT5720 can be abrogated by pre-treatment with the PP2B inhibitor cyclosporin A, or inclusion of a PP2B auto-inhibitory peptide in the pipette solution. Finally, we demonstrate that tonic PKA-mediated modulation of Kv requires intact caveolae. Pre-treatment of the cells with methyl-β-cyclodextrin to deplete cellular cholesterol, or adding caveolin-scaffolding domain peptide to the pipette solution to disrupt caveolae-dependent signalling each attenuated PKA-mediated modulation of the Kv current. These findings highlight a novel, caveolae-dependent, tonic modulatory role of PKA on Kv channels providing new insight into mechanisms and the potential for pharmacological manipulation of vascular tone

Organism-sediment interactions govern post-hypoxia recovery of ecosystem functioning

Hypoxia represents one of the major causes of biodiversity and ecosystem functioning loss for coastal waters. Since eutrophication-induced hypoxic events are becoming increasingly frequent and intense, understanding the response of ecosystems to hypoxia is of primary importance to understand and predict the stability of ecosystem functioning. Such ecological stability may greatly depend on the recovery patterns of communities and the return time of the system properties associated to these patterns. Here, we have examined how the reassembly of a benthic community contributed to the recovery of ecosystem functioning following experimentally-induced hypoxia in a tidal flat. We demonstrate that organism-sediment interactions that depend on organism size and relate to mobility traits and sediment reworking capacities are generally more important than recovering species richness to set the return time of the measured sediment processes and properties. Specifically, increasing macrofauna bioturbation potential during community reassembly significantly contributed to the recovery of sediment processes and properties such as denitrification, bedload sediment transport, primary production and deep pore water ammonium concentration. Such bioturbation potential was due to the replacement of the small-sized organisms that recolonised at early stages by large-sized bioturbating organisms, which had a disproportionately stronger influence on sediment. This study suggests that the complete recovery of organism-sediment interactions is a necessary condition for ecosystem functioning recovery, and that such process requires long periods after disturbance due to the slow growth of juveniles into adult stages involved in these interactions. Consequently, repeated episodes of disturbance at intervals smaller than the time needed for the system to fully recover organism-sediment interactions may greatly impair the resilience of ecosystem functioning.

Feasibility and willingness-to-pay for integrated community-based tuberculosis testing

BACKGROUND: Community-based screening for TB, combined with HIV and syphilis testing, faces a number of barriers. One significant barrier is the value that target communities place on such screening. METHODS: Integrated testing for TB, HIV, and syphilis was performed in neighborhoods identified using geographic information systems-based disease mapping. TB testing included skin testing and interferon gamma release assays. Subjects completed a survey describing disease risk factors, healthcare access, healthcare utilization, and willingness to pay for integrated testing. RESULTS: Behavioral and social risk factors among the 113 subjects were prevalent (71% prior incarceration, 27% prior or current crack cocaine use, 35% homelessness), and only 38% had a regular healthcare provider. The initial 24 subjects reported that they would be willing to pay a median $20 (IQR: 0-100) for HIV testing and$10 (IQR: 0-100) for TB testing when the question was asked in an open-ended fashion, but when the question was changed to a multiple-choice format, the next 89 subjects reported that they would pay a median $5 for testing, and 23$5 to get tested for TB, HIV, or syphilis. Among persons who received tuberculin skin testing, only 14/78 (18%) participants returned to have their skin tests read. Only 14/109 (13%) persons who underwent HIV testing returned to receive their HIV results. CONCLUSION: The relatively high-risk persons screened in this community outreach study placed low value on testing. Reported willingness to pay for such testing, while low, likely overestimated the true willingness to pay. Successful TB, HIV, and syphilis integrated testing programs in high risk populations will likely require one-visit diagnostic testing and incentives