Examining c-di-GMP and possible quorum sensing regulation in Pseudomonas fluorescens SBW25:links between intra and inter-cellular regulation benefits community cooperative activities such as biofilm formation

Bacterial success in colonizing complex environments requires individual response to micro-scale conditions as well as community-level cooperation to produce large-scale structures such as biofilms. Connecting individual and community responses could be achieved by linking the intracellular sensory and regulatory systems mediated by bis-(3′-5′)-cyclic dimeric guanosine monophosphate (c-di-GMP) and other compounds of individuals with intercellular quorum sensing (QS) regulation controlling populations. There is growing evidence to suggest that biofilm formation by many pseudomonads is regulated by both intra and intercellular systems, though in the case of the model Pseudomonas fluorescens SBW25 Wrinkly Spreader in which mutations increasing c-di-GMP levels result in the production of a robust cellulose-based air-liquid interface biofilm, no evidence for the involvement of QS regulation has been reported. However, our recent review of the P. fluorescens SBW25 genome has identified a potential QS regulatory pathway and other QS–associated genes linked to c-di-GMP homeostasis, and QS signal molecules have also been identified in culture supernatants. These findings suggest a possible link between c-di-GMP and QS regulation in P. fluorescens SBW25 which might allow a more sophisticated and responsive control of cellulose production and biofilm formation when colonising the soil and plant-associated environments P. fluorescens SBW25 normally inhabits.Анализ ц-ди-ГМФ и возможного чувства кворума у Pseudomonas fluorescens SBW 25: связь между внутри и межклеточной регуляцией способствует кооперативному поведению в сообществе и формированию биоплёнкиУспешность бактериальной колонизации сложных экониш требует индивидуального ответа на изменения условий на микроуровне равно как и кооперации на уровне сообщества для продукции таких крупно масштабных структур как биоплёнки. Координация индивидуальных ответ ов и ответов сообщества может быть достигнута путем связывания внутриклеточных сенсорных и регуляторных систем, опосредуемых бис-(3',5')-циклическим димерным гуанозинмонофосфатом (ц-ди-ГМФ) и другими соединениями индивидуумов с межклеточной регуляцией - чувством кворума (ЧК), контролирующем популяци ю. Накапливается всё больше доказательств того, что формирование биопленки многими псевдомонадами регулируется как внутри клеточными, так и меж клеточными регуляторными системами, хотя в случае модельной Pseudomonas fluorescens SBW25 Wrinkly Spreader, у которой мутации, повышающ ие уровни ц-ди-ГМФ, приводят к созданию прочной целлюлозной биоплёнки на границе раздела фаз воздух-жидкость, не было обнаружено ни ка кого свидетельства вовлечения кворум-зависимой регуляции. Однако наш недавний обзор генома P. fluorescens SBW25 выявил потенциальный ЧК-зависимый регуляторный пу ть и другие ЧК-зависимые гены, связанные с гомеостазом ц-ди-ГМФ, а молекулы ЧК-сигналинга были идентифицированы в культуре. Эти данные свидетельствуют о возможной связи между ц-ди-ГМФ-регуляцией и ЧК у P. fluorescens SBW25, что позволяет более сложный и гибкий контроль над продукцией целлюлозы и образовани ем биопленки при колонизации почв и экониш, aссоциированных с растениям и, - естественными средами обитания P. fluorescens SBW25

New metric reconstruction scheme for gravitational self-force calculations

Inspirals of stellar-mass objects into massive black holes will be important sources for the space-based gravitational-wave detector LISA. Modelling these systems requires calculating the metric perturbation due to a point particle orbiting a Kerr black hole. Currently, the linear perturbation is obtained with a metric reconstruction procedure that puts it in a "no-string" radiation gauge which is singular on a surface surrounding the central black hole. Calculating dynamical quantities in this gauge involves a subtle procedure of "gauge completion" as well as cancellations of very large numbers. The singularities in the gauge also lead to pathological field equations at second perturbative order. In this paper we re-analyze the point-particle problem in Kerr using the corrector-field reconstruction formalism of Green, Hollands, and Zimmerman (GHZ). We clarify the relationship between the GHZ formalism and previous reconstruction methods, showing that it provides a simple formula for the "gauge completion". We then use it to develop a new method of computing the metric in a more regular gauge: a Teukolsky puncture scheme. This scheme should ameliorate the problem of large cancellations, and by constructing the linear metric perturbation in a sufficiently regular gauge, it should provide a first step toward second-order self-force calculations in Kerr. Our methods are developed in generality in Kerr, but we illustrate some key ideas and demonstrate our puncture scheme in the simple setting of a static particle in Minkowski spacetime

Priming winter wheat seeds with the bacterial quorum sensing signal N-hexanoyl-L-homoserine lactone (C6-HSL) shows potential to improve plant growth and seed yield

Several model plants are known to respond to bacterial quorum sensing molecules with altered root growth and gene expression patterns and induced resistance to plant pathogens. These compounds may represent novel elicitors that could be applied as seed primers to enhance cereal crop resistance to pathogens and abiotic stress and to improve yields. We investigated whether the acyl-homoserine lactone N-hexanoyl-L-homoserine lactone (C6-HSL) impacted winter wheat (Triticum aestivum L.) seed germination, plant development and productivity, using two Ukrainian varieties, Volodarka and Yatran 60, in both in vitro experiments and field trials. In vitro germination experiments indicated that C6-HSL seed priming had a small but significant positive impact on germination levels (1.2x increase, p < 0.0001), coleoptile and radicle development (1.4x increase, p < 0.0001). Field trials over two growing seasons (2015-16 and 2016-17) also demonstrated significant improvements in biomass at the tillering stage (1.4x increase, p < 0.0001), and crop structure and productivity at maturity including grain yield (1.4 – 1.5x increase, p < 0.0007) and quality (1.3x increase in good grain, p < 0.0001). In some cases variety effects were observed (p ≤ 0.05) suggesting that the effect of C6-HSL seed priming might depend on plant genetics, and some benefits of priming were also evident in F1 plants grown from seeds collected the previous season (p ≤ 0.05). These field-scale findings suggest that bacterial acyl-homoserine lactones such as C6-HSL could be used to improve cereal crop growth and yield and reduce reliance on fungicides and fertilisers to combat pathogens and stress

Invalid party wall awards and how to avoid them

Considers the reasons for the invalidity of party wall awards. Examines decided cases under earlier party wall legislation in the context of the Party Wall etc. Act 1996. Explains invalidity on the basis of an excess of the surveyors’ statutory authority. Defines this authority in terms of jurisdiction and power. Demonstrates the limits of the surveyors’ authority and emphasises the importance of strict compliance with statutory procedures. Concludes that surveyors should adopt an inquisitive and analytical approach to the scope of their authority to avoid the possibility of invalid awards. Echoes John Anstey’s earlier warning that surveyors should avoid a broad-brush approach to their duties which will only leave them “covered in soot”

Chapter 3: Mental health treatment and services

In this chapter reported use of psychotropic medication and psychological therapy are examined, as well as the extent of use of health care services for a mental health reason (GP, inpatient and outpatient health care) and day and community service use. It should be noted that rates presented are based on participant self-reports, not health records. Misclassifications of type of treatment or service are possible, and which was the providing organisation was not established

Study of diffusion weighted MRI as a predictive biomarker of response during radiotherapy for high and intermediate risk squamous cell cancer of the oropharynx: The MeRInO study

Introduction and background: A significant proportion of patients with intermediate and high risk squamous cell cancer of the oropharynx (OPSCC) continue to relapse locally despite radical chemoradiotherapy (CRT). The toxicity of the current combination of intensified dose per fraction radiotherapy and platinum based chemotherapy limits further uniform intensification. If a predictive biomarker for outcomes from CRT can be identified during treatment then individualised and adaptive treatment strategies may be employed. Methods/design: The MeRInO study is a prospective observational imaging study of patients with intermediate and high risk, locally advanced OPSCC receiving radical RT or concurrent CRT Patients undergo diffusion weighted MRI prior to treatment (MRI_1) and during the third week of RT (MRI_2). Apparent diffusion coefficient (ADC) measurements will be made on each scan for previously specified target lesions (primary and lymph nodes) and change in ADC calculated. Patients will be followed up and disease status for each target lesion noted. The primary aim of the MeRInO study is to determine the threshold change in ADC from baseline to week 3 of RT that may identify the sub-group of non-responders during treatment. Discussion: The use of DW-MRI as a predictive biomarker during RT for SCC H&N is in its infancy but studies to date have found that response to treatment may indeed be predicted by comparison of DW-MRI carried out before and during treatment. However, previous studies have included all sub-sites and biological sub-types. Establishing ADC thresholds that predict for local failure is an essential step towards using DW-MRI to improve the therapeutic ratio in treating SCC H&N. This would be done most robustly in a specific H&N sub-site and in sub-types with similar biological behaviour. The MeRInO study will help establish these thresholds in OPSCC

A Phase IB open-label, dose-escalation study of NUC 1031 in combination with carboplatin for recurrent ovarian cancer

Funding: The study was funded and the investigational drug NUC-1031 was supplied by NuCana plc.Purpose: NUC-1031 is a first-in-class ProTide modification of gemcitabine. In PRO-002, NUC‑1031 was combined with carboplatin in recurrent ovarian cancer (OC). Experimental Design: NUC-1031 was administered on days 1 & 8 with carboplatin on day 1 every 3 weeks for up to 6 cycles. Four dose cohorts of NUC-1031 (500, 625 and 750 mg/m2) with carboplatin (AUC4 or 5) were investigated. Primary endpoint was RP2CD. Secondary endpoints included safety, investigator-assessed objective response rate (ORR), clinical benefit rate (CBR), progression-free survival (PFS) and pharmacokinetics (PK). Results: 25 women with recurrent OC, a mean of 3.8 prior lines of chemotherapy and a median platinum-free interval (PFI) of 5 months (range: 7 - 451 days) were enrolled, 15/25 (60%) platinum-resistant; 9 (36%) partially platinum-sensitive and 1 (4%) platinum-sensitive. Of the 23 response-evaluable: there was 1 confirmed complete response (CR, 4%), 5 partial responses (PR, 17%) and 8 (35%) stable disease (SD). The ORR was 26% and CBR was 74% across all doses and 100% in the RP2CD cohort. Median PFS was 27.1 weeks. NUC-1031 was stable in the plasma and rapidly generated high intracellular dFdCTP levels that were unaffected by carboplatin. Conclusions: NUC-1031 combined with carboplatin is well tolerated in recurrent OC. Highest efficacy was observed at the RP2CD of 500 mg/m2 NUC-1031 on days 1 & 8 with AUC5 carboplatin day 1, every 3 weeks for 6 cycles. The ability to deliver carboplatin at AUC5 and the efficacy of this schedule even in patients with platinum-resistant disease makes this an attractive therapeutic combination.PostprintPeer reviewe

Prevalence of psychosis in black ethnic minorities in Britain: analysis based on three national surveys

Purpose A considerable excess of psychosis in black ethnic minorities is apparent from clinical studies, in Britain, as in other developed economies with white majority populations. This excess is not so marked in population surveys. Equitable health service provision should be informed by the best estimates of the excess. We used national survey data to establish the difference in the prevalence of psychosis between black ethnic groups and the white majority in the British general population. Methods Analysis of the combined datasets (N = 26,091) from the British national mental health surveys of 1993, 2000 and 2007. Cases of psychosis were determined either by the use of the Schedules for Clinical Assessment in Neuropsychiatry (SCAN), or from a combination of screening items. We controlled for sex, age, social class, unemployment, design features and other putative confounders, using a Disease Risk Score. Results People from black ethnic minorities had an excess prevalence rate of psychosis compared with the white majority population. The OR, weighted for study design and response rate, was 2.72 (95 % CI 1.3–5.6, p = 0.002). This was marginally increased after controlling for potential confounders (OR = 2.90, 95 % CI 1.4–6.2, p = 0.006). Conclusions The excess of psychosis in black ethnic minority groups was similar to that in two previous British community surveys, and less than that based on clinical studies. Even so it confirms a considerable need for increased mental health service resources in areas with high proportions of black ethnic minority inhabitants

eDNA inactivation and biofilm inhibition by the polymeric biocide polyhexamethylene guanidine hydrochloride (PHMG-Cl)

The choice of effective biocides used for routine hospital practice should consider the role of disinfectants in the maintenance and development of local resistome and how they might affect antibiotic resistance gene transfer within the hospital microbial population. Currently, there is little understanding of how different biocides contribute to eDNA release that may contribute to gene transfer and subsequent environmental retention. Here, we investigated how different biocides affect the release of eDNA from mature biofilms of two opportunistic model strains Pseudomonas aeruginosa ATCC 27853 (PA) and Staphylococcus aureus ATCC 25923 (SA) and contribute to the hospital resistome in the form of surface and water contaminants and dust particles. The effect of four groups of biocides, alcohols, hydrogen peroxide, quaternary ammonium compounds, and the polymeric biocide polyhexamethylene guanidine hydrochloride (PHMG-Cl), was evaluated using PA and SA biofilms. Most biocides, except for PHMG-Cl and 70% ethanol, caused substantial eDNA release, and PHMG-Cl was found to block biofilm development when used at concentrations of 0.5% and 0.1%. This might be associated with the formation of DNA–PHMG-Cl complexes as PHMG-Cl is predicted to bind to AT base pairs by molecular docking assays. PHMG-Cl was found to bind high-molecular DNA and plasmid DNA and continued to inactivate DNA on surfaces even after 4 weeks. PHMG-Cl also effectively inactivated biofilm-associated antibiotic resistance gene eDNA released by a pan-drug-resistant Klebsiella strain, which demonstrates the potential of a polymeric biocide as a new surface-active agent to combat the spread of antibiotic resistance in hospital settings

Blood RNA analysis can increase clinical diagnostic rate and resolve variants of uncertain significance

Purpose Diagnosis of genetic disorders is hampered by large numbers of variants of uncertain significance (VUSs) identified through next-generation sequencing. Many such variants may disrupt normal RNA splicing. We examined effects on splicing of a large cohort of clinically identified variants and compared performance of bioinformatic splicing prediction tools commonly used in diagnostic laboratories. Methods Two hundred fifty-seven variants (coding and noncoding) were referred for analysis across three laboratories. Blood RNA samples underwent targeted reverse transcription polymerase chain reaction (RT-PCR) analysis with Sanger sequencing of PCR products and agarose gel electrophoresis. Seventeen samples also underwent transcriptome-wide RNA sequencing with targeted splicing analysis based on Sashimi plot visualization. Bioinformatic splicing predictions were obtained using Alamut, HSF 3.1, and SpliceAI software. Results Eighty-five variants (33%) were associated with abnormal splicing. The most frequent abnormality was upstream exon skipping (39/85 variants), which was most often associated with splice donor region variants. SpliceAI had greatest accuracy in predicting splicing abnormalities (0.91) and outperformed other tools in sensitivity and specificity. Conclusion Splicing analysis of blood RNA identifies diagnostically important splicing abnormalities and clarifies functional effects of a significant proportion of VUSs. Bioinformatic predictions are improving but still make significant errors. RNA analysis should therefore be routinely considered in genetic disease diagnostics