The stealth episome: suppression of gene expression on the excised genomic island PPHGI-1 from Pseudomonas syringae pv. phaseolicola

Pseudomonas syringae pv. phaseolicola is the causative agent of halo blight in the common bean, Phaseolus vulgaris. P. syringae pv. phaseolicola race 4 strain 1302A contains the avirulence gene avrPphB (syn. hopAR1), which resides on PPHGI-1, a 106 kb genomic island. Loss of PPHGI-1 from P. syringae pv. phaseolicola 1302A following exposure to the hypersensitive resistance response (HR) leads to the evolution of strains with altered virulence. Here we have used fluorescent protein reporter systems to gain insight into the mobility of PPHGI-1. Confocal imaging of dual-labelled P. syringae pv. phaseolicola 1302A strain, F532 (dsRFP in chromosome and eGFP in PPHGI-1), revealed loss of PPHGI-1::eGFP encoded fluorescence during plant infection and when grown in vitro on extracted leaf apoplastic fluids. Fluorescence-activated cell sorting (FACS) of fluorescent and non-fluorescent PPHGI-1::eGFP F532 populations showed that cells lost fluorescence not only when the GI was deleted, but also when it had excised and was present as a circular episome. In addition to reduced expression of eGFP, quantitative PCR on sub-populations separated by FACS showed that transcription of other genes on PPHGI-1 (avrPphB and xerC) was also greatly reduced in F532 cells harbouring the excised PPHGI-1::eGFP episome. Our results show how virulence determinants located on mobile pathogenicity islands may be hidden from detection by host surveillance systems through the suppression of gene expression in the episomal state

RNA secondary structure prediction from multi-aligned sequences

It has been well accepted that the RNA secondary structures of most functional non-coding RNAs (ncRNAs) are closely related to their functions and are conserved during evolution. Hence, prediction of conserved secondary structures from evolutionarily related sequences is one important task in RNA bioinformatics; the methods are useful not only to further functional analyses of ncRNAs but also to improve the accuracy of secondary structure predictions and to find novel functional RNAs from the genome. In this review, I focus on common secondary structure prediction from a given aligned RNA sequence, in which one secondary structure whose length is equal to that of the input alignment is predicted. I systematically review and classify existing tools and algorithms for the problem, by utilizing the information employed in the tools and by adopting a unified viewpoint based on maximum expected gain (MEG) estimators. I believe that this classification will allow a deeper understanding of each tool and provide users with useful information for selecting tools for common secondary structure predictions.Comment: A preprint of an invited review manuscript that will be published in a chapter of the book `Methods in Molecular Biology'. Note that this version of the manuscript may differ from the published versio

Сучасний стан і проблеми управління залізничним транспортом України

Проаналізовано стан і тенденції розвитку залізничного транспорту. Розглянуті основні завдання державного регулювання галузі.Проанализировано состояние и тенденции развития железнодорожного транспорта. Рассмотренны основные задания государственного регулирования отрасли.The condition and trends of railway transport has been anilized. The main tasks of state regulation of railway transport has been considereted

Display of probability densities for data from a continuous distribution

Based on cumulative distribution functions, Fourier series expansion and Kolmogorov tests, we present a simple method to display probability densities for data drawn from a continuous distribution. It is often more efficient than using histograms.Comment: 5 pages, 4 figures, presented at Computer Simulation Studies XXIV, Athens, GA, 201

Effectiveness of a parenting programme in a public health setting: a randomised controlled trial of the positive parenting programme (Triple P) level 3 versus care as usual provided by the preventive child healthcare (PCH)

BACKGROUND: Considering the high burden of disease of psychosocial problems in children and adolescents, early intervention regarding problem behaviour of young children is very important. The Preventive Child Healthcare (PCH) offers a good setting to detect such problem behaviour and to provide parenting support to the parents concerned. This paper aims to describe the design of an effectiveness study of a parenting programme for parents of children with mild psychosocial problems after an initial, evidence based screening in routine PCH. METHODS/DESIGN: The effects of the intervention will be studied in a randomised controlled trial. Prior to a routine PCH health examination, parents complete a screening questionnaire on psychosocial problems. Parents of children with increased but still subclinical levels of psychosocial problems will be assigned at random to the experimental group (Triple P, level 3) or to the control group (care as usual). Outcome measures, such as problem behaviour in the child and parenting behaviour, will be assessed before, directly after and 6 and 12 months after the intervention. DISCUSSION: Parenting support may be an effective intervention to reduce psychosocial problems in children but evidence-based parenting programmes that fit the needs of the PCH are not available as yet. Although the Triple P programme seems promising and suitable for a universal population approach, evidence on its effectiveness in routine PCH still lacks. TRIAL REGISTRATION: NTR1338

Arterivirus Nsp1 Modulates the Accumulation of Minus-Strand Templates to Control the Relative Abundance of Viral mRNAs

The gene expression of plus-strand RNA viruses with a polycistronic genome depends on translation and replication of the genomic mRNA, as well as synthesis of subgenomic (sg) mRNAs. Arteriviruses and coronaviruses, distantly related members of the nidovirus order, employ a unique mechanism of discontinuous minus-strand RNA synthesis to generate subgenome-length templates for the synthesis of a nested set of sg mRNAs. Non-structural protein 1 (nsp1) of the arterivirus equine arteritis virus (EAV), a multifunctional regulator of viral RNA synthesis and virion biogenesis, was previously implicated in controlling the balance between genome replication and sg mRNA synthesis. Here, we employed reverse and forward genetics to gain insight into the multiple regulatory roles of nsp1. Our analysis revealed that the relative abundance of viral mRNAs is tightly controlled by an intricate network of interactions involving all nsp1 subdomains. Distinct nsp1 mutations affected the quantitative balance among viral mRNA species, and our data implicate nsp1 in controlling the accumulation of full-length and subgenome-length minus-strand templates for viral mRNA synthesis. The moderate differential changes in viral mRNA abundance of nsp1 mutants resulted in similarly altered viral protein levels, but progeny virus yields were greatly reduced. Pseudorevertant analysis provided compelling genetic evidence that balanced EAV mRNA accumulation is critical for efficient virus production. This first report on protein-mediated, mRNA-specific control of nidovirus RNA synthesis reveals the existence of an integral control mechanism to fine-tune replication, sg mRNA synthesis, and virus production, and establishes a major role for nsp1 in coordinating the arterivirus replicative cycle

Modelling the impact of improving screening and treatment of chronic hepatitis C virus infection on future hepatocellular carcinoma rates and liver-related mortality.

BACKGROUND: The societal, clinical and economic burden imposed by the complications of chronic hepatitis C virus (HCV) infection - including cirrhosis and hepatocellular carcinoma (HCC) - is expected to increase over the coming decades. However, new therapies may improve sustained virological response (SVR) rates and shorten treatment duration. This study aimed to estimate the future burden of HCV-related disease in England if current management strategies remain the same and the impact of increasing diagnosis and treatment of HCV as new therapies become available. METHODS: A previously published model was adapted for England using published literature and government reports, and validated through an iterative process of three meetings of HCV experts. The impact of increasing diagnosis and treatment of HCV as new therapies become available was modelled and compared to the base-case scenario of continuing current management strategies. To assess the 'best case' clinical benefit of new therapies, the number of patients treated was increased by a total of 115% by 2018. RESULTS: In the base-case scenario, total viraemic (HCV RNA-positive) cases of HCV in England will decrease from 144,000 in 2013 to 76,300 in 2030. However, due to the slow progression of chronic HCV, the number of individuals with cirrhosis, decompensated cirrhosis and HCC will continue to increase over this period. The model suggests that the 'best case' substantially reduces HCV-related hepatic disease and HCV-related liver mortality by 2020 compared to the base-case scenario. The number of HCV-related HCC cases would decrease 50% by 2020 and the number progressing from infection to decompensated cirrhosis would decline by 65%. Therefore, compared to projections of current practices, increasing treatment numbers by 115% by 2018 would reduce HCV-related mortality by 50% by 2020. CONCLUSIONS: This analysis suggests that with current treatment practices the number of patients developing HCV-related cirrhosis, decompensated cirrhosis and HCC will increase substantially, with HCV-related liver deaths likely to double by 2030. However, increasing diagnosis and treatment rates could optimise the reduction in the burden of disease produced by the new therapies, potentially halving HCV-related liver mortality and HCV-related HCC by 2020

Differential lipid dependence of the function of bacterial sodium channels

The lipid bilayer is important for maintaining the integrity of cellular compartments and plays a vital role in providing the hydrophobic and charged interactions necessary for membrane protein structure, conformational flexibility and function. To directly assess the lipid dependence of activity for voltage-gated sodium channels, we compared the activity of three bacterial sodium channel homologues (NaChBac, NavMs, and NavSp) by cumulative 22Na+ uptake into proteoliposomes containing a 3:1 ratio of 1-palmitoyl 2-oleoyl phosphatidylethanolamine and different “guest” glycerophospholipids. We observed a unique lipid profile for each channel tested. NavMs and NavSp showed strong preference for different negatively-charged lipids (phosphatidylinositol and phosphatidylglycerol, respectively), whilst NaChBac exhibited a more modest variation with lipid type. To investigate the molecular bases of these differences we used synchrotron radiation circular dichroism spectroscopy to compare structures in liposomes of different composition, and molecular modeling and electrostatics calculations to rationalize the functional differences seen. We then examined pore-only constructs (with voltage sensor subdomains removed) and found that in these channels the lipid specificity was drastically reduced, suggesting that the specific lipid influences on voltage-gated sodium channels arise primarily from their abilities to interact with the voltage-sensing subdomains

Metabolomics of aging assessed in individual parasitoid wasps

Metabolomics studies of low-biomass organisms, such as small insects, have previously relied on the pooling of biological samples to overcome detection limits, particularly using NMR. We show that the differentiation of metabolite profiles of individual 1 mg parasitoid wasps of different ages is possible when using a modified sample preparation and a combination of untargeted NMR and LC-MS based metabolomics. Changes were observed between newly emerged and older wasps in glycerolipids, amino acids and circulatory sugars. This advance in chemical profiling has important implications for the study of the behaviour and ecology of parasitoids and many other species of small organisms because predictions and observations are typically made at the level of the individual. Thus, the metabolomic state of low-biomass individuals can now be related to their behaviour and ecological performance. We discuss specifically the utility of age-related metabolomic profiling but our new approach can be applied to a wide range of biological research