SnTox3 Acts in Effector Triggered Susceptibility to Induce Disease on Wheat Carrying the Snn3 Gene

The necrotrophic fungus Stagonospora nodorum produces multiple proteinaceous host-selective toxins (HSTs) which act in effector triggered susceptibility. Here, we report the molecular cloning and functional characterization of the SnTox3-encoding gene, designated SnTox3, as well as the initial characterization of the SnTox3 protein. SnTox3 is a 693 bp intron-free gene with little obvious homology to other known genes. The predicted immature SnTox3 protein is 25.8 kDa in size. A 20 amino acid signal sequence as well as a possible pro sequence are predicted. Six cysteine residues are predicted to form disulfide bonds and are shown to be important for SnTox3 activity. Using heterologous expression in Pichia pastoris and transformation into an avirulent S. nodorum isolate, we show that SnTox3 encodes the SnTox3 protein and that SnTox3 interacts with the wheat susceptibility gene Snn3. In addition, the avirulent S. nodorum isolate transformed with SnTox3 was virulent on host lines expressing the Snn3 gene. SnTox3-disrupted mutants were deficient in the production of SnTox3 and avirulent on the Snn3 differential wheat line BG220. An analysis of genetic diversity revealed that SnTox3 is present in 60.1% of a worldwide collection of 923 isolates and occurs as eleven nucleotide haplotypes resulting in four amino acid haplotypes. The cloning of SnTox3 provides a fundamental tool for the investigation of the S. nodorum–wheat interaction, as well as vital information for the general characterization of necrotroph–plant interactions

Genetic diversity and population structure of Ascochyta rabiei from the western Iranian Ilam and Kermanshah provinces using MAT and SSR markers

Knowledge of genetic diversity in A. rabiei provides different levels of information that are important in the management of crop germplasm resources. Gene flow on a regional level indicates a significant potential risk for the regional spread of novel alleles that might contribute to fungicide resistance or the breakdown of resistance genes. Simple sequence repeat (SSR) and mating type (MAT) markers were used to determine the genetic structure, and estimate genetic diversity and the prevalence of mating types in 103 Ascochyta rabiei isolates from seven counties in the Ilam and Kermanshah provinces of western Iran (Ilam, Aseman abad, Holaylan, Chardavol, Dareh shahr, Gilangharb, and Sarpul). A set of 3 microsatellite primer pairs revealed a total of 75 alleles; the number of alleles varied from 15 to 34 for each marker. A high level of genetic variability was observed among A. rabiei isolates in the region. Genetic diversity was high (He = 0.788) within populations with corresponding high average gene flow and low genetic distances between populations. The smallest genetic distance was observed between isolates from Ilam and Chardavol. Both mating types were present in all populations, with the majority of the isolates belonging to Mat1-1 (64%), but within populations the proportions of each mating type were not significantly different from 50%. Results from this study will be useful in breeding for Ascochyta blight-resistant cultivars and developing necessary control measures

A global perspective on marine photosynthetic picoeukaryote community structure

A central goal in ecology is to understand the factors affecting the temporal dynamics and spatial distribution of microorganisms and the underlying processes causing differences in community structure and composition. However, little is known in this respect for photosynthetic picoeukaryotes (PPEs), algae that are now recognised as major players in marine CO2 fixation. Here, we analysed dot blot hybridisation and cloning–sequencing data, using the plastid-encoded 16S rRNA gene, from seven research cruises that encompassed all four ocean biomes. We provide insights into global abundance, α- and β-diversity distribution and the environmental factors shaping PPE community structure and composition. At the class level, the most commonly encountered PPEs were Prymnesiophyceae and Chrysophyceae. These taxa displayed complementary distribution patterns, with peak abundances of Prymnesiophyceae and Chrysophyceae in waters of high (25:1) or low (12:1) nitrogen:phosphorus (N:P) ratio, respectively. Significant differences in phylogenetic composition of PPEs were demonstrated for higher taxonomic levels between ocean basins, using Unifrac analyses of clone library sequence data. Differences in composition were generally greater between basins (interbasins) than within a basin (intrabasin). These differences were primarily linked to taxonomic variation in the composition of Prymnesiophyceae and Prasinophyceae whereas Chrysophyceae were phylogenetically similar in all libraries. These data provide better knowledge of PPE community structure across the world ocean and are crucial in assessing their evolution and contribution to CO2 fixation, especially in the context of global climate change

Multi-centre parallel arm randomised controlled trial to assess the effectiveness and cost-effectiveness of a group-based cognitive behavioural approach to managing fatigue in people with multiple sclerosis

Abstract (provisional) Background Fatigue is one of the most commonly reported and debilitating symptoms of multiple sclerosis (MS); approximately two-thirds of people with MS consider it to be one of their three most troubling symptoms. It may limit or prevent participation in everyday activities, work, leisure, and social pursuits, reduce psychological well-being and is one of the key precipitants of early retirement. Energy effectiveness approaches have been shown to be effective in reducing MS-fatigue, increasing self-efficacy and improving quality of life. Cognitive behavioural approaches have been found to be effective for managing fatigue in other conditions, such as chronic fatigue syndrome, and more recently, in MS. The aim of this pragmatic trial is to evaluate the clinical and cost-effectiveness of a recently developed group-based fatigue management intervention (that blends cognitive behavioural and energy effectiveness approaches) compared with current local practice. Methods This is a multi-centre parallel arm block-randomised controlled trial (RCT) of a six session group-based fatigue management intervention, delivered by health professionals, compared with current local practice. 180 consenting adults with a confirmed diagnosis of MS and significant fatigue levels, recruited via secondary/primary care or newsletters/websites, will be randomised to receive the fatigue management intervention or current local practice. An economic evaluation will be undertaken alongside the trial. Primary outcomes are fatigue severity, self-efficacy and disease-specific quality of life. Secondary outcomes include fatigue impact, general quality of life, mood, activity patterns, and cost-effectiveness. Outcomes in those receiving the fatigue management intervention will be measured 1 week prior to, and 1, 4, and 12 months after the intervention (and at equivalent times in those receiving current local practice). A qualitative component will examine what aspects of the fatigue management intervention participants found helpful/unhelpful and barriers to change. Discussion This trial is the fourth stage of a research programme that has followed the Medical Research Council guidance for developing and evaluating complex interventions. What makes the intervention unique is that it blends cognitive behavioural and energy effectiveness approaches. A potential strength of the intervention is that it could be integrated into existing service delivery models as it has been designed to be delivered by staff already working with people with MS. Service users will be involved throughout this research. Trial registration: Current Controlled Trials ISRCTN7651747

The Cysteine Rich Necrotrophic Effector SnTox1 Produced by Stagonospora nodorum Triggers Susceptibility of Wheat Lines Harboring Snn1

The wheat pathogen Stagonospora nodorum produces multiple necrotrophic effectors (also called host-selective toxins) that promote disease by interacting with corresponding host sensitivity gene products. SnTox1 was the first necrotrophic effector identified in S. nodorum, and was shown to induce necrosis on wheat lines carrying Snn1. Here, we report the molecular cloning and validation of SnTox1 as well as the preliminary characterization of the mechanism underlying the SnTox1-Snn1 interaction which leads to susceptibility. SnTox1 was identified using bioinformatics tools and verified by heterologous expression in Pichia pastoris. SnTox1 encodes a 117 amino acid protein with the first 17 amino acids predicted as a signal peptide, and strikingly, the mature protein contains 16 cysteine residues, a common feature for some avirulence effectors. The transformation of SnTox1 into an avirulent S. nodorum isolate was sufficient to make the strain pathogenic. Additionally, the deletion of SnTox1 in virulent isolates rendered the SnTox1 mutated strains avirulent on the Snn1 differential wheat line. SnTox1 was present in 85% of a global collection of S. nodorum isolates. We identified a total of 11 protein isoforms and found evidence for strong diversifying selection operating on SnTox1. The SnTox1-Snn1 interaction results in an oxidative burst, DNA laddering, and pathogenesis related (PR) gene expression, all hallmarks of a defense response. In the absence of light, the development of SnTox1-induced necrosis and disease symptoms were completely blocked. By comparing the infection processes of a GFP-tagged avirulent isolate and the same isolate transformed with SnTox1, we conclude that SnTox1 may play a critical role during fungal penetration. This research further demonstrates that necrotrophic fungal pathogens utilize small effector proteins to exploit plant resistance pathways for their colonization, which provides important insights into the molecular basis of the wheat-S. nodorum interaction, an emerging model for necrotrophic pathosystems

Crystallographic Tomography and Molecular Modelling of Structured Organic Polycrystalline Powders

A fundamental understanding of the behaviour of polycrystalline materials, including pharmaceuticals, is vital for control of their physicochemical and crystalline properties, which in turn has the potential to improve drug product development for example. In this work, attenuation X-ray computed tomography (CT) and diffraction contrast tomography (DCT) are combined with molecular modelling to understand the powder packing behaviour and crystal interactions of the organic cubic compound hexamine (hexamethylenetetramine). It is the first application of DCT to polycrystalline organic materials. The crystal morphology is predicted through synthonic modelling, with fully 3D-resolved confirmation of the crystallography of the external {110} facets, edges and corner directions by DCT. Analysis of the powder-bed reveals agglomerate structures and orientational texture, with its chemical origins energetically predicted to be face-to-face in accordance with the experimental data. Finally, measurements of crystal & crystallite interactions provide evidence for different mechanisms of powder bed agglomeration

Expression of chemokine receptors on peripheral blood lymphocytes in multiple sclerosis and neuromyelitis optica

<p>Abstract</p> <p>Background</p> <p>The role of different chemokine receptors in the pathogenesis of multiple sclerosis (MS) has been extensively investigated; however, little is known about the difference in the role of chemokine receptors between the pathogenesis of neuromyelitis optica (NMO) and MS. Therefore, we examined the expression of chemokine receptors on peripheral blood lymphocytes (PBL) in MS and NMO.</p> <p>Methods</p> <p>We used flow cytometry to analyse lymphocyte subsets in 12 patients with relapsing NMO, 24 with relapsing-remitting MS during relapse, 3 with NMO and 5 with MS during remission.</p> <p>Results</p> <p>Compared with healthy controls (HC), the percentage of lymphocytes in white blood cells was significantly lower in NMO and MS patients. The percentage of T cells expressing CD4⁺CD25⁺and CD4⁺CD45RO⁺was higher, while that of CD4⁺CC chemokine receptor (CCR)3⁺(T helper 2, Th2) was significantly lower in MS patients than in HC. The ratios of CD4⁺CXC chemokine receptors (CXCR)3⁺/CD4⁺CCR3⁺(Th1/Th2) and CD8⁺CXCR3⁺/CD8⁺CCR4⁺(T cytotoxic 1, Tc1/Tc2) were higher in MS patients than in HC. The percentage of CD8⁺CXCR3⁺T cell (Tc1) and CD4⁺CXCR3⁺T cell (Th1) decreased significantly during remission in MS patients (<it>P <</it>0.05). No significant differences were identified in the expression of the chemokine receptors on PBL in NMO patients compared with MS patients and HC.</p> <p>Conclusions</p> <p>Th1 dominance of chemokine receptors on blood T cells and the correlation between CXCR3⁺T cell (Th1 and Tc1) and disease activity in MS patients were confirmed by analysing chemokines receptors on PBL. In contrast, deviation in the Th1/Th2 balance was not observed in NMO patients.</p

Organization and Biology of the Porcine Serum Amyloid A (SAA) Gene Cluster: Isoform Specific Responses to Bacterial Infection.

Serum amyloid A (SAA) is a prominent acute phase protein. Although its biological functions are debated, the wide species distribution of highly homologous SAA proteins and their uniform behavior in response to injury or inflammation in itself suggests a significant role for this protein. The pig is increasingly being used as a model for the study of inflammatory reactions, yet only little is known about how specific SAA genes are regulated in the pig during acute phase responses and other responses induced by pro-inflammatory host mediators. We designed SAA gene specific primers and quantified the gene expression of porcine SAA1, SAA2, SAA3, and SAA4 by reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) in liver, spleen, and lung tissue from pigs experimentally infected with the Gram-negative swine specific bacterium Actinobacillus pleuropneumoniae, as well as from pigs experimentally infected with the Gram-positive bacterium Staphylococcus aureus. Our results show that: 1) SAA1 may be a pseudogene in pigs; 2) we were able to detect two previously uncharacterized SAA transcripts, namely SAA2 and SAA4, of which the SAA2 transcript is primarily induced in the liver during acute infection and presumably contributes to circulating SAA in pigs; 3) Porcine SAA3 transcription is induced both hepatically and extrahepatically during acute infection, and may be correlated to local organ affection; 4) Hepatic transcription of SAA4 is markedly induced in pigs infected with A. pleuropneumoniae, but only weakly in pigs infected with S. aureus. These results for the first time establish the infection response patterns of the four porcine SAA genes which will be of importance for the use of the pig as a model for human inflammatory responses, e.g. within sepsis, cancer, and obesity research

To respond or not to respond - a personal perspective of intestinal tolerance

For many years, the intestine was one of the poor relations of the immunology world, being a realm inhabited mostly by specialists and those interested in unusual phenomena. However, this has changed dramatically in recent years with the realization of how important the microbiota is in shaping immune function throughout the body, and almost every major immunology institution now includes the intestine as an area of interest. One of the most important aspects of the intestinal immune system is how it discriminates carefully between harmless and harmful antigens, in particular, its ability to generate active tolerance to materials such as commensal bacteria and food proteins. This phenomenon has been recognized for more than 100 years, and it is essential for preventing inflammatory disease in the intestine, but its basis remains enigmatic. Here, I discuss the progress that has been made in understanding oral tolerance during my 40 years in the field and highlight the topics that will be the focus of future research