Development of RNA silencing as a novel disease control strategy to protect pines from Dothistroma needle blight : a thesis presented in the partial fulfilment of the requirements for the degree of Master of Science (MSc) in Genetics at Massey University, Manawatu, New Zealand

Permission was obtained for the re-use of Figures 1.1, 1.3 -1.8, 1.10, 1.11 & 3.1, and Tables 1.2 & 1.3. Figures 1.12 & 6.1 are re-used under a CC Attribution-NonCommercial-NoDerivatives 4.0 International (CC-BY-NC-ND 4.0) license. Figures 1.9, 2.9, 5.11 & 6.2 are re-used under a CC Attribution 4.0 International (CC BY 4.0) license.Pinus radiata, the main commercial forest species grown in New Zealand, is one of many pine species worldwide that are susceptible to Dothistroma needle blight (DNB), caused by the fungus Dothistroma septosporum. New methods are needed to help manage and control this disease over current control measures such as fungicide spraying, pruning, and thinning. RNA silencing is a radical new approach to directly combat pathogens. Due to the success of many studies in controlling agricultural and horticultural crops, by exogenously applying dsRNA molecules targeting virulence genes, this has raised the question of whether forest pathogens can also be controlled with this method. RNA silencing has the potential to silence genes specific to a fungal pathogen, rendering it less virulent, and reducing disease symptoms on affected host plants. The aims of this work were to create an RNA spray targeting individual genes specific to D. septosporum, and to determine if spray applications of the RNA can reduce pathogen virulence and protect pines from fungal infection. As proof of concept, a spray application of a 737 nt eGFP-dsRNA was used to target an enhanced green fluorescent protein gene in D. septosporum. Also, 509 nt and 408 nt DsAflR-dsRNAs were synthesised targeting two different regions of the dothistromin pathway regulatory protein gene, named DsAflR 1-dsRNA and DsAflR 2-dsRNA. The DsAflR gene is involved in the production of the virulence factor dothistromin. All three dsRNAs were labelled with fluorescein to detect its uptake into cells, which was successful. RNA silencing was detected by reduced gene expression levels in vitro for samples treated with DsAflR 1-, DsAflR 2-dsRNA, as well as the RNAi control eGFP. There was a statistically significant reduction in DsAflR gene expression by applying DsAflR 1-dsRNA to D. septosporum grown on agar; however not all the reductions seen for treatment with each respective dsRNA were statistically significant and a lot of variability was observed between replicates. In planta silencing trials, in which pine shoots were treated with dsRNA and inoculated with D. septosporum spores, revealed reductions in fungal biomass in dsRNA treated samples in some cases, although more replicates are needed to confirm these results. Nevertheless this study has contributed new knowledge for the development of spray applications of dsRNA to reduce DNB disease. It provides a starting point for more research in controlling forest pathogens and could ultimately help to replace existing chemical-based forest management practices. Furthermore, the knowledge gained is applicable to a diverse range of pathogens and plant hosts, for which this ground-breaking technology holds great promise

Hablamos Juntos (Together We Speak): Interpreters, Provider Communication, and Satisfaction with Care

The Hablamos Juntos—Together We Speak (HJ)—national demonstration project targeted the improvement of language access for Spanish-speaking Latinos in areas with rapidly growing Latino populations. The objective of HJ was to improve doctor-patient communication by increasing access to and quality of interpreter services for Spanish-speaking patients. To investigate how access to interpreters for adult Spanish-speaking Latinos is associated with ratings of doctor/office staff communication and satisfaction with care. Cross-sectional cohort study. A total of 1,590 Spanish-speaking Latino adults from eight sites across the United States who participated in the outpatient HJ evaluation. We analyzed two multi-item measures of doctor communication (4 items) and office staff helpfulness (2 items), and one global item of satisfaction with care by interpreter use. We performed regression analyses to control for patient sociodemographic characteristics, survey year, and clustering at the site of care. Ninety-five percent of participants were born outside the US, 81% were females, and survey response rates ranged from 45% to 85% across sites. In this cohort of Spanish-speaking patients, those who needed and always used interpreters reported better experiences with care than their counterparts who needed but had interpreters unavailable. Patients who always used an interpreter had better adjusted ratings of doctor communication [effect size (ES = 0.51)], office staff helpfulness (ES = 0.37), and satisfaction with care (ES = 0.37) than patients who needed but did not always use an interpreter. Patients who needed and always used interpreters also reported better experiences with care in all three domains measured [doctor communication (ES = 0.30), office staff helpfulness (ES = 0.21), and satisfaction with care (ES = 0.23)] than patients who did not need interpreters. Among adult Spanish-speaking Latinos, interpreter use is independently associated with higher satisfaction with doctor communication, office staff helpfulness, and ambulatory care. Increased attention to the need for effective interpreter services is warranted in areas with rapidly growing Spanish-speaking populations

Evaluation of a service intervention to improve awareness and uptake of bowel cancer screening in ethnically-diverse areas

The Policy Research Unit in Cancer Awareness, Screening and Early Diagnosis receives funding for a research programme from the UK Department of Health Policy Research Programme (grant no. 106/0001). It is a collaboration between researchers from seven institutions (the Queen Mary University of London, the UCL, the King’s College London, the London School of Hygiene and Tropical Medicine, the Hull York Medical School, the Durham University and the Peninsula Medical School)

Screening for colorectal cancer: possible improvements by risk assessment evaluation?

Emerging results indicate that screening improves survival of patients with colorectal cancer. Therefore, screening programs are already implemented or are being considered for implementation in Asia, Europe and North America. At present, a great variety of screening methods are available including colono- and sigmoidoscopy, CT- and MR-colonography, capsule endoscopy, DNA and occult blood in feces, and so on. The pros and cons of the various tests, including economic issues, are debated. Although a plethora of evaluated and validated tests even with high specificities and reasonable sensitivities are available, an international consensus on screening procedures is still not established. The rather limited compliance in present screening procedures is a significant drawback. Furthermore, some of the procedures are costly and, therefore, selection methods for these procedures are needed. Current research into improvements of screening for colorectal cancer includes blood-based biological markers, such as proteins, DNA and RNA in combination with various demographically and clinically parameters into a “risk assessment evaluation” (RAE) test. It is assumed that such a test may lead to higher acceptance among the screening populations, and thereby improve the compliances. Furthermore, the involvement of the media, including social media, may add even more individuals to the screening programs. Implementation of validated RAE and progressively improved screening methods may reform the cost/benefit of screening procedures for colorectal cancer. Therefore, results of present research, validating RAE tests, are awaited with interest

The effectiveness of computerized clinical guidelines in the process of care: a systematic review

<p>Abstract</p> <p>Background</p> <p>Clinical practice guidelines have been developed aiming to improve the quality of care. The implementation of the computerized clinical guidelines (CCG) has been supported by the development of computerized clinical decision support systems.</p> <p>This systematic review assesses the impact of CCG on the process of care compared with non-computerized clinical guidelines.</p> <p>Methods</p> <p>Specific features of CCG were studied through an extensive search of scientific literature, querying electronic databases: Pubmed/Medline, Embase and Cochrane Controlled Trials Register. A multivariable logistic regression was carried out to evaluate the association of CCG's features with positive effect on the process of care.</p> <p>Results</p> <p>Forty-five articles were selected. The logistic model showed that Automatic provision of recommendation in electronic version as part of clinician workflow (Odds Ratio [OR]= 17.5; 95% confidence interval [CI]: 1.6-193.7) and Publication Year (OR = 6.7; 95%CI: 1.3-34.3) were statistically significant predictors.</p> <p>Conclusions</p> <p>From the research that has been carried out, we can conclude that after implementation of CCG significant improvements in process of care are shown. Our findings also suggest clinicians, managers and other health care decision makers which features of CCG might improve the structure of computerized system.</p

Mobile Graphic-based Communication: Investigating Reminder Notifications to Support Tuberculosis Treatment in Africa

Visual communication is a method of communication using visual elements, which is suggested to be more effective than text or voice, and has the additional advantage that it, can also be used by who are unable to read. In this paper, the findings of a user requirements study, which was conducted at MnaziMmoja Hospital in Zanzibar, are presented. In a cross-sectional study, twenty nine people including TB patients and TB Health care workers were interviewed. The findings show that participants agreed that the use of mobile graphic-based communications could support TB patients in their treatment. The contribution of this work is the process to investigate and develop a new mobile graphic-based application for push notification services that are literacy-level and language agnostic

Living with diabetes: rationale, study design and baseline characteristics for an Australian prospective cohort study

Background: Diabetes mellitus is a major global public health threat. In Australia, as elsewhere, it is responsible for a sizeable portion of the overall burden of disease, and significant costs. The psychological and social impact of diabetes on individuals with the disease can be severe, and if not adequately addressed, can lead to the worsening of the overall disease picture. The Living With Diabetes Study aims to contribute to a holistic understanding of the psychological and social aspects of diabetes mellitus

Excessive Islet NO Generation in Type 2 Diabetic GK Rats Coincides with Abnormal Hormone Secretion and Is Counteracted by GLP-1

BACKGROUND: A distinctive feature of type 2 diabetes is inability of insulin-secreting beta-cells to properly respond to elevated glucose eventually leading to beta-cell failure. We have hypothesized that an abnormally increased NO production in the pancreatic islets might be an important factor in the pathogenesis of beta-cell dysfunction. PRINCIPAL FINDINGS: We show now that islets of type 2 spontaneous diabetes in GK rats display excessive NO generation associated with abnormal iNOS expression in insulin and glucagon cells, increased ncNOS activity, impaired glucose-stimulated insulin release, glucagon hypersecretion, and impaired glucose-induced glucagon suppression. Pharmacological blockade of islet NO production by the NOS inhibitor N(G)-nitro-L-arginine methyl ester (L-NAME) greatly improved hormone secretion from GK islets suggesting islet NOS activity being an important target to inactivate for amelioration of islet cell function. The incretin hormone GLP-1, which is used in clinical practice suppressed iNOS and ncNOS expression and activity with almost full restoration of insulin release and partial restoration of glucagon release. GLP-1 suppression of iNOS expression was reversed by PKA inhibition but unaffected by the proteasome inhibitor MG132. Injection of glucose plus GLP-1 in the diabetic rats showed that GLP-1 amplified the insulin response but induced a transient increase and then a poor depression of glucagon. CONCLUSION: The results suggest that abnormally increased NO production within islet cells is a significant player in the pathogenesis of type 2 diabetes being counteracted by GLP-1 through PKA-dependent, nonproteasomal mechanisms