A personalized biomedical risk assessment infographic for people who smoke with COPD: a qualitative study

Background Chronic obstructive pulmonary disease (COPD) causes 3 million deaths each year, yet 38% of COPD patients continue to smoke. Despite proof of effectiveness and universal guideline recommendations, smoking cessation interventions are underused in practice. We sought to develop an infographic featuring personalized biomedical risk assessment through future lung function decline prediction (with vs without ongoing smoking) to both prompt and enhance clinician delivery of smoking cessation advice and pharmacotherapy, and augment patient motivation to quit. Methods We recruited patients with COPD and pulmonologists from a quaternary care center in Toronto, Canada. Infographic prototype content and design was based on best evidence. After face validation, the prototype was optimized through rapid-cycle design. Each cycle consisted of: (1) infographic testing in a moderated focus group and a clinician interview (recorded/transcribed) (with questionnaire completion); (2) review of transcripts for emergent/critical findings; and (3) infographic modifications to address findings (until no new critical findings emerged). We performed iterative transcript analysis after each cycle and a summative qualitative transcript analysis with quantitative (descriptive) questionnaire analysis. Results Stopping criteria were met after 4 cycles, involving 20 patients (58% male) and 4 pulmonologists (50% male). The following qualitative themes emerged: Tool content (infographic content preferences); Tool Design (infographic design preferences); Advantages of Infographic Messaging (benefits of an infographic over other approaches); Impact of Tool on Determinants of Smoking Cessation Advice Delivery (impact on barriers and enablers to delivery of smoking cessation advice in practice); and Barriers and Enablers to Quitting (impact on barriers and enablers to quitting). Patient Likert scale ratings of infographic content and format/usability were highly positive, with improvements in scores for 20/21 questions through the design process. Providers scored the infographic at 77.8% (“superior”) on the Suitability Assessment of Materials questionnaire. Conclusions We developed a user preference-based personalized biomedical risk assessment infographic to drive smoking cessation in patients with COPD. Our findings suggest that this tool could impact behavioural determinants of provider smoking-cessation advice delivery, while increasing patient quit motivation. Impacts of the tool on provider care, patient motivation to quit, and smoking cessation success should now be evaluated in real-world settings.

Integrating Whole-Genome Sequencing in Clinical Genetics: A Novel Disruptive Structural Rearrangement Identified in the Dystrophin Gene (DMD)

While in most patients the identification of genetic alterations causing dystrophinopathies is a relatively straightforward task, a significant number require genomic and transcriptomic approaches that go beyond a routine diagnostic set-up. In this work, we present a Becker Muscular Dystrophy patient with elevated creatinine kinase levels, progressive muscle weakness, mild intellectual disability and a muscle biopsy showing dystrophic features and irregular dystrophin labelling. Routine molecular techniques (Southern-blot analysis, multiplex PCR, MLPA and genomic DNA sequencing) failed to detect a defect in the DMD gene. Muscle DMD transcript analysis (RT-PCR and cDNA-MLPA) showed the absence of exons 75 to 79, seen to be present at the genomic level. These results prompted the application of low-coverage linked-read whole-genome sequencing (WGS), revealing a possible rearrangement involving DMD intron 74 and a region located upstream of the PRDX4 gene. Breakpoint PCR and Sanger sequencing confirmed the presence of a ~8 Mb genomic inversion. Aberrant DMD transcripts were subsequently identified, some of which contained segments from the region upstream of PRDX4. Besides expanding the mutational spectrum of the disorder, this study reinforces the importance of transcript analysis in the diagnosis of dystrophinopathies and shows how WGS has a legitimate role in clinical laboratory genetics.

Error analysis of prospective mathematics teachers in solving of applying radian measurement problem in trigonometry courses

This research aims to describe the mistakes of the prospective teachers in solving the application of radian measurement problems and their causes. This type of research is qualitative descriptive research with a case study approach. The types of data collection in this research consisted of the results of the subject's work and transcripts of interviews with research subjects. By following the type of data, this research instrument consists of one question about the problem of applying radian measurement and interview guidelines developed by the researchers. Data analysis of the subject's work is carried out by classifying the types of errors to know the types of errors that arise in solving the problem of applying the radian measurements. While the transcript analysis of the interview results was carried out by coding the words to determine the factors causing the errors that appeared. The results of the research indicate that the error in solving the problem of applying the radian measurement are misconceptions and factual errors. This misconception is generally caused by intuitive thinking, while this factual error is generally caused by not paying careful attention to the information in the question. The solution to these errors is to analyze the elements of the circle that are interconnected in solving the problem of applying the radian measurements and to be careful in writing the information that is known in the question.

Characterisation of a putative M23-domain containing protein in Mycobacterium tuberculosis

Mycobacterium tuberculosis, the causative agent of tuberculosis remains a global health concern, further compounded by the high rates of HIV-TB co-infection and emergence of multi- and extensive drug resistant TB, all of which have hampered efforts to eradicate this disease. As a result, novel anti-tubercular interventions are urgently required, with the peptidoglycan component of the M. tuberculosis cell wall emerging as an attractive drug target. Peptidoglycan M23 endopeptidases can function as active cell wall hydrolases or degenerate activators of hydrolases in a variety of bacteria, contributing to important processes such as bacterial growth, division and virulence. Herein, we investigate the function of the Rv0950-encoded putative M23 endopeptidase in M. tuberculosis. In silico analysis revealed that this protein is conserved in mycobacteria, with a zinc-binding catalytic site predictive of hydrolytic activity. Transcript analysis indicated that expression of Rv0950c was elevated during lag and log phases of growth and reduced in stationary phase. Deletion of Rv0950c yielded no defects in growth, colony morphology, antibiotic susceptibility or intracellular survival but caused a reduction in cell length. Staining with a monopeptide-derived fluorescent D-amino acid, which spatially reports on sites of active PG biosynthesis or repair, revealed an overall reduction in uptake of the probe in ΔRv0950c. When stained with a dipeptide probe in the presence of cell wall damaging agents, the ΔRv0950c mutant displayed reduced sidewall labelling. As bacterial peptidoglycan metabolism is important for survival and pathogenesis, the role of Rv0950c and other putative M23 endopeptidases in M. tuberculosis should be explored further.

Transcript Profiling of MIKCc MADS-Box Genes Reveals Conserved and Novel Roles in Barley Inflorescence Development

MADS-box genes have a wide range of functions in plant reproductive development and grain production. The ABCDE model of floral organ development shows that MADS-box genes are central players in these events in dicotyledonous plants but the applicability of this model remains largely unknown in many grass crops. Here, we show that transcript analysis of all MIKCc MADS-box genes through barley (Hordeum vulgare L.) inflorescence development reveals co-expression groups that can be linked to developmental events. Thirty-four MIKCc MADS-box genes were identified in the barley genome and single-nucleotide polymorphism (SNP) scanning of 22,626 barley varieties revealed that the natural variation in the coding regions of these genes is low and the sequences have been extremely conserved during barley domestication. More detailed transcript analysis showed that MADS-box genes are generally expressed at key inflorescence developmental phases and across various floral organs in barley, as predicted by the ABCDE model. However, expression patterns of some MADS genes, for example HvMADS58 (AGAMOUS subfamily) and HvMADS34 (SEPALLATA subfamily), clearly deviate from predicted patterns. This places them outside the scope of the classical ABCDE model of floral development and demonstrates that the central tenet of antagonism between A- and C-class gene expression in the ABC model of other plants does not occur in barley. Co-expression across three correlation sets showed that specifically grouped members of the barley MIKCc MADS-box genes are likely to be involved in developmental events driving inflorescence meristem initiation, floral meristem identity and floral organ determination. Based on these observations, we propose a potential floral ABCDE working model in barley, where the classic model is generally upheld, but that also provides new insights into the role of MIKCc MADS-box genes in the developing barley inflorescence.

A Novel Liquid Biopsy (NETest) identifies Paragangliomas and Pheochromocytomas with High Accuracy

Pheochromocytomas and paragangliomas (PHEOs/PGLs) represent diagnostically challenging and complex neuroendocrine tumors (NETs). Current biomarker tests for PHEOs/PGLs are technically complex or limited. We assessed the diagnostic utility of a NET-specific 51-marker gene blood assay (NETest) in patients with PHEOs/PGLs (n=81), including 10 pediatric patients, and age-/gender-matched controls (n=142) using a prospective case:control (1:2) analysis. mRNA was measured (qPCR) and results scaled 0-100 (ULN<20). Receiver Operating Curve (ROC) and non-parametric (Mann-Whitney) were used for analyses (2-tailed). All data is presented as mean±SEM. NETest accuracy for PHEO/PGL diagnosis was 100%. PHEO/PGL scores were 70±3 versus 8.5±1 in controls (p<0.0001) and ROC analysis was 0.99±0.004 (p<0.0001). Diagnostic metrics were 94% accurate, 100% sensitive, and 92% specific. Imaging correlation with 68Ga-PET-SSA was 100%. NETest levels in PHEOs (n=26) were significantly (p<0.0001) elevated (83±4) versus 66±4 in PGLs (n=40) and mixed PHEOs/PGLs (n=5: 37±3). Adrenal-derived tumors (n=30) exhibited higher scores (76±5) than extra-adrenal-derived tumors (66±4, p<0.05). Cluster 2 tumors exhibited significantly (p=0.034) elevated NETest levels (n=4: 92±2) versus Cluster 1 tumors (n=35: 69±4). Regulatory pathway analysis identified elevated RAS-RAF, metastatic, pluripotential, neural and secretory gene cluster levels (p<0.05) in PHEOs compared to PGLs. Cluster 2 PPGLs exhibited elevated (p=0.046) levels of growth-factor signaling genes compared to cluster 1. The PHEOs/PGLs in the pediatric cohort (n=10) were all NETest-positive (81±8) and exhibited a gene expression profile spectrum analogous to adults. Circulating NET transcript analysis identifies PHEOs/PGLs with 100% efficacy and is likely to have clinical utility in the diagnosis and management of PHEO/PGL patients.

Osmo-priming in tomato seeds down-regulates genes associated with stress response and leads to reduction in longevity

Tomato seeds subjected to osmo-priming show fast and more uniform germination. However, osmo-priming reduces seed longevity, which is a complex seed physiological attribute influenced by several mechanisms, including response to stress. Thus, to have new insights as to why osmo-primed tomato seeds show a short life span, we performed a transcript analysis during their priming. For that, we performed gene expression studies of the heat-shock protein family genes that were previously reported to be associated with the enhancement of longevity in primed tomato seeds. Physiological assays of germination, vigour and longevity tests were used to support the data. The results show that the short life span of osmo-primed tomato seeds is related to the decrease in the expression of transcripts associated with response to stress during the priming treatment. These results are important because they add information regarding which seed longevity mechanisms are impacted by the priming treatment. In parallel, it will allow the use of these genes as markers to monitor longevity in osmo-primed tomato seeds.

Transcriptional insights into sugarcane aquaporin genes under water deficit conditions

Water deficit in soil during formative growth stage adversely hinders the crop productivity. Plant develop a key chain of mechanisms to cope these strains. Characterization of genotypes under water deficit will provide the basis for breeding new germplasm for efficient utilization of water and nutrients and adaptation to water stress. To achieve this, two tolerant (Co 98014 and Co 0118) and two sensitive (CoJ 85 and Co 89003) sugarcane genotypes were assessed for antioxidant responses followed by differential expression of three aquaporin genes (ShPIP2-1, ShPIP-5 and ShPIP2-6) under two water deficit conditions. The MDA and H2O2 contents were significantly higher (p<0.05) in sensitive genotypes as compared to tolerant ones, whereas SOD activity was higher (p<0.05) in tolerant than sensitive genotypes. The transcript analysis of AQPs reveals upregulation of ShPIP2;5, whereas down-regulation of ShPIP2;1 and ShPIP2;6 when plants were imposed to water deficit conditions. The findings under study suggested the role of PIP2 AQPs in regulation of plant water status under water deficit conditions.