Deep intronic variant causes aberrant splicing of ATP7A in a family with a variable occipital horn syndrome phenotype.

Genetic variants in ATP7A are associated with a spectrum of X-linked disorders. In descending order of severity, these are Menkes disease, occipital horn syndrome, and X-linked distal spinal muscular atrophy. After 30 years of diagnostic investigation, we identified a deep intronic ATP7A variant in four males from a family affected to variable degrees by a predominantly skeletal phenotype, featuring bowing of long bones, elbow joints with restricted mobility which dislocate frequently, coarse curly hair, chronic diarrhoea, and motor coordination difficulties. Analysis of whole genome sequencing data from the Genomics England 100,000 Genomes Project following clinical re-evaluation identified a deep intronic ATP7A variant, which was predicted by SpliceAI to have a modest splicing effect. Using a mini-gene splicing assay, we determined that the intronic variant results in aberrant splicing. Sanger sequencing of patient cDNA revealed ATP7A transcripts with exon 5 skipping, or inclusion of a novel intron 4 pseudoexon. In both instances, frameshift leading to premature termination are predicted. Quantification of ATP7A mRNA transcripts using a qPCR assay indicated that the majority of transcripts (86.1 %) have non-canonical splicing, with 68.0 % featuring exon 5 skipping, and 18.1 % featuring the novel pseudoexon. We suggest that the variability of the phenotypes within the affected males results from the stochastic effects of splicing. This deep intronic variant, resulting in aberrant ATP7A splicing, expands the understanding of intronic variation on the ATP7A-related disease spectrum.

Patient and ward related risk factors in a multi-ward nosocomial outbreak of COVID-19: Outbreak investigation and matched case-control study.

BACKGROUND: Risk factors for nosocomial COVID-19 outbreaks continue to evolve. The aim of this study was to investigate a multi-ward nosocomial outbreak of COVID-19 between 1st September and 15th November 2020, occurring in a setting without vaccination for any healthcare workers or patients. METHODS: Outbreak report and retrospective, matched case-control study using incidence density sampling in three cardiac wards in an 1100-bed tertiary teaching hospital in Calgary, Alberta, Canada. Patients were confirmed/probable COVID-19 cases and contemporaneous control patients without COVID-19. COVID-19 outbreak definitions were based on Public Health guidelines. Clinical and environmental specimens were tested by RT-PCR and as applicable quantitative viral cultures and whole genome sequencing were conducted. Controls were inpatients on the cardiac wards during the study period confirmed to be without COVID-19, matched to outbreak cases by time of symptom onset dates, age within ± 15 years and were admitted in hospital for at least 2 days. Demographics, Braden Score, baseline medications, laboratory measures, co-morbidities, and hospitalization characteristics were collected on cases and controls. Univariate and multivariate conditional logistical regression was used to identify independent risk factors for nosocomial COVID-19. RESULTS: The outbreak involved 42 healthcare workers and 39 patients. The strongest independent risk factor for nosocomial COVID-19 (IRR 3.21, 95% CI 1.47-7.02) was exposure in a multi-bedded room. Of 45 strains successfully sequenced, 44 (97.8%) were B.1.128 and differed from the most common circulating community lineages. SARS-CoV-2 positive cultures were detected in 56.7% (34/60) of clinical and environmental specimens. The multidisciplinary outbreak team observed eleven contributing events to transmission during the outbreak. CONCLUSIONS: Transmission routes of SARS-CoV-2 in hospital outbreaks are complex; however multi-bedded rooms play a significant role in the transmission of SARS-CoV-2.

Asymptomatic severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection in adults is uncommon using rigorous symptom characterization and follow-up in an acute-care adult hospital outbreak.

Asymptomatic coronavirus disease 2019 (COVID-19) has been reported as a significant driver of COVID-19 outbreaks. Our hospital ward outbreak analysis suggests that comprehensive symptoms and signs assessment, in combination with adequate follow-up, allows a more precise determination of COVID-19 symptoms. Asymptomatic infection was quite uncommon among adults in this setting.

Integration of molecular and physiological models to explain time of anthesis in wheat.

BACKGROUND AND AIMS: A model to predict anthesis time of a wheat plant from environmental and genetic information requires integration of current concepts in physiological and molecular biology. This paper describes the structure of an integrated model and quantifies its response mechanisms. METHODS: Literature was reviewed to formulate the components of the model. Detailed re-analysis of physiological observations are utilized from a previous publication by the second two authors. In this approach measurements of leaf number and leaf and primordia appearance of near isogenic lines of spring and winter wheat grown for different durations in different temperature and photoperiod conditions are used to quantify mechanisms and parameters to predict time of anthesis. KEY RESULTS: The model predicts the time of anthesis from the length of sequential phases: 1, embryo development; 2, dormant; 3, imbibed/emerging; 4, vegetative; 5, early reproductive; 6, pseudo-stem extension; and 7, ear development. Phase 4 ends with vernalization saturation (VS), Phase 5 with terminal spikelet (TS) and Phase 6 with flag leaf ligule appearance (FL). The durations of Phases 4 and 5 are linked to the expression of Vrn genes and are calculated in relation to change in Haun stage (HS) to account for the effects of temperature per se. Vrn1 must be expressed to sufficient levels for VS to occur. Vrn1 expression occurs at a base rate of 0·08/HS in winter 'Batten' and 0·17/HS in spring 'Batten' during Phases 1, 3 and 4. Low temperatures promote expression of Vrn1 and accelerate progress toward VS. Our hypothesis is that a repressor, Vrn4, must first be downregulated for this to occur. Rates of Vrn4 downregulation and Vrn1 upregulation have the same exponential response to temperature, but Vrn4 is quickly upregulated again at high temperatures, meaning short exposure to low temperature has no impact on the time of VS. VS occurs when Vrn1 reaches a relative expression of 0·76 and Vrn3 expression begins. However, Vrn2 represses Vrn3 expression so Vrn1 must be further upregulated to repress Vrn2 and enable Vrn3 expression. As a result, the target for Vrn1 to trigger VS was 0·76 in 8-h photoperiods (Pp) and increased at 0·026/HS under 16-h Pp as levels of Vrn2 increased. This provides a mechanism to model short-day vernalization. Vrn3 is expressed in Phase 5 (following VS), and apparent rates of Vrn3 expression increased from 0·15/HS at 8-h Pp to 0·33/HS at 16-h Pp. The final number of leaves is calculated as a function of the HS at which TS occurred (TS(HS)): 2·86 + 1·1 × TS(HS). The duration of Phase 6 is then dependent on the number of leaves left to emerge and how quickly they emerge. CONCLUSIONS: The analysis integrates molecular biology and crop physiology concepts into a model framework that links different developmental genes to quantitative predictions of wheat anthesis time in different field situations.

Designing new collaborative learning spaces in clinical environments: experiences from a children's hospital in Australia.

Hospitals are complex places that provide a rich learning environment for students, staff, patients and their families, professional groups and the community. The "new" Royal Children's Hospital opened in late 2011. Its mission is focused on improving health and well-being of children and adolescents through leadership in healthcare, research and education. Addressing the need to create "responsive learning environments" aligned with the shift to student-centred pedagogy, two distinct learning environments were developed within the new Royal Children's Hospital; (i) a dedicated education precinct providing a suite of physical environments to promote a more active, collaborative and social learning experience for education and training programs conducted on the Royal Children's Hospital campus and (ii) a suite of learning spaces embedded within clinical areas so that learning becomes an integral part of the daily activities of this busy Hospital environment. The aim of this article is to present the overarching educational principles that lead the design of these learning spaces and describe the opportunities and obstacles encountered in the development of collaborative learning spaces within a large hospital development.

Getting to 100%: a framework to define and reach target order entry rates.

Following a detailed review of orders entered into a clinical information system, we propose a framework to define computerized physician order entry types and a more useful formula for calculating order entry rate.

Optimizing standard patient management through order sets - impact on care (blood cultures).

We show that order set design and support must be thoughtful to result in improved quality of care and reduced waste and that order set use should be monitored to confirm expected impact and detect unanticipated consequences.

Physician adoption of an information system in an integrated health region.

We describe the strategies used to engage organizational and physician leadership through design, preparation, and support to achieve an inpatient Computerized Physician Order Entry (CPOE) rate over 70% by 1,700 physicians.

Post-endoscopy checklist reduces length of stay for non-variceal upper gastrointestinal bleeding.

OBJECTIVE: To examine the effect of improved gastroenterologist-to-admitting service communication on hospital stay for upper gastrointestinal bleeding. HYPOTHESIS: a detailed checklist addressing factors relevant to discharge planning would shorten hospital stay, when added to the procedure report. DESIGN: Pre-post intervention design, recording balance measures (potential confounders). SETTING: A Canadian university hospital. STUDY PARTICIPANTS: Intermittent 5- to 7-day batches of consecutive emergency patients presenting with non-variceal upper gastrointestinal bleeding as their primary problem. The durations of the background and intervention periods were 3 months (beginning 9 June 2003) and 4 weeks (beginning 8 September 2003), respectively. INTERVENTION: The gastrointestinal bleeding Quality Improvement and Health Information multidisciplinary team (quality improvement personnel; emergency physicians, hospitalists, gastroenterologists, in-patient and endoscopy nurses) developed a one-page checklist, outlining detailed recommendations (3-Ds-diet, drugs, discharge plan) to append to the procedure report. MAIN OUTCOME MEASURES: Difference in median length of hospital stay was the primary endpoint. As balance measures, demographics, bleeding severity, comorbidities, readmission rates, and various benchmark times were recorded prospectively. RESULTS: Thirty-nine patients met the criteria in the background period (4 months, intermittently sampled), and 22 in the intervention period (4 weeks, continuously sampled). There were no significant baseline differences. Median in-patient stay was 7.0 (95% interquartile range 2-24) versus 3.5 (95% interquartile range 1-12) days for the background and intervention periods, respectively (P = 0.003). This remained significant when outliers (stay > 10 days) were removed (P = 0.02). CONCLUSION: A checklist, with very specific recommendations to the admitting service, significantly reduced hospital stay for non-variceal gastrointestinal bleeding.

Gene organization and sequencing of the Choristoneura fumiferana defective nucleopolyhedrovirus genome.

Two distinct nucleopolyhedrovirus species of the eastern spruce budworm, Choristoneura fumiferana, exist in a symbiont-like relationship. C. fumiferana defective nucleopolyhedrovirus (CfDEFNPV) only infects C. fumiferana larvae per os in the presence of C. fumiferana nucleopolyhedrovirus Ireland strain (CfMNPV), but is infective when injected into the haemolymph. CfDEFNPV synergizes CfMNPV in per os infections and CfMNPV is always the predominant progeny. This study was undertaken to report the genomic makeup and organization of CfDEFNPV in an attempt to identify its defect and understand its synergistic role. The genome was mapped, sequenced, characterized and compared to other baculoviruses. The CfDEFNPV genome was 131,160 nt long with 149 putative open reading frames (ORFs) and a G + C content of 45.8 mol%. Homologues of all 62 conserved lepidopteran baculovirus genes were found including those implicated in per os infectivity, p74, per os infectivity factor (pif) and pif-2. Although no obvious deletions were observed to explain the defect, two ORFs, Cfdef79 and Cfdef99 (inhibitor of apoptosis-4), contained potential deletions. Cfdef50 (late expression factor-10)/Cfdef51 (vp1054) and Cfdef76/Cfdef77 (telokin-like protein) had large overlaps and a potential homologue to ac105/he65 was split. Four baculovirus repeat ORFs were present, as were two unique genes, but no enhancins were identified. CfDEFNPV contained 13 homologous regions, each with one to five palindromes. Comparison with fully sequenced baculovirus genomes identified CfDEFNPV as a group I NPV with the closest average amino acid identity to Epiphyas postvittana NPV, followed by Orgyia pseudotsugata MNPV and CfMNPV, with its closest matches being to individual Anticarsia gemmatalis MNPV gene sequences.

Modeling grain nitrogen accumulation and protein composition to understand the sink/source regulations of nitrogen remobilization for wheat.

A functional explanation for the regulation of grain nitrogen (N) accumulation in cereal by environmental and genetic factors remains elusive. Here, new mechanistic hypotheses of grain N accumulation are proposed and tested for wheat (Triticum aestivum). First, we tested experimentally the hypothesis that grain N accumulation is mostly source regulated. Four contrasting cultivars, in terms of their grain N concentrations and yield potentials, were grown with non-limiting N supply. Grain number per ear was reduced by removing the top part of the ear at anthesis. Reduction in grain number gave a significant increase in N content per grain for all cultivars, showing that grain N accumulation was source regulated. However, on a per ear basis, cultivars with a high grain number fully compensated their N accumulation for reduced grain number at anthesis. Cultivars with a lower grain number did not compensate completely, and grain N per ear was decreased by 16%. Second, new mechanistic hypotheses of the origins of grain N source regulation and its response to environment were tested by simulation. The hypotheses were: (a). The regulation by N sources of grain N accumulation applies only for the storage proteins (i.e. gliadin and glutenin fractions); (b). accumulation of structural and metabolic proteins (i.e. albumin-globulin and amphiphilic fractions) is sink-regulated; and (c). N partitioning between gliadins and glutenins is constant during grain development and unmodified by growing conditions. Comparison of experimental and simulation results of the accumulation of grain protein fractions under wide ranges of N fertilization, temperatures, and irrigation supported these hypotheses.