Letters to the Editor

Software Safety vs Software Reliability While looking back through Vol. 56, No. 1 (Summer 2020) of Journal of System Safety, I finally took the time to read Nathaniel Ozarin’s article “Lessons Learned in a Complex Software Safety Program.” The article is quite interesting and thought provoking, comparing what actually occurs while implementing a system safety program to the idealized descriptions found in documents such as MIL-STD-882, JSSSEH and AOP-52. While I found the article interesting and informative, I noted that the author consistently characterizes the “software safety problem” as a “reliability” problem, focused on finding and preventing “failures” and ensuring high “reliability.” Some Thoughts on the Probabilistic Criteria for Ensuring Safe Airplane-System Designs We have been employed in the risk sciences for a total of 86 years, including 62 years in reliability engineering and safety engineering positions at The Boeing Company. For many of those years, Yellman was the designated “Risk-Analysis Focal” (person) for Boeing’s 707, 727, 737 and 757 airplane models. For several decades, the United States government has published the same criteria, created by the U.S. Federal Aviation Administration (FAA), intended to ensure that the systems on large (transport-category) aircraft have been designed to be safe [Refs. 1 and 2]. But we believe that the criteria have failed to prevent certain aircraft accidents, and we think that the reasons for that should be better understood. We hope that this discussion will contribute to a better understanding by examining the part potentially played in those accidents by the FAA’s criteria that are defined probabilistically

Hemodynamics during the 10-minute NASA Lean Test: evidence of circulatory decompensation in a subset of ME/CFS patients.

BACKGROUND: Lightheadedness, fatigue, weakness, heart palpitations, cognitive dysfunction, muscle pain, and exercise intolerance are some of the symptoms of orthostatic intolerance (OI). There is substantial comorbidity of OI in ME/CFS (Myalgic Encephalomyelitis/Chronic Fatigue Syndrome). The 10-minute NASA Lean Test (NLT) is a simple, point-of-care method that can aid ME/CFS diagnosis and guide management and treatment of OI. The objective of this study was to understand the hemodynamic changes that occur in ME/CFS patients during the 10-minute NLT. METHODS: A total of 150 ME/CFS patients and 75 age, gender and race matched healthy controls (HCs) were enrolled. We recruited 75 ME/CFS patients who had been sick for less than 4 years (\u3c 4 ME/CFS) and 75 ME/CFS patients sick for more than 10 years (\u3e 10 ME/CFS). The 10-minute NLT involves measurement of blood pressure and heart rate while resting supine and every minute for 10 min while standing with shoulder-blades on the wall for a relaxed stance. Spontaneously reported symptoms are recorded during the test. ANOVA and regression analysis were used to test for differences and relationships in hemodynamics, symptoms and upright activity between groups. RESULTS: At least 5 min of the 10-minute NLT were required to detect hemodynamic changes. The \u3c 4 ME/CFS group had significantly higher heart rate and abnormally narrowed pulse pressure compared to \u3e 10 ME/CFS and HCs. The \u3c 4 ME/CFS group experienced significantly more OI symptoms compared to \u3e 10 ME/CFS and HCs. The circulatory decompensation observed in the \u3c 4 ME/CFS group was not related to age or medication use. CONCLUSIONS: Circulatory decompensation characterized by increased heart rate and abnormally narrow pulse pressure was identified in a subgroup of ME/CFS patients who have been sick for \u3c 4 years. This suggests inadequate ventricular filling from low venous pressure. The 10-minute NLT can be used to diagnose and treat the circulatory decompensation in this newly recognized subgroup of ME/CFS patients. The \u3e 10 ME/CFS group had less pronounced hemodynamic changes during the NLT possibly from adaptation and compensation that occurs over time. The 10-minute NLT is a simple and clinically useful point-of-care method that can be used for early diagnosis of ME/CFS and help guide OI treatment

Efficient yeast ChIP-Seq using multiplex short-read DNA sequencing

<p>Abstract</p> <p>Background</p> <p>Short-read high-throughput DNA sequencing technologies provide new tools to answer biological questions. However, high cost and low throughput limit their widespread use, particularly in organisms with smaller genomes such as <it>S. cerevisiae</it>. Although ChIP-Seq in mammalian cell lines is replacing array-based ChIP-chip as the standard for transcription factor binding studies, ChIP-Seq in yeast is still underutilized compared to ChIP-chip. We developed a multiplex barcoding system that allows simultaneous sequencing and analysis of multiple samples using Illumina's platform. We applied this method to analyze the chromosomal distributions of three yeast DNA binding proteins (Ste12, Cse4 and RNA PolII) and a reference sample (input DNA) in a single experiment and demonstrate its utility for rapid and accurate results at reduced costs.</p> <p>Results</p> <p>We developed a barcoding ChIP-Seq method for the concurrent analysis of transcription factor binding sites in yeast. Our multiplex strategy generated high quality data that was indistinguishable from data obtained with non-barcoded libraries. None of the barcoded adapters induced differences relative to a non-barcoded adapter when applied to the same DNA sample. We used this method to map the binding sites for Cse4, Ste12 and Pol II throughout the yeast genome and we found 148 binding targets for Cse4, 823 targets for Ste12 and 2508 targets for PolII. Cse4 was strongly bound to all yeast centromeres as expected and the remaining non-centromeric targets correspond to highly expressed genes in rich media. The presence of Cse4 non-centromeric binding sites was not reported previously.</p> <p>Conclusion</p> <p>We designed a multiplex short-read DNA sequencing method to perform efficient ChIP-Seq in yeast and other small genome model organisms. This method produces accurate results with higher throughput and reduced cost. Given constant improvements in high-throughput sequencing technologies, increasing multiplexing will be possible to further decrease costs per sample and to accelerate the completion of large consortium projects such as modENCODE.</p

CDK-dependent nuclear localization of B-Cyclin Clb1 promotes FEAR activation during meiosis I in budding yeast

Cyclin-dependent kinases (CDK) are master regulators of the cell cycle in eukaryotes. CDK activity is regulated by the presence, post-translational modification and spatial localization of its regulatory subunit cyclin. In budding yeast, the B-cyclin Clb1 is phosphorylated and localizes to the nucleus during meiosis I. However the functional significance of Clb1's phosphorylation and nuclear localization and their mutual dependency is unknown. In this paper, we demonstrate that meiosis-specific phosphorylation of Clb1 requires its import to the nucleus but not vice versa. While Clb1 phosphorylation is dependent on activity of both CDK and polo-like kinase Cdc5, its nuclear localization requires CDK but not Cdc5 activity. Furthermore we show that increased nuclear localization of Clb1 during meiosis enhances activation of FEAR (Cdc Fourteen Early Anaphase Release) pathway. We discuss the significance of our results in relation to regulation of exit from meiosis I

Cognitive impairment in post-acute sequelae of COVID-19 and short duration myalgic encephalomyelitis patients is mediated by orthostatic hemodynamic changes

IntroductionCognitive impairment is experienced by people with myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) and post-acute sequelae of COVID-19 (PASC). Patients report difficulty remembering, concentrating, and making decisions. Our objective was to determine whether orthostatic hemodynamic changes were causally linked to cognitive impairment in these diseases.MethodsThis prospective, observational cohort study enrolled PASC, ME/CFS, and healthy controls. All participants underwent clinical evaluation and assessment that included brief cognitive testing before and after an orthostatic challenge. Cognitive testing measured cognitive efficiency which is defined as the speed and accuracy of subject’s total correct responses per minute. General linear mixed models were used to analyze hemodynamics and cognitive efficiency during the orthostatic challenge. Additionally, mediation analysis was used to determine if hemodynamic instability induced during the orthostatic challenge mediated the relationship between disease status and cognitive impairment.ResultsOf the 276 participants enrolled, 256 were included in this study (34 PASC, 71 &lt; 4 year duration ME/CFS, 69 &gt; 10 year ME/CFS duration, and 82 healthy controls). Compared to healthy controls, the disease cohorts had significantly lower cognitive efficiency scores immediately following the orthostatic challenge. Cognitive efficiency remained low for the &gt;10 year ME/CFS 2 and 7 days after orthostatic challenge. Narrow pulse pressure less than 25% of systolic pressure occurred at 4 and 5 min into the orthostatic challenge for the PASC and ME/CFS cohorts, respectively. Abnormally narrow pulse pressure was associated with slowed information processing in PASC patients compared to healthy controls (−1.5, p = 0.04). Furthermore, increased heart rate during the orthostatic challenge was associated with a decreased procedural reaction time in PASC and &lt; 4 year ME/CFS patients who were 40 to 65 years of age.DiscussionFor PASC patients, both their disease state and hemodynamic changes during orthostatic challenge were associated with slower reaction time and decreased response accuracy during cognitive testing. Reduced cognitive efficiency in &lt;4 year ME/CFS patients was associated with higher heart rate in response to orthostatic stress. Hemodynamic changes did not correlate with cognitive impairment for &gt;10 year ME/CFS patients, but cognitive impairment remained. These findings underscore the need for early diagnosis to mitigate direct hemodynamic and other physiological effects on symptoms of cognitive impairment

Spatial regulation of Cdc55–PP2A by Zds1/Zds2 controls mitotic entry and mitotic exit in budding yeast

Zds1/2 regulate mitotic progression by directing the nucleocytoplasmic distribution of Cdc55–PP2A

Meiotic nuclear divisions in budding yeast require PP2ACdc55-mediated antagonism of Net1 phosphorylation by Cdk

During meiosis, one round of deoxyribonucleic acid replication is followed by two rounds of nuclear division. In Saccharomyces cerevisiae, activation of the Cdc14 early anaphase release (FEAR) network is required for exit from meiosis I but does not lead to the activation of origins of replication. The precise mechanism of how FEAR regulates meiosis is not understood. In this paper, we report that premature activation of FEAR during meiosis caused by loss of protein phosphatase PP2ACdc55 activity blocks bipolar spindle assembly and nuclear divisions. In cdc55 meiotic null (cdc55-mn) cells, the cyclin-dependent kinase (Cdk)–counteracting phosphatase Cdc14 was released prematurely from the nucleolus concomitant with hyperphosphorylation of its nucleolar anchor protein Net1. Crucially, a mutant form of Net1 that lacks six Cdk phosphorylation sites rescued the meiotic defect of cdc55-mn cells. Expression of a dominant mutant allele of CDC14 mimicked the cdc55-mn phenotype. We propose that phosphoregulation of Net1 by PP2ACdc55 is essential for preventing precocious exit from meiosis I

A Mathematical Model of Mitotic Exit in Budding Yeast: The Role of Polo Kinase

Cell cycle progression in eukaryotes is regulated by periodic activation and inactivation of a family of cyclin–dependent kinases (Cdk's). Entry into mitosis requires phosphorylation of many proteins targeted by mitotic Cdk, and exit from mitosis requires proteolysis of mitotic cyclins and dephosphorylation of their targeted proteins. Mitotic exit in budding yeast is known to involve the interplay of mitotic kinases (Cdk and Polo kinases) and phosphatases (Cdc55/PP2A and Cdc14), as well as the action of the anaphase promoting complex (APC) in degrading specific proteins in anaphase and telophase. To understand the intricacies of this mechanism, we propose a mathematical model for the molecular events during mitotic exit in budding yeast. The model captures the dynamics of this network in wild-type yeast cells and 110 mutant strains. The model clarifies the roles of Polo-like kinase (Cdc5) in the Cdc14 early anaphase release pathway and in the G-protein regulated mitotic exit network

Elevated Levels of the Vesicular Monoamine Transporter and a Novel Repetitive Behavior in the Drosophila Model of Fragile X Syndrome

Fragile X Syndrome (FXS) is characterized by mental impairment and autism in humans, and it often features hyperactivity and repetitive behaviors. The mechanisms for the disease, however, remain poorly understood. Here we report that the dfmr1 mutant in the Drosophila model of FXS grooms excessively, which may be regulated differentially by two signaling pathways. Blocking metabotropic glutamate receptor signaling enhances grooming in dfmr1 mutant flies, whereas blocking the vesicular monoamine transporter (VMAT) suppresses excessive grooming. dfmr1 mutant flies also exhibit elevated levels of VMAT mRNA and protein. These results suggest that enhanced monoamine signaling correlates with repetitive behaviors and hyperactivity associated with FXS

A case study in estimating avionics availability from field reliability data

Under incentivized contractual mechanisms such as availability-based contracts the support service provider and its customer must share a common understanding of equipment reliability baselines. Emphasis is typically placed on the Information Technology-related solutions for capturing, processing and sharing vast amounts of data. In the case of repairable fielded items scant attention is paid to the pitfalls within the modelling assumptions that are often endorsed uncritically, and seldom made explicit during field reliability data analysis. This paper presents a case study in which good practices in reliability data analysis are identified and applied to real-world data with the aim of supporting the effective execution of a defence avionics availability-based contract. The work provides practical guidance on how to make a reasoned choice between available models and methods based on the intelligent exploration of the data available in practical industrial applications