Myosin-X facilitates Shigella-induced membrane protrusions and cell-to-cell spread.

The intracellular pathogen Shigella flexneri forms membrane protrusions to spread from cell to cell. As protrusions form, myosin-X (Myo10) localizes to Shigella. Electron micrographs of immunogold-labelled Shigella-infected HeLa cells reveal that Myo10 concentrates at the bases and along the sides of bacteria within membrane protrusions. Time-lapse video microscopy shows that a full-length Myo10 GFP-construct cycles along the sides of Shigella within the membrane protrusions as these structures progressively lengthen. RNAi knock-down of Myo10 is associated with shorter protrusions with thicker stalks, and causes a >80% decrease in confluent cell plaque formation. Myo10 also concentrates in membrane protrusions formed by another intracellular bacteria, Listeria, and knock-down of Myo10 also impairs Listeria plaque formation. In Cos7 cells (contain low concentrations of Myo10), the expression of full-length Myo10 nearly doubles Shigella-induced protrusion length, and lengthening requires the head domain, as well as the tail-PH domain, but not the FERM domain. The GFP-Myo10-HMM domain localizes to the sides of Shigella within membrane protrusions and the GFP-Myo10-PH domain localizes to host cell membranes. We conclude thatMyo10 generates the force to enhance bacterial-induced protrusions by binding its head region to actin filaments and its PH tail domain to the peripheral membrane.

Anthrax lethal and edema toxins fail to directly impair human platelet function.

Hemorrhage is a prominent clinical manifestation of systemic anthrax. Therefore, we have examined the effects of anthrax lethal and edema toxins on human platelets. We find that anthrax lethal toxin fails to cleave its target, mitogen-activated protein kinase 1, and anthrax edema toxin fails to increase intracellular cyclic adenosine monophosphate. Surface expression of toxin receptors tumor endothelial marker 8 and capillary morphogenesis gene 2, as well as coreceptor low density lipoprotein receptor-related protein 6 (LRP6), are markedly reduced, preventing toxin binding to platelets. Our studies suggest that the hemorrhagic clinical manifestations of systemic anthrax are unlikely to be caused by the direct binding and entry of anthrax toxins into human platelets.

Home Monitoring for Fever: An Inexpensive Screening Method to Prevent Household Spread of COVID-19.

The COVID-19 pandemic surge has exceeded testing capacities in many parts of the world. We investigated the effectiveness of home temperature monitoring for early identification of COVID-19 patients. Study Design ­: We compared home temperature measurements from a convenience sample of 1180 individuals who reported being test positive for SARS-CoV-2 to an age, sex, and location matched control group of 1249 individuals who had not tested positive. Methods ­: All individuals monitored their temperature at home using an electronic smartphone thermometer that relayed temperature measurements and symptoms to a centralized cloud based, de-identified data bank. Results -: Individuals varied in the number of times they monitored their temperature. When temperature was monitored for over 72 hours fever (≥ 37.6°C or 99.7°F or a change in temperature of ≥ 1°C or 1.8°F) was detected in 73% of test positive individuals, a sensitivity comparable to rapid SARS-CoV-2 antigen tests. When compared to our control group the specificity of fever for COVID-19 was 0.70. However, when fever was combined with complaints of loss of taste and smell, difficulty breathing, fatigue, chills, diarrhea, or stuffy nose the odds ratio of having COVID-19 was sufficiently high as to obviate the need to employ RTPCR or antigen testing to screen for and isolate coronavirus infected cases. Conclusions -: Our findings suggest that home temperature monitoring could serve as an inexpensive convenient screen for the onset of COVID-19, encourage earlier isolation of potentially infected individuals, and more effectively reduce the spread of infection in closed spaces.

Lessons in clinical reasoning - pitfalls, myths, and pearls: a case of crushing, substernal chest pain.

OBJECTIVES: Diagnostic error is not uncommon and diagnostic accuracy can be improved with the use of problem representation, pre-test probability, and Bayesian analysis for improved clinical reasoning. CASE PRESENTATION: A 48-year-old female presented as a transfer from another Emergency Department (ED) to our ED with crushing, substernal pain associated with dyspnea, diaphoresis, nausea, and a tingling sensation down both arms with radiation to the back and neck. Troponins were elevated along with an abnormal electrocardiogram. A negative myocardial perfusion scan led to the patient's discharge. The patient presented to the ED 10 days later with an anterior ST-elevation myocardial infarction. CONCLUSIONS: An overemphasis on a single testing modality led to diagnostic error and a severe event. The use of pre-test probabilities guided by history-taking can lead to improved interpretation of test results, ultimately improving diagnostic accuracy and preventing serious medical errors.

Bacillus anthracis' lethal toxin induces broad transcriptional responses in human peripheral monocytes.

BACKGROUND: Anthrax lethal toxin (LT), produced by the Gram-positive bacterium Bacillus anthracis, is a highly effective zinc dependent metalloprotease that cleaves the N-terminus of mitogen-activated protein kinase kinases (MAPKK or MEKs) and is known to play a role in impairing the host immune system during an inhalation anthrax infection. Here, we present the transcriptional responses of LT treated human monocytes in order to further elucidate the mechanisms of LT inhibition on the host immune system. RESULTS: Western Blot analysis demonstrated cleavage of endogenous MEK1 and MEK3 when human monocytes were treated with 500 ng/mL LT for four hours, proving their susceptibility to anthrax lethal toxin. Furthermore, staining with annexin V and propidium iodide revealed that LT treatment did not induce human peripheral monocyte apoptosis or necrosis. Using Affymetrix Human Genome U133 Plus 2.0 Arrays, we identified over 820 probe sets differentially regulated after LT treatment at the p <0.001 significance level, interrupting the normal transduction of over 60 known pathways. As expected, the MAPKK signaling pathway was most drastically affected by LT, but numerous genes outside the well-recognized pathways were also influenced by LT including the IL-18 signaling pathway, Toll-like receptor pathway and the IFN alpha signaling pathway. Multiple genes involved in actin regulation, signal transduction, transcriptional regulation and cytokine signaling were identified after treatment with anthrax LT. CONCLUSION: We conclude LT directly targets human peripheral monocytes and causes multiple aberrant gene responses that would be expected to be associated with defects in human monocyte's normal signaling transduction pathways and function. This study provides further insights into the mechanisms associated with the host immune system collapse during an anthrax infection, and suggests that anthrax LT may have additional downstream targets outside the well-known MAPK pathway.

Applying Lean principles to create a high throughput mass COVID-19 vaccination site.

A high throughput COVID-19 vaccination site was created using Lean principles and tools. Mass-vaccination sites can achieve high output by creating a standard physical design for workspaces and standardised work protocols, and by timing each step in the vaccination process to create a value stream map that can identify and remove all wasteful steps. Reliability of the vaccination process can be assured by creating a visual checklist that monitors the individual steps as well as by building in second checks by downstream personnel. Finally, productivity can be closely monitored by recording the start and completion time for each vaccination and plotting run charts. With 78 personnel working efficiently and effectively together, a maximum throughput of 5024 injections over 10 hours was achieved. As compared with other published COVID-19 mass-vaccination sites, our site attained threefold-fourfold higher productivity. We share our approach to encourage others to reproduce our vaccination system.

Prospective application of the interdisciplinary bedside rounding checklist 'TEMP' is associated with reduced infections and length of hospital stay.

Protocols that enhance communication between nurses, physicians and patients have had a variable impact on the quality and safety of patient care. We combined standardised nursing and physician interdisciplinary bedside rounds with a mnemonic checklist to assure all key nursing care components were modified daily. The mnemonic TEMP allowed the rapid review of 11 elements. T stands for tubes assuring proper management of intravenous lines and foleys; E stands for eating, exercise, excretion and sleep encouraging a review of orders for diet, exercise, laxatives to assure regular bowel movements, and inquiry about sleep; M stands for monitoring reminding the team to review the need for telemetry and the frequency of vital sign monitoring as well as the need for daily blood tests; and P stands for pain and plans reminding the team to discuss pain medications and to review the management plan for the day with the patient and family. Faithful implementation eliminated central line-associated bloodstream infections and catheter-associated urinary tract infections and resulted in a statistically significant reduction in average hospital length of stay of 13.3 hours, one unit achieving a 23-hour reduction. Trends towards reduced 30-day readmissions (20% down to 10%-11%) were observed. One unit improved the percentage of patients who reported nurses and doctors always worked together as a team from a 56% baseline to 75%. However, the combining of both units failed to demonstrate statistically significant improvement. Psychologists well versed in implementing behavioural change were recruiting to improve adherence to our protocols. Following training physicians and nurses achieved adherence levels of over 70%. A high correlation (r2=0.69) between adherence and reductions in length of stay was observed emphasising the importance of rigorous training and monitoring of performance to bring about meaningful and reliable improvements in the efficiency and quality of patient care.

Neuraxial and peripheral misconnection events leading to wrong-route medication errors: a comprehensive literature review.

We conducted a search of the literature to identify case reports of neuraxial and peripheral nervous system misconnection events leading to wrong-route medication errors. This narrative review covers a 20-year period (1999-2019; English-language publications and abstracts) and included the published medical literature (PubMed and Embase) and public access documents. Seventy-two documents representing 133 case studies and 42 unique drugs were determined relevant. The most commonly reported event involved administering an epidural medication by an intravenous line (29.2% of events); a similar proportion of events (27.7%) involved administering an intravenous medication by an epidural line. Medication intended for intravenous administration, but delivered intrathecally, accounted for 25.4% of events. In the most serious cases, outcomes were directly related to the toxicity of the drug that was unintentionally administered. Patient deaths were reported due to the erroneous administration of chemotherapies (n=16), muscle relaxants (n=4), local anesthetics (n=4), opioids (n=1), and antifibrinolytics (n=1). Severe outcomes, including paraplegia, paraparesis, spinal cord injury, and seizures were reported with the following medications: vincristine, gadolinium, diatrizoate meglumine, doxorubicin, mercurochrome, paracetamol, and potassium chloride. These case reports confirm that misconnection events leading to wrong-route errors can occur and may cause serious injury. This comprehensive characterization of events was conducted to better inform clinicians and policymakers, and to describe an emergent strategy designed to mitigate patient risk.

Increasing adherence to a standardized rounding procedure in two hospital in-patient units.

Standardized bedside rounds can improve communication and the quality of care for patients in hospitals. However, it can be challenging to change previously established provider practices to adhere to new procedures. This study evaluated 2 packaged interventions, derived from a modified Performance Diagnostic Checklist interview, to increase adherence to standardized rounding practices in 2 hospital units. Researchers observed physicians at a university hospital on rounds 2-3 times per week, and 2 phases of intervention were implemented to improve adherence. The interventions included task clarification, feedback, and weekly huddles. Compared to baseline, phases 1 and 2 of the intervention improved clinician adherence to the standardized bedside rounding checklist by 24.94% and 30.94% in unit 1 and 26.76% and 44.06% in unit 2, respectively. The standardized rounds did not require additional time following the intervention. These results indicate that physician adherence can be improved through behavioral interventions.

Bacillus anthracis edema toxin impairs neutrophil actin-based motility.

Inhalation anthrax results in high-grade bacteremia and is accompanied by a delay in the rise of the peripheral polymorphonuclear neutrophil (PMN) count and a paucity of PMNs in the infected pleural fluid and mediastinum. Edema toxin (ET) is one of the major Bacillus anthracis virulence factors and consists of the adenylate cyclase edema factor (EF) and protective antigen (PA). Relatively low concentrations of ET (100 to 500 ng/ml of PA and EF) significantly impair human PMN chemokinesis, chemotaxis, and ability to polarize. These changes are accompanied by a reduction in chemoattractant-stimulated PMN actin assembly. ET also causes a significant decrease in Listeria monocytogenes intracellular actin-based motility within HeLa cells. These defects in actin assembly are accompanied by a >50-fold increase in intracellular cyclic AMP and a >4-fold increase in the phosphorylation of protein kinase A. We have previously shown that anthrax lethal toxin (LT) also impairs neutrophil actin-based motility (R. L. During, W. Li, B. Hao, J. M. Koenig, D. S. Stephens, C. P. Quinn, and F. S. Southwick, J. Infect. Dis. 192:837-845, 2005), and we now find that LT combined with ET causes an additive inhibition of PMN chemokinesis, polarization, chemotaxis, and FMLP (N-formyl-met-leu-phe)-induced actin assembly. We conclude that ET alone or combined with LT impairs PMN actin assembly, resulting in paralysis of PMN chemotaxis.

Teaching novice clinicians how to reduce diagnostic waste and errors by applying the Toyota Production System.

Background Diagnostic waste, defined as the ordering of low value tests, increases cost, causes delays, increases complexity, and reduces reliability. The Toyota Production System (TPS) is a powerful approach for process improvement that has not been applied to the diagnostic process. We describe a curriculum based on tools and principles of TPS that provides medical students with an approach for reducing diagnostic waste and improving patient management. Methods A 2-day elective course "Fixing Healthcare Delivery" was offered to medical students at the University of Florida, Gainesville. A section within the course had three learning objectives related to TPS: (1) define value in health care; (2) describe how diagnostic waste leads to time delays and diagnostic errors; and (3) apply sequential and iterative value streams for patient management. Instruction methods included videos, readings, and online quizzes followed by a 2-h seminar with facilitated discussion and active problem solving. Results During the 3 years the course was offered students (n = 25) achieved average scores of 95% on a pre-seminar test of manufacturing principles applied to the diagnostic and management process. Course evaluations averaged 4.94 out of 5 (n = 31). Conclusions Students appreciated the application of the TPS principles to the diagnostic process and expressed the desire to apply these manufacturing principles in their future diagnostic and management decision-making.

Learning From Patients' Experiences Related To Diagnostic Errors Is Essential For Progress In Patient Safety.

Diagnostic error research has largely focused on individual clinicians' decision making and system design, while overlooking information from patients. We analyzed a unique new data source of patient- and family-reported error narratives to explore factors that contribute to diagnostic errors. From reports of adverse medical events submitted in the period January 2010-February 2016, we identified 184 unique patient narratives of diagnostic error. Problems related to patient-physician interactions emerged as major contributors. Our analysis identified 224 instances of behavioral and interpersonal factors that reflected unprofessional clinician behavior, including ignoring patients' knowledge, disrespecting patients, failing to communicate, and manipulation or deception. Patients' perspectives can lead to a more comprehensive understanding of why diagnostic errors occur and help develop strategies for mitigation. Health systems should develop and implement formal programs to collect patients' experiences with the diagnostic process and use these data to promote an organizational culture that strives to reduce harm from diagnostic error.

Theodore E. Woodward Award: spare me the PowerPoint and bring back the medical textbook.

A tutorial for 4(th) year medical students revealed absent long-term retention of microbiology and infectious disease facts taught during the 2(nd) year. Students were suffering from the Ziegarnik effect, the loss of memory after completion of a task. PowerPoint lectures and PowerPoint notes combined with multiple-choice questions may have encouraged this outcome; this teaching format was also associated with minimal use of the course textbook. During the subsequent year, active learning techniques, Just-in-Time Teaching (JiTT) and Peer Instruction (PI) were used, and instructors specifically taught from the textbook. Essays and short answer questions were combined with multiple-choice questions to encourage understanding and recall. Performance on the National Board Shelf exam improved from the 59(th) percentile (2002-2004) to the 83(rd) percentile (2005), and textbook use increased from 1.6% to 79%. This experience demonstrates that strategies incorporating active learning and textbook use correlate with striking improvement in medical student performance.

A patient-initiated voluntary online survey of adverse medical events: the perspective of 696 injured patients and families.

BACKGROUND: Preventable medical errors continue to be a major cause of death in the USA and throughout the world. Many patients have written about their experiences on websites and in published books. METHODS: As patients and family members who have experienced medical harm, we have created a nationwide voluntary survey in order to more broadly and systematically capture the perspective of patients and patient families experiencing adverse medical events and have used quantitative and qualitative analysis to summarise the responses of 696 patients and their families. RESULTS: Harm was most commonly associated with diagnostic and therapeutic errors, followed by surgical or procedural complications, hospital-associated infections and medication errors, and our quantitative results match those of previous provider-initiated patient surveys. Qualitative analysis of 450 narratives revealed a lack of perceived provider and system accountability, deficient and disrespectful communication and a failure of providers to listen as major themes. The consequences of adverse events included death, post-traumatic stress, financial hardship and permanent disability. These conditions and consequences led to a loss of patients' trust in both the health system and providers. Patients and family members offered suggestions for preventing future adverse events and emphasised the importance of shared decision-making. CONCLUSIONS: This large voluntary survey of medical harm highlights the potential efficacy of patient-initiated surveys for providing meaningful feedback and for guiding improvements in patient care.

Exserohilum rostratum: characterization of a cross-kingdom pathogen of plants and humans.

Pathogen host shifts represent a major source of new infectious diseases. There are several examples of cross-genus host jumps that have caused catastrophic epidemics in animal and plant species worldwide. Cross-kingdom jumps are rare, and are often associated with nosocomial infections. Here we provide an example of human-mediated cross-kingdom jumping of Exserohilum rostratum isolated from a patient who had received a corticosteroid injection and died of fungal meningitis in a Florida hospital in 2012. The clinical isolate of E. rostratum was compared with two plant pathogenic isolates of E. rostratum and an isolate of the closely related genus Bipolaris in terms of morphology, phylogeny, and pathogenicity on one C3 grass, Gulf annual rye grass (Lolium multiflorum), and two C4 grasses, Japanese stilt grass (Microstegium vimineum) and bahia grass (Paspalum notatum). Colony growth and color, as well as conidia shape and size were the same for the clinical and plant isolates of E. rostratum, while these characteristics differed slightly for the Bipolaris sp. isolate. The plant pathogenic and clinical isolates of E. rostratum were indistinguishable based on morphology and ITS and 28S rDNA sequence analysis. The clinical isolate was as pathogenic to all grass species tested as the plant pathogenic strains that were originally isolated from plant hosts. The clinical isolate induced more severe symptoms on stilt grass than on rye grass, while this was the reverse for the plant isolates of E. rostratum. The phylogenetic similarity between the clinical and plant-associated E. rostratum isolates and the ability of the clinical isolate to infect plants suggests that a plant pathogenic strain of E. rostratum contaminated the corticosteroid injection fluid and was able to cause systemic disease in the affected patient. This is the first proof that a clinical isolate of E. rostratum is also an effective plant pathogen.

Perspective: a business school view of medical interprofessional rounds: transforming rounding groups into rounding teams.

An effective interprofessional medical team can efficiently coordinate health care providers to achieve the collective outcome of improving each patient's health. To determine how current teams function, four groups of business students independently observed interprofessional work rounds on four different internal medicine services in a typical academic hospital and also interviewed the participants. In all instances, caregivers had formed working groups rather than working teams. Participants consistently exhibited parallel interdependence (individuals working alone and assuming their work would be coordinated with other caregivers) rather than reciprocal interdependence (individuals working together to actively coordinate patient care), the hallmark of effective teams. With one exception, the organization was hierarchical, with the senior attending physician possessing the authority. The interns exclusively communicated with the attending physician in one-on-one conversations that excluded all other members of the team. Although nurses and pharmacists were often present, they never contributed their ideas and rarely spoke.The authors draw on these observations to form recommendations for enhancing interprofessional rounding teams. These are to include the bedside nurse, pharmacist, and case manager as team members, begin with a formal team launch that encourages active participation by all team members, use succinct communication protocols, conduct work rounds in a quiet, distraction-free environment, have teams remain together for longer durations, and receive teamwork training and periodic coaching. High-performing businesses have effectively used teams for decades to achieve their goals, and health care professionals should follow this example.