Four-dimensional imaging of E. coli nucleoid organization and dynamics in living cells.

Visualization of living E. coli nucleoids, defined by HupA-mCherry, reveals a discrete, dynamic helical ellipsoid. Three basic features emerge. (1) Nucleoid density coalesces into longitudinal bundles, giving a stiff, low-DNA-density ellipsoid. (2) This ellipsoid is radially confined within the cell cylinder. Radial confinement gives helical shape and directs global nucleoid dynamics, including sister segregation. (3) Longitudinal density waves flux back and forth along the nucleoid, with 5%-10% of density shifting within 5 s, enhancing internal nucleoid mobility. Furthermore, sisters separate end-to-end in sequential discontinuous pulses, each elongating the nucleoid by 5%-15%. Pulses occur at 20 min intervals, at defined cell-cycle times. This progression includes sequential installation and release of programmed tethers, implying cyclic accumulation and relief of intranucleoid mechanical stress. These effects could comprise a chromosome-based cell-cycle engine. Overall, the presented results suggest a general conceptual framework for bacterial nucleoid morphogenesis and dynamics.

The 3D Topography of Mitotic Chromosomes.

A long-standing conundrum is how mitotic chromosomes can compact, as required for clean separation to daughter cells, while maintaining close parallel alignment of sister chromatids. Pursuit of this question, by high resolution 3D fluorescence imaging of living and fixed mammalian cells, has led to three discoveries. First, we show that the structural axes of separated sister chromatids are linked by evenly spaced "mini-axis" bridges. Second, when chromosomes first emerge as discrete units, at prophase, they are organized as co-oriented sister linear loop arrays emanating from a conjoined axis. We show that this same basic organization persists throughout mitosis, without helical coiling. Third, from prophase onward, chromosomes are deformed into sequential arrays of half-helical segments of alternating handedness (perversions), accompanied by correlated kinks. These arrays fluctuate dynamically over <15 s timescales. Together these discoveries redefine the foundation for thinking about the evolution of mitotic chromosomes as they prepare for anaphase segregation.

One-dimensional spatial patterning along mitotic chromosomes: A mechanical basis for macroscopic morphogenesis.

Spatial patterns are ubiquitous in both physical and biological systems. We have recently discovered that mitotic chromosomes sequentially acquire two interesting morphological patterns along their structural axes [L. Chu et al., Mol. Cell, 10.1016/j.molcel.2020.07.002 (2020)]. First, axes of closely conjoined sister chromosomes acquire regular undulations comprising nearly planar arrays of sequential half-helices of similar size and alternating handedness, accompanied by periodic kinks. This pattern, which persists through all later stages, provides a case of the geometric form known as a "perversion." Next, as sister chromosomes become distinct parallel units, their individual axes become linked by bridges, which are themselves miniature axes. These bridges are dramatically evenly spaced. Together, these effects comprise a unique instance of spatial patterning in a subcellular biological system. We present evidence that axis undulations and bridge arrays arise by a single continuous mechanically promoted progression, driven by stress within the chromosome axes. We further suggest that, after sister individualization, this same stress also promotes chromosome compaction by rendering the axes susceptible to the requisite molecular remodeling. Thus, by this scenario, the continuous presence of mechanical stress within the chromosome axes could potentially underlie the entire morphogenetic chromosomal program. Direct analogies with meiotic chromosomes suggest that the same effects could underlie interactions between homologous chromosomes as required for gametogenesis. Possible mechanical bases for generation of axis stress and resultant deformations are discussed. Together, these findings provide a perspective on the macroscopic changes of organized chromosomes.

Adolescent abdominal pain due to rare mullerian duct anomaly.

Abdominal pain is a common presenting complaint to the Emergency Department (ED). Often, rare etiologies can be discovered in the work up of this common complaint. Here we present the case of an adolescent female who presented with abdominal pain and was found to have obstructed hemivagina and ipsilateral renal anomaly (OHVIRA) or Herlyn-Werner-Wunderlich Syndrome. A 12 year old female with known renal agenesis presented with 5 days of left sided abdominal pain that then developed into right lower quadrant pain. She had regular menses for the last 2 years. Ultrasound (US) showed a fluid collection in the lower uterine segment and a complex cystic structure anterior to the uterus. Magnetic resonance imaging (MRI) showed the patient to have didelphys uterus with "severe dilatation of the cervix/vaginal canal… extending from the right uterine horn" and left-sided ovarian and Fallopian tube torsion. She was taken to the operating room where she underwent vaginal septum excision and a left salpingo-oopherectomy. OHVIRA includes the triad of obstructed hemivagina, uterine didelphys, and ipsilateral renal agenesis. This occurs due to embryologic arrest of the mullerian and mesonephric ducts at 8 weeks of gestation. Most abnormalities are right sided which leads to right lower abdominal and pelvic pain approximately 4 months post-menarche. Diagnosis of OHVIRA is made utilizing US and CT scans. MRI can also be useful to further delineate specific anatomy. It is important for the emergency physician to be aware of this entity as most patients don't present to care until acute, severe symptoms develop. This makes it more likely for them to seek care in the ED as opposed to the outpatient setting.

Clinical Spectrum of Pediatric Heat Illness and Heatstroke in a North American Desert Climate.

ABSTRACT: The spectrum of historical features and clinical presentations of heat illness and heatstroke in the pediatric population has received limited focus in the emergency medicine literature. The majority of published cases involve children trapped in closed spaces and adolescent athletes undergoing high-intensity training regimens in geographical regions with moderately high ambient temperatures and high humidity. There has been less research on the potential impact of extreme temperatures and radiant heat that are the hallmarks of the US southwest region. We performed a retrospective review of pediatric heat illness at our facility located in a North American desert climate.

Rapid and severe neurologic deterioration due to influenza associated encephalopathy in a healthy child.

Rapid neurologic deterioration is rare in healthy school age children, particularly in the absence of trauma or toxic exposure. Hemorrhagic or ischemic stroke, infectious etiologies and metabolic causes must be emergently considered. We present the clinical details of a previously well child with two days of mild viral symptoms who progressed from playful and active to severe neurologic injury over the course of eight hours.

Critical procedure performance in pediatric patients: Results from a national emergency medicine group.

STUDY OBJECTIVE: We sought to examine the frequency of pediatric critical procedures performed in a national group of emergency physicians. METHODS: We performed a retrospective analysis of an administrative billing and coding dataset for procedural performance documentation verification from 2014 to 2018. We describe and compare incident rates of pediatric (age <18 years) patient critical procedure performance by emergency physicians in general emergency departments (EDs), pediatric EDs, and freestanding ED/urgent care centers. Critical procedures were endotracheal intubation, electrical cardioversion, central venous placement, intraosseous access, and chest tube insertion. RESULTS: Among 2290 emergency physicians working in 186 EDs (1844 working in 129 general EDs, 125 in 8 pediatric EDs, and 321 in 49 freestanding EDs/urgent cares), a total of 2233 pediatric critical procedures were performed during the study period. Many physicians at general EDs and freestanding EDs/urgent cares performed zero pediatric procedures per year (53.9% and 89% respectively). Per 1000 ED visits seen (All patient ages), physicians working in general EDs performed fewer pediatric critical procedures than physicians in pediatric EDs (0.12/1000 visits vs 0.68/1000 visits; rate difference = 0.56, 95% confidence interval [CI] 0.51-0.61). Per 1000 clinical hours worked, physicians working in general EDs performed 0.26 procedures compared to 1.66 for physicians in pediatric EDs (rate difference = 1.39; 95% CI 1.27-1.52). CONCLUSION: Pediatric critical procedures are rarely performed by emergency physicians and are exceedingly rare in general EDs and freestanding EDs/urgent cares. The rarity of performance of these skills has implications for ED pediatric readiness.

Emergency department presentation of 'delusional parasitosis by proxy'. Delusional parent, injured child.

We report a case of 'delusional parasitosis by proxy'. A sixyear old child was brought to the emergency department by a mother with concerns that her son had a skin and scalp infestation. Despite the absence of any clinical findings being found on exam, the mother remained disproportionately concerned. Follow up care was recommended with the child's primary care. The mother returned to the ED with her child three weeks later with concerns that her son had an inflamed scalp and eyes. The mother remained insistent that the child was infested with bugs and she had sought care at two other locations where the child was prescribed permethrin on both visits. She had been applying the medication repeatedly. On exam the boy's scalp had been shaved and was erythematous and irritated; his eyebrows and eyelashes had also been shaved off and likely contributed to an irritant conjunctivitis from repeated applications of topical permethrin lotion. No evidence of infestation was identified. We recruited the assistance of the maternal grandparents, child protective services and primary care pediatrics and the child was removed from the mother's custody and placed into the custody of the grandparents. Six weeks later with basic skin care and erythromycin ophthalmic ointment for the eyes, the child's hair, eyebrows and eyelashes grew had grown in, and the scalp irritation had resolved. The mother had sought and received psychiatric care and was improving.

Clinical spectrum of previously undiagnosed pediatric cardiac disease.

Previously undiagnosed pediatric cardiac disease represents a clinical challenge for the emergency physician. The clinical presentation of these disorders can mimic other conditions of the respiratory, gastrointestinal and neurologic systems at a time when the need for early identification and treatment is at a premium. A high index of suspicion and superb clinical acumen is required to make a timely diagnosis and initiate optimal care. METHODS: A retrospective chart review using explicit criteria and a structured data collection process was performed on all children presenting with previously undiagnosed cardiac disease over a five and half year period. RESULTS: Thirty-six patients were identified over a five and a half year period representing one patient per 4838 pediatric ED presentations. A diverse set of chief complaints, triage categories, clinical presentations and diagnoses were identified. Undiagnosed congenital lesions, acquired cardiac disease, dysrhythmias and infectious diseases of the heart were represented. The need for surgical intervention (22%) and mortality (6%) was substantial. CONCLUSION: In the aggregate, these conditions occur at an important rate and represent a high risk subset of pediatric patients presenting to the emergency department. Individually, the conditions occur infrequently and 'pattern recognition' may not aid the clinician. Early diagnosis and prompt intervention is important in this population.

Initial ED oxygen saturation ≤90% increases the risk of a complicated hospital course in pediatric asthmatics requiring admission.

Emergency physicians are responsible for admitting children with asthma who do not respond to initial therapy. We examined the hypothesis that an initial room air pulse oximetry ≤90% elevates the risk of a complicated hospital course in children who require admission with acute asthma. METHODS: Charts of all patients ages 2 years-17 years admitted for asthma from January 2017 to December 2017 were reviewed. An explicit chart review was performed by trained data extractors using a standardized form. RESULTS: A total of 244 children meeting inclusion criteria were admitted for asthma from the ED during the study period. All patients had an initial room air pulse oximetry documented. Sixty-five were admitted to PICU status (27%), and 179 (73%) were admitted to floor status. The relative risk of a complicated course in those patients presenting with a saturation of ≤90% was 11.3 (95% CI 3.9-32.6). The mean initial pulse oximetry on patients with a complicated course was 85% versus 93% for those without a complicated course (p < 0.005). CONCLUSION: Our data suggest that in pediatric asthmatics that require admission from the ED, those with pulse oximetry readings less than or equal to 90% on presentation are at higher risk of a complicated hospital course.

Bilateral Upper Extremity Hyperesthesia and Absence of Neck Tenderness in Four Adolescent Athletes With Cervical Spine Injuries.

Cervical spine injury in the pediatric athlete is an uncommon but potentially devastating condition that can require a heightened index of suspicion to correctly diagnose. Although some cervical spine injuries present overtly with paraplegia due to cord transection, others can present more subtly with symptoms suggestive of bilateral peripheral neuropathy. Patients with traumatic brachial neuropraxia or "stinger" physiology can present similarly, but symptoms are exclusively unilateral. We present 4 patients with bilateral upper extremity hyperesthesias and absence of tenderness over the cervical vertebral landmarks who were subsequently diagnosed with cervical spine injuries.

The 100-year evolution of the isolated internal iliac artery aneurysm.

BACKGROUND: Isolated internal iliac artery aneurysms (IIIAA) are a rare form of aneurysm. The incidence increases with age, and the prevalence is higher in men. The clinical presentation can vary, and standard treatment protocols are not established. The first case of an IIIAA was described more than 100 years ago. The purpose of the study is to summarize the various clinical presentations and treatment options that have been reported in the literature in the past 100 years. METHODS: Literature about IIIAA was reviewed using the electronic databank PubMed. All case reports and case series were analyzed, and we included our own data with 2 case reports. RESULTS: Over time, IIIAA diagnosis increasingly resulted from asymptomatic incidental findings on radiologic studies. Various clinical presentations included abdominal pain, back pain, rectal bleeding, hydronephrosis with renal failure, hematuria, and free rupture with shock. Rupture has a mortality rate of 53%. IIIAAs were more common on the left (61.8% left, 27.3% right, 10.9% bilateral). Treatments include open surgical repair and endovascular repair using a variety of methods. One article reported a hybrid method using both endovascular and open surgical technique. CONCLUSIONS: Since its first description 100 years ago, we have gained knowledge about the natural history of IIIAA. Multiple treatment options have been described, but long-term outcome needs further investigation.

Escherichia coli sister chromosome separation includes an abrupt global transition with concomitant release of late-splitting intersister snaps.

The basis for segregation of sister chromosomes in bacteria is not established. We show here that two discrete ~150-kb regions, both located early in the right replichore, exhibit prolonged juxtaposition of sister loci, for 20 and 30 min, respectively, after replication. Flanking regions, meanwhile, separate. Thus, the two identified regions comprise specialized late-splitting intersister connections or snaps. Sister snap loci separate simultaneously in both snap regions, concomitant with a major global nucleoid reorganization that results in emergence of a bilobed nucleoid morphology. Split snap loci move rapidly apart to a separation distance comparable with one-half the length of the nucleoid. Concomitantly, at already split positions, sister loci undergo further separation to a comparable distance. The overall consequence of these and other effects is that thus far replicated sister chromosomes become spatially separated (individualized) into the two nucleoid lobes, while the terminus region (and likely, all unreplicated portions of the chromosome) moves to midcell. These and other findings imply that segregation of Escherichia coli sister chromosomes is not a smooth continuous process but involves at least one and likely, two major global transition(s). The presented patterns further suggest that accumulation of internal intranucleoid forces and constraining of these forces by snaps play central roles in global chromosome dynamics. They are consistent with and supportive of our previous proposals that individualization of sisters in E. coli is driven primarily by internally generated pushing forces and is directly analogous to sister individualization at the prophase to prometaphase transition of the eukaryotic cell cycle.

Mortality after discharge in clinically stable infants admitted with a first-time apparent life-threatening event.

OBJECTIVE: The objective of this study is to review the mortality after discharge in clinically stable infants admitted with a first apparent life-threatening event. DESIGN: Retrospective chart review of all infants 0 to 6 months presenting with a first apparent life-threatening event (ALTE) over a 5-year period using explicit criteria. Patients with an emergency department (ED) diagnosis of ALTE, seizure, choking spell, or cyanosis were reviewed by 2 of 3 physicians. Level of agreement between reviewers was monitored. Mortalities were identified by a review of the county death record database and hospital records. RESULTS: Three hundred sixty-six charts were reviewed; 176 cases met inclusion criteria. All apparent life-threatening event (ALTE) cases were admitted; 1 signed out against medical advice. Blood cultures were obtained in 111 patients (63%)-no pathogens were identified. Cerebrospinal fluid analysis and culture was performed in 65 patients (37%)-no pathogens were identified. One patient had pleocytosis. Chest radiographs were obtained in 115 patients (65%); 12 patients had infiltrates. Respiratory syncytial virus nasal washings were obtained in 32% of patients and were positive in 9 patients. The average length of follow-up was 34 months; 2 patients (1.1%) had died at the time of follow-up. Both deaths occurred after hospital discharge and within 2 weeks of the ED visit. Neither of the fatalities had a positive diagnostic evaluation in the ED. The cause of death by coroner report was pneumonia in both instances. CONCLUSIONS: The risk of subsequent mortality in infants admitted from our pediatric ED with an ALTE is substantial. Emergency physicians should consider routine admission for patients with ALTE.

A modular microfluidic device that uses magnetically actuatable microposts for enhanced magnetic bead-based workflows.

Magnetic beads have been widely and successfully used for target enrichment in life science assays. There exists a large variety of commercially available magnetic beads functionalized for specific target capture, as well as options that enable simple surface modifications for custom applications. While magnetic beads are ideal for use in the macrofluidic context of typical laboratory workflows, their performance drops in microfluidic contexts, such as consumables for point-of-care diagnostics. A primary cause is the diffusion-limited analyte transport in these low Reynolds number environments. A new method, BeadPak, uses magnetically actuatable microposts to enhance analyte transport, improving yield of the desired targets. Critical parameters were defined for the operation of this technology and its performance characterized in canonical life-science assays. BeadPak achieved up to 1000× faster capture than a microfluidic chamber relying on diffusion alone, enabled a significant specimen concentration via volume reduction, and demonstrated compatibility with a range of biological specimens. The results shown in this work can be extended to other systems that utilize magnetic beads for target capture, concentration, and/or purification.

Pediatric chest pain: using evidence to reduce diagnostic testing in the emergency department.

Pediatric chest pain is a relatively common presenting complaint, but identifying serious pathologies without overtesting patients with less-serious pathologies can be a challenge for emergency clinicians. This issue reviews the available literature to provide evidence-based recommendations to support a more standardized approach to the evaluation and management of pediatric patients with chest pain. This issue will help the emergency clinician identify red flags associated with cardiac causes of pediatric chest pain, recognize life-threatening causes of cardiac and non- cardiac chest pain, clinically diagnose the most common causes of non-cardiac chest pain, and appropriately utilize diagnostic tests in the evaluation of chest pain patients.

Spontaneous isolated celiac and splenic artery dissection with splenic infarction.

Spontaneous isolated celiac artery dissection is considered an uncommon clinical condition. Rarer still is spontaneous isolated celiac and splenic artery dissection, with a total of 42 reported cases. There is no known definitive cause of visceral artery dissections, but risk factors include male sex, age in 5th or 6th decade, hypertension, and connective tissue disorders. The presentation varies, diagnosis is principally radiographic, and the mainstay of treatment is anticoagulation or antiplatelet therapy. Splenic infarction is a common finding with splenic artery dissection, although the strength of this association has not previously been reported. Herein we present a case of spontaneous isolated celiac and splenic artery dissection with splenic infarction that was successfully managed with blood pressure control and antiplatelet therapy. We review previous literature, principles of diagnosis and management, and incidence and outcomes of splenic infarction as it related to splenic artery dissection.

Successful resuscitation from cardiopulmonary arrest due to profound hypothermia using noninvasive techniques.

Profound hypothermia is defined as a core body temperature of 20.0 °C or less. Successful resuscitation from this degree of hypothermia is rare. We present a case of successful resuscitation in a 2-year-old boy found in cardiac arrest due to profound hypothermia. Invasive techniques such as cavity lavage, extracorporeal membrane oxygenation, and cardiopulmonary bypass were not used.

Mixing and oxygen transfer characteristics of a microplate bioreactor with surface-attached microposts.

Bioprocess optimization for cell-based therapies is a resource heavy activity. To reduce the associated cost and time, process development may be carried out in small volume systems, with the caveat that such systems be predictive for process scale-up. The transport of oxygen from the gas phase into the culture medium, characterized using the volumetric mass transfer coefficient, kL a, has been identified as a critical parameter for predictive process scale-up. Here, we describe the development of a 96-well microplate with integrated Redbud Posts to provide mixing and enhanced kL a. Mixing in the microplate is characterized by observation of dyes and analyzed using the relative mixing index (RMI). The kL a is measured via dynamic gassing out method. Actuating Redbud Posts are shown to increase rate of planar homogeneity (2 min) verse diffusion alone (120 min) and increase oxygenation, with increasing stirrer speed (3500-9000 rpm) and decreasing fill volume (150-350 µL) leading to an increase in kL a (4-88 h-1 ). Significant increase in Chinese Hamster Ovary growth in Redbud Labs vessel (580,000 cells mL-1 ) versus the control (420,000 cells mL-1 ); t(12.814) = 8.3678, p ≤ .001), and CD4+ Naïve cell growth in the microbioreactor indicates the potential for this technology in early stage bioprocess development and optimization.

Clinical spectrum of shock in the pediatric emergency department.

OBJECTIVE: The objective of this study was to describe the clinical spectrum of patients presenting with shock or developing shock in a pediatric emergency department (ED) during an 8-year period. METHODS: An observational study of all pediatric ED patients with shock between September 1998 and September 2006 was performed. Trauma activations were excluded. A structured, explicit chart review using a standardized abstraction form and case definition was completed by 3 physicians board certified in pediatric emergency medicine. Interrater reliability was monitored. RESULTS: A total of 147 cases of shock were identified. Septic shock was the underlying physiology in 57% of cases. A pathogen was identified in 45% of these cases. Hypovolemic shock due to gastroenteritis, metabolic disease, surgical emergencies, or hemorrhage was the cause in 24% of cases. Distributive shock represented 14% of cases. Cardiogenic shock contributed to 5% of cases. Patients with septic shock received a mean of 58 mL/kg of crystalloid or colloid versus 50 mL/kg in patients with other causes. Intubation and vasopressor use was required in 41% and 21% of cases, respectively. Clinical signs of shock developed in the ED after initially presenting without clinical signs of shock in 14% of study subjects. Nearly half of these episodes occurred after the administration of antimicrobials or performance of a lumbar puncture. Mortality was 6% overall and 5% in septic shock patients. CONCLUSIONS: Pediatric ED patients with shock represent a diverse population with substantial mortality. Of 147 patients, 21 presented without clinical signs of shock and deteriorated to a clinical condition meeting the definition of shock during the ED course.