Rapid IDH1-R132 genotyping panel utilizing locked nucleic acid loop-mediated isothermal amplification.

While the detection of single-nucleotide variants (SNVs) is important for evaluating human health and disease, most genotyping methods require a nucleic acid extraction step and lengthy analytical times. Here, we present a protocol which utilizes the integration of locked nucleic acids (LNAs) into self-annealing loop primers for the allelic discrimination of five isocitrate dehydrogenase 1 R132 (IDH1-R132) variants using loop-mediated isothermal amplification (LAMP). This genotyping panel was initially evaluated using purified synthetic DNA to show proof of specific SNV discrimination. Additional evaluation using glioma tumor lysates with known IDH1-R132 mutational status demonstrated specificity in approximately 35 min without the need for a nucleic acid extraction purification step. This LNA-LAMP-based genotyping assay can detect single base differences in purified nucleic acids or tissue homogenates, including instances where the variant of interest is present in an excess of background wild-type DNA. The pH-based colorimetric indicator of LNA-LAMP facilitates convenient visual interpretation of reactions, and we demonstrate successful translation to an end-point format using absorbance ratio, allowing for an alternative and objective approach for differentiating between positive and negative reactions. Importantly, the LNA-LAMP genotyping panel is highly reproducible, with no false-positive or false-negative results observed.

Cerastecins inhibit membrane lipooligosaccharide transport in drug-resistant Acinetobacter baumannii.

Carbapenem-resistant Acinetobacter baumannii infections have limited treatment options. Synthesis, transport and placement of lipopolysaccharide or lipooligosaccharide (LOS) in the outer membrane of Gram-negative bacteria are important for bacterial virulence and survival. Here we describe the cerastecins, inhibitors of the A. baumannii transporter MsbA, an LOS flippase. These molecules are potent and bactericidal against A. baumannii, including clinical carbapenem-resistant Acinetobacter baumannii isolates. Using cryo-electron microscopy and biochemical analysis, we show that the cerastecins adopt a serpentine configuration in the central vault of the MsbA dimer, stalling the enzyme and uncoupling ATP hydrolysis from substrate flipping. A derivative with optimized potency and pharmacokinetic properties showed efficacy in murine models of bloodstream or pulmonary A. baumannii infection. While resistance development is inevitable, targeting a clinically unexploited mechanism avoids existing antibiotic resistance mechanisms. Although clinical validation of LOS transport remains undetermined, the cerastecins may open a path to narrow-spectrum treatment modalities for important nosocomial infections.

The American Association for Thoracic Surgery (AATS) 2023 Expert Consensus Document: Recommendation for the care of children with trisomy 13 or trisomy 18 and a congenital heart defect.

OBJECTIVES: Recommendations for surgical repair of a congenital heart defect in children with trisomy 13 or trisomy 18 remain controversial, are subject to biases, and are largely unsupported with limited empirical data. This has created significant distrust and uncertainty among parents and could potentially lead to suboptimal care for patients. A working group, representing several clinical specialties involved with the care of these children, developed recommendations to assist in the decision-making process for congenital heart defect care in this population. The goal of these recommendations is to provide families and their health care teams with a framework for clinical decision making based on the literature and expert opinions. METHODS: This project was performed under the auspices of the AATS Congenital Heart Surgery Evidence-Based Medicine Taskforce. A Patient/Population, Intervention, Comparison/Control, Outcome process was used to generate preliminary statements and recommendations to address various aspects related to cardiac surgery in children with trisomy 13 or trisomy 18. Delphi methodology was then used iteratively to generate consensus among the group using a structured communication process. RESULTS: Nine recommendations were developed from a set of initial statements that arose from the Patient/Population, Intervention, Comparison/Control, Outcome process methodology following the groups' review of more than 500 articles. These recommendations were adjudicated by this group of experts using a modified Delphi process in a reproducible fashion and make up the current publication. The Class (strength) of recommendations was usually Class IIa (moderate benefit), and the overall level (quality) of evidence was level C-limited data. CONCLUSIONS: This is the first set of recommendations collated by an expert multidisciplinary group to address specific issues around indications for surgical intervention in children with trisomy 13 or trisomy 18 with congenital heart defect. Based on our analysis of recent data, we recommend that decisions should not be based solely on the presence of trisomy but, instead, should be made on a case-by-case basis, considering both the severity of the baby's heart disease as well as the presence of other anomalies. These recommendations offer a framework to assist parents and clinicians in surgical decision making for children who have trisomy 13 or trisomy 18 with congenital heart defect.

Rapid extraction-free detection of the R132H isocitrate dehydrogenase mutation in glioma using colorimetric peptide nucleic acid-loop mediated isothermal amplification (CPNA-LAMP).

The R132H isocitrate dehydrogenase one (IDH1) mutation is a prognostic biomarker present in a subset of gliomas and is associated with heightened survival when paired with aggressive surgical resection. In this study, we establish proof-of-principle for rapid colorimetric detection of the IDH1-R132H mutation in tumor samples in under 1 hour without the need for a nucleic acid extraction. Colorimetric peptide nucleic acid loop-mediated isothermal amplification (CPNA-LAMP) utilizes 4 conventional LAMP primers, a blocking PNA probe complementary to the wild-type sequence, and a self-annealing loop primer complementary to the single nucleotide variant to only amplify the DNA sequence containing the mutation. This assay was evaluated using IDH1-WT or IDH1-R132H mutant synthetic DNA, wild-type or IDH1-R132H mutant U87MG cell lysates, and tumor lysates from archived patient samples in which the IDH1 status was previously determined using immunohistochemistry (IHC). Reactions were performed using a hot water bath and visually interpreted as positive by a pink-to-yellow color change. Results were subsequently verified using agarose gel electrophoresis. CPNA-LAMP successfully detected the R132H single nucleotide variant, and results from tumor lysates yielded 100% concordance with IHC results, including instances when the single nucleotide variant was limited to a portion of the tumor. Importantly, when testing the tumor lysates, there were no false positive or false negative results.

Sustained year-round oceanographic measurements from Rothera Research Station, Antarctica, 1997-2017.

Oceanographic changes adjacent to Antarctica have global climatic and ecological impacts. However, this is the most challenging place in the world to obtain marine data due to its remoteness and inhospitable nature, especially in winter. Here, we present more than 2000 Conductivity-Temperature-Depth (CTD) profiles and associated water sample data collected with (almost uniquely) full year-round coverage from the British Antarctic Survey Rothera Research Station at the west Antarctic Peninsula. Sampling is conducted from a small boat or a sled, depending on the sea ice conditions. When conditions allow, sampling is twice weekly in summer and weekly in winter, with profiling to nominally 500 m and with discrete water samples taken at 15 m water depth. Daily observations are made of the sea ice conditions in the area. This paper presents the first 20 years of data collection, 1997-2017. This time series represents a unique and valuable resource for investigations of the high-latitude ocean's role in climate change, ocean/ice interactions, and marine biogeochemistry and carbon drawdown.

Late-Onset Hyponatremia in Premature Infants.

Late-onset hyponatremia (LOH) frequently affects premature infants 2 or more weeks of age due to inadequate sodium intake and excessive kidney loss. Late-onset hyponatremia typically occurs in infants who are physiologically stable and is defined as serum sodium of 132 mEq/L or less or between 133 and 135 mEq/L if receiving sodium supplementation. Recent evidence suggests that spot urine sodium levels may improve the recognition of LOH, as low levels of excreted urine reflect a total body sodium deficit and negative balance. Untreated LOH may result in poor somatic growth, neurodevelopmental delay, higher incidence of bronchopulmonary dysplasia, and more severe retinopathy of prematurity. The primary prevention of LOH is to maintain serum sodium between 135 and 145 mEq/L; however, there are currently no formal protocols guiding sodium supplementation. The purpose of this article is to present on overview of LOH pathophysiology and its effect on somatic growth, neurodevelopment outcomes, and other related sequelae. We further discuss general management strategies and describe a protocol for sodium supplementation that is presently undergoing an evaluation for effectiveness.

Comparison of refractive and visual acuity results after Contoura® Vision topography-guided LASIK planned with the Phorcides Analytic Engine to results after wavefront-optimized LASIK in eyes with oblique astigmatism.

PURPOSE: To compare visual acuity and refractive results between topography-guided laser in situ keratomileusis (LASIK) planned with the Phorcides Analytic Engine (PAE) to results after wavefront-optimized (WFO) LASIK in subjects with preoperative oblique astigmatism in their manifest refraction. METHODS: This was a retrospective chart review of clinical results from eyes treated with topography-guided LASIK planned with PAE compared to eyes treated with WFO LASIK using the same Wavelight® excimer laser system. All included subjects had preoperative oblique astigmatism. Residual refractive error and visual acuity (uncorrected and corrected) were the measures of interest, at the visit closest to 90 days postoperative. RESULTS: A matched data set from 100 WFO and 97 PAE eyes was extracted from clinical records. At the postoperative visit the PAE group showed lower residual refractive cylinder (p = 0.04), uncorrected distance visual acuity (UDVA) (-0.06 PAE vs. -0.02 WFO, p < 0.01) and distance corrected visual acuity (CDVA) (p < 0.01). The percentage of eyes with a mean refraction spherical equivalent (MRSE) magnitude within 0.25 D and 0.50 D of plano was statistically significantly higher in the PAE group (p = 0.04 and 0.01, respectively). A statistically significantly higher percentage of eyes in the PAE group had UDVA better than or equal to -0.10 logMAR (20/16 Snellen, 36% vs 22%, p = 0.04). More eyes gained CDVA after surgery in the PAE group (53% vs 32%, p < 0.01). There were five enhancements in the WFO group versus none in the PAE group, a statistically significant difference (p = 0.03). CONCLUSIONS: Visual acuity and refractive outcomes after LASIK using PAE in eyes with oblique astigmatism in their preoperative refraction were statistically significantly better than those obtained when WFO treatment was used. The number of refractive outliers and the number of retreatments were also significantly lower with PAE treatment.

Clinical and refractive outcomes after topography-guided refractive surgery planned using Phorcides surgery planning software.

PURPOSE: To evaluate prospectively the clinical outcomes of topography-guided laser in situ keratomileusis (LASIK) surgery performed with Contoura Vision using the Wavelight excimer laser and planned with the Phorcides Analytical Engine. SETTING: Four clinical practices in the U.S. DESIGN: Prospective single-arm interventional study. METHODS: 130 eyes of 65 patients with myopia or myopic astigmatism were enrolled in a prospective study of visual and refractive outcomes after treatment with Contoura Vision using the Wavelight EX500 excimer laser to achieve optimal distance vision. At 3 months postoperatively, uncorrected distance visual acuity (UDVA), manifest refraction, and corrected distance visual acuity (CDVA) were measured and compared with previously published studies. RESULTS: At 3 months postoperatively, 100%, 89%, and 28% of eyes achieved UDVA of 20/20, 20/15, and 20/12.5 or better, respectively. 92% of eyes had postoperative UDVA equal to or better than their preoperative CDVA. Postoperative CDVA was equal to, 1 line better, or 2 lines better than the preoperative CDVA in 53%, 40%, and 6% of eyes, respectively. Only 1 eye lost 1 line of CDVA, and no eyes lost more than 1 line of CDVA. A statistically significant decrease was noted in subjective visual complaints, including glare, halos, difficulty driving at night, reading difficulty, starbursts, fluctuation in vision, and light sensitivity. CONCLUSIONS: The Phorcides Analytical Engine can be used to optimize visual outcomes for the correction of myopia and myopic astigmatism.

Renal Oxygenation (rSO 2 ) Population Parameter Estimates in Premature Infants Routinely Monitored With Near-Infrared Spectroscopy.

BACKGROUND: Currently, reference ranges for renal oxygenation measured by near-infrared spectroscopy (NIRS) in preterm infants beyond the first days of life are lacking, especially those born prior to 29 weeks' gestation. Population estimates of renal oxygenation (rSO 2 ) levels among preterm infants over time have yet to be established, leading to reluctance in clinical application. PURPOSE: To characterize the distribution and estimate population parameters for renal oxygenation measured by NIRS during the first 14 days of life among preterm infants. METHODS: We prospectively observed rSO 2 trends of 37 infants before 34 weeks' gestation and 1800-g or less birth weight for the first 14 days of life. Analyses included distribution fit tests, ordinary least squares (OLS) regression, and t tests. RESULTS: Average daily rSO 2 variation steadily increased with 42% difference through the first 14 days of life. For all infants, renal rSO 2 means peaked during the first 3 days of life and plateaued around 7 days. Daily rSO 2 slopes were significantly lower among males and infants 29 weeks' or less gestation. IMPLICATIONS FOR PRACTICE: Renal rSO 2 during the first 14 days of life reflects normal extrauterine transition reaching stabilization around 7 days of life. Gestational age, birth weight, and gender may predict the early trajectory of rSO 2 patterns. Population estimates provide parameters for renal rSO 2 that may indicate early-onset tissue hypoxia when acute or significant drops from baseline occur. IMPLICATIONS FOR RESEARCH: We present a framework to guide future research using renal NIRS technology in preterm infants to determine deviations from expected trends that may precede renal injury.

Degrading permafrost river catchments and their impact on Arctic Ocean nearshore processes.

Arctic warming is causing ancient perennially frozen ground (permafrost) to thaw, resulting in ground collapse, and reshaping of landscapes. This threatens Arctic peoples' infrastructure, cultural sites, and land-based natural resources. Terrestrial permafrost thaw and ongoing intensification of hydrological cycles also enhance the amount and alter the type of organic carbon (OC) delivered from land to Arctic nearshore environments. These changes may affect coastal processes, food web dynamics and marine resources on which many traditional ways of life rely. Here, we examine how future projected increases in runoff and permafrost thaw from two permafrost-dominated Siberian watersheds-the Kolyma and Lena, may alter carbon turnover rates and OC distributions through river networks. We demonstrate that the unique composition of terrestrial permafrost-derived OC can cause significant increases to aquatic carbon degradation rates (20 to 60% faster rates with 1% permafrost OC). We compile results on aquatic OC degradation and examine how strengthening Arctic hydrological cycles may increase the connectivity between terrestrial landscapes and receiving nearshore ecosystems, with potential ramifications for coastal carbon budgets and ecosystem structure. To address the future challenges Arctic coastal communities will face, we argue that it will become essential to consider how nearshore ecosystems will respond to changing coastal inputs and identify how these may affect the resiliency and availability of essential food resources.

Long COVID Syndrome and Takotsubo Cardiomyopathy: An Unwelcome Combination.

Since the report of the first case from China in late 2019, the coronavirus disease (COVID-19) has spread very rapidly through the countries and regions leaving a trail of devastation in its path, everywhere. Although COVID-19 is primarily a respiratory illness mainly affecting the lungs; involvement of other organs including the cardiovascular system has been widely recognized. Whilst COVID-19 is an acute illness for a majority of cases; some of the debilitating virus-related symptoms can last for weeks and months, and are collectively termed as long COVID syndrome. Several published reports have described an association between acute COVID-19 illness and cardiac complications such as myocarditis and Takotsubo cardiomyopathy. However, little is known about any link between long COVID syndrome and the cardiac disease. We describe the case of a middle-aged woman with long COVID syndrome who presented with central chest pain and breathlessness. Her initial investigations showed an elevated cardiac troponin I and ischemic changes on 12 lead ECG. She was initially treated for non-ST elevation myocardial infarction. A subsequent coronary angiogram showed unobstructed coronary vessels and left ventricle (LV) gram demonstrated apical LV ballooning. She was managed conservatively and was discharged home following her clinical improvement. This case highlights the importance of holistic assessment of patients presenting with chest pain with the background of long COVID syndrome. It also outlines an emergent need to better understand pathophysiological mechanisms that underpin the development of cardiac complications in those with COVID-19 and long COVID syndrome.

The Secret of Quality Is Love: A Qualitative Study Exploring Physician and Nurse Perspectives on What It Means to Love Their Patients.

Little research systematically explores healthcare professionals' understanding of what it means to love their patients. The authors hypothesized that nurses and physicians would describe a language of love relevant to health care. Researchers conducted structured interviews with 29 physicians and 32 nurses at an academic medical center through a combination of purposive, convenience, and snowball sampling. Interviews were transcribed verbatim and analyzed using grounded theory, identifying major themes from qualitative data. Most nurses and physicians declared they should love their patients. Characteristics of loving physicians and nurses included caring, clinical excellence, advocating, meeting needs, compassion, sacrifice, and tough love. Moral imperatives included the duty to act on behalf of the patient's best interest, respect the patient's wishes, treat patients as you would want your family treated, and recognize limits of life. Many physicians and nurses commented that loving patients could transform health care. Physicians and nurses described characteristics and imperatives of love that may serve as an ethical standard for healthcare professionals.

A Gay Epidemiologist and the DC Commission of Public Health AIDS Advisory Committee.

Based upon the lead author's deep personal and professional experience, this case narrative illustrates the importance of engagement between public health practitioners and members of affected populations and their advocates. The case underscores the need to build strong coalitions to address serious public health and social issues. It also illustrates how decisions about control groups in research raise ethical issues. In addition, the case illustrates the reality that public health and social services are sometimes inadequate in the face of dire circumstances. Justice in public health has both a distributive aspect (how to allocate limited resources and distribute potential benefits as fairly as possible) and a procedural dimension (ensuring public participation, especially of those most affected). Frameworks for public health ethics, which post-date the events detailed in the autobiographical case narrative, highlight both distributive justice and procedural justice.

The Impact of Image Registration for Ablation Orientation on Clinical Outcomes After Wavefront-Optimized Refractive Surgery in Eyes with Myopia and Astigmatism.

PURPOSE: To compare the clinical outcomes from laser refractive surgery performed with the same laser with and without incorporating iris registration technology to compensate for ocular cyclotorsion. DESIGN: Single-site, two-arm, retrospective chart review. METHODS: Clinical outcomes at a single site after wavefront-optimized LASIK using the Wavelight excimer laser with and without the Vario imaging system for iris registration (IR) were evaluated. Eligible subjects were those that received on-label wavefront-optimized treatment of myopia with astigmatism >1.5 D. Measures of interest were the amount of residual refractive cylinder after surgery, the refractive error, and the best-corrected (BCVA) and uncorrected (UCVA) visual acuities, with a target follow-up of around 90 days. RESULTS: A total of 112 eligible eyes that were treated with IR and 126 similar eyes treated without IR (NO IR) were included. The refractive sphere and spherical equivalent refractions were statistically significantly different between groups (p < 0.05), but the mean differences were <0.1 D in both cases. Refractive cylinder averaged around 0.12 D and was not statistically significantly different between groups. The number of eyes with residual cylinder >0.50 D was higher in the NO IR group vs the IR group (6% vs 1%, respectively, p = 0.04). The mean logMAR UCVA and BCVA were statistically significantly better in the IR group, with a difference of 1.5 letters for UCVA and 1.0 letters for BCVA (p < 0.001 for both). Significantly more eyes in the IR group had a UCVA (p = 0.01) and a BCVA of 20/15 or better (p = 0.003). Overall, 96% of eyes in the IR group and 91% of eyes in the NO IR group had uncorrected visual acuity of 20/20 or better. CONCLUSION: Iris registration with the VARIO imaging device demonstrably reduced the overall variability in clinical outcomes.

Clinical Outcomes After Topography-Guided Refractive Surgery in Eyes with Myopia and Astigmatism - Comparing Results with New Planning Software to Those Obtained Using the Manifest Refraction.

PURPOSE: To compare clinical outcomes from topography-guided laser refractive surgery based on new planning software to outcomes based on using the manifest refraction. DESIGN: Single site, two-arm, retrospective chart review. METHODS: Clinical outcomes at a single site after topography-guided LASIK using the Wavelight excimer laser were evaluated, with a target postoperative follow-up time of 90 days. Eligible eyes were those that received on-label topography-guided treatment of myopia or myopic astigmatism with correction based on either the manifest refraction or results from the Phorcides Analytical Engine (PAE). Measures analyzed included the uncorrected (UDVA) and best-corrected (CDVA) distance visual acuity, the magnitude of refractive cylinder after surgery, the refractive error and changes from preoperative CDVA. RESULTS: The study included 115 eyes in the PAE group and 133 eyes in the Manifest group. Significantly more eyes in the PAE group had a CDVA of 20/15 or better (p = 0.05) and a UDVA of 20/15 or better (p = 0.05). Significantly more eyes in the Manifest group had a UDVA of 20/25 or worse (13/133 vs 1/115 in the PAE group, p = 0.002). There were significantly more eyes in the PAE group with no postoperative refractive cylinder (90% vs 77% in the Manifest group, p = 0.004). No eye in either group had a postoperative CDVA a line or worse than their preoperative CDVA. Three eyes in the Manifest group and no PAE eyes have had subsequent enhancement surgery. CONCLUSION: Mean results for postoperative refractive astigmatism, CDVA and UDVA were similar between the groups, but the clinical outcomes for the PAE group appeared less variable, with more eyes having no refractive astigmatism and a higher percentage of eyes having 20/15 or better CDVA and UCVA. The objective nature of the PAE is an advantage.

Implementation and evaluation of a near-peer-facilitated medical ethics curriculum for first-year medical students: a pilot study.

INTRODUCTION: The primary objectives of this study were to implement a novel near-peer-facilitated case-based medical ethics curriculum intended for the audience of a large cohort of first-year medical students (n = 193) and to objectively evaluate the immediate efficacy of the curriculum based on pre- and post-session survey responses to ethical quandaries. METHODS: Two near-peer-facilitated medical ethics case discussion sessions were included in the first-year curriculum during the 2017-2018 academic year. The sessions were designed and led by second-year medical student facilitators under the direction of a faculty mentor and were presented as a year-long curricular thread. First-year students were asked to complete pre- and post-session surveys with ethical questions relevant to each case and session. Students were additionally asked to measure the contribution of discussion sessions to their development as a future physician. RESULTS: Post-session survey results showed that students had a better understanding of specific ethical issues immediately following discussion sessions (p<0.0001). Over three-quarters of students indicated that the near-peer-led medical ethics case discussions contributed somewhat or very much to their development as a future physician. Anecdotal feedback from second-year medical students also suggested that their involvement as facilitators was beneficial to their educational development. CONCLUSION: Near-peer-facilitated case discussions were an effective strategy for teaching medical ethics to first-year medical students with demonstrated objective improvements in ethical decision-making. Additionally, near-peer discussions of ethical cases and principles with first-year medical students aided in subjective measures of professional development.

A logic framework for evaluating social determinants of health interventions in primary care.

BACKGROUND: Increasing efforts have been made in primary care settings to screen for a broad array of social determinants of health including inadequate food and nutrition, lack of education, unemployment, and inadequate housing, and to refer patients to community resources. Core tenets of primary care include integration with community resources. METHODS: In the course of designing a randomized controlled trial of the effectiveness of a social determinants of health intervention aimed at adult, at-risk, African American primary care clinic patients, our research team developed a logic model to assist with the evaluation of the intervention. RESULTS: In this article, we describe the logic model including elements of the intervention, mediator variables, and outcome variables. CONCLUSIONS: The proposed logic framework is likely to be helpful for planning, conducting, and evaluating social determinants of health interventions in primary care settings.