Tn-seq of the Candida glabrata reference strain CBS138 reveals epigenetic plasticity, structural variation, and intrinsic mechanisms of resistance to micafungin.

C. glabrata is an opportunistic pathogen that can resist common antifungals and rapidly acquire multidrug resistance. A large amount of genetic variation exists between isolates, which complicates generalizations. Portable Tn-seq methods can efficiently provide genome-wide information on strain differences and genetic mechanisms. Using the Hermes transposon, the CBS138 reference strain and a commonly studied derivative termed 2001 were subjected to Tn-seq in control conditions and after exposure to varying doses of the clinical antifungal micafungin. The approach revealed large differences between these strains, including a 131 kb tandem duplication and a variety of fitness differences. Additionally, both strains exhibited up to 1000-fold increased transposon accessibility in subtelomeric regions relative to the BG2 strain, indicative of open subtelomeric chromatin in these isolates and large epigenetic variation within the species. Unexpectedly, the Pdr1 transcription factor conferred resistance to micafungin through targets other than CDR1 . Other micafungin resistance pathways were also revealed including mannosyltransferase activity and biosynthesis of the lipid precursor sphingosine, the drugging of which by SDZ 90-215 or myriocin enhanced the potency of micafungin in vitro . These findings provide insights into complexity of the C. glabrata species as well as strategies for improving antifungal efficacy. Summary: Candida glabrata is an emerging pathogen with large genetic diversity and genome plasticity. The type strain CBS138 and a laboratory derivative were mutagenized with the Hermes transposon and profiled using Tn-seq. Numerous genes that regulate innate and acquired resistance to an important clinical antifungal were uncovered, including a pleiotropic drug resistance gene (PDR1) and a duplication of part of one chromosome. Compounds that target PDR1 and other genes may augment the potency of existing antifungals.

Handoffs and Transitions of Care: A Systematic Review, Meta-Analysis, and Practice Management Guideline from the Eastern Association for the Surgery of Trauma.

BACKGROUND: The Joint Commission reports at least half of communication breakdowns occur during handovers or transitions of care. There is no consensus on how best to approach the transfer of care within Acute Care Surgery (ACS). We conduct a systematic review and meta-analysis of the current data on handoffs and transitions of care in ACS patients and evaluate the impact of standardization and formalized communication processes. METHODS: Clinically relevant questions regarding handoffs and transitions of care with clearly defined patient Population(s), Intervention(s), Comparison(s), and appropriately selected Outcomes (PICO) were determined. These centered around specific transitions of care within the setting of ACS - specifically perioperative interactions, EMS and trauma team interactions, and intra/inter floor and ICU interactions. A systematic literature review and meta-analysis was conducted utilizing the Grading of Recommendations Assessment, Development and Evaluation (GRADE) methodology. RESULTS: A total of 10 studies were identified for analysis. These included 5,113 patients in the standardized handoff group and 5,293 in the current process group. Standardized handoffs reduced handover errors for perioperative interactions and preventable adverse events for intra/inter floor and ICU interactions. There was insufficient data to evaluate outcomes of clinical complications and medical errors. CONCLUSION: We conditionally recommend a standardized handoff in in the field of ACS, including perioperative interactions, EMS and trauma team interactions, as well as intra-inter floor and ICU interactions. LEVEL OF EVIDENCE: Guideline; Systematic review/meta-analysis, Level III.

Calcineurin-dependent contributions to fitness in the opportunistic pathogen Candida glabrata.

The protein phosphatase calcineurin is vital for the virulence of the opportunistic fungal pathogen Candida glabrata. The host-induced stresses that activate calcineurin signaling are unknown, as are the targets of calcineurin relevant to virulence. To potentially shed light on these processes, millions of transposon insertion mutants throughout the genome of C. glabrata were profiled en masse for fitness defects in the presence of FK506, a specific inhibitor of calcineurin. Eighty-seven specific gene deficiencies depended on calcineurin signaling for full viability in vitro both in wild-type and pdr1∆ null strains lacking pleiotropic drug resistance. Three genes involved in cell wall biosynthesis (FKS1, DCW1, FLC1) possess co-essential paralogs whose expression depended on calcineurin and Crz1 in response to micafungin, a clinical antifungal that interferes with cell wall biogenesis. Interestingly, 80% of the FK506-sensitive mutants were deficient in different aspects of vesicular trafficking, such as endocytosis, exocytosis, sorting, and biogenesis of secretory proteins in the endoplasmic reticulum (ER). In response to the experimental antifungal manogepix that blocks GPI-anchor biosynthesis in the ER, calcineurin signaling increased and strongly prevented cell death independent of Crz1, one of its major targets. Comparisons between manogepix, micafungin, and the ER-stressing tunicamycin reveal a correlation between the degree of calcineurin signaling and the degree of cell survival. These findings suggest that calcineurin plays major roles in mitigating stresses of vesicular trafficking. Such stresses may arise during host infection and in response to antifungal therapies.IMPORTANCECalcineurin plays critical roles in the virulence of most pathogenic fungi. This study sheds light on those roles in the opportunistic pathogen Candida glabrata using a genome-wide analysis in vitro. The findings could lead to antifungal developments that also avoid immunosuppression.

Early warning: End-tidal carbon dioxide is associated with central venous oxygenation under continuous cardiorespiratory monitoring in a porcine model of hemorrhagic shock and resuscitation.

BACKGROUND: End-tidal carbon dioxide (ETCO2) has previously shown promise as a predictor of shock severity and mortality in trauma. ETCO2 monitoring is non-invasive, real-time, and readily available in prehospital settings, but the temporal relationship of ETCO2 to systemic oxygen transport has not been thoroughly investigated in the context of hemorrhagic shock. METHODS: A validated porcine model of hemorrhagic shock and resuscitation was used in male Yorkshire swine (N â€‹= â€‹7). Both ETCO2 and central venous oxygenation (SCVO2) were monitored and recorded continuously in addition to other traditional hemodynamic variables. RESULTS: Linear regression analysis showed that ETCO2 was associated with ScvO2 both throughout the experiment (ߠ​= â€‹1.783, 95% confidence interval (CI) [1.552-2.014], p â€‹< â€‹0.001) and during the period of most rapid hemorrhage (ߠ​= â€‹4.896, 95% CI [2.416-7.377], p â€‹< â€‹0.001) when there was a marked decrease in ETCO2. CONCLUSIONS: ETCO2 and ScvO2 were closely associated during rapid hemorrhage and continued to be temporally associated throughout shock and resuscitation.

Redefining pleiotropic drug resistance in a pathogenic yeast: Pdr1 functions as a sensor of cellular stresses in Candida glabrata.

Candida glabrata is a prominent opportunistic fungal pathogen of humans. The increasing incidence of C. glabrata infections is attributed to both innate and acquired resistance to antifungals. Previous studies suggest the transcription factor Pdr1 and several target genes encoding ABC transporters are critical elements of pleiotropic defense against azoles and other antifungals. This study utilizes Hermes transposon insertion profiling to investigate Pdr1-independent and Pdr1-dependent mechanisms that alter susceptibility to the frontline antifungal fluconazole. Several new genes were found to alter fluconazole susceptibility independent of Pdr1 (CYB5, SSK1, SSK2, HOG1, TRP1). A bZIP transcription repressor of mitochondrial function (CIN5) positively regulated Pdr1 while hundreds of genes encoding mitochondrial proteins were confirmed as negative regulators of Pdr1. The antibiotic oligomycin activated Pdr1 and antagonized fluconazole efficacy likely by interfering with mitochondrial processes in C. glabrata. Unexpectedly, disruption of many 60S ribosomal proteins also activated Pdr1, thus mimicking the effects of the mRNA translation inhibitors. Cycloheximide failed to fully activate Pdr1 in a cycloheximide-resistant Rpl28-Q38E mutant. Similarly, fluconazole failed to fully activate Pdr1 in a strain expressing a low-affinity variant of Erg11. Fluconazole activated Pdr1 with very slow kinetics that correlated with the delayed onset of cellular stress. These findings are inconsistent with the idea that Pdr1 directly senses xenobiotics and support an alternative hypothesis where Pdr1 senses cellular stresses that arise only after engagement of xenobiotics with their targets. IMPORTANCE Candida glabrata is an opportunistic pathogenic yeast that causes discomfort and death. Its incidence has been increasing because of natural defenses to our common antifungal medications. This study explores the entire genome for impacts on resistance to fluconazole. We find several new and unexpected genes can impact susceptibility to fluconazole. Several antibiotics can also alter the efficacy of fluconazole. Most importantly, we find that Pdr1-a key determinant of fluconazole resistance-is not regulated directly through binding of fluconazole and instead is regulated indirectly by sensing the cellular stresses caused by fluconazole blockage of sterol biosynthesis. This new understanding of drug resistance mechanisms could improve the outcomes of current antifungals and accelerate the development of novel therapeutics.

Intercostal nerve cryoablation during surgical stabilization of rib fractures decreases post-operative opioid use, ventilation days, and intensive care days.

BACKGROUND: Intercostal nerve cryoablation is an adjunctive measure that has demonstrated pain control, decrease in opioid consumption, and decrease in hospital length of stay (LOS) in patients who undergo surgical stabilization of rib fractures (SSRF). METHODS: SSRF patients from January 2015 to September 2021 were retrospectively compared. All patients received multimodal pain regimens post-operatively and the independent variable was intraoperative cryoablation. RESULTS: 241 patients met inclusion criteria. 51 (21%) underwent intra-operative cryoablation during SSRF and 191 (79%) did not. Patients with standard treatment consumed 9.4 more daily MME (p = 0.035), consumed 73 percent more post-operative total MME (p = 0.001), spent 1.55 times as many days in the intensive care unit (p = 0.013), and spent 3.8 times as many days on the ventilator than patients treated with cryoablation, respectively. Overall hospital LOS, operative case time, pulmonary complications, MME at discharge, and numeric pain scores at discharge were no different (all p>0.05). CONCLUSION: Intercostal nerve cryoablation during SSRF is associated with fewer ventilator days, ICU LOS, total post-operative, and daily opioid use without increasing time in the operating room or perioperative pulmonary complications.

Redefining Pleiotropic Drug Resistance in a Pathogenic Yeast: Pdr1 Functions as a Sensor of Cellular Stresses in Candida glabrata.

Candida glabrata is a prominent opportunistic fungal pathogen of humans. The increasing incidence of C. glabrata infections is attributed to both innate and acquired resistance to antifungals. Previous studies suggest the transcription factor Pdr1 and several target genes encoding ABC transporters are critical elements of pleiotropic defense against azoles and other antifungals. This study utilizes Hermes transposon insertion profiling to investigate Pdr1-independent and Pdr1-dependent mechanisms that alter susceptibility to the frontline antifungal fluconazole. Several new genes were found to alter fluconazole susceptibility independent of Pdr1 ( CYB5 , SSK1 , SSK2 , HOG1 , TRP1 ). A bZIP transcription repressor of mitochondrial function ( CIN5 ) positively regulated Pdr1 while hundreds of genes encoding mitochondrial proteins were confirmed as negative regulators of Pdr1. The antibiotic oligomycin activated Pdr1 and antagonized fluconazole efficacy likely by interfering with mitochondrial processes in C. glabrata . Unexpectedly, disruption of many 60S ribosomal proteins also activated Pdr1, thus mimicking the effects of the mRNA translation inhibitors. Cycloheximide failed to fully activate Pdr1 in a cycloheximide-resistant Rpl28-Q38E mutant. Similarly, fluconazole failed to fully activate Pdr1 in a strain expressing a low-affinity variant of Erg11. Fluconazole activated Pdr1 with very slow kinetics that correlated with the delayed onset of cellular stress. These findings are inconsistent with the idea that Pdr1 directly senses xenobiotics and support an alternative hypothesis where Pdr1 senses cellular stresses that arise only after engagement of xenobiotics with their targets. Importance: Candida glabrata is an opportunistic pathogenic yeast that causes discomfort and death. Its incidence has been increasing because of natural defenses to our common antifungal medications. This study explores the entire genome for impacts on resistance to fluconazole. We find several new and unexpected genes can impact susceptibility to fluconazole. Several antibiotics can also alter the efficacy of fluconazole. Most importantly, we find that Pdr1 - a key determinant of fluconazole resistance - is not regulated directly through binding of fluconazole and instead is regulated indirectly by sensing the cellular stresses caused by fluconazole blockage of sterol biosynthesis. This new understanding of drug resistance mechanisms could improve the outcomes of current antifungals and accelerate the development of novel therapeutics.

Timing of venous thromboembolism chemoprophylaxis using objective hemoglobin criteria in blunt solid organ injury.

BACKGROUND: The purpose of this study was to evaluate the safety and efficacy of early venous thromboembolism (VTE) chemoprophylaxis following blunt solid organ injury. METHODS: A retrospective review of patients was performed for patients with blunt solid organ injury between 2009-2019. Enoxaparin was initiated when patients had <1g/dl Hemoglobin decline over a 24 h period. These patients were then categorized by initiation: ≤48 h and >48 h. RESULTS: There were 653 patients: 328 (50.2%) <48 h and 325 (49.8%) ≥48 h. Twenty-nine (4.4%) developed VTE. Patients in ≥48 h group suffered more frequent VTE events (6.5% vs 2.4%, p = 0.021). Non-operative failure occurred in 6 patients (1.9%) in ≥48 h group, and 5 patients (1.5%) < 48 h group. Blood transfusion following chemophrophylaxis initiation was required in 69 (21.3%) in ≥48 h group, and 46 (14.0%) in < 48 h group, occurring similarly between groups (p=0.021). CONCLUSION: Stable hemoglobin in the first 24 h is an efficacious, objective measure that allows early initiation of VTE chemoprophylaxis in solid organ injury. This practice is associated with earlier initiation of and fewer VTE events.

Emergency General Surgery Regionalization: Retrospective Cohort Study of Emergency General Surgery Patients at a Tertiary Care Center.

BACKGROUND: Emergency general surgery (EGS) patients presenting at tertiary care hospitals may bypass local hospitals with adequate resources. However, many tertiary care hospitals frequently operate at capacity. We hypothesized that understanding patient geographic origin could identify opportunities for enhanced system triage and optimization and be an important first step for EGS regionalization and care coordination that could potentially lead to improved utilization of resources. METHODS: We analyzed patient zip code and categorized EGS patients who were cared for at our tertiary care hospital as potentially divertible if the southern region hospital was geographically closer to their home, regional hospital admission (RHA) patients, or local admission (LA) patients if the tertiary care facility was closer. Baseline characteristics and outcomes were compared for RHA and LA patients. RESULTS: Of 14 714 EGS patients presenting to the tertiary care hospital, 30.2% were categorized as RHA patients. Overall, 1526 (10.4%) patients required an operation including 527 (34.5%) patients who were potentially divertible. Appendectomy and cholecystectomy comprised 66% of the operations for potentially divertible patients. Length of stay was not significantly different (P = .06) for RHA patients, but they did have lower measured short-term and long-term mortality when compared to their LA counterparts (P < .05). CONCLUSIONS: EGS diagnoses and patient geocode analysis can identify opportunities to optimize regional operating room and bed utilization. Understanding where EGS patients are cared for and factors that influenced care facility will be critical for next steps in developing EGS regionalization within our system.

Atrial Cannulation During Resuscitative Clamshell Thoracotomy.

BACKGROUND: Resuscitative thoracotomy and clamshell thoracotomy are performed in the setting of traumatic arrest with the intent of controlling hemorrhage, relieving tamponade, and providing open chest cardiopulmonary resuscitation. Historically, return of spontaneous circulation rates for penetrating traumatic arrest as well as out of hospital survival have been reported as low as 40% and 10%. Vascular access can be challenging in patients who have undergone a traumatic arrest and can be a limiting step to effective resuscitation. Atrial cannulation is a well-established surgical technique in cardiac surgery. Herein, we present a case series detailing our application of this technique in the context of acute trauma resuscitation during clamshell thoracotomy for traumatic arrest in the emergency department. METHODS: A retrospective case series of atrial cannulation during traumatic arrest was conducted in Charlotte, NC at Carolinas Medical Center an urban level 1 trauma center. RESULTS: The mean rate of return of spontaneous circulation in our series, 60%, was greater than previously published upper limit of return of spontaneous circulation for penetrating causes of traumatic arrest. DISCUSSION: Intravenous access can be difficult to establish in the hypovolemic and exsanguinating patient. Traditional methods of vascular access may be insufficient in the setting of central vascular injury. Atrial appendage cannulation during atrial cannulation is a quick and reliable technique to achieve vascular access that employs common methods from cardiac surgery to improve resuscitation of traumatic arrest.

The Association of the Affordable Care Act With Medicaid Enrollment Status and Costs of Care at a Level I Trauma Center in a Medicaid Non-expansion State.

INTRODUCTION: The intended purpose of the Patient Protection and Affordable Care Act (ACA) was to expand access to health care insurance for all Americans. In our study, we examine the association of Medicaid enrollment status, health care outcomes, and financial outcomes for trauma patients at a level I urban trauma center in a state that did not expand Medicaid coverage under the ACA. METHODS: We retrospectively reviewed trauma admissions from 2011 to 2016, via the trauma registry (n = 36,250). A subgroup of Medicaid patients (n = 8840) was identified and compared for changes in selected variables and demographics following ACA implementation. The association of Medicaid payor status, by 3 year average pre-ACA (n = 3516) and post-ACA (n = 3324), on patient outcomes, payments collected, and accrued costs of care were analyzed. RESULTS: Three-year Medicaid median actual payments decreased 7.5% following implementation of the ACA ($4072 vs. $3767, P < .01). In contrast, the Medicaid median total cost of care increased 23% ($3964 vs. $4882, P < .01). The rate of patients insured by Medicaid decreased (24.0% vs. 16.2%, P<.001). Patients were admitted longer (1 d vs. 2 d, P < .01), and more injured (ISS 5 vs. 6, P < .01). DISCUSSION: Medicaid payor status under the ACA was associated with a decrease in actual payments and an increase in total cost of care. Moreover, the divergence in actual payments collected with the increased total cost of care warrants examination to ascertain the root cause in efforts to reduce this widening gap.

Emergencies do not shut down during a pandemic: COVID pandemic impact on Acute Care Surgery volume and mortality at a level I trauma center.

BACKGROUND: The aim of this study was to evaluate the impact of the COVID-19 pandemic on volume and outcomes of Acute Care Surgery patients, and we hypothesized that inpatient mortality would increase due to COVID+ and resource constraints. METHODS: An American College of Surgeons verified Level I Trauma Center's trauma and operative emergency general surgery (EGS) registries were queried for all patients from Jan. 2019 to Dec. 2020. April 1st, 2020, was the demarcation date for pre- and during COVID pandemic. Primary outcome was inpatient mortality. RESULTS: There were 14,460 trauma and 3091 EGS patients, and month-over-month volumes of both remained similar (p > 0.05). Blunt trauma decreased by 7.4% and penetrating increased by 31%, with a concomitant 25% increase in initial operative management (p < 0.001). Despite this, trauma (3.7%) and EGS (2.9-3.0%) mortality rates remained stable which was confirmed on multivariate analysis; p > 0.05. COVID + mortality was 8.8% and 3.7% in trauma and EGS patients, respectively. CONCLUSION: Acute Care Surgeons provided high quality care to trauma and EGS patients during the pandemic without allowing excess mortality despite many hardships and resource constraints.

Yeast cell death pathway requiring AP-3 vesicle trafficking leads to vacuole/lysosome membrane permeabilization.

Unicellular eukaryotes have been suggested as undergoing self-inflicted destruction. However, molecular details are sparse compared with the mechanisms of programmed/regulated cell death known for human cells and animal models. Here, we report a molecular cell death pathway in Saccharomyces cerevisiae leading to vacuole/lysosome membrane permeabilization. Following a transient cell death stimulus, yeast cells die slowly over several hours, consistent with an ongoing molecular dying process. A genome-wide screen for death-promoting factors identified all subunits of the AP-3 complex, a vesicle trafficking adapter known to transport and install newly synthesized proteins on the vacuole/lysosome membrane. To promote cell death, AP-3 requires its Arf1-GTPase-dependent vesicle trafficking function and the kinase Yck3, which is selectively transported to the vacuole membrane by AP-3. Video microscopy revealed a sequence of events where vacuole permeability precedes the loss of plasma membrane integrity. AP-3-dependent death appears to be conserved in the human pathogenic yeast Cryptococcus neoformans.

Prehospital End-Tidal CO2: A Superior Marker for Mortality Risk in the Acutely Injured Patient.

BACKGROUND: Emergency medical personnel must expeditiously triage acutely injured patients to the appropriate medical facility. Efficient and objective variables to facilitate this process and provide information to the receiving trauma center are needed. Currently, multiple variables are used to prognosticate injury severity and risk of mortality including vital signs, mental status, lactate, and base excess. We investigated the prehospital use of end-tidal carbon dioxide (ETCO2) as a noninvasive physiologic measure that can be obtained in the acutely injured patient. METHODS: We performed a retrospective analysis of 557 acutely injured patients over 2 years at a Level 1 trauma center. All patients arriving as trauma activations with ETCO2 measurements were included in analysis. End-tidal carbon dioxide measurements were categorized as low, normal, and high based on reference levels. Mortality was the primary outcome. Secondary receiver operator curves (ROC) for base excess, venous lactate, blood pressure, and venous pH were compared. We hypothesized ETCO2 levels would be able to predict mortality. RESULTS: End-tidal carbon dioxide levels conferred a mortality rate of 38%, 17.3%, and 2.9% for low, normal, and high, respectively (P < .001). Receiver operator curve analysis produced an area under the curve predictive value for ETCO2 (.748) which was superior to lactate (.660), SBP (.578), pH (.560), and base excess (.497). DISCUSSION: End-tidal carbon dioxide is a more sensitive and specific predictor of mortality in the acutely injured patient compared to venous lactate, base deficit, blood pressure, or venous pH. Additional studies are needed to determine if ETCO2 can be used as an effective prehospital adjunct to prevent mortality in acutely injured patients.

Attention to detail: A dedicated rib fracture consultation service leads to earlier operation and improved clinical outcomes.

BACKGROUND: Surgical stabilization of rib fractures (SSRF) has been correlated with improved outcomes, including decreased length of stay (LOS). We hypothesized that an SSRF consultation service would increase the frequency of SSRF and improve outcomes. METHODS: A prospective observational study was performed to compare outcomes before and after implementing an SSRF service. Primary outcome was time from admission to surgery; secondary outcomes included LOS, mortality and morphine milligram equivalents (MME) prescribed at discharge. RESULTS: 1865 patients met consultation criteria and 128 patients underwent SSRF. Mortality decreased (6.3% vs. 3%) and patients were prescribed fewer MME at discharge (328 MME vs. 124 MME) following implementation. For the operative cohort, time from admission to surgery decreased by 1.72 days and ICU LOS decreased by 2.6 days. CONCLUSION: Establishment of an SSRF service provides a mechanism to maximize capture and evaluation of operative candidates, provide earlier intervention, and improve patient outcomes. Additional study to determine which elements and techniques are most beneficial is warranted. LEVEL OF EVIDENCE: III.

Emergency medicine residents spend over 7.5 months of their 3-year residency on the electronic health record.

BACKGROUND: Use of the electronic health record (EHR) is a standard component of modern patient care. Although EHRs have improved since inception, cumbersome workflows decrease the time for residents to spend on clinical and educational activities. This study aims to quantify the time spent interacting with the EHR during a 3-year emergency medicine (EM) residency. METHODS: System records of time spent actively engaged in EHR use were analyzed for 98 unique EM residents over a period of 5 years from July 2015 to June 2020. Time spent on the EHR was totaled to give a career time, with a "work month" defined as a 4-week period of 70.5 h per week, based on Accreditation Council for Graduate Medical Education work hour restrictions for EM residents. Engagement in specific activities such as chart review, documentation preparation, and order entry were separately analyzed. RESULTS: Over their 3-year training, a resident interacted with the EHR for 2,171 continuous hours. This amounts to 30.8 work weeks or 7.7 work months. Chart review was the most time-intensive activity at 11.42 weeks. Documentation accounted for 9.91 weeks, with an average career total of 7,280 notes created. Additionally, each resident spent 4.57 weeks on order entry, with 46,347 orders entered during training. While the number of charts opened increased after first year of residency, average time spent on each activity per patient decreased. CONCLUSIONS: This unique study quantifies the total time an EM resident spends on the EHR during a 3-year residency. Use of the EHR accounted for over 7.5 work months or nearly 21% of their training. Residents spend a substantial portion of their training interacting with the EHR and workflow improvements to reduce EHR time are critical for maximizing training time.

SARS-CoV-2 Seroprevalence and Drug Use in Trauma Patients from Six Sites in the United States.

In comparison to the general patient population, trauma patients show higher level detections of bloodborne infectious diseases, such as Hepatitis and Human Immunodeficiency Virus. In comparison to bloodborne pathogens, the prevalence of respiratory infections such as SARS-CoV-2 and how that relates with other variables, such as drug usage and trauma type, is currently unknown in trauma populations. Here, we evaluated SARS-CoV-2 seropositivity and antibody isotype profile in 2,542 trauma patients from six Level-1 trauma centers between April and October of 2020 during the first wave of the COVID-19 pandemic. We found that the seroprevalence in trauma victims 18-44 years old (9.79%, 95% confidence interval/CI: 8.33 - 11.47) was much higher in comparison to older patients (45-69 years old: 6.03%, 4.59-5.88; 70+ years old: 4.33%, 2.54 - 7.20). Black/African American (9.54%, 7.77 - 11.65) and Hispanic/Latino patients (14.95%, 11.80 - 18.75) also had higher seroprevalence in comparison, respectively, to White (5.72%, 4.62 - 7.05) and Non-Latino patients (6.55%, 5.57 - 7.69). More than half (55.54%) of those tested for drug toxicology had at least one drug present in their system. Those that tested positive for narcotics or sedatives had a significant negative correlation with seropositivity, while those on anti-depressants trended positive. These findings represent an important consideration for both the patients and first responders that treat trauma patients facing potential risk of respiratory infectious diseases like SARS-CoV-2.

Platelet Dysfunction after Traumatic Brain Injury: A Review.

Coagulopathy is a known sequela of traumatic brain injury (TBI) and can lead to increased morbidity and mortality. Platelet dysfunction has been identified as one of several etiologies of coagulopathy following TBI and has been associated with poor outcomes. Regardless of whether the platelet dysfunction occurs as a direct consequence of the injury or because of pre-existing medical comorbidities or medication use, accurate detection and monitoring of response to therapy is key to optimal patient care. Platelet transfusion has been proposed as a potential therapeutic intervention to treat platelet dysfunction, with several studies using platelet function assays to monitor response. The development of increasingly precise diagnostic testing is providing enhanced understanding of the specific derangement in the hemostatic process, allowing clinicians to provide patient-specific treatment plans. There is wide variability in the currently available literature on the incidence and clinical significance of platelet dysfunction following TBI, which creates challenges with developing evidence-based management guidelines. The relatively high prevalence of platelet inhibitor therapy serves as an additional confounding factor. In addition, the data are largely retrospective in nature. We performed a literature review to provide clarity on this clinical issue. We reviewed 348 abstracts, and included 97 manuscripts in our final literature review. Based on the currently available research, platelet dysfunction has been consistently demonstrated in patients with moderate-severe TBI. We recommend the use of platelet functional assays to evaluate patients with TBI. Platelet transfusion directed at platelet dysfunction may lead to improved clinical outcome. A randomized trial guided by implementation science could improve the applicability of these practices.

Identification of Essential Genes and Fluconazole Susceptibility Genes in Candida glabrata by Profiling Hermes Transposon Insertions.

Within the budding yeasts, the opportunistic pathogen Candida glabrata and other members of the Nakaseomyces clade have developed virulence traits independently from C. albicans and C. auris To begin exploring the genetic basis of C. glabrata virulence and its innate resistance to antifungals, we launched the Hermes transposon from a plasmid and sequenced more than 500,000 different semi-random insertions throughout the genome. With machine learning, we identified 1278 protein-encoding genes (25% of total) that could not tolerate transposon insertions and are likely essential for C. glabrata fitness in vitro Interestingly, genes involved in mRNA splicing were less likely to be essential in C. glabrata than their orthologs in S. cerevisiae, whereas the opposite is true for genes involved in kinetochore function and chromosome segregation. When a pool of insertion mutants was challenged with the first-line antifungal fluconazole, insertions in several known resistance genes (e.g., PDR1, CDR1, PDR16, PDR17, UPC2A, DAP1, STV1) and 15 additional genes (including KGD1, KGD2, YHR045W) became hypersensitive to fluconazole. Insertions in 200 other genes conferred significant resistance to fluconazole, two-thirds of which function in mitochondria and likely down-regulate Pdr1 expression or function. Knockout mutants of KGD2 and IDH2, which consume and generate alpha-ketoglutarate in mitochondria, exhibited increased and decreased resistance to fluconazole through a process that depended on Pdr1. These findings establish the utility of transposon insertion profiling in forward genetic investigations of this important pathogen of humans.

Comparing the utility of in vivo transposon mutagenesis approaches in yeast species to infer gene essentiality.

In vivo transposon mutagenesis, coupled with deep sequencing, enables large-scale genome-wide mutant screens for genes essential in different growth conditions. We analyzed six large-scale studies performed on haploid strains of three yeast species (Saccharomyces cerevisiae, Schizosaccaromyces pombe, and Candida albicans), each mutagenized with two of three different heterologous transposons (AcDs, Hermes, and PiggyBac). Using a machine-learning approach, we evaluated the ability of the data to predict gene essentiality. Important data features included sufficient numbers and distribution of independent insertion events. All transposons showed some bias in insertion site preference because of jackpot events, and preferences for specific insertion sequences and short-distance vs long-distance insertions. For PiggyBac, a stringent target sequence limited the ability to predict essentiality in genes with few or no target sequences. The machine learning approach also robustly predicted gene function in less well-studied species by leveraging cross-species orthologs. Finally, comparisons of isogenic diploid versus haploid S. cerevisiae isolates identified several genes that are haplo-insufficient, while most essential genes, as expected, were recessive. We provide recommendations for the choice of transposons and the inference of gene essentiality in genome-wide studies of eukaryotic haploid microbes such as yeasts, including species that have been less amenable to classical genetic studies.