We are what we eat: macrophages and efferocytosis.

Liebold et al. recently revealed how the identity of dying cells drives distinct changes to the macrophages which engulf and clear them, a process known as efferocytosis. During infection with the helminth Schistosoma mansoni, liver macrophages recapitulate these phenotypes, mediated by Axl/MerTK receptors and regulating egg burdens.

Surgical stabilization of rib fractures in the geriatric trauma population is associated with equivalent outcomes to a younger cohort: A propensity matched analysis.

BACKGROUND: Surgical stabilization of rib fractures (SSRF) improves outcomes in chest wall trauma. Geriatric patients are particularly vulnerable to poor outcomes; yet, this population is often excluded from SSRF studies. Further delineating patient outcomes by age is necessary to optimize care for the aging trauma population. METHODS: A retrospective cohort study was conducted examining outcomes among patients aged 40+ for whom an SSRF consult was placed between 2017 and 2022 at a level 1 trauma center. Patients were categorized into geriatric (65+) and adult (40-64), as well as 80 years and older (80+) and 79 and younger (40-79). Patient outcomes were assessed comparing non-operative and operative management of chest wall trauma. Propensity matched analysis was performed to evaluate mortality differences between adult and geriatric patients who did and did not undergo SSRF. RESULTS: A total of 543 patients had an SSRF consult. Of these, 227 were 65+, and 73 were 80+. A total of 129 patients underwent SSRF (24 %). The percentage of patients undergoing SSRF did not vary between 40 and 64 and 65+ (23.7 % and 23.6 %, respectively, p = 0.97) or 40-79 and 80+ (24.0 vs 21.9, p = 0.69). Patients undergoing SSRF had higher chest injury burden and were more likely to require mechanical ventilation and ICU level care on admission. Overall, in-hospital mortality rate was 4.6 %. Among patients who underwent SSRF, mortality rate did not significantly differ between 65+ and 40-64 (7.8% vs 2.7 %, p = 0.18) or 80+ and 40-79 (6.3% vs 4.6 %, p = 0.77). This remained true in propensity matched analysis. CONCLUSION: Geriatric and octogenarian patients with rib fractures underwent SSRF at similar rates and achieved equivalent outcomes to their younger counterparts. SSRF did not differentially affect mortality outcomes based on age group in propensity matched analysis. SSRF is safe for geriatric patients including octogenarians.

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.

Eastern Association for the Surgery of Trauma (EAST) System Wellness White Paper: An Evaluation of Wellness from a Systems Perspective.

ABSTRACT: Providers are charged with responsibility to maintain their own health and wellness [1,2]. Wellness or wellbeing is multifactorial and complex, but the construct lacks clarity [3]. Wellness can impact patient safety and it has significant financial implications. Both financial and healthcare industries have recognized this multifactorial issue, and have applied research and resources to the issue of employee wellness and wellbeing [4]. Thus, solutions to sustainable change must be multi-tiered and intentional [1,5].We reviewed the wellness literature with a focus on systems to provide a framework for consensus-building for a quality Acute Care Surgery system. Within this review we highlight several categories for consideration: 1) provider wellness 2) culture of safety, 3)learning health systems, and 4) organizational perspectives, Figure 1. Finally, we provide specific system recommendations for the Acute Care Surgery practice. We aim to support personal safety, longevity, and preserve our workforce by creating a system that works for its providers.

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.

Development of Good Manufacturing Practice-Compatible Isolation and Culture Methods for Human Olfactory Mucosa-Derived Mesenchymal Stromal Cells.

Demyelination in the central nervous system (CNS) resulting from injury or disease can cause loss of nerve function and paralysis. Cell therapies intended to promote remyelination of axons are a promising avenue of treatment, with mesenchymal stromal cells (MSCs) a prominent candidate. We have previously demonstrated that MSCs derived from human olfactory mucosa (hOM-MSCs) promote myelination to a greater extent than bone marrow-derived MSCs (hBM-MSCs). However, hOM-MSCs were developed using methods and materials that were not good manufacturing practice (GMP)-compliant. Before considering these cells for clinical use, it is necessary to develop a method for their isolation and expansion that is readily adaptable to a GMP-compliant environment. We demonstrate here that hOM-MSCs can be derived without enzymatic tissue digestion or cell sorting and without culture antibiotics. They grow readily in GMP-compliant media and express typical MSC surface markers. They robustly produce CXCL12 (a key secretory factor in promoting myelination) and are pro-myelinating in in vitro rodent CNS cultures. GMP-compliant hOM-MSCs are comparable in this respect to those grown in non-GMP conditions. However, when assessed in an in vivo model of demyelinating disease (experimental autoimmune encephalitis, EAE), they do not significantly improve disease scores compared with controls, indicating further pre-clinical evaluation is necessary before their advancement to clinical trials.

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.

Extracorporeal Membrane Oxygenation During Surgical Stabilization of Rib Fractures in Successful Management of Severe Blunt Chest Trauma.

Early surgical stabilization of rib fracture (SSRF) improves outcomes in patients with flail physiology and severely displaced fractures. We present two cases of patients with severe chest injury and large flail segment who underwent SSRF while on veno-venous extracorporeal membrane oxygenation (VV-ECMO). The patients developed respiratory failure within 24 hours of admission requiring VV-ECMO. The extent of their chest wall injury limited pulmonary mechanics prohibiting transition off VV-ECMO. Therefore, SSRF was performed on hospital days 2 and 3 and while on VV-ECMO support. Stabilizing the chest wall allowed for improved ventilation and successful decannulation from VV-ECMO on postoperative days 3 and 4. Ultimately, both achieved a functional recovery and were discharged home. These cases demonstrate a unique thoracic damage control strategy wherein SSRF is performed while on VV-ECMO. Improving chest stability and pulmonary mechanics with SSRF allowed for safe transition off VV-ECMO and achieved a favorable long-term outcome.

The helminth TGF-ß mimic TGM4 is a modular ligand that binds CD44, CD49d and TGF-ß receptors to preferentially target myeloid cells.

The murine helminth parasite Heligmosomoides polygyrus expresses a family of modular proteins which, replicating the functional activity of the immunomodulatory cytokine TGF-ß, have been named TGM (TGF-ß Μimic). Multiple domains bind to different receptors, including TGF-ß receptors TßRI (ALK5) and TßRII through domains 1-3, and prototypic family member TGM1 binds the cell surface co-receptor CD44 through domains 4-5. This allows TGM1 to induce T lymphocyte Foxp3 expression, characteristic of regulatory (Treg) cells, and to activate a range of TGF-ß-responsive cell types. In contrast, a related protein, TGM4, targets a much more restricted cell repertoire, primarily acting on myeloid cells, with less potent effects on T cells and lacking activity on other TGF-ß-responsive cell types. TGM4 binds avidly to myeloid cells by flow cytometry, and can outcompete TGM1 for cell binding. Analysis of receptor binding in comparison to TGM1 reveals a 10-fold higher affinity than TGM1 for TGFßR-I (TßRI), but a 100-fold lower affinity for TßRII through Domain 3. Consequently, TGM4 is more dependent on co-receptor binding; in addition to CD44, TGM4 also engages CD49d (Itga4) through Domains 1-3, as well as CD206 and Neuropilin-1 through Domains 4 and 5. TGM4 was found to effectively modulate macrophage populations, inhibiting lipopolysaccharide-driven inflammatory cytokine production and boosting interleukin (IL)-4-stimulated responses such as Arginase-1 in vitro and in vivo. These results reveal that the modular nature of TGMs has allowed the fine tuning of the binding affinities of the TßR- and co-receptor binding domains to establish cell specificity for TGF-ß signalling in a manner that cannot be attained by the mammalian cytokine.

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.

CD44 acts as a coreceptor for cell-specific enhancement of signaling and regulatory T cell induction by TGM1, a parasite TGF-ß mimic.

Long-lived parasites evade host immunity through highly evolved molecular strategies. The murine intestinal helminth, Heligmosomoides polygyrus, down-modulates the host immune system through release of an immunosuppressive TGF-ß mimic, TGM1, which is a divergent member of the CCP (Sushi) protein family. TGM1 comprises 5 domains, of which domains 1-3 (D1/2/3) bind mammalian TGF-ß receptors, acting on T cells to induce Foxp3+ regulatory T cells; however, the roles of domains 4 and 5 (D4/5) remain unknown. We noted that truncated TGM1, lacking D4/5, showed reduced potency. Combination of D1/2/3 and D4/5 as separate proteins did not alter potency, suggesting that a physical linkage is required and that these domains do not deliver an independent signal. Coprecipitation from cells treated with biotinylated D4/5, followed by mass spectrometry, identified the cell surface protein CD44 as a coreceptor for TGM1. Both full-length and D4/5 bound strongly to a range of primary cells and cell lines, to a greater degree than D1/2/3 alone, although some cell lines did not respond to TGM1. Ectopic expression of CD44 in nonresponding cells conferred responsiveness, while genetic depletion of CD44 abolished enhancement by D4/5 and ablated the ability of full-length TGM1 to bind to cell surfaces. Moreover, CD44-deficient T cells showed attenuated induction of Foxp3 by full-length TGM1, to levels similar to those induced by D1/2/3. Hence, a parasite protein known to bind two host cytokine receptor subunits has evolved a third receptor specificity, which serves to raise the avidity and cell type-specific potency of TGF-ß signaling in mammalian cells.

How do we PI? Results of an EAST quality, patient safety, and outcomes survey.

Background: Quality improvement is a cornerstone for any verified trauma center. Conducting effective quality and performance improvement, however, remains a challenge. In this study, we sought to better explore the landscape and challenges facing the members of the Eastern Association for the Surgery of Trauma (EAST) through a survey. Methods: A survey was designed by the EAST Quality Patient Safety and Outcomes Committee. It was reviewed by the EAST Research and Scholarship Committee and then distributed to 2511 EAST members. The questions were designed to understand the frequency, content, and perceptions surrounding quality improvement processes. Results: There were 151 respondents of the 2511 surveys sent (6.0%). The majority were trauma faculty (55%) or trauma medical directors (TMDs) (37%) at American College of Surgeons level I (62%) or II (17%) trauma centers. We found a wide variety of resources being used across hospitals with the majority of cases being identified by a TMD or attending (81%) for a multidisciplinary peer review (70.2%). There was a statistically significant difference in the perception of the effectiveness of the quality improvement process with TMDs being more likely to describe their process as moderately or very effective compared with their peers (77.5% vs. 57.7%, p=0.026). The 'Just Culture' model appeared to have a positive effect on the process improvement environment, with providers less likely to report a non-conducive environment (10.9% vs. 27.6%, p=0.012) and less feelings of assigning blame (3.1% vs. 13.8%, p=0.026). Conclusion: Case review remains an essential but challenging process. Our survey reveals a need to continue to advocate for appropriate time and resources to conduct strong quality improvement processes. Level of evidence: Epidemiological study, level III.

Does lower extremity fracture fixation technique influence neurologic outcomes in patients with traumatic brain injury? The EAST Brain vs. Bone multicenter trial.

OBJECTIVE: This study aimed to determine whether lower extremity fracture fixation technique and timing (≤24 vs. >24 hours) impact neurologic outcomes in TBI patients. METHODS: A prospective observational study was conducted across 30 trauma centers. Inclusion criteria were age 18 years and older, head Abbreviated Injury Scale (AIS) score of >2, and a diaphyseal femur or tibia fracture requiring external fixation (Ex-Fix), intramedullary nailing (IMN), or open reduction and internal fixation (ORIF). The analysis was conducted using analysis of variamce, Kruskal-Wallis, and multivariable regression models. Neurologic outcomes were measured by discharge Ranchos Los Amigos Revised Scale (RLAS-R). RESULTS: Of the 520 patients enrolled, 358 underwent Ex-Fix, IMN, or ORIF as definitive management. Head AIS was similar among cohorts. The Ex-Fix group experienced more severe lower extremity injuries (AIS score, 4-5) compared with the IMN group (16% vs. 3%, p = 0.01) but not the ORIF group (16% vs. 6%, p = 0.1). Time to operative intervention varied between the cohorts with the longest time to intervention for the IMN group (median hours: Ex-Fix, 15 [8-24] vs. ORIF, 26 [12-85] vs. IMN, 31 [12-70]; p < 0.001). The discharge RLAS-R score distribution was similar across the groups. After adjusting for confounders, neither method nor timing of lower extremity fixation influenced the discharge RLAS-R. Instead, increasing age and head AIS score were associated with a lower discharge RLAS-R score (odds ratio [OR], 1.02; 95% confidence interval [CI], 1.002-1.03 and OR, 2.37; 95% CI, 1.75-3.22), and a higher Glasgow Coma Scale motor score on admission (OR, 0.84; 95% CI, 0.73-0.97) was associated with higher RLAS-R score at discharge. CONCLUSION: Neurologic outcomes in TBI are impacted by severity of the head injury and not the fracture fixation technique or timing. Therefore, the strategy of definitive fixation of lower extremity fractures should be dictated by patient physiology and the anatomy of the injured extremity and not by the concern for worsening neurologic outcomes in TBI patients. LEVEL OF EVIDENCE: Prognostic and Epidemiological; Level III.

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.

Implementation of a Gastrografin Order Set for Small Bowel Obstruction Across a Health System: A Cohort Study.

BACKGROUND: The use of Gastrografin (GG) in the management of adhesive small bowel obstruction (SBO) has been shown to decrease the length of stay and operative intervention. METHODS: This retrospective cohort study examined patients with an SBO diagnosis prior to implementation (PRE, January 2017-January 2019) and following implementation (POST, January 2019-May 2021) of a GG challenge order set made available across 9 hospitals within a health care system. Primary outcomes were utilization of the order set across facilities and over time. Secondary outcomes included time to surgery for operative patients, rate of surgery, nonoperative length of stay, and 30-day readmission. Standard descriptive, univariate, and multivariable regression analyses were performed. RESULTS: PRE cohort had 1746 patients and POST had 1889. The utilization of GG increased from 14% to 49.5% following implementation. Significant variability existed within the hospital system with utilization at each individual hospital from 11.5% to 60%. There was an increase in surgical intervention (13.9% vs 16.4%, P = .04) and decrease in nonoperative LOS (65.6 vs 59.9 hours, P < .001) following implementation. For POST patients, multivariable linear regression showed significant reduction in nonoperative length of stay (-23.1 hours, P < .001) but no significant difference in time to surgery (-19.6 hours, P = .08). DISCUSSION: The availability of a standardized order set for SBO can result in increased Gastrografin administration across hospital settings. The implementation of a Gastrografin order set was associated with decreased length of stay in nonoperative patients.

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.

Modulation of haematopoiesis by protozoal and helminth parasites.

During inflammation, haematopoietic stem cells (HSCs) in the bone marrow (BM) and periphery rapidly expand and preferentially differentiate into myeloid cells that mediate innate immune responses. HSCs can be directed into quiescence or differentiation by sensing alterations to the haematopoietic niche, including cytokines, chemokines, and pathogen-derived products. Most studies attempting to identify the mechanisms of haematopoiesis have focused on bacterial and viral infections. From intracellular protozoan infections to large multicellular worms, parasites are a global health burden and represent major immunological challenges that remain poorly defined in the context of haematopoiesis. Immune responses to parasites vary drastically, and parasites have developed sophisticated immunomodulatory mechanisms that allow development of chronic infections. Recent advances in imaging, genomic sequencing, and mouse models have shed new light on how parasites induce unique forms of emergency haematopoiesis. In addition, parasites can modify the haematopoiesis in the BM and periphery to improve their survival in the host. Parasites can also induce long-lasting modifications to HSCs, altering future immune responses to infection, inflammation or transplantation, a term sometimes referred to as central trained immunity. In this review, we highlight the current understanding of parasite-induced haematopoiesis and how parasites target this process to promote chronic infections.

Acute Care Surgeons Spend More Time than General Surgeons on the Electronic Health Record (EHR).

BACKGROUND: The paradigm of Acute Care Surgery (ACS) emerged in response to decreasing operative opportunities for trauma surgeons and increasing need for surgical coverage in disciplines to which the expertise of trauma surgeons naturally extends. While the continued evolution of this specialty remains largely beneficial, unintended consequences may have arisen along the way. One aspect of ACS that remains to be thoroughly investigated is the impact of the electronic health record (EHR). The purpose of this study is to objectively quantify EHR usage for ACS and compare it to other general surgery specialties. METHODS: EHR user data were collected for fifteen ACS attendings and thirty-seven general surgery attendings from October 2014 to September 2019. Comparative analysis was conducted using two-tailed t-tests. Subgroup analysis was conducted for subspecialties included in the general surgery group. RESULTS: ACS attendings opened almost 3 times as many charts as general surgery attendings per month (180 vs 64 charts/month, P < .0001), and ultimately spent more time on the EHR as a result (10 vs 6.4 hours/month, P < .0001). Documentation was the most time consuming EHR task for both groups. Although ACS attendings spent less overall time per patient chart, the proportion of time spent on certain EHR tasks was similar to that of general surgery colleagues. DISCUSSION: The EHR imposes a disproportionate burden on ACS attendings compared to their general surgery counterparts, and additional study is warranted to improve usage. EHR usage burden has workforce implications for trainees considering a career in ACS.