The electrostatic landscape of MHC-peptide binding revealed using inception networks.

Predictive modeling of macromolecular recognition and protein-protein complementarity represents one of the cornerstones of biophysical sciences. However, such models are often hindered by the combinatorial complexity of interactions at the molecular interfaces. Exemplary of this problem is peptide presentation by the highly polymorphic major histocompatibility complex class I (MHC-I) molecule, a principal component of immune recognition. We developed human leukocyte antigen (HLA)-Inception, a deep biophysical convolutional neural network, which integrates molecular electrostatics to capture non-bonded interactions for predicting peptide binding motifs across 5,821 MHC-I alleles. These predictions of generated motifs correlate strongly with experimental peptide binding and presentation data. Beyond molecular interactions, the study demonstrates the application of predicted motifs in analyzing MHC-I allele associations with HIV disease progression and patient response to immune checkpoint inhibitors. A record of this paper's transparent peer review process is included in the supplemental information.

Debridement, Antibiotics, and Implant Retention in Unicompartmental Knee Arthroplasty Infection.

BACKGROUND: This multicenter study sought to further investigate the method and outcome of debridement, antibiotics, and implant retention (DAIR) for the management of unicompartmental knee periprosthetic joint infection (PJI). METHODS: This retrospective study was performed on 52 patients who underwent DAIR for PJI of a unicompartmental knee arthroplasty (UKA) across 4 academic medical centers, all performed by fellowship-trained arthroplasty surgeons. Patient demographics, American Society of Anesthesiologists score, infecting organism, operative data, antibiotic data, and success in infection control at 1 year were collected. RESULTS: The average time from index surgery to diagnosis of PJI was 11.1 weeks (range, 1.4 to 48). There was no correlation between time of diagnosis and success at 1 year (R = 0.09, P = .46). There was an association between surgical synovectomy and the eradication of infection (R = 0.28, P = .04). Overall, there was an 80.8% (42 of 52) infection-controlled success rate at 1 year from the DAIR procedure. All DAIR failures went on to require another procedure, either 1-stage (2 of 10) or 2-stage (8 of 10) revision to total knee arthroplasty (TKA). Of the DAIR successes, 6 (14.3%) went on to require conversion to TKA for progression of arthritis within 5 years. CONCLUSIONS: This study demonstrates that DAIR is a safe and moderately effective procedure in the setting of acute PJI of UKA across institutions, with a success rate consistent with DAIR for TKA. The data suggest that a wide exposure and thorough synovectomy be incorporated during the DAIR UKA to improve the likelihood of successful eradication of PJI at the 1-year mark. LEVEL OF EVIDENCE: Level III.

An immunogenetic basis for lung cancer risk.

Cancer risk is influenced by inherited mutations, DNA replication errors, and environmental factors. However, the influence of genetic variation in immunosurveillance on cancer risk is not well understood. Leveraging population-level data from the UK Biobank and FinnGen, we show that heterozygosity at the human leukocyte antigen (HLA)-II loci is associated with reduced lung cancer risk in smokers. Fine-mapping implicated amino acid heterozygosity in the HLA-II peptide binding groove in reduced lung cancer risk, and single-cell analyses showed that smoking drives enrichment of proinflammatory lung macrophages and HLA-II+ epithelial cells. In lung cancer, widespread loss of HLA-II heterozygosity (LOH) favored loss of alleles with larger neopeptide repertoires. Thus, our findings nominate genetic variation in immunosurveillance as a critical risk factor for lung cancer.

Outcomes Following Intraoperative Calcar Fractures During Cementless Total Hip Arthroplasty.

BACKGROUND: Intraoperative calcar fractures (IOCFs) are an established complication of cementless total hip arthroplasty (THA). Prompt recognition and management may prevent subsequent postoperative complications. This study aimed to evaluate the outcomes and revision rates of THAs with IOCFs identified and managed intraoperatively. METHODS: There were 11,438 primary cementless THAs performed at a single institution from 2009 to 2022. Prospectively collected data on cases with an IOCF was compared to cases without the complication. The fracture group had a lower body mass index (26.9 versus 28.9 kg/m2; P = .01). Patient age, sex, and mean follow-up (3.2 (0 to 12.8) versus 3.5 years (0 to 14); P = .45) were similar between groups. RESULTS: An IOCF occurred in 62 of 11,438 (0.54%) cases. The THAs done via a direct anterior approach experienced the lowest rate of fractures (31 of 7,505, 0.4%) compared to postero-lateral (27 of 3,759, 0.7%; P = .03) and lateral (4 of 165, 2.4%; P < .01) approaches. Of the IOCFs, 48 of 62 (77%) were managed with cerclage cabling, 4 of 62 (6.5%) with intraoperative stem design change and cabling, 4 of 62 (6.5%) with restricted weight-bearing, and 6 of 62 (9.7%) with no modification to the standard postoperative protocol. The IOCF group experienced one case of postoperative component subsidence. No subjects in the IOCF cohort required revision, and rates were similar between groups (0 of 62, 0% versus 215 of 11,376, 1.9%; P = .63). Postoperative Hip dysfunction and Osteoarthritis Outcome Score for Joint Replacement scores were comparable (85.7 versus 86.4; P = .80). CONCLUSIONS: Cementless THA complicated by IOCF had similar postoperative revision rates and patient-reported outcome measures at early follow-up when compared to patients not experiencing this complication. Surgeons may use these data to provide postoperative counseling on expectations and outcomes following these rare intraoperative events.

Performance of Endoscopic Sleeve Gastroplasty by Obesity Class in the United States Clinical Setting.

INTRODUCTION: Endoscopic sleeve gastroplasty (ESG) has gained popularity over the past decade and has been adopted in both academic and private institutions globally. We present outcomes of the largest cohort of patients from the United States undergoing ESG and evaluate these according to obesity class. METHODS: We performed a retrospective analysis of adult patients who underwent ESG. Medical information was abstracted from the electronic record with weight records up to 2 years after ESG. Percent total body weight loss (%TBWL) at 6, 12, 18, and 24 months was calculated based on baseline weight at the procedure. SPSS (version 29.0) was used for all statistical analyses. RESULTS: A total of 1,506 patients from 7 sites were included (501 Class I obesity, 546 Class II, and 459 Class III). Baseline demographics differed according to obesity class due to differences in age, body mass index (BMI), height, sex distribution, and race. As early as 6 months post-ESG, mean BMI for each class dropped to the next lower class and remained there through 2 years. %TWBL achieved in the Class III group was significantly greater when compared with other classes at all time points. At 12 months, 83.2% and 60.9% of patients had ≥10% and ≥15% TBWL for all classes. There were no differences in adverse events between classes. DISCUSSION: Real-world data from a large cohort of patients of all BMI classes across the United States shows significant and sustained weight loss with ESG. ESG is safe to perform in a higher obesity class with acceptable midterm efficacy.

Structural insights into the modulation of coronavirus spike tilting and infectivity by hinge glycans.

Coronavirus spike glycoproteins presented on the virion surface mediate receptor binding, and membrane fusion during virus entry and constitute the primary target for vaccine and drug development. How the structure dynamics of the full-length spikes incorporated in viral lipid envelope correlates with the virus infectivity remains poorly understood. Here we present structures and distributions of native spike conformations on vitrified human coronavirus NL63 (HCoV-NL63) virions without chemical fixation by cryogenic electron tomography (cryoET) and subtomogram averaging, along with site-specific glycan composition and occupancy determined by mass spectrometry. The higher oligomannose glycan shield on HCoV-NL63 spikes than on SARS-CoV-2 spikes correlates with stronger immune evasion of HCoV-NL63. Incorporation of cryoET-derived native spike conformations into all-atom molecular dynamic simulations elucidate the conformational landscape of the glycosylated, full-length spike that reveals a role of hinge glycans in modulating spike bending. We show that glycosylation at N1242 at the upper portion of the stalk is responsible for the extensive orientational freedom of the spike crown. Subsequent infectivity assays implicated involvement of N1242-glyan in virus entry. Our results suggest a potential therapeutic target site for HCoV-NL63.

Outpatient Total Knee Arthroplasty From a Stand-Alone Surgery Center: Safe as the Hospital?

BACKGROUND: Literature suggests that outpatient arthroplasty may result in low rates of complications and readmissions. There is, however, a dearth of information on the relative safety of total knee arthroplasty (TKA) performed at stand-alone ambulatory surgery centers (ASCs) versus hospital outpatient (HOP) settings. We aimed to compare safety profiles and 90-day adverse events of these 2 cohorts. METHODS: Prospectively collected data were reviewed on all patients who underwent outpatient TKA from 2015 to 2022. The ASC and HOP groups were compared, and differences in demographics, complications, reoperations, revisions, readmissions, and emergency department (ED) visits within 90 days of surgery were analyzed. There were 4 surgeons who performed 4,307 TKAs during the study period, including 740 outpatient cases (ASC = 157; HOP = 583). The ASC patients were younger than HOP patients (ASC = 61 versus HOP = 65; P < .001). Body mass index and sex did not differ significantly between groups. RESULTS: Within 90 days, 44 (6%) complications occurred. No differences were observed between groups in rates of 90-day complications (ASC = 9 of 157, 5.7% versus HOP = 35 of 583, 6.0%; P = .899), reoperations (ASC = 2 of 157, 1.3% versus HOP = 3 of 583, 0.5%; P = .303), revisions (ASC = 0 of 157 versus HOP = 3 of 583, 0.5%; P = 1), readmissions (ASC = 3 of 157, 1.9% versus HOP = 8 of 583, 1.4%; P = .625), and ED visits (ASC = 1 of 157, 0.6% versus HOP = 3 of 583, 0.5%; P = .853). CONCLUSION: These results suggest that in appropriately selected patients, outpatient TKA can be safely performed in both ASC and HOP settings with similar low rates of 90-day complications, reoperations, revisions, readmissions, and ED visits.

Early Practice All-Cause Complications for Fellowship-Trained Anterior Hip Surgeons Are Not Increased When Compared to "Gold Standard" Experienced Posterior Approach Surgeons.

BACKGROUND: Increased complication rates have been reported during the learning curve for direct anterior approach (DAA) total hip arthroplasty (THA). However, emerging literature suggests that complications associated with the learning curve may be substantially reduced with fellowship training. METHODS: Our institutional database was queried to identify 2 groups: (1) 600 THAs comprised of the first 300 consecutive cases performed by 2 DAA fellowship-trained surgeons; and (2) 600 posterolateral approach (PA) THAs, including the most recent 300 primary cases performed by 2 experienced PA surgeons. All-cause complications, revision rates, reoperations, operative times, and transfusion rates were evaluated. RESULTS: Comparing DAA and PA cases, there were no significant differences in rates of all-cause complications (DAA = 18, 3.0% versus PA = 23, 3.8%; P = .43), periprosthetic fractures (DAA = 5, 0.8% versus PA = 10, 1.7%; P = .19), wound complications (DAA = 7, 1.2% versus PA = 2, 0.3%; P = .09), dislocations (DAA = 2, 0.3% versus PA = 8, 1.3%, P = .06), or revisions (DAA = 2, 0.3% versus PL = 5, 0.8%; P = .45) at 120 days postoperatively. There were 4 patients who required reoperation for wound complications, all within the DAA group (DAA = 4, 0.67% versus PA = 0; P = .045). Operative times were shorter in the DAA group (DAA <1.5 hours = 93% versus PA <1.5 hours = 86%; P < .01). No blood transfusions were given in either group. CONCLUSION: In this retrospective study, DAA THAs performed by fellowship-trained surgeons early in practice were not associated with higher complication rates compared to THAs performed by experienced PA surgeons. These results suggest that fellowship training may allow DAA surgeons to complete their learning curve period with complication rates similar to experienced PA surgeons.

High-resolution structures with bound Mn2+ and Cd2+ map the metal import pathway in an Nramp transporter.

Transporters of the Nramp (Natural resistance-associated macrophage protein) family import divalent transition metal ions into cells of most organisms. By supporting metal homeostasis, Nramps prevent diseases and disorders related to metal insufficiency or overload. Previous studies revealed that Nramps take on a LeuT fold and identified the metal-binding site. We present high-resolution structures of Deinococcus radiodurans (Dra)Nramp in three stable conformations of the transport cycle revealing that global conformational changes are supported by distinct coordination geometries of its physiological substrate, Mn2+, across conformations, and by conserved networks of polar residues lining the inner and outer gates. In addition, a high-resolution Cd2+-bound structure highlights differences in how Cd2+ and Mn2+ are coordinated by DraNramp. Complementary metal binding studies using isothermal titration calorimetry with a series of mutated DraNramp proteins indicate that the thermodynamic landscape for binding and transporting physiological metals like Mn2+ is different and more robust to perturbation than for transporting the toxic Cd2+ metal. Overall, the affinity measurements and high-resolution structural information on metal substrate binding provide a foundation for understanding the substrate selectivity of essential metal ion transporters like Nramps.

Timing and Factors Associated With Revision for Infection After Primary Total Knee Arthroplasty Based on American Joint Replacement Registry Data.

BACKGROUND: Infection following total knee arthroplasty (TKA) remains a challenging clinical problem. Using American Joint Replacement Registry data, this study examined factors related to the incidence and timing of infection. METHODS: Primary TKAs performed from January 2012 through December 2018 among patients ≥65 years of age at surgery were queried from the American Joint Replacement Registry and merged with Medicare data to enhance capture of revisions for infection. Multivariate Cox regressions incorporating patient, surgical, and institutional factors were used to produce hazard ratios (HRs) associated with revision for infection and mortality after revision for infection. RESULTS: Among 525,887 TKAs, 2,821 (0.54%) were revised for infection. Men had an increased risk of revision for infection at all-time intervals (≤90 days, HR = 2.06, 95% CI: 1.75-2.43, P < .0001; >90 days to 1 year, HR = 1.90, 95% CI: 1.58-2.28, P < .0001; >1 year, HR = 1.57, 95% CI: 1.37-1.79, P < .0001). TKAs performed for osteoarthritis had an increased risk of revision for infection at ≤90 days (HR = 2.01, 95% CI: 1.45-2.78, P < .0001) but not at later times. Mortality was more likely among patients who had a Charlson Comorbidity Index (CCI) ≥ 5 compared to those who had a CCI ≤ 2 (HR = 3.21, 95% CI: 1.35-7.63, P = .008). Mortality was also more likely among older patients (HR = 1.61 for each decade, 95% CI: 1.04-2.49, P = .03). CONCLUSION: Based on primary TKAs performed in the United States, men were found to have a persistently higher risk of revision for infection, while a diagnosis of osteoarthritis was associated with a significantly higher risk only during the first 90 days after surgery.

Integrated analyses reveal a hinge glycan regulates coronavirus spike tilting and virus infectivity.

Coronavirus spike glycoproteins presented on the virion surface mediate receptor binding, and membrane fusion during virus entry and constitute the primary target for vaccine and drug development. How the structure dynamics of the full-length spikes incorporated in viral lipid envelope correlates with the virus infectivity remains poorly understood. Here we present structures and distributions of native spike conformations on vitrified human coronavirus NL63 (HCoV-NL63) virions without chemical fixation by cryogenic electron tomography (cryoET) and subtomogram averaging, along with site-specific glycan composition and occupancy determined by mass spectroscopy. The higher oligomannose glycan shield on HCoV-NL63 spikes than on SARS-CoV-2 spikes correlates with stronger immune evasion of HCoV-NL63. Incorporation of cryoET-derived native spike conformations into all-atom molecular dynamic simulations elucidate the conformational landscape of the glycosylated, full-length spike that reveals a novel role of stalk glycans in modulating spike bending. We show that glycosylation at N1242 at the upper portion of the stalk is responsible for the extensive orientational freedom of the spike crown. Subsequent infectivity assays support the hypothesis that this glycan-dependent motion impacts virus entry. Our results suggest a potential therapeutic target site for HCoV-NL63.

A scoping review of distributed cognition in acute care clinical decision-making.

OBJECTIVES: In acute care settings, interactions between providers and tools drive clinical decision-making. Most studies of decision-making focus on individual cognition and fail to capture critical collaborations. Distributed Cognition (DCog) theory provides a framework for examining the dispersal of tasks among agents and artifacts, enhancing the investigation of decision-making and error. CONTENT: This scoping review maps the evidence collected in empiric studies applying DCog to clinical decision-making in acute care settings and identifies gaps in the existing literature. SUMMARY AND OUTLOOK: Thirty-seven articles were included. The majority (n=30) used qualitative methods (observations, interviews, artifact analysis) to examine the work of physicians (n=28), nurses (n=27), residents (n=16), and advanced practice providers (n=12) in intensive care units (n=18), operating rooms (n=7), inpatient units (n=7) and emergency departments (n=5). Information flow (n=30) and task coordination (n=30) were the most frequently investigated elements of DCog. Provider-artifact (n=35) and provider-provider (n=30) interactions were most explored. Electronic (n=18) and paper (n=15) medical records were frequently described artifacts. Seven prominent themes were identified. DCog is an underutilized framework for examining how information is obtained, represented, and transmitted through complex clinical systems. DCog offers mechanisms for exploring how technologies, like EMRs, and workspaces can help or hinder clinical decision-making.

Increased Microbial Diversity and Decreased Prevalence of Common Pathogens in the Gut Microbiomes of Wild Turkeys Compared to Domestic Turkeys.

Turkeys (Meleagris gallopavo) provide a globally important source of protein and constitute the second most important source of poultry meat in the world. Bacterial diseases are common in commercial poultry production, causing significant production losses for farmers. Due to the increasingly recognized problems associated with large-scale/indiscriminate antibiotic use in agricultural settings, poultry producers need alternative methods to control common bacterial pathogens. In this study, we compared the cecal microbiota of wild and domestic turkeys, hypothesizing that environmental pressures faced by wild birds may select for a disease-resistant microbial community. Sequence analyses of 16S rRNA genes amplified from cecal samples indicate that free-roaming wild turkeys carry a rich and variable microbiota compared to domestic turkeys raised on large-scale poultry farms. Wild turkeys also had very low levels of Staphylococcus, Salmonella, and Escherichia coli compared to domestic turkeys. E. coli strains isolated from wild and domestic turkey cecal samples also belong to distinct phylogenetic backgrounds and differ in their propensity to carry virulence genes. E. coli strains isolated from factory-raised turkeys were far more likely to carry genes for capsule (kpsII and kpsIII) or siderophore (iroN and fyuA) synthesis than were those isolated from wild turkeys. These results suggest that the microbiota of wild turkeys may provide colonization resistance against common poultry pathogens. IMPORTANCE Due to the increasingly recognized problems associated with antibiotic use in agricultural settings, poultry producers need alternative methods to control common bacterial pathogens. In this study, we compare the microbiota of wild and domestic turkeys. The results suggest that free-ranging wild turkeys carry a distinct microbiome compared to farm-raised turkeys. The microbiome of wild birds contains very low levels of poultry pathogens compared to that of farm-raised birds. The microbiomes of wild turkeys may be used to guide the development of new ways to control disease in large-scale poultry production.

Histology education in an integrated, time-restricted medical curriculum: Academic outcomes and students' study adaptations.

In an ever-changing medical curricular environment, time dedicated for anatomical education has been progressively reduced. This happened at the University of Michigan Medical School starting in 2016-2017 when preclinical medical education was condensed to one year. Histology instruction remained integrated in organ system courses but reduced to a lecture-only format without scheduling time for laboratory exercises, requiring students to study virtual histology slides on their own time. In accordance with the shortened instructional time, the number of histology examination questions was reduced more than twofold. This study analyzed students' histology examination results and assessed their motivation to learn histology and use of educational opportunities before and after these curricular changes were implemented. Students' motivation to learn histology and their evaluation of histology lectures increased in the new curriculum. However, students devoted less study time to studying histology. Students' cumulative histology examination scores were significantly lower in the new curriculum and the number of students with overall scores <75%, defined as a substandard performance, increased more than 15-fold. Academically weaker students' histology scores were disproportionately more affected. As medical educational strategies, priorities, and curricular frameworks continue to evolve, traditional didactic topics like histology will need to adapt to continue providing educational value to future health care providers.

ChAdOx1 interacts with CAR and PF4 with implications for thrombosis with thrombocytopenia syndrome.

Vaccines derived from chimpanzee adenovirus Y25 (ChAdOx1), human adenovirus type 26 (HAdV-D26), and human adenovirus type 5 (HAdV-C5) are critical in combatting the severe acute respiratory coronavirus 2 (SARS-CoV-2) pandemic. As part of the largest vaccination campaign in history, ultrarare side effects not seen in phase 3 trials, including thrombosis with thrombocytopenia syndrome (TTS), a rare condition resembling heparin-induced thrombocytopenia (HIT), have been observed. This study demonstrates that all three adenoviruses deployed as vaccination vectors versus SARS-CoV-2 bind to platelet factor 4 (PF4), a protein implicated in the pathogenesis of HIT. We have determined the structure of the ChAdOx1 viral vector and used it in state-of-the-art computational simulations to demonstrate an electrostatic interaction mechanism with PF4, which was confirmed experimentally by surface plasmon resonance. These data confirm that PF4 is capable of forming stable complexes with clinically relevant adenoviruses, an important step in unraveling the mechanisms underlying TTS.

Utility of a Telephone Triage Hotline in Response to the COVID-19 Pandemic: Longitudinal Observational Study.

BACKGROUND: During the initial months of the COVID-19 pandemic, rapidly rising disease prevalence in the United States created a demand for patient-facing information exchanges that addressed questions and concerns about the disease. One approach to managing increased patient volumes during a pandemic involves the implementation of telephone-based triage systems. During a pandemic, telephone triage hotlines can be employed in innovative ways to conserve medical resources and offer useful population-level data about disease symptomatology and risk factor profiles. OBJECTIVE: The aim of this study is to describe and evaluate the COVID-19 telephone triage hotline used by a large academic medical center in the midwestern United States. METHODS: Michigan Medicine established a telephone hotline to triage inbound patient calls related to COVID-19. For calls received between March 24, 2020, and May 5, 2020, we described total call volume, data reported by callers including COVID-19 risk factors and symptomatology, and distribution of callers to triage algorithm endpoints. We also described symptomatology reported by callers who were directed to the institutional patient portal (online medical visit questionnaire). RESULTS: A total of 3929 calls (average 91 calls per day) were received by the call center during the study period. The maximum total number of daily calls peaked at 211 on March 24, 2020. Call volumes were the highest from 6 AM to 11 AM and during evening hours. Callers were most often directed to the online patient portal (1654/3929, 42%), nursing hotlines (1338/3929, 34%), or employee health services (709/3929, 18%). Cough (126/370 of callers, 34%), shortness of breath (101/370, 27%), upper respiratory infection (28/111, 25%), and fever (89/370, 24%) were the most commonly reported symptoms. Immunocompromised state (23/370, 6%) and age >65 years (18/370, 5%) were the most commonly reported risk factors. CONCLUSIONS: The triage algorithm successfully diverted low-risk patients to suitable algorithm endpoints, while directing high-risk patients onward for immediate assessment. Data collected from hotline calls also enhanced knowledge of symptoms and risk factors that typified community members, demonstrating that pandemic hotlines can aid in the clinical characterization of novel diseases.

Understanding diagnostic processes in emergency departments: a mixed methods case study protocol.

INTRODUCTION: Diagnostic processes in the emergency department (ED) involve multiple interactions among individuals who interface with information systems to access and record information. A better understanding of diagnostic processes is needed to mitigate errors. This paper describes a study protocol to map diagnostic processes in the ED as a foundation for developing future error mitigation strategies. METHODS AND ANALYSIS: This study of an adult and a paediatric academic ED uses a prospective mixed methods case study design informed by an ED-specific diagnostic decision-making model (the modified ED-National Academies of Sciences, Engineering and Medicine (NASEM) model) and two cognitive theories (dual process theory and distributed cognition). Data sources include audio recordings of patient and care team interactions, electronic health record data, observer field notes and stakeholder interviews. Multiple qualitative analysis methods will be used to explore diagnostic processes in situ, including systems information flow, human-human and human-system interactions and contextual factors influencing cognition. The study has three parts. Part 1 involves prospective field observations of patients with undifferentiated symptoms at high risk for diagnostic error, where each patient is followed throughout the entire care delivery process. Part 2 involves observing individual care team providers over a 4-hour window to capture their diagnostic workflow, team coordination and communication across multiple patients. Part 3 uses interviews with key stakeholders to understand different perspectives on the diagnostic process, as well as perceived strengths and vulnerabilities, in order to enrich the ED-NASEM diagnostic model. ETHICS AND DISSEMINATION: The University of Michigan Institutional Review Board approved this study, HUM00156261. This foundational work will help identify strengths and vulnerabilities in diagnostic processes. Further, it will inform the future development and testing of patient, provider and systems-level interventions for mitigating error and improving patient safety in these and other EDs. The work will be disseminated through journal publications and presentations at national and international meetings.

Bile Salts Regulate Zinc Uptake and Capsule Synthesis in a Mastitis-Associated Extraintestinal Pathogenic Escherichia coli Strain.

Extraintestinal pathogenic Escherichia coli (ExPEC) strains are major causes of urinary and bloodstream infections. ExPEC reservoirs are not completely understood. Some mastitis-associated E. coli (MAEC) strains carry genes associated with ExPEC virulence, including metal scavenging, immune avoidance, and host attachment functions. In this study, we investigated the role of the high-affinity zinc uptake (znuABC) system in the MAEC strain M12. Elimination of znuABC moderately decreased fitness during mouse mammary gland infections. The ΔznuABC mutant strain exhibited an unexpected growth delay in the presence of bile salts, which was alleviated by the addition of excess zinc. We isolated suppressor mutants with improved growth in bile salts, several of which no longer produced the K96 capsule made by strain M12. The addition of bile salts also reduced capsule production by strain M12 and ExPEC strain CP9, suggesting that capsule synthesis may be detrimental when bile salts are present. To better understand the role of the capsule, we compared the virulence of mastitis strain M12 with that of its unencapsulated ΔkpsCS mutant in two models of ExPEC disease. The wild-type strain successfully colonized mouse bladders and kidneys and was highly virulent in intraperitoneal infections. Conversely, the ΔkpsCS mutant was unable to colonize kidneys and was unable to cause sepsis. These results demonstrate that some MAEC strains may be capable of causing human ExPEC illness. The virulence of strain M12 in these infections is dependent on its capsule. However, capsule may interfere with zinc homeostasis in the presence of bile salts while in the digestive tract.

Large-Scale Molecular Dynamics Simulations of Cellular Compartments.

Molecular dynamics or MD simulation is gradually maturing into a tool for constructing in vivo models of living cells in atomistic details. The feasibility of such models is bolstered by integrating the simulations with data from microscopic, tomographic and spectroscopic experiments on exascale supercomputers, facilitated by the use of deep learning technologies. Over time, MD simulation has evolved from tens of thousands of atoms to over 100 million atoms comprising an entire cell organelle, a photosynthetic chromatophore vesicle from a purple bacterium. In this chapter, we present a step-by-step outline for preparing, executing and analyzing such large-scale MD simulations of biological systems that are essential to life processes. All scripts are provided via GitHub.