Increasing the Soluble Expression and Whole-Cell Activity of the Plastic-Degrading Enzyme MHETase through Consensus Design.

The mono(2-hydroxyethyl) terephthalate hydrolase (MHETase) from Ideonella sakaiensis carries out the second step in the enzymatic depolymerization of poly(ethylene terephthalate) (PET) plastic into the monomers terephthalic acid (TPA) and ethylene glycol (EG). Despite its potential industrial and environmental applications, poor recombinant expression of MHETase has been an obstacle to its industrial application. To overcome this barrier, we developed an assay allowing for the medium-throughput quantification of MHETase activity in cell lysates and whole-cell suspensions, which allowed us to screen a library of engineered variants. Using consensus design, we generated several improved variants that exhibit over 10-fold greater whole-cell activity than wild-type (WT) MHETase. This is revealed to be largely due to increased soluble expression, which biochemical and structural analysis indicates is due to improved protein folding.

A retrospective chart review assessing antibiotic treatment of hospitalized patients with discordant Clostridioides difficile assays in an urban hospitalized setting.

Clostridioides difficile infection (CDI) threatens vulnerable populations in health care. Two-step testing improves specificity, avoiding over-treatment. This study analyzed inpatient records to estimate diagnostic outcomes and identify characteristics associated with treatment after discordant testing. Among discordant patients, those aged 65+ years were significantly more likely to be prescribed antibiotics (67% vs 39%).

Rugged fitness landscapes minimize promiscuity in the evolution of transcriptional repressors.

How a protein's function influences the shape of its fitness landscape, smooth or rugged, is a fundamental question in evolutionary biochemistry. Smooth landscapes arise when incremental mutational steps lead to a progressive change in function, as commonly seen in enzymes and binding proteins. On the other hand, rugged landscapes are poorly understood because of the inherent unpredictability of how sequence changes affect function. Here, we experimentally characterize the entire sequence phylogeny, comprising 1,158 extant and ancestral sequences, of the DNA-binding domain (DBD) of the LacI/GalR transcriptional repressor family. Our analysis revealed an extremely rugged landscape with rapid switching of specificity, even between adjacent nodes. Further, the ruggedness arises due to the necessity of the repressor to simultaneously evolve specificity for asymmetric operators and disfavors potentially adverse regulatory crosstalk. Our study provides fundamental insight into evolutionary, molecular, and biophysical rules of genetic regulation through the lens of fitness landscapes.

Academic Half Day Improves Resident Perception of Education Without Compromising Patient Safety.

BACKGROUND: Residency programs are required to offer a didactic curriculum and protect resident time for education. Our institution implemented an academic half day (AHD) in the 2021-2022 academic year to address issues related to the standard noon conference series. OBJECTIVE: Determine the impact of AHD implementation on education, patient safety, and workflow. METHODS: This was a prospective, single-site educational intervention study. Pre- and post-implementation surveys and Accreditation Council for Graduate Medical Education (ACGME) surveys assessed changes in trainee and faculty attitudes and behaviors. Patient safety and workflow were evaluated by comparing the number of safety event reports, rapid response team activations, time to admission from the ED, and time of discharge on AHD days compared to other weekdays. RESULTS: Survey response rates were: residents 68%/48%, fellows 42%/35%, and faculty 59%/29%. AHD was associated with a significant, positive change in resident attitudes and experiences and on ACGME survey items. On AHDs compared with other weekdays, there were no significant differences in safety event report rates (P = .98), nor in rapid response team activation rates (P = .99). There was not a clinically meaningful difference in median admission time from the ED on AHD weekdays (125 minutes) compared to other weekdays (130 minutes, P = .04). There was no significant difference in median discharge time on AHD vs other weekdays (P = .13). CONCLUSIONS: This study suggests that there is no significant difference in patient safety or workflow with the implementation of AHD. This study supports prior studies that residents strongly prefer AHD. AHD may be a useful framework for resident education without compromising patient care.

The more you know: Insights from integrated pre-visit surveys in a pediatric environmental health center.

The Pediatric Environmental Health Center (PEHC) at Boston Children's Hospital is a specialty referral clinic that provides consultation for approximately 250 patients annually. Identifying environmental hazards is key for clinical management. Exposure concerns include lead, mold, pesticides, perfluoroalkyl substances (PFAS), impaired air quality, and more. Our goal was to identify concerns and visit priorities of our patient population to guide visits. A 47-question pre-visit survey was created exploring potential environmental hazards and administered prior to visits using a platform integrated into the electronic medical record (EMR). The study group was a convenience sample of patients from June 2021 to June 2022. Of 204 total visits, 101 surveys were submitted, yielding a response rate of 49.5%. 66/101 (65.3%) were surveys from initial consultations used for descriptive analysis. The majority of patients were seen for a chief complaint of lead exposure (90.1%). Most respondents had concerns about peeling paint (40.0%), and those reporting peeling paint were more likely to report additional concerns [75.0%, p < 0.001]. Other concerns highlighted were mold (15.2%), pests (15.2%), asbestos (10.6%), air pollution (9.1%), temperature regulation (7.6%), pesticides (6.1%), PFAS (4.5%), and formaldehyde (4.5%). A knowledge gap was identified; 45.5% (30/66) respondents responded "no" to the question asking if the Poison Control Center phone number was stored in their phone. This study illustrates how the implementation of a pre-visit EMR integrated survey engages families, informs clinical care, and serves as a point-of-care education tool for specific knowledge gaps. Findings will guide development of future environmental health screeners.