Galleria mellonella as an Antimicrobial Screening Model.

To combat the rising global issue of antibiotic resistance, the accelerated development of novel antibiotics is essential. Current preclinical antimicrobial development yields a significant number of leads that prove unsuitable either prior to or during clinical trials. To increase the efficiency of preclinical development, relevant, standardized, accessible, and cost-effective models must be developed. Galleria mellonella (greater wax moth) larvae are widely used as an infection model to assess microbial virulence, conduct drug toxicity testing, and serve as a preliminary means of evaluating the in vivo efficacy of novel antimicrobial compounds. These infection models have greater biological relevance than many in vitro screens of comparable throughput and decrease reliance on mammalian models when used as a pre-screen for antimicrobial testing. This protocol describes a standardized methodology for the optimization of G. mellonella infection models, which can be applied to bacterial species and antimicrobial therapeutics of choice. Using the WHO priority pathogen Pseudomonas aeruginosa as an exemplar, we outline steps that can be undertaken to develop a reproducible model of infection and therapeutic testing. This includes recommendations on experimental setup, sample preparation, and infection and treatment protocols. Integration of this model within preclinical antimicrobial development pipelines would decrease reliance on mammalian models, reduce the number of ineffective compounds reaching clinical trials, and ultimately increase the efficiency of preclinical antimicrobial development.

Defining splicing factor requirements for androgen receptor variant synthesis in advanced prostate cancer.

Resistance to androgen receptor (AR)-targeted therapies represent a major challenge in prostate cancer (PC). A key mechanism of treatment resistance in patients who progress to castrate-resistant PC (CRPC) is the generation of alternatively spliced androgen receptor variants (AR-Vs). Unlike full-length AR (FL-AR) isoforms, AR-Vs are constitutively active and refractory to current receptor-targeting agents hence drive tumour progression. Identifying regulators of AR-V synthesis may therefore provide new therapeutic opportunities in combination with conventional AR-targeting agents. Our understanding of AR transcript splicing, and the factors that control the synthesis of AR-Vs, remains limited. While candidate-based approaches have identified a small number of AR-V splicing regulators, an unbiased analysis of splicing factors important for AR-V generation is required to fill an important knowledge gap and furnish the field with novel and tractable targets for PC treatment. To that end, we conducted a bespoke CRISPR screen to profile splicing factor requirements for AR-V synthesis. MFAP1 and CWC22 were shown to be required for the generation of AR-V mRNA transcripts and their depletion resulted in reduced AR-V protein abundance and cell proliferation in several CRPC models. Global transcriptomic analysis of MFAP1-depleted cells revealed both AR-dependent and -independent transcriptional impact, including genes associated with DDR. As such, MFAP1 downregulation sensitised PC cells to ionising radiation suggesting therapeutically targeting AR-V splicing could provide novel cellular vulnerabilities which can be exploited in CRPC. Implications: We have utilised a CRISPR screening approach to identify key regulators of pathogenic AR splicing in prostate cancer.

Post-meal exercise under ecological conditions improves post-prandial glucose levels but not 24-hour glucose control.

We investigated whether post-meal walking (PMW) improved post-prandial glucose and 24h glucose control under free-living conditions among physically inactive young women. METHODS: Young women (Age: 20±1years; percent body fat: 28.2 ± 12%; BMI: 23.8 ± 4.2kg·m-1) completed a randomised crossover study to assess if PMW confers benefit. On the PMW day, women completed three bouts of brisk walks, and on the Control day they were instructed to follow normal habitual activities. Continuous glucose monitors captured post-prandial and 24h glucose, and physical activity monitors tracked physical activity throughout the study. RESULTS: PMW walking increased total daily step count (Control = 9,159 ± 2,962 steps vs. PMW = 14,611±3,891 steps, p<0.001) and activity scores (Control=33.87±1.16 METs·h vs. PMW = 36.11±1.58 METs·h, p < 0.001). PMW led to lower 3h average post-prandial glucose (main effect of condition, p=0.011) and 3h post-prandial area under curve glucose responses (main effect of condition, p = 0.027) compared to the control condition. Post hoc analysis revealed the largest decline occurred after dinner (3h average glucose Control = 7.55±1.21 mmol/L vs. PMW = 6.71 ± 0.80mmol/L, p = 0.039), when insulin sensitivity is typically diminished. Despite improvements in post-prandial glucose control, this did not translate to improvements in 24h glucose control (p > 0.05). CONCLUSION: Physically inactive and metabolically healthy young women, PMW improves post-prandial glucose but not 24h glucose control.

Overcoming Osimertinib Resistance with AKT Inhibition in EGFRm-Driven Non-Small Cell Lung Cancer with PIK3CA/PTEN Alterations.

PURPOSE: Osimertinib is an EGFR tyrosine kinase inhibitor indicated for the treatment of EGFR-mutated (EGFRm)-driven lung adenocarcinomas. Osimertinib significantly improves progression-free survival in first-line-treated patients with EGFRm advanced non-small cell lung cancer (NSCLC). Despite the durable disease control, the majority of patients receiving osimertinib eventually develop disease progression. EXPERIMENTAL DESIGN: ctDNA profiling analysis of on-progression plasma samples from patients treated with osimertinib in both first- (phase III, FLAURA trial) and second-line trials (phase III, AURA3 trial) revealed a high prevalence of PIK3CA/AKT/PTEN alterations. In vitro and in vivo evidence using CRISPR-engineered NSCLC cell lines and patient-derived xenograft (PDX) models supports a functional role for PIK3CA and PTEN mutations in the development of osimertinib resistance. RESULTS: These alterations are functionally relevant as EGFRm NSCLC cells with engineered PIK3CA/AKT/PTEN alterations develop resistance to osimertinib and can be resensitized by treatment with the combination of osimertinib and the AKT inhibitor capivasertib. Moreover, xenograft and PDX in vivo models with PIK3CA/AKT/PTEN alterations display limited sensitivity to osimertinib relative to models without alterations, and in these double-mutant models, capivasertib and osimertinib combination elicits an improved antitumor effect versus osimertinib alone. CONCLUSIONS: Together, this approach offers a potential treatment strategy for patients with EGFRm-driven NSCLC who have a suboptimal response or develop resistance to osimertinib through PIK3CA/AKT/PTEN alterations. See related commentary by Vokes et al., p. 3968.

Genome-wide CRISPR screens identify the YAP/TEAD axis as a driver of persister cells in EGFR mutant lung cancer.

Most lung cancer patients with metastatic cancer eventually relapse with drug-resistant disease following treatment and EGFR mutant lung cancer is no exception. Genome-wide CRISPR screens, to either knock out or overexpress all protein-coding genes in cancer cell lines, revealed the landscape of pathways that cause resistance to the EGFR inhibitors osimertinib or gefitinib in EGFR mutant lung cancer. Among the most recurrent resistance genes were those that regulate the Hippo pathway. Following osimertinib treatment a subpopulation of cancer cells are able to survive and over time develop stable resistance. These 'persister' cells can exploit non-genetic (transcriptional) programs that enable cancer cells to survive drug treatment. Using genetic and pharmacologic tools we identified Hippo signalling as an important non-genetic mechanism of cell survival following osimertinib treatment. Further, we show that combinatorial targeting of the Hippo pathway and EGFR is highly effective in EGFR mutant lung cancer cells and patient-derived organoids, suggesting a new therapeutic strategy for EGFR mutant lung cancer patients.

Secondary messenger signalling influences Pseudomonas aeruginosa adaptation to sinus and lung environments.

Pseudomonas aeruginosa is a cause of chronic respiratory tract infections in people with cystic fibrosis (CF), non-CF bronchiectasis, and chronic obstructive pulmonary disease. Prolonged infection allows the accumulation of mutations and horizontal gene transfer, increasing the likelihood of adaptive phenotypic traits. Adaptation is proposed to arise first in bacterial populations colonizing upper airway environments. Here, we model this process using an experimental evolution approach. Pseudomonas aeruginosa PAO1, which is not airway adapted, was serially passaged, separately, in media chemically reflective of upper or lower airway environments. To explore whether the CF environment selects for unique traits, we separately passaged PAO1 in airway-mimicking media with or without CF-specific factors. Our findings demonstrated that all airway environments-sinus and lungs, under CF and non-CF conditions-selected for loss of twitching motility, increased resistance to multiple antibiotic classes, and a hyper-biofilm phenotype. These traits conferred increased airway colonization potential in an in vivo model. CF-like conditions exerted stronger selective pressures, leading to emergence of more pronounced phenotypes. Loss of twitching was associated with mutations in type IV pili genes. Type IV pili mediate surface attachment, twitching, and induction of cAMP signalling. We additionally identified multiple evolutionary routes to increased biofilm formation involving regulation of cyclic-di-GMP signalling. These included the loss of function mutations in bifA and dipA phosphodiesterase genes and activating mutations in the siaA phosphatase. These data highlight that airway environments select for traits associated with sessile lifestyles and suggest upper airway niches support emergence of phenotypes that promote establishment of lung infection.

Housing-Sensitive Health Conditions Can Predict Poor-Quality Housing.

Improving housing quality may improve residents' health, but identifying buildings in poor repair is challenging. We developed a method to improve health-related building inspection targeting. Linking New York City Medicaid claims data to Landlord Watchlist data, we used machine learning to identify housing-sensitive health conditions correlated with a building's presence on the Watchlist. We identified twenty-three specific housing-sensitive health conditions in five broad categories consistent with the existing literature on housing and health. We used these results to generate a housing health index from building-level claims data that can be used to rank buildings by the likelihood that their poor quality is affecting residents' health. We found that buildings in the highest decile of the housing health index (controlling for building size, community district, and subsidization status) scored worse across a variety of housing quality indicators, validating our approach. We discuss how the housing health index could be used by local governments to target building inspections with a focus on improving health.

PROVIDENT: Development and Validation of a Machine Learning Model to Predict Neighborhood-level Overdose Risk in Rhode Island.

BACKGROUND: Drug overdose persists as a leading cause of death in the United States, but resources to address it remain limited. As a result, health authorities must consider where to allocate scarce resources within their jurisdictions. Machine learning offers a strategy to identify areas with increased future overdose risk to proactively allocate overdose prevention resources. This modeling study is embedded in a randomized trial to measure the effect of proactive resource allocation on statewide overdose rates in Rhode Island (RI). METHODS: We used statewide data from RI from 2016 to 2020 to develop an ensemble machine learning model predicting neighborhood-level fatal overdose risk. Our ensemble model integrated gradient boosting machine and super learner base models in a moving window framework to make predictions in 6-month intervals. Our performance target, developed a priori with the RI Department of Health, was to identify the 20% of RI neighborhoods containing at least 40% of statewide overdose deaths, including at least one neighborhood per municipality. The model was validated after trial launch. RESULTS: Our model selected priority neighborhoods capturing 40.2% of statewide overdose deaths during the test periods and 44.1% of statewide overdose deaths during validation periods. Our ensemble outperformed the base models during the test periods and performed comparably to the best-performing base model during the validation periods. CONCLUSIONS: We demonstrated the capacity for machine learning models to predict neighborhood-level fatal overdose risk to a degree of accuracy suitable for practitioners. Jurisdictions may consider predictive modeling as a tool to guide allocation of scarce resources.

Utilizing a Dual Human Transporter MDCKII-MDR1-BCRP Cell Line to Assess Efflux at the Blood Brain Barrier.

To facilitate the design of drugs readily able to cross the blood brain barrier (BBB), a Madin-Darby canine kidney (MDCK) cell line was established that over expresses both P-glycoprotein (Pgp) and breast cancer resistance protein (BCRP), the main human efflux transporters of the BBB. Proteomics analyses indicate BCRP is expressed at a higher level than Pgp in this cell line. This cell line shows good activity for both transporters [BCRP substrate dantrolene efflux ratio (ER) 16.3 ± 0.9, Pgp substrate quinidine ER 27.5 ± 1.2], and use of selective transporter inhibitors enables an assessment of the relative contributions to overall ERs. The MDCKII-MDR1-BCRP ER negatively correlates with rat unbound brain/unbound plasma ratio, Kpuu Highly brain penetrant compounds with rat Kpuu ≥ 0.3 show ERs ≤ 2 in the MDCKII-MDR1-BCRP assay while compounds predominantly excluded from the brain, Kpuu ≤ 0.05, demonstrate ERs ≥ 20. A subset of compounds with MDCKII-MDR1-BCRP ER < 2 and rat Kpuu < 0.3 were shown to be substrates of rat Pgp using a rat transfected cell line, MDCKII-rMdr1a. These compounds also showed ERs > 2 in the human National Institutes of Health (NIH) MDCKI-MDR1 (high Pgp expression) cell line, which suggests that they are weak human Pgp substrates. Characterization of 37 drugs targeting the central nervous system in the MDCKII-MDR1-BCRP efflux assay show 36 have ERs < 2. In drug discovery, use of the MDCKII-MDR1-BCRP in parallel with the NIH MDCKI-MDR1 cell line is useful for identification of compounds with high brain penetration. SIGNIFICANCE STATEMENT: A single cell line that includes both the major human efflux transporters of the blood brain barrier (MDCKII-MDR1-BCRP) has been established facilitating the rapid identification of efflux substrates and enabling the design of brain penetrant molecules. Efflux ratios using this cell line demonstrate a clear relationship with brain penetration as defined by rat brain Kpuu.

Emerging strategies to target virulence in Pseudomonas aeruginosa respiratory infections.

Pseudomonas aeruginosa is an opportunistic pathogen that is responsible for infections in people living with chronic respiratory conditions, such as cystic fibrosis (CF) and non-CF bronchiectasis (NCFB). Traditionally, in people with chronic respiratory disorders, P. aeruginosa infection has been managed with a combination of inhaled and intravenous antibiotic therapies. However, due in part to the prolonged use of antibiotics in these people, the emergence of multi-drug resistant P. aeruginosa strains is a growing concern. The development of anti-virulence therapeutics may provide a new means of treating P. aeruginosa lung infections whilst also combatting the AMR crisis, as these agents are presumed to exert reduced pressure for the emergence of drug resistance as compared to antibiotics. However, the pipeline for developing anti-virulence therapeutics is poorly defined, and it is currently unclear as to whether in vivo and in vitro models effectively replicate the complex pulmonary environment sufficiently to enable development and testing of such therapies for future clinical use. Here, we discuss potential targets for P. aeruginosa anti-virulence therapeutics and the effectiveness of the current models used to study them. Focus is given to the difficulty of replicating the virulence gene expression patterns of P. aeruginosa in the CF and NCFB lung under laboratory conditions and to the challenges this poses for anti-virulence therapeutic development.

A study of the impact of the COVID-19 pandemic on equine veterinary care in the UK.

Background: During the COVID-19 pandemic, equine health care in the UK may have been adversely affected due to mandated changes in the delivery of veterinary healthcare and the potential for reduced health-seeking behaviour. Methods: Electronic patient records (EPRs) were analysed to describe veterinary activity for all equids under the active care of 20 veterinary practices in the UK in the 12 months before and after the introduction of the first UK lockdown. Pre-pandemic and pandemic levels of clinical activity were compared. Further comparisons of care, including immediate management and treatment, were made following a detailed review of EPRs from randomly selected subsets of equids under care in four time periods. Results: All measures of activity and face-to-face interaction were lower in the early pandemic period than in the equivalent pre-pandemic period. Compared to pre-pandemic, the early pandemic was associated with a decrease in prophylactic care and non-urgent diagnostic imaging and an increase in systemic non-steroid anti-inflammatory prescription. Convenience sampling of veterinary practices may have limited the generalisability of the findings. The quality of EPRs was variable. Conclusions: While equine veterinary activity was significantly disrupted in the early pandemic period, there was a rapid return to pre-pandemic levels of activity. Subsequent lockdowns appeared to have had little effect on veterinary care.

Airway metabolic profiling during Streptococcus pneumoniae infection identifies branched chain amino acids as signatures of upper airway colonisation.

Streptococcus pneumoniae is a leading cause of community-acquired pneumonia and bacteraemia and is capable of remarkable phenotypic plasticity, responding rapidly to environmental change. Pneumococcus is a nasopharyngeal commensal, but is responsible for severe, acute infections following dissemination within-host. Pneumococcus is adept at utilising host resources, but the airways are compartmentalised and those resources are not evenly distributed. Challenges and opportunities in metabolite acquisition within different airway niches may contribute to the commensal-pathogen switch when pneumococcus moves from nasopharynx into lungs. We used NMR to characterise the metabolic landscape of the mouse airways, in health and during infection. Using paired nasopharynx and lung samples from naïve animals, we identified fundamental differences in metabolite bioavailability between airway niches. Pneumococcal pneumonia was associated with rapid and dramatic shifts in the lung metabolic environment, whilst nasopharyngeal carriage led to only modest change in upper airway metabolite profiles. NMR spectra derived from the nasopharynx of mice infected with closely-related pneumococcal strains that differ in their colonisation potential could be distinguished from one another using multivariate dimensionality reduction methods. The resulting models highlighted that increased branched-chain amino acid (BCAA) bioavailability in nasopharynx is a feature of infection with the high colonisation potential strain. Subsequent analysis revealed increased expression of BCAA transport genes and increased intracellular concentrations of BCAA in that same strain. Movement from upper to lower airway environments is associated with shifting challenges in metabolic resource allocation for pneumococci. Efficient biosynthesis, liberation or acquisition of BCAA is a feature of adaptation to nasopharyngeal colonisation.

Neighborhood-Level Risk Factors for Severe Hyperglycemia among Emergency Department Patients without a Prior Diabetes Diagnosis.

A person's place of residence is a strong risk factor for important diagnosed chronic diseases such as diabetes. It is unclear whether neighborhood-level risk factors also predict the probability of undiagnosed disease. The objective of this study was to identify neighborhood-level variables associated with severe hyperglycemia among emergency department (ED) patients without a history of diabetes. We analyzed patients without previously diagnosed diabetes for whom a random serum glucose value was obtained in the ED. We defined random glucose values ≥ 200 mg/dL as severe hyperglycemia, indicating probable undiagnosed diabetes. Patient addresses were geocoded and matched with neighborhood-level socioeconomic measures from the American Community Survey and claims-based surveillance estimates of diabetes prevalence. Neighborhood-level exposure variables were standardized based on z-scores, and a series of logistic regression models were used to assess the association of selected exposures and hyperglycemia adjusting for biological and social individual-level risk factors for diabetes. Of 77,882 ED patients without a history of diabetes presenting in 2021, 1,715 (2.2%) had severe hyperglycemia. Many geospatial exposures were associated with uncontrolled hyperglycemia, even after controlling for individual-level risk factors. The most strongly associated neighborhood-level variables included lower markers of educational attainment, higher percentage of households where limited English is spoken, lower rates of white-collar employment, and higher rates of Medicaid insurance. Including these geospatial factors in risk assessment models may help identify important subgroups of patients with undiagnosed disease.

Identification of two distinct phylogenomic lineages and model strains for the understudied cystic fibrosis lung pathogen Burkholderia multivorans.

Burkholderia multivorans is the dominant Burkholderia pathogen recovered from lung infection in people with cystic fibrosis. However, as an understudied pathogen there are knowledge gaps in relation to its population biology, phenotypic traits and useful model strains. A phylogenomic study of B. multivorans was undertaken using a total of 283 genomes, of which 73 were sequenced and 49 phenotypically characterized as part of this study. Average nucleotide identity analysis (ANI) and phylogenetic alignment of core genes demonstrated that the B. multivorans population separated into two distinct evolutionary clades, defined as lineage 1 (n=58 genomes) and lineage 2 (n=221 genomes). To examine the population biology of B. multivorans, a representative subgroup of 77 B. multivorans genomes (28 from the reference databases and the 49 novel short-read genome sequences) were selected based on multilocus sequence typing (MLST), isolation source and phylogenetic placement criteria. Comparative genomics was used to identify B. multivorans lineage-specific genes - ghrB_1 in lineage 1 and glnM_2 in lineage 2 - and diagnostic PCRs targeting them were successfully developed. Phenotypic analysis of 49 representative B. multivorans strains showed considerable inter-strain variance, but the majority of the isolates tested were motile and capable of biofilm formation. A striking absence of B. multivorans protease activity in vitro was observed, but no lineage-specific phenotypic differences were demonstrated. Using phylogenomic and phenotypic criteria, three model B. multivorans CF strains were identified, BCC0084 (lineage 1), BCC1272 (lineage 2a) and BCC0033 lineage 2b, and their complete genome sequences determined. B. multivorans CF strains BCC0033 and BCC0084, and the environmental reference strain, ATCC 17616, were all capable of short-term survival within a murine lung infection model. By mapping the population biology, identifying lineage-specific PCRs and model strains, we provide much needed baseline resources for future studies of B. multivorans.

"Bringing you the best": John Player & Sons, Cricket, and the Politics of Tobacco Sport Sponsorship in Britain, 1969-1986.

This article explores some of the marketing strategies associated with the British tobacco industry's sponsorship of sport during the 1960s and 1970s. It focuses on the British cigarette and tobacco manufacturer John Player & Sons and the firm's pioneering initiative to sponsor one-day cricket, which began with the John Player League in 1969. The league was enormously popular and gained significant broadcast coverage, becoming an invaluable means of increasing public exposure for the company, in the context of the ban of cigarette advertising from British television. At a time when the link between smoking and disease was making headlines, John Player & Sons nimbly deflected attention away from the health issue, and instead consciously repositioned the tobacco company as a generous benefactor of the nation's sport and leisure. Less conspicuously, but even more powerfully, spokespeople for the tobacco industry actively mobilised influential opinion behind the scenes in political circles. We show particularly how Denis Howell, Minister for Sport from 1964 to 1969 and from 1974 to 1979, became a valuable ally, acting as a bulwark against more restrictive government interventions into the sponsorship of sports by the tobacco industry. This alliance exposes changing industry-government relations and presents new historical context to better understand the way British tobacco manufacturers proactively sought to elide restrictions on their advertising activities from the 1980s onwards.

Discovery and Optimization of the First ATP Competitive Type-III c-MET Inhibitor.

Recent clinical reports have highlighted the need for wild-type (WT) and mutant dual inhibitors of c-MET kinase for the treatment of cancer. We report herein a novel chemical series of ATP competitive type-III inhibitors of WT and D1228V mutant c-MET. Using a combination of structure-based drug design and computational analyses, ligand 2 was optimized to a highly selective chemical series with nanomolar activities in biochemical and cellular settings. Representatives of the series demonstrate excellent pharmacokinetic profiles in rat in vivo studies with promising free-brain exposures, paving the way for the design of brain permeable drugs for the treatment of c-MET driven cancers.

Excess adiposity contributes to higher ambulatory central blood pressure and arterial stiffness in physically inactive young adults.

BACKGROUND: It remains unknown if physical inactivity and excess adiposity increases 24-h central blood pressure and arterial stiffness in young adults. This study examined 24-h central blood pressure and indirect measures of arterial stiffness (e.g., central pulse pressure) in physically inactive young adults with and without excess adiposity. METHODS: Body fat and ambulatory 24-h blood pressure were measured in 31 young adults (men: 22±4 years, N.=15; women: 22±5 years, N=16). Multi-frequency bioelectrical impedance measured body fat. Normal adiposity was defined as <20% body fat in men and <32% body fat in women, whereas excess adiposity was defined as ≥20% and ≥32% in men and women, respectively. Ambulatory 24-h central blood pressure was calculated based on brachial blood pressure and volumetric displacement waveforms. RESULTS: By design, the normal adiposity group had a lower body fat percentage (men: 15.5±4.6%; women: 20.8±2.5%) compared to the physically inactive excess adiposity group (men: 29.8±5.4%; women: 34.3±7.5%). Men and women with excess adiposity group had elevated central blood pressure (central systolic, P<0.05 vs. normal adiposity groups). Central pulse pressure was elevated in the excess adiposity group (men: 45±5 mmHg; women: 41±9 mmHg) compared to normal adiposity groups (men: 36±4 mmHg; women: 32±3 mmHg, P<0.05 for both), while other arterial stiffness (augmentation index and ambulatory arterial stiffness index) measures trended toward significance only in men with excess adiposity. CONCLUSIONS: Physically inactive men and women with excess adiposity have increased 24h central blood pressure and pulse pressure compared to physically inactive young adults with normal adiposity.

Translating Predictive Analytics for Public Health Practice: A Case Study of Overdose Prevention in Rhode Island.

Prior applications of machine learning to population health have relied on conventional model assessment criteria, limiting the utility of models as decision support tools for public health practitioners. To facilitate practitioners' use of machine learning as a decision support tool for area-level intervention, we developed and applied 4 practice-based predictive model evaluation criteria (implementation capacity, preventive potential, health equity, and jurisdictional practicalities). We used a case study of overdose prevention in Rhode Island to illustrate how these criteria could inform public health practice and health equity promotion. We used Rhode Island overdose mortality records from January 2016-June 2020 (n = 1,408) and neighborhood-level US Census data. We employed 2 disparate machine learning models, Gaussian process and random forest, to illustrate the comparative utility of our criteria to guide interventions. Our models predicted 7.5%-36.4% of overdose deaths during the test period, illustrating the preventive potential of overdose interventions assuming 5%-20% statewide implementation capacities for neighborhood-level resource deployment. We describe the health equity implications of use of predictive modeling to guide interventions along the lines of urbanicity, racial/ethnic composition, and poverty. We then discuss considerations to complement predictive model evaluation criteria and inform the prevention and mitigation of spatially dynamic public health problems across the breadth of practice. This article is part of a Special Collection on Mental Health.

Sulfur-Polymer Nanoparticles: Preparation and Antibacterial Activity.

High sulfur content polymers prepared by inverse vulcanization have many reported potential applications, including as novel antimicrobial materials. High sulfur content polymers usually have limited water-solubility and dispersibility due to their hydrophobic nature, which could limit the development of their applications. Herein, we report the formulation of high sulfur content polymeric nanoparticles by a nanoprecipitation and emulsion-based method. High sulfur content polymeric nanoparticles were found to have an inhibitory effect against important bacterial pathogens, including Gram-positive methicillin-resistant Staphylococcus aureus and Gram-negative Pseudomonas aeruginosa. Salt-stable particles were formulated with the addition of a surfactant, which did not inhibit the antibacterial activity of the polymeric particles. Furthermore, the polymeric nanoparticles were found to inhibit S. aureus biofilm formation and exhibited low cytotoxicity against mammalian liver cells. Interaction of the polymeric particles with cellular thiols could be a potential mechanism of action against bacterial cells, as demonstrated by reaction with cysteine as a model thiol. The findings presented demonstrate methods of preparing aqueous dispersions of high sulfur content polymeric nanoparticles that could have useful biological applications.

An innate pathogen sensing strategy involving ubiquitination of bacterial surface proteins.

Sensing of pathogens by ubiquitination is a critical arm of cellular immunity. However, universal ubiquitination targets on microbes remain unidentified. Here, using in vitro, ex vivo, and in vivo studies, we identify the first protein-based ubiquitination substrates on phylogenetically diverse bacteria by unveiling a strategy that uses recognition of degron-like motifs. Such motifs form a new class of intra-cytosolic pathogen-associated molecular patterns (PAMPs). Their incorporation enabled recognition of nonubiquitin targets by host ubiquitin ligases. We find that SCFFBW7 E3 ligase, supported by the regulatory kinase, glycogen synthase kinase 3ß, is crucial for effective pathogen detection and clearance. This provides a mechanistic explanation for enhanced risk of infections in patients with chronic lymphocytic leukemia bearing mutations in F-box and WD repeat domain containing 7 protein. We conclude that exploitation of this generic pathogen sensing strategy allows conservation of host resources and boosts antimicrobial immunity.