Deciphering the complexity of the chemicals in food packaging materials using molecular networks.

Chemicals from packaging materials might be transferred into food resulting in consumer exposure. Identifying these migrated chemicals is highly challenging and crucial to perform their safety assessment, usually starting by the understanding of the chemical composition of the packaging material itself. This study explores the use of the Molecular Networking (MN) approach to support identification of the extracted chemicals. Two formulations of bioplastics were analyzed using Liquid Chromatography hyphenated to High-Resolution Mass Spectrometry. Data processing and interpretation using a conventional manual method was performed as a point of comparison to understand the power of MN. Interestingly, only the MN approach facilitated the identification of unknown chemicals belonging to a novel oligomer series containing the azelaic acid monomer. The MN approach provided a faster visualization of chemical families in addition to the highlight of unrelated chemicals enabling to prioritize chemicals for further investigation improving the safety assessment of packaging materials.

Consistent Delivery of Adeno-Associated Virus via Lateral Tail-Vein Injection in Adult Mice.

Many disorders affect multiple organs or involve different regions of the body, so it is critical to deliver therapeutics systemically to target the affected cells located in different sites. Intravenous injection is a widely used systemic delivery route in preclinical studies that assess treatments intended for body-wide administration. In adult mice, it involves the intravenous administration of the therapeutic agent into the mouse's lateral tail veins. When mastered, tail-vein injections are safe and fast, and only require simple and commonly available tools. However, tail-vein injections are technically challenging and require extensive training and continuous practice to ensure the accurate delivery of the intended dose. Here we describe a detailed, optimized, lateral tail-vein injection protocol that we have developed based on our experience and on recommendations that had been previously reported by other groups. Other than the mouse restrainers and insulin syringes, this protocol requires only reagents and equipment that are readily available in most labs. We found that following this protocol results in consistently successful intravenous delivery of adeno-associated virus (AAV) into the tail veins of unsedated 7-9 week-old mice. Additionally, we describe the optimized protocols for the histological detection of fluorescent reporter proteins and vector genome per diploid genome (vg/dg) quantification used to assess AAV transduction and biodistribution. The goal of this protocol is to aid experimenters in easily performing tail-vein injections successfully and consistently, which can reduce the practice time needed to master the technique.

Data integrity in an online world: Demonstration of multimodal bot screening tools and considerations for preserving data integrity in two online social and behavioral research studies with marginalized populations.

Internet-based studies are widely used in social and behavioral health research, yet bots and fraud from "survey farming" bring significant threats to data integrity. For research centering marginalized communities, data integrity is an ethical imperative, as fraudulent data at a minimum poses a threat to scientific integrity, and worse could even promulgate false, negative stereotypes about the population of interest. Using data from two online surveys of sexual and gender minority populations (young men who have sex with men and transgender women of color), we (a) demonstrate the use of online survey techniques to identify and mitigate internet-based fraud, (b) differentiate techniques for and identify two different types of "survey farming" (i.e., bots and false responders), and (c) demonstrate the consequences of those distinct types of fraud on sample characteristics and statistical inferences, if fraud goes unaddressed. We provide practical recommendations for internet-based studies in psychological, social, and behavioral health research to ensure data integrity and discuss implications for future research testing data integrity techniques. (PsycInfo Database Record (c) 2024 APA, all rights reserved).

Trying to outrun causality with machine learning: Limitations of model explainability techniques for exploratory research.

Machine learning explainability techniques have been proposed as a means for psychologists to "explain" or interrogate a model in order to gain an understanding of a phenomenon of interest. Researchers concerned with imposing overly restrictive functional form (e.g., as would be the case in a linear regression) may be motivated to use machine learning algorithms in conjunction with explainability techniques, as part of exploratory research, with the goal of identifying important variables that are associated with/predictive of an outcome of interest. However, and as we demonstrate, machine learning algorithms are highly sensitive to the underlying causal structure in the data. The consequences of this are that predictors which are deemed by the explainability technique to be unrelated/unimportant/unpredictive, may actually be highly associated with the outcome. Rather than this being a limitation of explainability techniques per se, we show that it is rather a consequence of the mathematical implications of regression, and the interaction of these implications with the associated conditional independencies of the underlying causal structure. We provide some alternative recommendations for psychologists wanting to explore the data for important variables. (PsycInfo Database Record (c) 2024 APA, all rights reserved).

Advanced Practice Providers in Pediatric Surgery: A 10-Year Analysis of Clinical Productivity.

INTRODUCTION: Advanced practice providers (APPs) are integral to nonsurgical and surgical fields. In surgical settings, APPs work autonomously alongside surgeons to enhance continuity of care. While small-scale studies across various surgical specialties have explored APP utilization, literature specific to the pediatric population is limited. This study assesses the impact of increased APP utilization on productivity within one of the largest pediatric surgery departments in the United States. METHODS: Data was sourced from the Surgery Department Executive Committee (SDEC) annual reports. Descriptive analyses were performed to delineate trends in clinical volumes, OR cases, gross charges, work relative value units (wRVUs), and provider full-time equivalents (FTEs) between October 2011 and September 2023. Secondary analyses were performed from 2016 to 2023, after APP roles were refined in each division. RESULTS: Since 2012, an upward trend is observed in total clinic visits, total OR cases, and OR cases per MD. In 2016, APPs saw 1 clinic patient for every 2.2 seen by MDs, decreasing to a 1:1 ratio by 2023. The MD surgical conversion rate has increased by 20.5% since 2016, while the conversion rate per total clinic volume has decreased by 7.3% over the same period. Total wRVUs and gross charges increased across the department and per provider, despite fewer overall providers since 2021. CONCLUSION: Increased utilization of APPs within the department of surgery may enhance procedural productivity for surgeons while ensuring continued patient access to care. This study highlights the importance of developing care models that optimize the full skillset of APPs.

The genetic basis of the kakapo structural color polymorphism suggests balancing selection by an extinct apex predator.

The information contained in population genomic data can tell us much about the past ecology and evolution of species. We leveraged detailed phenotypic and genomic data of nearly all living kakapo to understand the evolution of its feather color polymorphism. The kakapo is an endangered and culturally significant parrot endemic to Aotearoa New Zealand, and the green and olive feather colorations are present at similar frequencies in the population. The presence of such a neatly balanced color polymorphism is remarkable because the entire population currently numbers less than 250 birds, which means it has been exposed to severe genetic drift. We dissected the color phenotype, demonstrating that the two colors differ in their light reflectance patterns due to differential feather structure. We used quantitative genomics methods to identify two genetic variants whose epistatic interaction can fully explain the species' color phenotype. Our genomic forward simulations show that balancing selection might have been pivotal to establish the polymorphism in the ancestrally large population, and to maintain it during population declines that involved a severe bottleneck. We hypothesize that an extinct apex predator was the likely agent of balancing selection, making the color polymorphism in the kakapo a "ghost of selection past."

Association of in vitro fertilization with severe maternal morbidity in low-risk patients without comorbidities.

OBJECTIVE: To determine whether an association exists between in vitro fertility (IVF) and severe maternal morbidity among low-risk pregnant patients. DESIGN: Retrospective cohort study SUBJECTS: Low-risk pregnant patients who delivered between 1/2019 and 12/2022. Low-risk was defined as having an Obstetric Comorbidity Index (OB-CMI) score of 0. EXPOSURE: IVF MAIN OUTCOME MEASURES: The primary outcome (dependent variable) was any severe maternal morbidity. The secondary outcome was the need for a cesarean delivery. A modified Poisson regression with robust error variance was used to model the probability of severe maternal morbidity as a function of IVF. Risk ratios (RR) and their associated 95% confidence intervals (CI) were computed. An alpha value of 0.05 was considered statistically significant. RESULTS: A total of 39,668 pregnancies were included for analysis, and 454 (1.1%) were conceived by IVF. The overall severe maternal morbidity rate was 2.4% (n=949), with the most common indicator being blood transfusion. Overall cesarean delivery rate was 18.8% (n=7,459). On modified Poisson regression, IVF-conceived pregnancies were associated with 2.56 times the risk of severe maternal morbidity (95% CI 1.73 - 3.79) and 1.54 times the risk of having a cesarean delivery (95% CI, 1.37-1.74) compared to non-IVF pregnancies. CONCLUSION: IVF is associated with higher rates of severe maternal morbidity, primarily the need for a blood transfusion, and cesarean delivery in low-risk pregnancies without major comorbidities. Recognizing this association allows healthcare providers to implement proactive measures for better monitoring and tailored postpartum care.

Drosophila are hosts to the first described parasitoid wasp of adult flies.

Parasitoid wasps are exceptionally diverse and use specialized adaptations capable of manipulating the physiology and behaviour of host organisms1. In more than two centuries since the first records of Drosophila-parasitizing wasps, nearly 200 described and provisional parasitoid species of drosophilids have been identified2. These include endoparasitoids and ectoparasitoids, as well as species attacking larval and pupal hosts3. Despite a deep history of research attention and remarkable biodiversity, a wasp species that attacks and develops inside the adult stage of a fly host has not been described previously. Here we report the discovery of a wasp species that infects the adult stage of fruit flies in the genus Drosophila, including one of the most deeply studied model organisms in biology, Drosophila melanogaster. Notably, this wasp can be easily collected from backyard fly baits and has a broad geographic distribution throughout the eastern USA. We document its life history and unique host interactions, including egg-laying into and larval emergence from adult flies, and provide protocols to raise wasps from wild-caught host flies. Our results emphasize the need for ongoing research investment in insect biodiversity and systematics. As parasitoid research continues to uncover unusual biology and supports fundamental mechanistic insights into immunity4, metabolism5, ecology6, evolution7-9 and behaviour10-12, we anticipate that this wasp's association with the laboratory model organism, D. melanogaster, will provide new research opportunities across the life sciences.

Prediction of biochemical recurrence after radical prostatectomy from primary tumour characteristics.

OBJECTIVES: To construct and externally calibrate a predictive model for early biochemical recurrence (BCR) after radical prostatectomy (RP) incorporating clinical and modern imaging characteristics of the primary tumour. PATIENTS AND METHODS: Patients who underwent RP following multiparametric magnetic resonance imaging, prostate biopsy and prostate-specific membrane antigen-positron emission tomography/computed tomography (PSMA-PET/CT), from two centres in Australia and the Netherlands. The primary outcome was biochemical recurrence-free survival (BRFS), where BCR was defined as a rising PSA level of ≥0.2 ng/mL or initiation of postoperative treatment per clinician discretion. Proportional hazards models to predict time to event were developed in the Australian sample using relevant pre- and post-surgical parameters and primary tumour maximum standardised uptake value (SUVmax) on diagnostic PSMA-PET/CT. Calibration was assessed in an external dataset from the Netherlands with the same inclusion criteria. RESULTS: Data from 846 patients were used to develop the models. Tumour SUVmax was associated with worse predicted 3-year BRFS for both pre- and post-surgical models. SUVmax change from 4 to 16 lessened the predicted 3-year BRFS from 66% to 42% for a patient aged 65 years with typical pre-surgical parameters (PSA level 8 ng/mL, Prostate Imaging-Reporting and Data System score 4/5 and biopsy Gleason score ≥4 + 5). Considering post-surgical variables, a patient with the same age and PSA level but pathological stage pT3a, RP Gleason score ≥4 + 5 and negative margins, SUVmax change from 4 to 16 lessened the predicted 3-year BRFS from 76% to 61%. Calibration on an external sample (n = 464) showed reasonable performance; however, a tendency to overestimate survival in patients with good prognostic factors was observed. CONCLUSION: Tumour SUVmax on diagnostic PSMA-PET/CT has utility additional to commonly recognised variables for prediction of BRFS after RP.

Delaying post-exercise carbohydrate intake impairs next-day exercise capacity but not muscle glycogen or molecular responses.

AIM: To investigate how delayed post-exercise carbohydrate intake affects muscle glycogen, metabolic- and mitochondrial-related molecular responses, and subsequent high-intensity interval exercise (HIIE) capacity. METHODS: In a double-blind cross-over design, nine recreationally active men performed HIIE (10 × 2-min cycling, ~94% W˙peak) in the fed state, on two occasions. During 0-3 h post-HIIE, participants drank either carbohydrates ("Immediate Carbohydrate" [IC], providing 2.4 g/kg) or water ("Delayed Carbohydrate" [DC]); total carbohydrate intake over 24 h post-HIIE was matched (~7 g/kg/d). Skeletal muscle (sampled pre-HIIE, post-HIIE, +3 h, +8 h, +24 h) was analyzed for whole-muscle glycogen and mRNA content, plus signaling proteins in cytoplasmic- and nuclear-enriched fractions. After 24 h, participants repeated the HIIE protocol until failure, to test subsequent HIIE capacity; blood lactate, heart rate, and ratings of perceived effort (RPE) were measured throughout. RESULTS: Muscle glycogen concentrations, and relative changes, were similar between conditions throughout (p > 0.05). Muscle glycogen was reduced from baseline (mean ± SD mmol/kg dm; IC: 409 ± 166; DC: 352 ± 76) at post-HIIE (IC: 253 ± 96; DC: 214 ± 82), +3 h (IC: 276 ± 62; DC: 269 ± 116) and + 8 h (IC: 321 ± 56; DC: 269 ± 116), returning to near-baseline by +24 h. Several genes (PGC-1ɑ, p53) and proteins (p-ACCSer79, p-P38 MAPKThr180/Tyr182) elicited typical exercise-induced changes irrespective of condition. Delaying carbohydrate intake reduced next-day HIIE capacity (5 ± 3 intervals) and increased RPE (~2 ratings), despite similar physiological responses between conditions. CONCLUSION: Molecular responses to HIIE (performed in the fed state) were not enhanced by delayed post-exercise carbohydrate intake. Our findings support immediate post-exercise refueling if the goal is to maximize next-day HIIE capacity and recovery time is ≤24 h.

Antibody sequence determinants of viral antigen specificity.

Throughout life, humans experience repeated exposure to viral antigens through infection and vaccination, resulting in the generation of diverse, antigen-specific antibody repertoires. A paramount feature of antibodies that enables their critical contributions in counteracting recurrent and novel pathogens, and consequently fostering their utility as valuable targets for therapeutic and vaccine development, is the exquisite specificity displayed against their target antigens. Yet, there is still limited understanding of the determinants of antibody-antigen specificity, particularly as a function of antibody sequence. In recent years, experimental characterization of antibody repertoires has led to novel insights into fundamental properties of antibody sequences but has been largely decoupled from at-scale antigen specificity analysis. Here, using the LIBRA-seq technology, we generated a large data set mapping antibody sequence to antigen specificity for thousands of B cells, by screening the repertoires of a set of healthy individuals against 20 viral antigens representing diverse pathogens of biomedical significance. Analysis uncovered virus-specific patterns in variable gene usage, gene pairing, somatic hypermutation, as well as the presence of convergent antiviral signatures across multiple individuals, including the presence of public antibody clonotypes. Notably, our results showed that, for B-cell receptors originating from different individuals but leveraging an identical combination of heavy and light chain variable genes, there is a specific CDRH3 identity threshold above which B cells appear to exclusively share the same antigen specificity. This finding provides a quantifiable measure of the relationship between antibody sequence and antigen specificity and further defines experimentally grounded criteria for defining public antibody clonality.IMPORTANCEThe B-cell compartment of the humoral immune system plays a critical role in the generation of antibodies upon new and repeated pathogen exposure. This study provides an unprecedented level of detail on the molecular characteristics of antibody repertoires that are specific to each of the different target pathogens studied here and provides empirical evidence in support of a 70% CDRH3 amino acid identity threshold in pairs of B cells encoded by identical IGHV:IGL(K)V genes, as a means of defining public clonality and therefore predicting B-cell antigen specificity in different individuals. This is of exceptional importance when leveraging public clonality as a method to annotate B-cell receptor data otherwise lacking antigen specificity information. Understanding the fundamental rules of antibody-antigen interactions can lead to transformative new approaches for the development of antibody therapeutics and vaccines against current and emerging viruses.

An Efficient Fluorescence-Activated Protoplast Sorting (FAPS) and Regeneration Protocol for Canola (Brassica napus).

Protoplast sorting and purification methods are powerful tools enabling the enrichment of cellular subpopulations for basic and applied studies in plant sciences. Fluorescence-activated protoplast sorting (FAPS) is an efficient method to isolate specific protoplast populations based on innate features (size and autofluorescence) or expression of fluorescent proteins. FAPS-based methods have recently been deployed in single-cell purification for single-cell RNA sequencing-based transcriptional profiling studies. Protoplast sorting methods integrated with the ability to culture and recover whole plants add value to functional genomics and gene editing applications. Enriching cells expressing nucleases linked to fluorescent proteins can maximize knockout or knockin editing efficiencies and minimize toxic and off-target effects. Here, we report the protocol for protoplast preparation, sterile cell sorting, culture, and downstream regeneration of plants from canola protoplasts. This protocol can be successfully applied to all totipotent protoplast methods that can regenerate into whole plants. © 2024 Wiley Periodicals LLC. Basic Protocol 1: Preparation of transfected canola protoplasts for sorting Basic Protocol 2: Fluorescence-activated protoplast sorting Basic Protocol 3: Bead culture of sorted protoplasts and recovery of plantlets.

Real-world impact of differences in the WHO and ICC classifications of non-Hodgkin lymphoma: a LEO Cohort Study analysis.

Recent introduction of two different lymphoma classifications has raised concerns about consistency in diagnosis, management, and clinical trial enrollment. Data from a large cohort reflecting real-world clinical practice suggest that differences between the classifications will impact <1% of non-Hodgkin lymphomas.

Whole nervous system expression of glutamate receptors reveals distinct receptor roles in sensorimotor circuits.

A goal of connectomics is to reveal the links between neural circuits and behavior. Larvae of the primitive chordate Ciona are well-suited to make contributions in this area. In addition to having a described connectome, Ciona larvae have a range of readily-quantified behaviors. Moreover, the small number of neurons in the larval CNS (∼180) holds the promise of a comprehensive characterization of individual neurons. We present single-neuron predictions for glutamate receptor (GlutR) expression based on in situ hybridization. Included are both ionotropic receptors (AMPA, NMDA, and Kainate), and metabotropic receptors. The predicted glutamate receptor expression dataset is discussed in the context of known circuits driving behaviors such as phototaxis, mechanosensation, and looming shadow response. The predicted expression of AMPA and NMDA receptors may help to resolve issues regarding the co-production of GABA and glutamate by a subset of photoreceptors. The targets of these photoreceptors in the midbrain appear to express NMDA receptors, but not AMPA receptors. This is in agreement with previous results indicating that GABA is the primary neurotransmitter from the photoreceptors evoking a swimming response through a disinhibition mechanism, and that glutamate may, therefore, have only a modulatory action in this circuit. Other findings reported here are more unexpected. For example, many of the targets of glutamatergic epidermal sensory neurons (ESNs) do not express any of the ionotropic receptors, yet the ESNs themselves express metabotropic receptors. Thus, we speculate that their production of glutamate may be for communication with neighboring ESNs, rather than to their interneuron targets.Significance Statement Simple invertebrates offer a tractable alternative to complex vertebrate brains, facilitating holistic understanding of brain function. One such invertebrate is the marine chordate Ciona, which has the benefit of a complete synaptic wiring diagram for its swimming larva. This "connectome" allowed identification of putative neural circuits driving defined behaviors. Fuller understanding of neural circuits, however, requires a description of the attributes of individual neurons. This study focuses on the excitatory neurotransmitter glutamate, which signals via a complex set of both ionotropic and metabotropic receptors. Here, we present a nervous system-wide prediction of GlutR expression in Ciona at the individual neuron level, considered in the context of neural circuits, with emphasis on how GlutR expression accounts for function of neural circuits.