Antagonism between neuropeptides and monoamines in a distributed circuit for pathogen avoidance.

Pathogenic infection elicits behaviors that promote recovery and survival of the host. After exposure to the pathogenic bacterium Pseudomonas aeruginosa PA14, the nematode Caenorhabditis elegans modifies its sensory preferences to avoid the pathogen. Here, we identify antagonistic neuromodulators that shape this acquired avoidance behavior. Using an unbiased cell-directed neuropeptide screen, we show that AVK neurons upregulate and release RF/RYamide FLP-1 neuropeptides during infection to drive pathogen avoidance. Manipulations that increase or decrease AVK activity accelerate or delay pathogen avoidance, respectively, implicating AVK in the dynamics of avoidance behavior. FLP-1 neuropeptides drive pathogen avoidance through the G protein-coupled receptor DMSR-7, as well as other receptors. DMSR-7 in turn acts in multiple neurons, including tyraminergic/octopaminergic neurons that receive convergent avoidance signals from the cytokine DAF-7/transforming growth factor ß. Neuromodulators shape pathogen avoidance through multiple mechanisms and targets, in agreement with the distributed neuromodulatory connectome of C. elegans.

Acromegaly Type 1: A Representative Patient.

A 46-year-old woman was troubled by a 3-year history of constant headaches and arthralgias. She was treated with paracetamol with no symptom resolution. An abnormal fasting glucose level prompted endocrine evaluation. On physical examination, she casually mentioned that her wedding ring no longer fit, and she also confirmed an increase in shoe size. There were no characteristic facial features for acromegaly and there was no evidence of acral enlargement. Biochemical evaluation, including insulin-like growth factor type 1 (IGF-1) measurement and oral glucose loading with growth hormone (GH) measurement confirmed excess GH production and a diagnosis of acromegaly. Pituitary magnetic resonance imaging showed a central pituitary microadenoma. After transsphenoidal surgical resection, tissue immunohistochemistry revealed a densely granulated somatotroph adenoma. Currently, the patient is asymptomatic with biochemical disease control, normal fasting glucose levels, and no pituitary hormone deficiencies. This patient is illustrative of a type 1 acromegaly with mild clinical manifestations. Clinicians should be aware of acromegaly subtypes to avoid delay in diagnosis and to individualize therapy.

An overview of log likelihood ratio cost in forensic science - Where is it used and what values can we expect?

There is increasing support for reporting evidential strength as a likelihood ratio (LR) and increasing interest in (semi-)automated LR systems. The log-likelihood ratio cost (Cllr) is a popular metric for such systems, penalizing misleading LRs further from 1 more. Cllr = 0 indicates perfection while Cllr = 1 indicates an uninformative system. However, beyond this, what constitutes a "good" Cllr is unclear. Aiming to provide handles on when a Cllr is "good", we studied 136 publications on (semi-)automated LR systems. Results show Cllr use heavily depends on the field, e.g., being absent in DNA analysis. Despite more publications on automated LR systems over time, the proportion reporting Cllr remains stable. Noticeably, Cllr values lack clear patterns and depend on the area, analysis and dataset. As LR systems become more prevalent, comparing them becomes crucial. This is hampered by different studies using different datasets. We advocate using public benchmark datasets to advance the field.

Local and dynamic regulation of neuronal glycolysis in vivo.

Energy metabolism supports neuronal function. While it is well established that changes in energy metabolism underpin brain plasticity and function, less is known about how individual neurons modulate their metabolic states to meet varying energy demands. This is because most approaches used to examine metabolism in living organisms lack the resolution to visualize energy metabolism within individual circuits, cells, or subcellular regions. Here, we adapted a biosensor for glycolysis, HYlight, for use in Caenorhabditis elegans to image dynamic changes in glycolysis within individual neurons and in vivo. We determined that neurons cell-autonomously perform glycolysis and modulate glycolytic states upon energy stress. By examining glycolysis in specific neurons, we documented a neuronal energy landscape comprising three general observations: 1) glycolytic states in neurons are diverse across individual cell types; 2) for a given condition, glycolytic states within individual neurons are reproducible across animals; and 3) for varying conditions of energy stress, glycolytic states are plastic and adapt to energy demands. Through genetic analyses, we uncovered roles for regulatory enzymes and mitochondrial localization in the cellular and subcellular dynamic regulation of glycolysis. Our study demonstrates the use of a single-cell glycolytic biosensor to examine how energy metabolism is distributed across cells and coupled to dynamic states of neuronal function and uncovers unique relationships between neuronal identities and metabolic landscapes in vivo.

Understanding neural circuit function through synaptic engineering.

Synapses are a key component of neural circuits, facilitating rapid and specific signalling between neurons. Synaptic engineering - the synthetic insertion of new synaptic connections into in vivo neural circuits - is an emerging approach for neural circuit interrogation. This approach is especially powerful for establishing causality in neural circuit structure-function relationships, for emulating synaptic plasticity and for exploring novel patterns of circuit connectivity. Contrary to other approaches for neural circuit manipulation, synaptic engineering targets specific connections between neurons and functions autonomously with no user-controlled external activation. Synaptic engineering has been successfully implemented in several systems and in different forms, including electrical synapses constructed from ectopically expressed connexin gap junction proteins, synthetic optical synapses composed of presynaptic photon-emitting luciferase coupled with postsynaptic light-gated channels, and artificial neuropeptide signalling pathways. This Perspective describes these different methods and how they have been applied, and examines how the field may advance.

Mis-spliced transcripts generate de novo proteins in TDP-43-related ALS/FTD.

Functional loss of TDP-43, an RNA binding protein genetically and pathologically linked to amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD), leads to the inclusion of cryptic exons in hundreds of transcripts during disease. Cryptic exons can promote the degradation of affected transcripts, deleteriously altering cellular function through loss-of-function mechanisms. Here, we show that mRNA transcripts harboring cryptic exons generated de novo proteins in TDP-43-depleted human iPSC-derived neurons in vitro, and de novo peptides were found in cerebrospinal fluid (CSF) samples from patients with ALS or FTD. Using coordinated transcriptomic and proteomic studies of TDP-43-depleted human iPSC-derived neurons, we identified 65 peptides that mapped to 12 cryptic exons. Cryptic exons identified in TDP-43-depleted human iPSC-derived neurons were predictive of cryptic exons expressed in postmortem brain tissue from patients with TDP-43 proteinopathy. These cryptic exons produced transcript variants that generated de novo proteins. We found that the inclusion of cryptic peptide sequences in proteins altered their interactions with other proteins, thereby likely altering their function. Last, we showed that 18 de novo peptides across 13 genes were present in CSF samples from patients with ALS/FTD spectrum disorders. The demonstration of cryptic exon translation suggests new mechanisms for ALS/FTD pathophysiology downstream of TDP-43 dysfunction and may provide a potential strategy to assay TDP-43 function in patient CSF.

The American Society of Pain and Neuroscience (ASPN) Guidelines for Radiofrequency Ablative Procedures in Patients with Implanted Devices.

Radiofrequency ablation (RFA) is a treatment modality used in interventional pain management to treat several conditions including chronic neck or back pain, sacroiliac joint pain, major joint pain, and pain from sites that can be isolated to a sensory nerve amenable to RFA. The goals of such procedures are to reduce pain, improve function, delay need for surgical intervention, and reduce pain medication consumption. As applications for RFA expand through novel techniques and nerve targets, there is concern with how RFA may impact patients with implanted medical devices. Specifically, the electrical currents used in RFA produce electromagnetic interference, which can result in unintentional energy transfer to implanted devices. This may also interfere with device function or cause damage to the device itself. As the number of patients with implanted devices increases, it is imperative to establish guidelines for the management of implanted devices during RFA procedures. This review aims to establish guidelines to assist physicians in the preoperative, intraoperative, and postoperative management of implanted devices in patients undergoing procedures using radiofrequency energy. Here, we provide physicians with background knowledge and a summary of current evidence to allow safe utilization of RFA treatment in patients with implanted devices such as cardiac implantable electronic devices, spinal cord stimulators, intrathecal pumps, and deep brain stimulators. While these guidelines are intended to be comprehensive, each patient should be assessed on an individual basis to optimize outcomes.

Deep learning-based aberration compensation improves contrast and resolution in fluorescence microscopy.

Optical aberrations hinder fluorescence microscopy of thick samples, reducing image signal, contrast, and resolution. Here we introduce a deep learning-based strategy for aberration compensation, improving image quality without slowing image acquisition, applying additional dose, or introducing more optics into the imaging path. Our method (i) introduces synthetic aberrations to images acquired on the shallow side of image stacks, making them resemble those acquired deeper into the volume and (ii) trains neural networks to reverse the effect of these aberrations. We use simulations to show that applying the trained 'de-aberration' networks outperforms alternative methods, and subsequently apply the networks to diverse datasets captured with confocal, light-sheet, multi-photon, and super-resolution microscopy. In all cases, the improved quality of the restored data facilitates qualitative image inspection and improves downstream image quantitation, including orientational analysis of blood vessels in mouse tissue and improved membrane and nuclear segmentation in C. elegans embryos.

An ANXA11 P93S variant dysregulates TDP-43 and causes corticobasal syndrome.

As genetic testing has become more accessible and affordable, variants of uncertain significance (VUS) are increasingly identified, and determining whether these variants play causal roles in disease is a major challenge. The known disease-associated Annexin A11 (ANXA11) mutations result in ANXA11 aggregation, alterations in lysosomal-RNA granule co-trafficking, and TDP-43 mis-localization and present as amyotrophic lateral sclerosis or frontotemporal dementia. We identified a novel VUS in ANXA11 (P93S) in a kindred with corticobasal syndrome and unique radiographic features that segregated with disease. We then queried neurodegenerative disorder clinic databases to identify the phenotypic spread of ANXA11 mutations. Multi-modal computational analysis of this variant was performed and the effect of this VUS on ANXA11 function and TDP-43 biology was characterized in iPSC-derived neurons. Single-cell sequencing and proteomic analysis of iPSC-derived neurons and microglia were used to determine the multiomic signature of this VUS. Mutations in ANXA11 were found in association with clinically diagnosed corticobasal syndrome, thereby establishing corticobasal syndrome as part of ANXA11 clinical spectrum. In iPSC-derived neurons expressing mutant ANXA11, we found decreased colocalization of lysosomes and decreased neuritic RNA as well as decreased nuclear TDP-43 and increased formation of cryptic exons compared to controls. Multiomic assessment of the P93S variant in iPSC-derived neurons and microglia indicates that the pathogenic omic signature in neurons is modest compared to microglia. Additionally, omic studies reveal that immune dysregulation and interferon signaling pathways in microglia are central to disease. Collectively, these findings identify a new pathogenic variant in ANXA11, expand the range of clinical syndromes caused by ANXA11 mutations, and implicate both neuronal and microglia dysfunction in ANXA11 pathophysiology. This work illustrates the potential for iPSC-derived cellular models to revolutionize the variant annotation process and provides a generalizable approach to determining causality of novel variants across genes.

[Septic shock originating from a pulmonary focus caused by Pasteurella multocida. Report of one case]. / Shock séptico de foco respiratorio por Pasteurella multocida en un paciente inmunocompetente. Caso Clínico.

Pasteurella multocida is a gram-negative coccobacillus bacterium found as a commensal in the oropharynx of domestic animals such as cats and dogs and some farm animals. Soft tissue infections and occasionally bacteremia in immunocompromised patients with direct contact with animals are described. We report a 61 year old male with a history of scratches and close contact with domestic cats, with a septic shock originating from a pulmonary focus, requiring mechanical ventilation and vasopressors. Blood cultures disclosed the presence of Pasteurella multocida. He responded successfully to antimicrobials.

Local and dynamic regulation of neuronal glycolysis in vivo.

Energy metabolism supports neuronal function. While it is well established that changes in energy metabolism underpin brain plasticity and function, less is known about how individual neurons modulate their metabolic states to meet varying energy demands. This is because most approaches used to examine metabolism in living organisms lack the resolution to visualize energy metabolism within individual circuits, cells, or subcellular regions. Here we adapted a biosensor for glycolysis, HYlight, for use in C. elegans to image dynamic changes in glycolysis within individual neurons and in vivo. We determined that neurons perform glycolysis cell-autonomously, and modulate glycolytic states upon energy stress. By examining glycolysis in specific neurons, we documented a neuronal energy landscape comprising three general observations: 1) glycolytic states in neurons are diverse across individual cell types; 2) for a given condition, glycolytic states within individual neurons are reproducible across animals; and 3) for varying conditions of energy stress, glycolytic states are plastic and adapt to energy demands. Through genetic analyses, we uncovered roles for regulatory enzymes and mitochondrial localization in the cellular and subcellular dynamic regulation of glycolysis. Our study demonstrates the use of a single-cell glycolytic biosensor to examine how energy metabolism is distributed across cells and coupled to dynamic states of neuronal function, and uncovers new relationships between neuronal identities and metabolic landscapes in vivo.

Programmed Cell Death Modifies Neural Circuits and Tunes Intrinsic Behavior.

Programmed cell death is a common feature of animal development. During development of the C. elegans hermaphrodite, programmed cell death (PCD) removes 131 cells from stereotyped positions in the cell lineage, mostly in neuronal lineages. Blocking cell death results in supernumerary "undead" neurons. We find that undead neurons can be wired into circuits, can display activity, and can modify specific behaviors. The two undead RIM-like neurons participate in the RIM-containing circuit that computes movement. The addition of these two extra neurons results in animals that initiate fewer reversals and lengthens the duration of those reversals that do occur. We describe additional behavioral alterations of cell-death mutants, including in turning angle and pharyngeal pumping. These findings reveal that, like too much PCD, too little PCD can modify nervous system function and animal behavior.

Case report: Urbanized non-human primates as sentinels for human zoonotic diseases: a case of acute fatal toxoplasmosis in a free-ranging marmoset in coinfection with yellow fever virus.

Free-ranging non-human primates (NHP) can live in anthropized areas or urban environments in close contact with human populations. This condition can enable the emergence and transmission of high-impact zoonotic pathogens. For the first time, we detected a coinfection of the yellow fever (YF) virus with Toxoplasma gondii in a free-ranging NHP in a highly urbanized area of a metropolis in Brazil. Specifically, we observed this coinfection in a black-tufted marmoset found dead and taken for a necropsy by the local health surveillance service. After conducting an epidemiological investigation, characterizing the pathological features, and performing molecular assays, we confirmed that the marmoset developed an acute fatal infection caused by T. gondii in coinfection with a new YF virus South American-1 sub-lineage. As a result, we have raised concerns about the public health implications of these findings and discussed the importance of diagnosis and surveillance of zoonotic agents in urbanized NHPs. As competent hosts of zoonotic diseases such as YF and environmental sentinels for toxoplasmosis, NHPs play a crucial role in the One Health framework to predict and prevent the emergence of dangerous human pathogens.

A fully automated FAIMS-DIA mass spectrometry-based proteomic pipeline.

Here, we present a standardized, "off-the-shelf" proteomics pipeline working in a single 96-well plate to achieve deep coverage of cellular proteomes with high throughput and scalability. This integrated pipeline streamlines a fully automated sample preparation platform, a data-independent acquisition (DIA) coupled with high-field asymmetric waveform ion mobility spectrometer (FAIMS) interface, and an optimized library-free DIA database search strategy. Our systematic evaluation of FAIMS-DIA showing single compensation voltage (CV) at -35 V not only yields the deepest proteome coverage but also best correlates with DIA without FAIMS. Our in-depth comparison of direct-DIA database search engines shows that Spectronaut outperforms others, providing the highest quantifiable proteins. Next, we apply three common DIA strategies in characterizing human induced pluripotent stem cell (iPSC)-derived neurons and show single-shot mass spectrometry (MS) using single-CV (-35 V)-FAIMS-DIA results in >9,000 quantifiable proteins with <10% missing values, as well as superior reproducibility and accuracy compared with other existing DIA methods.

Specific configurations of electrical synapses filter sensory information to drive choices in behavior.

Synaptic configurations in precisely wired circuits underpin how sensory information is processed by the nervous system, and the emerging animal behavior. This is best understood for chemical synapses, but far less is known about how electrical synaptic configurations modulate, in vivo and in specific neurons, sensory information processing and context-specific behaviors. We discovered that INX-1, a gap junction protein that forms electrical synapses, is required to deploy context-specific behavioral strategies during C. elegans thermotaxis behavior. INX-1 couples two bilaterally symmetric interneurons, and this configuration is required for the integration of sensory information during migration of animals across temperature gradients. In inx-1 mutants, uncoupled interneurons display increased excitability and responses to subthreshold temperature stimuli, resulting in abnormally longer run durations and context-irrelevant tracking of isotherms. Our study uncovers a conserved configuration of electrical synapses that, by increasing neuronal capacitance, enables differential processing of sensory information and the deployment of context-specific behavioral strategies.

Application of Aligned-UMAP to longitudinal biomedical studies.

High-dimensional data analysis starts with projecting the data to low dimensions to visualize and understand the underlying data structure. Several methods have been developed for dimensionality reduction, but they are limited to cross-sectional datasets. The recently proposed Aligned-UMAP, an extension of the uniform manifold approximation and projection (UMAP) algorithm, can visualize high-dimensional longitudinal datasets. We demonstrated its utility for researchers to identify exciting patterns and trajectories within enormous datasets in biological sciences. We found that the algorithm parameters also play a crucial role and must be tuned carefully to utilize the algorithm's potential fully. We also discussed key points to remember and directions for future extensions of Aligned-UMAP. Further, we made our code open source to enhance the reproducibility and applicability of our work. We believe our benchmarking study becomes more important as more and more high-dimensional longitudinal data in biomedical research become available.

Polarized localization of kinesin-1 and RIC-7 drives axonal mitochondria anterograde transport.

Mitochondria transport is crucial for mitochondria distribution in axons and is mediated by kinesin-1-based anterograde and dynein-based retrograde motor complexes. While Miro and Milton/TRAK were identified as key adaptors between mitochondria and kinesin-1, recent studies suggest the presence of additional mechanisms. In C. elegans, ric-7 is the only single gene described so far, other than kinesin-1, that is absolutely required for axonal mitochondria localization. Using CRISPR engineering in C. elegans, we find that Miro is important but is not essential for anterograde traffic, whereas it is required for retrograde traffic. Both the endogenous RIC-7 and kinesin-1 act at the leading end to transport mitochondria anterogradely. RIC-7 recruitment to mitochondria requires its N-terminal domain and partially relies on MIRO-1, whereas RIC-7 accumulation at the leading end depends on its disordered region, kinesin-1 and metaxin2. We conclude that polarized transport complexes containing kinesin-1 and RIC-7 form at the leading edge of mitochondria, and that these complexes are required for anterograde axonal transport.

Impact of lymph node ratio and number of lymph node metastases on survival and recurrence in laryngeal squamous cell carcinoma.

INTRODUCTION: The aim of this study is to assess the impact of lymph node ratio (LNR) and number of positive lymph nodes (NPLN) on mortality and recurrence rates in patients with laryngeal squamous cell carcinoma. MATERIALS AND METHODS: We conducted a retrospective multicenter international study involving 24 Otorhinolaryngology-Head and Neck Surgery divisions. Disease-specific survival (DSS) and disease-free survival (DFS) were evaluated as the main outcomes. The curves for DSS and DFS according to NPLN and LNR were analyzed to identify significant variations and establish specific cut-off values. RESULTS: 2507 patients met the inclusion criteria. DSS and DFS were significantly different in the groups of patients stratified according to LNR and NPLN. The 5-year DSS and DFS based on LNR and NPLN demonstrated an improved ability to stratify patients when compared to pN staging. CONCLUSION: Our data demonstrate the potential prognostic value of NPLN and LNR in laryngeal squamous cell carcinoma.

Gaussianization of LA-ICP-MS features to improve calibration in forensic glass comparison.

The forensic comparison of glass aims to compare a glass sample of an unknown source with a control glass sample of a known source. In this work, we use multi-elemental features from Laser Ablation Inductively Coupled Plasma with Mass Spectrometry (LA-ICP-MS) to compute a likelihood ratio. This calculation is a complex procedure that generally requires a probabilistic model including the within-source and between-source variabilities of the features. Assuming the within-source variability to be normally distributed is a practical premise with the available data. However, the between-source variability is generally assumed to follow a much more complex distribution, typically described with a kernel density function. In this work, instead of modeling distributions with complex densities, we propose the use of simpler models and the introduction of a data pre-processing step consisting on the Gaussianization of the glass features. In this context, to obtain a better fit of the features with the Gaussian model assumptions, we explore the use of different normalization techniques of the LA-ICP-MS glass features, namely marginal Gaussianization based on histogram matching, marginal Gaussianization based on Yeo-Johnson transformation and a more complex joint Gaussianization using normalizing flows. We report an improvement in the performance of the Likelihood Ratios computed with the previously Gaussianized feature vectors, particularly relevant in their calibration, which implies a more reliable forensic glass comparison.

The active zone protein CLA-1 (Clarinet) bridges two subsynaptic domains to regulate presynaptic sorting of ATG-9.

In neuronal synapses, autophagosome biogenesis is coupled with the activity-dependent synaptic vesicle cycle via ATG-9. How vesicles containing ATG-9 are sorted at the presynapse is unknown. We performed forward genetic screens at single synapses of C. elegans neurons for mutants that disrupt ATG-9 presynaptic localization, and identified the long isoform of the active zone protein CLA-1 (Clarinet; CLA-1 L). We find that disrupting CLA-1 L results in abnormal accumulation of ATG-9-containing vesicles enriched with clathrin. The adaptor protein complexes and proteins at the periactive zone genetically interact with CLA-1 L in ATG-9 sorting. Moreover, the phenotype of the ATG-9 protein in cla-1(L) mutants was not observed for integral synaptic vesicle proteins, suggesting distinct mechanisms that regulate sorting of ATG-9-containing vesicles and synaptic vesicles. Our findings reveal novel roles for active zone proteins in the sorting of ATG-9 and in presynaptic macroautophagy/autophagy.