Biology subjects

Huh, H.

Publications and source records attributed to Huh, H..

5 recordsLinked to original sources

bioRxiv2026

Development of an Exploratory Taxonomy for Veterinary Professionals' AI Query Patterns Across Clinical Stages: An Expert Panel Study

Background/ObjectivesThe integration of large language model (LLM)-based AI tools into veterinary clinical practice is rapidly increasing; however, no systematically derived taxonomy of veterinary AI query patterns has been established. This study aimed to develop and refine an exploratory taxonomy of veterinary AI query patterns across clinical stages through a structured expert-panel review process. MethodsAn exploratory cross-sectional expert panel study was conducted. 5,372 real-world query logs from a veterinary clinical AI chatbot deployed over eight months were analyzed using AI-assisted inductive coding to derive an initial taxonomy. The taxonomy was refined through literature review and subsequently reviewed by an expert panel of 38 veterinary professionals via structured online survey across five clinical stages. ResultsA taxonomy of 3 categories and 21 subtypes was established: Clinical Support Queries (Types A-H), Evidence-Based Research Queries (Types I-L), and Terminology and Drug Reference Queries (Types M-U). Type B (Differential Reasoning) had the highest overall frequency (57/188 first-choice responses), while Type D (Clinical Decision Support) was dominant immediately post-consultation (55.3%). Veterinary professionals with [≥]10 years of experience showed a higher frequency of Type G (Evidence Search) preference than those with <10 years of experience (18 vs. 4), while university-affiliated professionals demonstrated a distinct pattern dominated by Type G. ConclusionsTo our knowledge, no published study has previously established a veterinary-specific, clinical-stage-sensitive exploratory taxonomy of AI query patterns; this study addresses that gap. The findings provide a foundational framework for designing context-aware, stage-adaptive veterinary AI systems and benchmark evaluation tools.

scientific communication and education↗

bioRxiv2024

Time-resolved tracking of cellulose biosynthesis and microfibril network assembly during cell wall regeneration in live Arabidopsis protoplasts

Significance StatementCellulose is a major extracellular matrix component of cells that is critical for plant development and has applications to bioenergy, agricultural food/feed, textile, and wood production. Cellulose is thought to be assembled by the closely coordinated motion of plasma membrane-embedded cellulose synthase enzyme complexes. To date, however, it has not been possible to visualize de novo plant cell wall synthesis at the single cell level with the necessary spatiotemporal resolution to derive a data-driven model of how plant cells can resynthesize and assemble cell wall after its removal. Based on our time-resolved data, we propose a new model for cellulose biosynthesis after successfully performing live protoplast time-lapse imaging to visualize for the first time the complex dynamics of de novo cellulose biosynthesis and assembly into an intertwined microfibril network. Plant cell walls are composed of polysaccharides among which cellulose is the most abundant component. Cellulose is processively synthesized as bundles of linear {beta}-1,4-glucan homopolymer chains via the coordinated action of multiple enzymes in cellulose synthase complexes (CSCs) embedded within the plasma cell membrane. Plant cell walls are composed of multiple layers of cellulose fibrils that form highly intertwined extracellular matrix networks. However, it is not yet clear as to how cellulose fibrils synthesized by multiple CSCs are assembled into the intricate cellulose network deposited on plant cell surfaces. Herein, we have established an in vivo time-resolved imaging platform for visualizing cellulose during its biosynthesis and assembly into a complex fibrillar network on the surface of Arabidopsis thaliana mesophyll protoplasts as the primary cell wall regenerates. We performed total internal reflection fluorescence microscopy (TIRFM) with fluorophore-conjugated tandem carbohydrate binding modules (tdCBMs) that were engineered to specifically bind to nascent cellulose fibrils. Together with a well-controlled environment, it was possible to monitor in vivo cellulose fibril synthesis dynamics in a time-resolved manner for nearly one day of continuous cell wall regeneration on protoplast cell surfaces. Our observations provide the basis for a novel model of cellulose fibril network development in protoplasts driven by complex interplay of multi-scale dynamics that include: rapid diffusion and coalescence of short nascently synthesized cellulose fibrils; processive elongation of single fibrils; and cellulose fibrillar network rearrangement during cell wall maturation. This platform is valuable for exploring mechanistic aspects of cell wall synthesis while visualizing cellulose microfibrils assembly.

plant biology↗

bioRxiv2024

Bioadhesive Hydrogel-Coupled and Miniaturized Ultrasound Transducer System for Long-Term, Wearable Neuromodulation

Transcranial focused ultrasound has become a promising non-invasive approach for neuromodulation applications, particularly for neurodegenerative diseases and psychiatric illnesses. However, its implementation in wearable neuromodulation has thus far been limited due to the devices large size, which needs external supporting systems for the neuromodulation process. Furthermore, the need for ultrasound gel for acoustic coupling between the device and skin limits the viability for long-term use, due to its inherent susceptibility to dehydration and lack of adhesiveness to form a stable interface. Here, we report a wearable miniaturized ultrasound device with size comparable to standard EEG/ECG electrodes integrated with bioadhesive hydrogel to achieve efficient acoustic intensity upon ultrasound stimulation for long-term, wearable primary somatosensory cortical stimulation. Specifically, air-cavity Fresnel lens (ACFAL) based self-focusing acoustic transducer (SFAT) was fabricated using a lithography-free microfabrication process. Our transducer was able to achieve an acoustic intensity of up to 30.7 W/cm2 (1.92 MPa) in free-field with a focal depth of 10 mm. Bioadhesive hydrogel was developed to address the need for long-term stability of acoustic couplant for ultrasound application. The hydrogel demonstrated less than 13% attenuation in acoustic intensity and stable adhesion force of 0.961 N/cm over 35 days. Leveraging our bioadhesive hydrogel-integrated wearable ultrasound transducer, we were able to suppress somatosensory evoked potentials elicited by median nerve stimulation via functional electrical stimulation over 28 days, demonstrating the efficacy of our transducer for long-term, wearable neuromodulation in the brain.

bioengineering↗

bioRxiv2020

Multiscale Characterization of Complex Binding Interactions of Cellulolytic Enzymes Highlights Limitations of Classical Approaches

Cellulolytic microorganisms, like Trichoderma reesei or Clostridium thermocellum, frequently have non-catalytic carbohydrate-binding modules (CBMs) associated with secreted or cell surface bound multidomain carbohydrate-active enzymes (CAZymes) like cellulases. Mostly type-A family CBMs are known to promote cellulose deconstruction by increasing the substrate-bound concentration of cognate cellulase catalytic domains. However, due to the interfacial nature of cellulose hydrolysis and the structural heterogeneity of cellulose, it has been challenging to fully understand the role of CBMs on cellulase activity using classical protein-ligand binding assays. Here, we report a single-molecule CAZyme assay for an industrially relevant processive cellulase Cel7A (from T. reesei) to reveal how subtle CBM1 binding differences can drastically impact cellulase motility/velocity and commitment to initial processive motion for deconstruction of two well-studied crystalline cellulose allomorphs (namely cellulose I and III). We take a multifaceted approach to characterize the complex binding interactions of all major type-A family representative CBMs including CBM1, using an optical-tweezers based single-molecule CBM-cellulose bond rupture assay to complement several classical bulk ensemble protein-ligand binding characterization methods. While our work provides a basis for the cautious use of Langmuir-type adsorption models to characterize classical protein-ligand binding assay data, we highlight the critical limitations of using such overly simplistic models to gain a truly molecular-level understanding of interfacial protein binding interactions at heterogeneous solid-liquid interfaces. Finally, molecular dynamics simulations provided a theoretical basis for the complex binding behavior seen for CBM1 towards two distinct cellulose allomorphs reconciling experimental findings from multiscale analytical methods. Significance StatementMultimodal biomolecular binding interactions involving carbohydrate polymers (e.g., cellulose, starch, chitin, glycosaminoglycans) are fundamental molecular processes relevant to the recognition, biosynthesis, and degradation of all major terrestrial and aquatic biomass. Protein-carbohydrate binding interactions are also critical to industrial biotechnology operations such as enzymatically-catalyzed bioconversion of starch and lignocellulose into biochemicals like ethanol. However, despite the ubiquitous importance of such interfacial processes, we have a poor molecular-level understanding of protein-polysaccharide binding interactions. Here, we provide a comprehensive experimental and theoretical analysis of bulk ensemble versus single-molecule binding interactions of enzyme motors and associated non-catalytic binding domains with cellulosic polysaccharides to highlight the critical limitations of applying classical biochemical assay techniques alone to understanding protein adsorption or biological activity at solid-liquid interfaces.

biochemistry↗

bioRxiv2020

Developmental differentiation in a cytoplasm-dwelling obligate intracellular bacterium

Orientia tsutsugamushi (Ot) is an obligate intracellular bacterium in the family Rickettsiaceae that causes scrub typhus, a severe mite-borne human disease. Its mechanism of cell exit is unusual amongst Rickettsiaceae, as Ot buds off the surface of infected cells enveloped in plasma membrane. Here, we show that Ot bacteria that have budded out of host cells are in a distinct developmental stage compared with intracellular bacteria. We refer to these two stages as intracellular and extracellular bacteria (IB and EB, respectively). These two forms differ in physical properties: IB is elongated, and EB is round. Additionally, IB has higher levels of peptidoglycan and is physically robust compared with EB. The two bacterial forms differentially express proteins involved in bacterial physiology and host-pathogen interactions, specifically those involved in bacterial dormancy and stress response, secreted bacterial effectors, and outer membrane autotransporter proteins ScaA and ScaC. Whilst both populations are infectious, entry of IB Ot is sensitive to inhibitors of both clathrin-mediated endocytosis and macropinocytosis, whereas entry of EB Ot is only sensitive to a macropinocytosis inhibitor. Our identification and detailed characterization of two developmental forms of Ot significantly advances our understanding of the intracellular lifecycle of an important human pathogen. Author SummaryOrientia tsutsugamushi (Ot) is a bacterial pathogen that causes scrub typhus, a mite-transmitted human disease. This illness is traditionally known to be endemic in the Asia-Pacific, but recent reports of Orientia-like organisms from the Middle East, Africa, and Latin America suggest that it may be globally distributed. Scrub typhus is associated with high mortality if not treated promptly with appropriate antibiotics. Ot is a highly specialized bacterium that can only replicate within living cells, either within the mite vector or in mammalian or human hosts. Ot exits infected cells using a unique mechanism that involves budding off the surface of infected cells. We have discovered that this unusual aspect of its lifecycle involves the bacteria themselves differentiating into a distinct growth form. Different growth forms have not been described in other members of the family Rickettsiaceae, and no other family members have been shown to bud out of host cells in a manner similar to Ot. We find that the two forms of Ot, which we refer to as intracellular and extracellular bacteria (IB and EB respectively), differ in physical properties and protein expression and infect cells through different mechanisms. The identification of structurally and functionally distinct forms of Ot elucidates a vital aspect of this pathogens intracellular life cycle. The two forms are likely to have different antibiotic susceptibilities, therefore our findings may advance the development of novel interventions aimed at inhibiting Ot growth in scrub typhus patients.

microbiology↗