Biology subjects

Lee, K.

Publications and source records attributed to Lee, K..

At least 19 recordsLinked to original sources

bioRxiv2026

Human Saliva Induces Acetoin-Dependent Metabolic Adaptation that Promotes Stress Tolerance in Streptococcus mutans

Oral microbes are continually exposed to host-derived saliva, yet its effects on bacterial metabolism and stress adaptation remain incompletely defined. We investigated how saliva supplementation alters the physiology of the caries-associated bacterium Streptococcus mutans using transcriptomics (RNA-Seq), targeted metabolite measurements, specific gene deletions, and transposon sequencing (Tn-Seq). Growth in a chemically defined medium supplemented with pooled saliva altered expression of genes involved in carbohydrate transport, purine biosynthesis, and pyruvate-associated metabolism, including induction of the acetoin dehydrogenase operon. Saliva significantly increased extracellular acetoin accumulation across all S. mutans isolates tested, whereas little accumulation occurred in two commensal streptococcal species examined. Deletion of acetolactate synthase/decarboxylase (alsSaldB) abolished the saliva-associated increase, while deletion of individual acetoin dehydrogenase components (adhC / lplA) enhanced accumulation, supporting the predicted roles of genes in acetoin synthesis and catabolism. Increasing the available carbohydrate concentrations within the media further increased acetoin accumulation, alongside declining terminal culture pH that was significantly lower in cultures supplemented with saliva. Following growth with saliva, S. mutans exhibited approximately 10,000-fold greater survival during a hydrogen peroxide challenge than cells grown without saliva. Mutants within the acetoin pathway retained saliva-associated protection but showed approximately 100-fold lower survival than the wild type. Serial passage of a pooled transposon library identified previously unrecognized saliva-associated fitness determinants, including depletion of mutants carrying insertions in the acetoin dehydrogenase operon. Together, these findings identify acetoin metabolism as a contributor to saliva-associated metabolic remodeling and oxidative stress tolerance in S. mutans and establish saliva as a critical modulator of S. mutans physiology.

microbiology↗

bioRxiv2026

Differential value processing in the rostral and causal subthalamic nucleus during flexible and habitual decision-making in primates: Implications for cognitive dysfunction in Parkinson's disease

The subthalamic nucleus (STN) is a major target region of deep brain stimulation (DBS) for Parkinson's disease (PD), effectively alleviating motor symptoms. However, STN-DBS can also induce cognitive impairments, suggesting that the STN contributes to cognitive functions beyond simple motor control. Furthermore, anatomical organization supports functional segregation of cognitive processes, as basal ganglia subregions encoding flexible and stable values topographically project to distinct STN subregions. Yet, whether the STN contributes to cognitive functions and, if so, how subregions of the STN differentially process cognitive flexibility and habitual stability remains unknown. Here, we reveal a rostral-caudal functional organization of the STN that differentially supports flexible value updating and long-term value memory, enabling adaptive and habitual decision-making, respectively. Neurons in the rostral STN (rSTN) preferentially encode flexibly updated values, whereas neurons in the caudal STN (cSTN) preferentially encode stable long-term value memories. Notably, inactivation of the STN along the rostral-caudal axis produced dissociable behavioral deficits: rSTN inactivation selectively impaired goal-directed behavior requiring flexible value updating, whereas cSTN inactivation selectively disrupted habitual behavior guided by long-term value memories. Together, these results reveal a rostral-caudal functional segregation of the STN, with the rSTN supporting goal-directed behavior and the cSTN supporting habitual behavior. These findings further provide a framework for understanding how STN dysfunction or STN-DBS in Parkinson's disease may differentially affect cognitive processes beyond motor control.

neuroscience↗

bioRxiv2026

Genomic characterization of patient derived xenograft models of multiple myeloma reveals selection of multiple pre-existing high risk clones

Multiple myeloma (MM) is a genomically complex malignancy of terminally differentiated plasma cells. Here, we have utilized the SCID-rab patient derived xenograft (PDX) model as a platform to expand scarce primary MM samples across disease stages and major cytogenetic subgroups for comprehensive molecular analysis. The model generated abundant CD138+ cells while retaining key genomic, transcriptomic and proteomic features of corresponding patient tumors. Compared with human myeloma cell lines, PDX cells more closely resembled primary MM in proliferative state and expression of therapeutically relevant targets. WGS and single cell WGS showed that PDXs arose from the selection of pre-existing subclones, including clones harboring high-risk alterations and convergent mechanisms of tumor suppressor inactivation. These findings establish SCID-rab PDX models as a platform for multiomic profiling, studies of clonal evolution, and determining therapeutic response in MM.

cancer biology↗

bioRxiv2026

Ultrastructurally planar growth cones during neocortical axon navigation

Neurons extend axons toward spatially defined but often distant targets to establish brain circuits. Callosal projection neurons exemplify this organization, linking corresponding cortical territories between the two hemispheres. Growing axons rely on growth cones to regulate direction and speed through densely packed tissues. Although growth cone morphology and dynamics have been extensively characterized in cultured neurons and invertebrate models, how growth cones navigate the three-dimensional environment of the developing mammalian brain remains poorly explored. Here, we combined a genetically encoded electron microscopy (EM)-labeling strategy and zero-shot deep-learning-based tracing with high-resolution volume EM to reconstruct growth cones of identified callosal projection neurons in the developing mouse brain. Mediolaterally extending growth cones exhibited a planar, fan-shaped morphology, preferentially expanded along the dorsoventral axis. Filopodia extended from the edges of this plane and aligned predominantly with the same axis but showed no directional bias toward the growing direction. Longitudinal in vivo imaging revealed that increased growth cone complexity associated with filopodial extension coincided with lower instantaneous velocity but preceded accelerated extension. Together, these findings reveal a dorsoventrally oriented planar growth cone architecture in which filopodial extension precedes accelerated axon growth, suggesting a structural constraint that may restrict axon navigation to the callosal plane.

neuroscience↗

bioRxiv2026

Growth-state as a determinant of in vivo virulence of Mycoplasma pneumoniae

Mycoplasma pneumoniae is a leading cause of community acquired pneumonia and bronchitis in people, particularly children and young adults. Like many bacterial pathogens, M. pneumoniae can grow in biofilm towers on both biotic and abiotic surfaces. Current models indicate that planktonic M. pneumoniae are simply individual cells that are dispersed from biofilm towers due to physical shear forces, and do not represent a bona fide planktonic growth state. However recent data indicate that these fractions may be physiologically distinct and may differentially influence disease outcome. We observed that experimental infection of mice with inoculum containing both adherent and non-adherent fractions of M. pneumoniae produced more severe lung pathology than inoculum prepared from M. pneumoniae adherent biofilms alone. Direct comparison of experimental infection with titer matched non-adherent (P-Mp) and adherent (B-Mp) fractions revealed that P-Mp produces more severe lung pathology compared to infection with B-Mp. Furthermore, experimental infection with a lower dose of P-Mp resulted in similar lung pathology relative to B-Mp infected mice. This increase in pathology was not associated with differences in bacterial burden, bacterial dispersal during inoculum preparation, nor due to differences in the phenotype of host immune responses. Comparative proteomics revealed that certain known and putative virulence factors associated with immune evasion are upregulated in P-Mp, potentially explaining the difference in virulence between the two fractions. Furthermore, comparative proteomics revealed distinct metabolic phenotypes between the two fractions, consistent with distinct planktonic and biofilm growth states, indicating that P-Mp likely represents a bona fide planktonic growth state. Collectively, these data implicate the growth state of M. pneumoniae as a determinant of in vivo virulence for this atypical human pathogen.

microbiology↗

bioRxiv2026

Assessing the Functional-Structural Reorganization of Connector Hubs in Temporal Lobe Epilepsy

Temporal lobe epilepsy (TLE) is associated with altered functional connectivity, yet its effects on connector hubs, regions supporting long-range and inter-network communication, remain poorly understood. We used SParsity-based Analysis of Reliable k-hubness to characterize connector hub reorganization in unilateral TLE compared with healthy controls. Patients exhibited significant widespread reductions in k-hubness involving the occipital, cingulate, and precuneus regions, ipsilateral hippocampus, and the bilateral pallidum. Compared to the distribution of k-hubness in healthy controls, the hub disruption index revealed rapid loss of k-hubness alongside emergence of new hubs in contralateral homologous regions. To investigate microstructural substrates of these functional changes, apparent diffusion coefficient and fractional anisotropy from diffusion magnetic resonance imaging were examined. The strongest associations with k-hubness were observed in subcortical structures and the hippocampus. These findings demonstrate that TLE is characterized by widespread, asymmetric connector hub reorganization extending beyond the epileptogenic temporal lobe, revealing distinct patterns of network disruption and reorganization across the brain.

neuroscience↗

bioRxiv2026

Real Science Is Harder Than Benchmarks: Evaluating Advanced AI Frameworks on Published Studies. II. Antibody Properties, Lipid-RNA Interactions

Artificial Intelligence (AI) frameworks for automating scientific research have shown strong performance on benchmarks, but their utility for real-world industrial research remains insufficiently characterized. Extending the analysis presented in the first paper of this series, we evaluated the same five advanced AI research frameworks (Kosmos, K-Dense, ToolUniverse, BioAgents from bio.xyz, and the AI Scientist-v2 from Sakana AI) on two more projects of high practical importance for biopharmaceutical development: predicting antibody developability properties with the use of pretrained protein language model embeddings, and modeling non-covalent lipid-RNA interactions in lipid nanoparticles with all-atom molecular dynamics (MD) simulations. The AI frameworks again showed genuine strengths, including unprompted identification of subtle methodological issues, successful use of pretrained protein embeddings, and consistent reporting of p-values and confidence intervals often absent from the original papers. However, no framework approached the scope of the original studies, and severe failures and hallucinations were observed. Our results confirm and extend the conclusion of the first paper that real published research from pharmaceutical companies that we tried to reproduce proved to be considerably harder for current AI frameworks than standard benchmarks suggest.

bioinformatics↗

bioRxiv2026

Strain-Level Diversity Decouples Biofilm Architecture, Acidogenic and Aciduric Traits, and Antimicrobial Tolerance in Streptococcus mutans

Streptococcus mutans is a key contributor to dental caries, with its capacity to form structured biofilm microcolonies being a principal component of its cariogenic potential. Yet, most mechanistic studies rely on a limited number of laboratory strains and may not capture the functional diversity present across the species. Here, we characterized a panel of phenotypically and genomically diverse S. mutans isolates to determine how strain background influences biofilm architecture, extracellular matrix accumulation, acid-associated physiology, environmental responsiveness, and antimicrobial susceptibility. Quantitative high-resolution imaging revealed extensive heterogeneity in produced biofilm microcolony size, structure, and matrix composition, demonstrating that biofilm architecture is not a uniform species-level trait. Interestingly, the commonly used reference strain UA159 displayed an intermediate phenotype related to microcolony size and biofilm organization. Human saliva further altered biofilm structure and matrix accumulation in a strain-dependent manner rather than producing a standard species-wide response. Isolates also differed in growth and retained biofilm biomass under acidic conditions, while acid accumulation within mature biofilms varied independently of average microcolony volume, demonstrating that strains that produce larger microcolonies on average were not necessarily associated with greater acid accumulation. Susceptibility to the antiseptics chlorhexidine and cetylpyridinium chloride likewise differed among isolates and could not be predicted from formed biofilm architecture alone. Together, these findings demonstrate that disease-relevant traits commonly attributed to S. mutans are distributed unevenly and only partially coupled across strain backgrounds, with biofilm spatial organization failing to serve as a dominant phenotype linking acid accumulation, acid tolerance, and antimicrobial susceptibility.

microbiology↗

bioRxiv2026

Multiplex Genome Editing Overcomes Photoperiod Sensitivity in Tropical Maize

Tropical maize is a rich source of genetic diversity that could enhance temperate maize breeding programs, but its sensitivity to long-day photoperiods, resulting in delayed flowering, limits its widespread use. To overcome this barrier, the Genome Engineering to Sustain Crop Improvement (GETSCI) project used CRISPR/Cas9 to mutate three flowering repressor genes, ZmCCT9, ZmCCT10, and ZmRAP2.7, in the tropical inbred Tzi8. A single sgRNA targeting the first exon of each target gene was combined with an excision cassette carrying the morphogenic genes Babyboom (Bbm) and Wuschel2 (Wus2) to enable efficient transgenic plant regeneration. Transgenic plants carrying frameshift edits in each target gene were recovered, and subsequent crosses produced two non-transgenic genotypes: a double-edited zmcct10, zmrap2.7 line and a triple-edited zmcct9, zmcct10, zmrap2.7 line. Multiple flowering traits were measured for the edited genotypes and unedited Tzi8 inbred in short-day (Hawai i) and long-day (Iowa) field conditions. Both edited genotypes flowered significantly earlier than Tzi8 in both environments. Notably, under long-day conditions, flowering of the two edited lines overlapped with that of the temperate inbred B73, whereas Tzi8 did not. Together, these results demonstrate that targeted, multiplex gene editing can reduce photoperiod sensitivity in a tropical inbred, expanding access to previously untapped genetic diversity for temperate maize improvement.

plant biology↗

bioRxiv2026

Hypothalamic oligodendrocytes regulate systemic energy balance through Notch-dependent state transitions

The central mechanisms through which glial cells regulate whole-body metabolism remain poorly understood. Here, we identify Notch signaling in hypothalamic oligodendrocyte lineage cells as a previously unrecognized regulator of systemic energy homeostasis. Pharmacological inhibition of the Notch ligands Jagged1 (Jag1) and Jagged2 (Jag2) induces rapid and reversible weight loss across diverse physiological and metabolic contexts independently of toxicity or caloric intake. Single-nucleus transcriptomic analyses identify hypothalamic oligodendrocyte precursor cells (OPCs) as the principal Notch-responsive population following systemic Jag1/2 inhibition and reveal expansion of a metabolically specialized GPR17 intermediate state characterized by enhanced oxidative metabolism and increased predicted communication with hypothalamic neurons. This glial remodeling is accompanied by fasting-like transcriptional reprogramming of AgRP neurons, reorganization of melanocortin-autonomic circuit activity, and activation of peripheral catabolic programs. Importantly, selective deletion of Notch1/2 in hypothalamic OPCs recapitulates the major physiological and metabolic effects of systemic Jag1/2 inhibition, establishing oligodendrocyte Notch signaling as a causal regulator of whole-body metabolism. Together, our findings establish Notch-dependent oligodendrocyte state transitions as a previously unrecognized mechanism linking glial plasticity to systemic energy homeostasis.

physiology↗

bioRxiv2026

NPC1 deficiency engages a lysosome - genome - immune program linked to neurodegeneration and cellular aging signatures

Lysosomal dysfunction is a prominent feature of neurodegeneration and aging, yet how primary defects in lysosomal trafficking are converted into progressive cellular decline remains poorly understood. Niemann Pick disease type C (NPC), caused by impaired NPC1 dependent cholesterol export, provides a genetically defined model to address this question. Here, we show that NPC1 deficiency activates a lysosome, genome, immune axis linking cholesterol trafficking failure to neurodegeneration and hallmarks of cellular aging. In Npc1 mutant mice, NPC1 loss triggered DNA damage, neuroinflammation, microglial and astrocytic activation, Purkinje neuron degeneration, and motor dysfunction. Consistently, NPC patient-derived fibroblasts exhibited mitochondrial abnormalities and widespread DNA double-strand breaks. Genome-wide DNA break mapping and transcriptomic analyses revealed extensive genomic instability at regulatory regions, including enrichment of DNA breaks at transcription start sites and G quadruplex associated loci, accompanied by widespread transcriptional reprogramming, activation of innate immune pathways, disruption of fibroblast identity, and induction of cellular aging signatures. We further identify Fingolimod, an FDA approved sphingosine - 1 phosphate receptor modulator, as a potent modifier of this disease network. Fingolimod improved lysosomal cholesterol trafficking, increased LAMP1 abundance, attenuated STING associated inflammatory signaling, normalized mitochondrial function, reduced neuroinflammatory and neurodegenerative phenotypes in Npc1 mutant mice, and broadly shifted disease-associated transcriptional programs toward a healthier state. Extending these findings beyond NPC, Fingolimod improved age-associated phenotypes in C. elegans and prolonged lifespan in aged male mice. Together, these findings identify genome instability and chronic innate immune activation as major downstream consequences of lysosomal cholesterol trafficking failure and establish Fingolimod as a clinically actionable modulator of lysosomal dysfunction, neurodegeneration, and aging-related decline.

molecular biology↗

bioRxiv2026

Schistosoma mansoni Granulin binds the human neutrophil receptor CD177 and modulates neutrophil activation

To establish chronic infection in the vasculature of their infected host, schistosomes have developed multifaceted strategies of immune subversion. Extracellular parasite proteins are believed to play immunomodulatory functions, but their mode of action remains largely elusive. To investigate whether proteins secreted by the Schistosoma mansoni parasite have the potential to directly interact with host immune receptors, we performed a large-scale protein:protein interaction study between selected parasite proteins sharing structural similarities with known host immune effectors and a protein array of over 750 full-length human ectodomains mostly expressed by immune cells. We identified CD177 as a neutrophil receptor for S. mansoni Granulin (SmGrn). SmGrn exclusively bound the surface of CD177+ human neutrophils and led to cellular hyporesponsiveness following stimulation with LPS as evidenced by decreases in surface markers of activation, delayed reactive oxygen species production and reduced IL-8 release. In addition, human neutrophils exposed to SmGrn showed delayed apoptosis and morphological changes compatible with a more quiescent state as well as transcriptional upregulation of negative regulators of interferon signalling. These data suggest that SmGrn dampens human neutrophil response to stimulation and may lead to suboptimal function during schistosome infection.

immunology↗

bioRxiv2026

Late-Stage Large Extracellular Vesicles Reprogram CHO Cell Metabolism in a Glutamine-Dependent Mode and Promote Antibody-Productivity to Cell-Growth Tradeoff

Extracellular vesicles (EVs) are mediators of intercellular communication, yet their impact on Chinese Hamster Ovary (CHO) cell physiology and bioprocess performance remains poorly understood. Here, we investigated whether small EVs (sEVs) and large EVs (LgEVs) that accumulate during fed-batch and perfusion cultures modulate CHO cell growth, metabolism, apoptosis, and monoclonal antibody (mAb) production. EVs isolated from early- and late-stage cultures were added to fresh CHO cultures grown with or without glutamine supplementation. Only LgEVs had a significant impact. Late-stage LgEVs markedly altered CHO-cell behavior, reducing cell proliferation, increasing apoptosis under glutamine-limited conditions, and substantially enhancing mAb productivity in a dose-dependent manner. Glutamine supplementation largely alleviated the growth-inhibitory and pro-apoptotic effects of LgEVs while preserving their positive impact on productivity, suggesting that glutamine decouples EV-mediated stress from productivity enhancement. Metabolic analyses revealed increased glucose consumption, a glutamine-dependent shift between glycine and alanine overflow metabolism, and remodeling of amino-acid utilization. Metabolic flux analysis further demonstrated enhanced glycolytic overflow and increased reliance on amino acid-supported anaplerosis. Conversely, selective removal of LgEVs from perfusion medium significantly improved cell expansion without reducing antibody production, supporting an inhibitory role for late-stage LgEVs. These LgEVs were enriched in let-7 family miRNAs and miR-21, consistent with RNAseq analyses demonstrating stress-associated enrichment of these miRNAs in CHO EVs and with functional studies showing that let-7a and miR-21reduce CHO-cell growth. Together, these observations suggest that selective miRNA loading contributes to the growth, metabolic, and productivity phenotypes elicited by late-stage LgEVs. Our findings identify LgEVs as endogenous regulators of CHO-cell physiology and potential targets for optimizing high-density fed-batch and perfusion biomanufacturing processes. HighlightsO_LIEndogenous late-stage Large Extracellular Vesicles (LgEVs) reduce CHO cell growth but boost specific mAb productivity. C_LIO_LIGlutamine supplementation rescues LgEV-mediated growth inhibition and apoptosis. C_LIO_LIMetabolic Flux Analysis (MFA) based on the dynamic behavior of amino acid and other metabolite and substrate concentrations reveals the pyruvate node as a metabolic bottleneck and the associated lactate overflow metabolism as resulting from LgEV exposure. C_LIO_LIStress-associated let-7 and miR-21 microRNAs are highly enriched on a per-EV basis in late-stage LgEVs. C_LIO_LISelective removal of LgEVs improves perfusion cell growth without impacting antibody titer. C_LI

Cell Biology↗

bioRxiv2026

Real Science Is Harder Than Benchmarks: Evaluating Advanced AI Frameworks on Published Studies. I. Uncertainty Quantification, ML on Therapeutic Data Commons, and Agent-Based Modeling

Artificial Intelligence (AI) frameworks for automating scientific research have shown strong performance on benchmarks, but their capacity to routinely reproduce results from multiple real-life published studies remains largely untested. We evaluated five advanced AI research frameworks (Kosmos, K-Dense, ToolUniverse, BioAgents from bio.xyz, and the AI Scientist-v2 from Sakana AI) on three real-life tasks (including two recently published papers) spanning uncertainty quantification for molecular property predictions, machine learning on Therapeutic Data Commons benchmarks, and agent-based modeling. AI frameworks demonstrated genuine strengths: generating original hypotheses, competently executing routine data acquisition and coding tasks, providing statistical measures of confidence often absent from the original papers, and producing well-formatted final reports. At the same time, our experiments revealed that real-world scientific tasks remain considerably harder than current benchmarks suggest. No AI framework matched the scope or depth of the original studies, results varied across multiple runs of the same framework with the same prompt, and we documented cases of severe hallucinations in final reports, gaps in literature coverage, and overconfident conclusions. Verification of AI outputs required substantial domain expertise. While these three tasks are only partially representative of the broader scientific landscape, they offer a starting point for developing a more rigorous methodology for evaluation of AI performance than what is currently practiced. We conclude that AI frameworks are already valuable for prototyping research directions and stress-testing completed studies, and some of the limitations documented here appear largely tractable through infrastructure improvements and continued development.

bioinformatics↗

bioRxiv2026

SMARCA4, STK11, and KEAP1 co-inactivation associates with poor prognosis and upregulation of the TGF-β pathway in lung adenocarcinoma

BackgroundLung adenocarcinoma (LUAD) is clinically and molecularly defined by oncogenic driver mutations, identification of which has led to the development of driver-targeted therapies and substantial improvements in prognosis for subsets of LUAD patients. Recent studies assessing clinical outcomes in the context of multigenic alterations have identified secondary mutations that might explain differential responses to targeted therapies, chemotherapies and immunotherapies. Genetic inactivation or loss of SMARCA4, which frequently co-occurs with loss-of-function mutations in STK11 and KEAP1, is especially predictive of poor prognosis and shorter overall survival in LUAD patients, regardless of driver status. We sought to examine the clinical and functional associations of SMARCA4 deficiency in LUAD, with or without co-associated STK11/KEAP1 loss-of-function. MethodsWe examined correlation between SMARCA4 loss, gene expression and prognosis through genomic and transcriptomic profiling of clinically annotated LUAD samples. We generated isogenic cell line models with genetic knockouts of SMARCA4 with or without concomitant STK11 and KEAP1 to profile mutationally-defined genotypes of interest in vitro and in vivo. Lastly, we interrogated the functional dependency of SMARCA4/STK11/KEAP1 triple mutant models on TGF-{beta} signaling to assess its potential as a therapeutic target. ResultsSMARCA4/STK11/KEAP1 triple mutant LUAD is associated with poor survival and high frequency of multisite metastasis. SMARCA4/STK11/KEAP1 triple knockout models showed enhanced migration and invasion in vitro, and diversified organotropism in an in vivo intracardiac xenograft metastasis assay. RNA-Seq and DNaseI-Seq of these in vitro models and clinical samples identified upregulation of TGF-{beta} signaling and EMT gene expression signatures, and corresponding changes in chromatin accessibility, in SMARCA4/STK11/KEAP1 triple mutant LUAD. ConclusionsWe identify SMARCA4/STK11/KEAP1 triple mutant LUAD as a prognostically significant disease subset and nominate TGF-{beta} signaling as a potential therapeutic target.

cancer biology↗

bioRxiv2026

Slow Oscillations Gate Interictal Spikes Across the Human Thalamocortical-Epileptogenic Network

BackgroundSlow oscillations (SOs; 0.5-1.5 Hz), a hallmark of non-rapid eye movement (NREM) sleep, are associated with a marked amplification of interictal epileptiform spike (IIS) activity in focal epilepsy. However, the network-level organization of this effect across the thalamocortical-epileptogenic system, and whether IIS-permissive SOs can be predicted from pre-onset brain states, remain unclear. MethodsWe analyzed simultaneous scalp EEG and stereo-EEG (SEEG) recordings from 6 patients with drug-resistant focal epilepsy across 24 full-day recording days, sampling prefrontal cortex (PFC), thalamus, and seizure onset zone (SOZ). SO-IIS coupling was characterized across vigilance states using peri-event and phase-based analyses, with a gamma-based validation step to reduce contamination by IIS-related slow potentials. Pre-onset phase-amplitude coupling (PAC) was compared between IIS-permissive and non-permissive SOs. ResultsSO-IIS coupling was observed across all regions, with the strongest and most temporally precise pre-trough IIS clustering in the SOZ (peak 4.4% in NREM), exceeding PFC (1.7%) and thalamic coupling. Thalamic coupling was preserved across wakefulness and NREM and was significant in 7/11 nuclei, with nucleus-specific phase preferences. SO morphological features, particularly up-slope and peak-to-peak amplitude at PFC contacts, predicted IIS occurrence in the SOZ. Pre-onset PAC differed significantly between permissive and non-permissive SOs across regions. ConclusionsSO-IIS coupling is a distributed, network-level phenomenon with region- and state-specific characteristics, and pre-onset PAC provides a predictive signature of IIS-permissive brain states. These findings support the feasibility of developing personalized, closed-loop neuromodulatory strategies targeting SO-gated IIS suppression in focal epilepsy.

neuroscience↗

bioRxiv2026

Student-Driven Microbiome Exploration: A Low-Cost 16S rRNA Sequencing Curriculum for Undergraduate Biology Education

Microbiome science is increasingly important in modern biology education because microbial communities influence human health, ecosystems, and environmental processes. However, undergraduate microbiome instruction is often limited by the high cost and technical complexity of sequencing-based workflows, restricting opportunities for authentic student-driven research. To address this challenge, we developed a low-cost, inquiry-based curriculum that enables undergraduate students to conduct complete microbiome studies using 16S rRNA gene sequencing. The module integrates project design, environmental sample collection, microbial cell processing, PCR amplification, sequencing, and bioinformatic analysis using open-source tools such as QIIME 2. Cost-reduction strategies included centrifugation-based cell collection and a surfactant-assisted direct PCR workflow that eliminated the need for commercial DNA extraction kits. Students designed independent research projects investigating microbial communities in local environments, including campus water sources and gym equipment surfaces. Assessment data from post-course surveys, knowledge checks, and student research products demonstrated strong learning gains in microbiome concepts, molecular biology techniques, scientific communication, and computational analysis. Students reported high confidence in PCR, experimental design, and microbiome interpretation, while also identifying bioinformatics as the most challenging yet rewarding component of the curriculum. All participants expressed increased interest in future research in microbiology or bioinformatics. Overall, this curriculum provides an accessible, scalable framework for integrating next-generation sequencing into undergraduate education while promoting inquiry-driven learning, student ownership, and engagement in authentic scientific research.

microbiology↗

bioRxiv2026

Mannosylated nanoparticle immunogens enhance the circumsporozoite protein-specific B cell response and improve protection against sporozoite challenge

Underprocessed oligomannose glycans on protein nanoparticle immunogens engage the innate immune system through mannose-binding lectin and complement, enhancing immunogen trafficking and B cell responses. However, the extent to which oligomannose glycans directly improve protective immunity has remained unclear. Here we generate a series of CSP-bearing I53-50 nanoparticle malaria vaccine candidates with defined numbers and types of engineered N-linked glycans and systematically evaluate their immunogenicity and protective efficacy. Oligomannose display enhanced early plasmablast and germinal center B cell responses, leading to increased CSP-specific memory B cells, long-lived plasma cells, and durable serum antibody titers. Furthermore, nanoparticles bearing oligomannose glycans conferred the strongest protection against sporozoite challenge. By comparing immunogens with defined glycoforms, we attribute improved immune responses and protection specifically to oligomannose rather than complex or truncated glycans. These results will help guide the development of general strategies for glycan engineering aimed at enhancing the protective efficacy of nanoparticle vaccines.

immunology↗