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Abstract Hemophilia A (HA) is caused by mutations in coagulation factor VIII (FVIII). Genome editing in conjunction with patient-derived induced pluripotent stem cells (iPSCs) is a promising cell therapy strategy, as it replaces dysfunctional proteins resulting from genetic mutations with normal proteins. However, the low expression level and short half-life of FVIII still remain significant limiting factors in the efficacy of these approaches in HA. Here, we constructed a functionally enhanced FVIII variant, F309S/E1984V-mutated B domain-deleted (BDD)-FVIII (FE-FVIII), with increased activity and stability. We inserted FE-FVIII with a human elongation factor-1 alpha (EF1α) promoter into the AAVS1 locus of HA patient-derived iPSCs via CRISPR/Cas9 (D10A) nickase to ensure expression in any cell type. FE-FVIII was expressed not only in undifferentiated FE-FVIII-inserted (FE-KI) iPSCs but also in endothelial cells (ECs) differentiated from them in vitro. Compared with mice transplanted with wild-type BDD-FVIII-containing ECs, immunocompetent HA mice intravenously transplanted with FE-KI ECs presented a 2.12-fold increase in FVIII activity in the blood and an approximately 20% greater survival rate after hemorrhagic tail injury. For sustained efficacy, FE-KI ECs were subcutaneously transplanted into immunodeficient HA mice, resulting in amelioration of the hemophilia phenotype for more than 3 months. This strategy can improve FVIII function and may provide a universal therapeutic approach for treating HA.

BackgroundArtificial intelligence (AI) social chatbots represent a major advancement in merging technology with mental health, offering benefits through natural and emotional communication. Unlike task-oriented chatbots, social chatbots build relationships and provide social support, which can positively impact mental health outcomes like loneliness and social anxiety. However, the specific effects and mechanisms through which these chatbots influence mental health remain underexplored. ObjectiveThis study explores the mental health potential of AI social chatbots, focusing on their impact on loneliness and social anxiety among university students. The study seeks to (i) assess the impact of engaging with an AI social chatbot in South Korea, "Luda Lee," on these mental health outcomes over a 4-week period and (ii) analyze user experiences to identify perceived strengths and weaknesses, as well as the applicability of social chatbots in therapeutic contexts. MethodsA single-group pre-post study was conducted with university students who interacted with the chatbot for 4 weeks. Measures included loneliness, social anxiety, and mood-related symptoms such as depression, assessed at baseline, week 2, and week 4. Quantitative measures were analyzed using analysis of variance and stepwise linear regression to identify the factors affecting change. Thematic analysis was used to analyze user experiences and assess the perceived benefits and challenges of chatbots. ResultsA total of 176 participants (88 males, average age=22.6 (SD 2.92)) took part in the study. Baseline measures indicated slightly elevated levels of loneliness (UCLA Loneliness Scale, mean 27.97, SD (11.07)) and social anxiety (Liebowitz Social Anxiety Scale, mean 25.3, SD (14.19)) compared to typical university students. Significant reductions were observed as loneliness decreasing by week 2 (t175=2.55, P=.02) and social anxiety decreasing by week 4 (t175=2.67, P=.01). Stepwise linear regression identified baseline loneliness (β=0.78, 95% CI 0.67 to 0.89), self-disclosure (β=–0.65, 95% CI –1.07 to –0.23) and resilience (β=0.07, 95% CI 0.01 to 0.13) as significant predictors of week 4 loneliness (R2=0.64). Baseline social anxiety (β=0.92, 95% CI 0.81 to 1.03) significantly predicted week 4 anxiety (R2=0.65). These findings indicate higher baseline loneliness, lower self-disclosure to the chatbot, and higher resilience significantly predicted higher loneliness at week 4. Additionally, higher baseline social anxiety significantly predicted higher social anxiety at week 4. Qualitative analysis highlighted the chatbot's empathy and support as features for reliability, though issues such as inconsistent responses and excessive enthusiasm occasionally disrupted user immersion. ConclusionsSocial chatbots may have the potential to mitigate feelings of loneliness and social anxiety, indicating their possible utility as complementary resources in mental health interventions. User insights emphasize the importance of empathy, accessibility, and structured conversations in achieving therapeutic goals. Trial RegistrationClinical Research Information Service (CRIS) KCT0009288; https://tinyurl.com/hxrznt3t

Fusarium basal rot (FBR), caused by the ascomycete fungus Fusarium oxysporum, is an economically important disease of onion worldwide. The most economical and effective way to manage FBR would be the use of FBR resistant onion cultivars. This study was carried out to develop a rapid screening method for resistant onion cultivars in seedling stage. We used the F. oxysporum 19-385 isolate, which causes damping-off in onion seedlings and basal rot in onion bulbs. We optimized broth incubation and medium composition for the production of inoculum, and determined conidial concentration for the preparation of F. oxysporum infected soil. Ten commercial cultivars of onion were evaluated the seedling survival rates and heights by infected soil inoculation methods. As a result, ‘K-force’ was the most resistant cultivar with 97.4% of relative seedling survival rate against the pathogen, whereas ‘Sunpower’ was the most susceptible cultivar with 20.0% of relative seedling survival rate.

In September 2023, a leaf spot disease occurred on chrysanthemum (Chrysanthemum morifolium) in Imsil-gun Agricultural Technology Center, Korea. A fungal species was isolated from the diseased plant and identified as an ascomycete Epicoccum sorghinum based on morphological characteristics and phylogenetic analyses using the sequences of the ITS, TUB2, and ACT genes. A pathogenicity test revealed that the isolate was the causal agent of leaf spot disease on chrysanthemum. This is the first report of leaf spot disease caused by E. sorghinum on chrysanthemum in Korea.

BackgroundChronic Insomnia Disorder (CID) is highly prevalent among older adults and impairs cognitive function. Insomnia accelerates the progression of mild cognitive impairment (MCI) and increases the risk of developing dementia. Acupuncture has been demonstrated in improving sleep quality and cognitive function. This study aims to explore the functional brain characteristics of CID with MCI patients and to assess the effects of acupuncture therapy using functional near-infrared spectroscopy (fNIRS).Methods and designThis study is a single-center randomized controlled trial. Participants will be randomly assigned to the manual acupuncture group or the placebo acupuncture group for an 8-week intervention period. fNIRS data will be collected during resting test and working memory test at baseline and at end of the intervention. The primary outcome is the change of the Montreal Cognitive Assessment (MoCA) score, secondary outcomes include the change of Mini-Mental State Examination (MMSE), Insomnia Severity Index (ISI), Patient Health Questionnaire-9 (PHQ-9), Generalized Anxiety Disorder Scale (GAD-7), and Apathy Evaluation Scale-Informant (AES-I).DiscussionThe results of the study will provide insights into the effects of acupuncture on sleep quality and cognitive performance in CID with MCI patients. By utilizing fNIRS technology, we will elucidate the neural functional characteristic underlying the therapeutic benefits of acupuncture.Clinical trial registrationhttps://ClinicalTrials.gov, identifier ChiCTR2300076182.

ABSTRACT Sexual reproduction is crucial for increasing the genetic diversity of populations and providing overwintering structures, such as perithecia and associated tissue, in the destructive plant pathogenic fungus Fusarium graminearum. While mating-type genes serve as master regulators in fungal sexual reproduction, the molecular mechanisms underlying this process remain elusive. Winged-helix DNA-binding proteins are key regulators of embryogenesis and cell differentiation in higher eukaryotes. These proteins are implicated in the morphogenesis and development of several fungal species. However, their involvement in sexual reproduction remains largely unexplored in F. graminearum. Here, we investigated the function of winged-helix DNA-binding proteins in vegetative growth, conidiation, and sexual reproduction, with a specific focus on the FgWING27, which is highly conserved among Fusarium species. Deletion of FgWING27 resulted in an abnormal pattern characterized by a gradual increase in the expression of mating-type genes during sexual development, indicating its crucial role in the stage-specific genetic regulation of MAT genes in the late stages of sexual development. Furthermore, using chromatin immunoprecipitation followed by sequencing analysis, we identified Fg17056 as a downstream gene of Fgwing27, which is essential for sexual reproduction. These findings underscore the significance of winged-helix DNA-binding proteins in fungal development and reproduction in F. graminearum, and highlight the pivotal role of Fgwing27 as a core genetic factor in the intricate genetic regulatory network governing sexual reproduction.IMPORTANCEFusarium graminearum is a devastating plant pathogenic fungus causing significant economic losses due to reduced crop yields. In Fusarium Head Blight epidemics, spores produced through sexual and asexual reproduction serve as inoculum, making it essential to understand the fungal reproduction process. Here, we focus on winged-helix DNA-binding proteins, which have been reported to play crucial roles in cell cycle regulation and differentiation, and address their requirement in the sexual reproduction of F. graminearum. Furthermore, we identified a highly conserved protein in Fusarium as a key factor in self-fertility, along with the discovery of its direct downstream genes. This provides crucial information for constructing the complex genetic regulatory network of sexual reproduction and significantly contribute to further research on sexual reproduction in Fusarium species.

Abstract Background Epilepsy, characterized by recurrent unprovoked seizures, poses significant challenges to affected individuals globally. While several established risk factors for epilepsy exist, the association with cigarette smoking remains debated. This study aims to conduct systematic review and meta-analysis to elucidate the potential association between smoking and the likelihood of epilepsy. Methods The search was performed on March 31st, 2023, using the Medline, Embase, Web of Science, Scopus, and ScienceDirect. We included cohort, cross-sectional, and case–control studies in our meta-analysis, conducting subgroup analyses based on smoking history, sex, and epilepsy type to yield specific insights. Results We identified 2550 studies, of which 17 studies were finally included in this study. The pooled odds ratio of epilepsy was 1.14 (0.96–1.36) in smokers compared to non-smokers. In current smokers compared to non-smokers, the odds ratio was 1.46 (1.13–1.89), while, in former smokers compared to non-smokers, the odds ratio was 1.14 (0.83–1.56). Conclusions While the overall association between smoking and epilepsy did not reach statistical significance, a notable association was found among current smokers. The study emphasizes the importance of smoking cessation as a potential preventive measure against epilepsy, especially given the proconvulsive effects of nicotine. Future research should address limitations and explore specific clinical scenarios to enhance our understanding of the complex relationship between cigarette use and epilepsy. Systematic review registration CRD42022342510.

AbstractFungi are cosmopolitan and they occupy diverse niches as consumers, producers, and decomposers. They play critical roles in the environment by enabling nutrient cycling and generating a plethora of secondary metabolites. This study aimed to identify and characterize fungal strains isolated from diverse sources on Muui Island, Republic of Korea. In 2023, a total of 86 fungal strains were collected and examined. Investigation of the morphological features and phylogenetic analyses of multiple barcode loci identified one putative novel species and two species previously unrecorded in the Republic of Korea: Colletotrichum sp., Colletotrichum guizhouense, and Fusarium brachygibbosum. This study provides a comprehensive description of their molecular phylogenies and morphological characteristics. These findings will contribute to the existing knowledge about fungal species in the Republic of Korea and future research on the fungal diversity on Muui Island.

ABSTRACTThe oxidative stress response is required for plant pathogens to endure host-derived oxidative stress during infection. Previously, we identified the eight transcription factors (TFs) involved in the oxidative stress response in the plant pathogenic fungus Fusarium graminearum and found that of these TFs, the deletion of FgbZIP007 caused hypersensitivity to oxidative stress. However, the underlying mechanisms of Fgbzip007 are not fully understood. Based on chromatin immunoprecipitation followed by sequencing (ChIP-seq) analysis, we found the regulons of Fgbzip007, and further genetic studies demonstrated that Fgbzip007 is a key regulator for sulfur assimilation. The deletion strains of FgbZIP007 and its regulons exhibited low level of glutathione biosynthesis, which led to characterize glutathione biosynthesis. Fgbzip007-mediated sulfur assimilation is required for glutathione biosynthesis, which is essential for oxidative stress resistance and pathogenicity in F. graminearum. Although the reduced resistance of glutathione-deficient mutants against oxidative stress was restored by overexpression of FCA7, encoding a core peroxidase, but not on pathogenicity, suggesting that glutathione in pathogenesis is independent of antioxidant properties. This study characterized the function of genes of glutathione biosynthesis, provides specific insight into how Fgbzip007 regulates pathogenesis in F. graminearum, and establishes a genetic framework for the molecular dissection of a TF Fgbzip007 with the integration of pathogen responses to oxidative stress.IMPORTANCEFusarium graminearum is a destructive fungal pathogen that causes Fusarium head blight (FHB) on a wide range of cereal crops. To control fungal diseases, it is essential to comprehend the pathogenic mechanisms that enable fungi to overcome host defenses during infection. Pathogens require an oxidative stress response to overcome host-derived oxidative stress. Here, we identify the underlying mechanisms of the Fgbzip007-mediated oxidative stress response in F. graminearum. ChIP-seq and subsequent genetic analyses revealed that the role of glutathione in pathogenesis is not dependent on antioxidant functions in F. graminearum. Altogether, this study establishes a comprehensive framework for the Fgbzip007 regulon on pathogenicity and oxidative stress responses, offering a new perspective on the role of glutathione in pathogenicity.

ABSTRACT In plant-pathogen interactions, oxidative bursts are crucial for plants to defend themselves against pathogen infections. Rapid production and accumulation of reactive oxygen species kill pathogens directly and cause local cell death, preventing pathogens from spreading to adjacent cells. Meanwhile, the pathogens have developed several mechanisms to tolerate oxidative stress and successfully colonize plant tissues. In this study, we investigated the mechanisms responsible for resistance to oxidative stress by analyzing the transcriptomes of six oxidative stress-sensitive strains of the plant pathogenic fungus Fusarium graminearum. Weighted gene co-expression network analysis identified several pathways related to oxidative stress responses, including the DNA repair system, autophagy, and ubiquitin-mediated proteolysis. We also identified hub genes with high intramodular connectivity in key modules and generated deletion or conditional suppression mutants. Phenotypic characterization of those mutants showed that the deletion of FgHGG4, FgHGG10, and FgHGG13 caused sensitivity to oxidative stress, and further investigation on those genes revealed that transcriptional elongation and DNA damage responses play roles in oxidative stress response and pathogenicity. The suppression of FgHGL7 also led to hypersensitivity to oxidative stress, and we demonstrated that FgHGL7 plays a crucial role in heme biosynthesis and is essential for peroxidase activity. This study increases the understanding of the adaptive mechanisms to cope with oxidative stress in plant pathogenic fungi. IMPORTANCE Fungal pathogens have evolved various mechanisms to overcome host-derived stresses for successful infection. Oxidative stress is a representative defense system induced by the host plant, and fungi have complex response systems to cope with it. Fusarium graminearum is one of the devastating plant pathogenic fungi, and understanding its pathosystem is crucial for disease control. In this study, we investigated adaptive mechanisms for coping with oxidative stress at the transcriptome level using oxidative stress-sensitive strains. In addition, by introducing genetic modification technique such as CRISPR-Cas9 and the conditional gene expression system, we identified pathways/genes required for resistance to oxidative stress and also for virulence. Overall, this study advances the understanding of the oxidative stress response and related mechanisms in plant pathogenic fungi.

RNA lariat debranching enzyme Dbr1 is required for intron turnover in the fungal plant pathogen Fusarium graminearum , and accumulation of lariat RNA affects its development and pathogenesis.

ABSTRACT Lipases, which catalyze the hydrolysis of long-chain triglycerides, diglycerides, and monoglycerides into free fatty acids and glycerol, participate in various biological pathways in fungi. In this study, we examined the biological functions and regulatory mechanisms of fungal lipases via two approaches. First, we performed a systemic functional characterization of 86 putative lipase-encoding genes in the plant-pathogenic fungus Fusarium graminearum. The phenotypes were assayed for vegetative growth, asexual and sexual reproduction, stress responses, pathogenicity, mycotoxin production, and lipase activity. Most mutants were normal in the assessed phenotypes, implying overlapping roles for lipases in F. graminearum. In particular, FgLip1 and Fgl1 were revealed as core extracellular lipases in F. graminearum. Second, we examined the lipase activity of previously constructed transcription factor (TF) mutants of F. graminearum and identified three TFs and one histone acetyltransferase that significantly affect lipase activity. The relative transcript levels of FgLIP1 and FGL1 were markedly reduced or enhanced in these TF mutants. Among them, Gzzc258 was identified as a key lipase regulator that is also involved in the induction of lipase activity during sexual reproduction. To our knowledge, this study is the first comprehensive functional analysis of fungal lipases and provides significant insights into the genetic and regulatory mechanisms underlying lipases in fungi. IMPORTANCE Fusarium graminearum is an economically important plant-pathogenic fungus that causes Fusarium head blight (FHB) on wheat and barley. Here, we constructed a gene knockout mutant library of 86 putative lipase-encoding genes and established a comprehensive phenotypic database of the mutants. Among them, we found that FgLip1 and Fgl1 act as core extracellular lipases in this pathogen. Moreover, several putative transcription factors (TFs) that regulate the lipase activities in F. graminearum were identified. The disruption mutants of F. graminearum-lipase regulatory TFs all showed defects in sexual reproduction, which implies a strong relationship between sexual development and lipase activity in this fungus. These findings provide valuable insights into the genetic mechanisms regulating lipase activity as well as its importance to the developmental stages of this plant-pathogenic fungus.