Multi-omics blood atlas reveals unique features of immune and platelet responses to SARS-CoV-2 Omicron breakthrough infection.

Although host responses to the ancestral SARS-CoV-2 strain are well described, those to the new Omicron variants are less resolved. We profiled the clinical phenomes, transcriptomes, proteomes, metabolomes, and immune repertoires of >1,000 blood cell or plasma specimens from SARS-CoV-2 Omicron patients. Using in-depth integrated multi-omics, we dissected the host response dynamics during multiple disease phases to reveal the molecular and cellular landscapes in the blood. Specifically, we detected enhanced interferon-mediated antiviral signatures of platelets in Omicron-infected patients, and platelets preferentially formed widespread aggregates with leukocytes to modulate immune cell functions. In addition, patients who were re-tested positive for viral RNA showed marked reductions in B cell receptor clones, antibody generation, and neutralizing capacity against Omicron. Finally, we developed a machine learning model that accurately predicted the probability of re-positivity in Omicron patients. Our study may inspire a paradigm shift in studying systemic diseases and emerging public health concerns.

Epitranscriptomic cytidine methylation of the hepatitis B viral RNA is essential for viral reverse transcription and particle production.

Epitranscriptomic RNA modifications have emerged as important regulators of the fate and function of viral RNAs. One prominent modification, the cytidine methylation 5-methylcytidine (m5C), is found on the RNA of HIV-1, where m5C enhances the translation of HIV-1 RNA. However, whether m5C functionally enhances the RNA of other pathogenic viruses remains elusive. Here, we surveyed a panel of commonly found RNA modifications on the RNA of hepatitis B virus (HBV) and found that HBV RNA is enriched with m5C as well as ten other modifications, at stoichiometries much higher than host messenger RNA (mRNA). Intriguingly, m5C is mostly found on the epsilon hairpin, an RNA element required for viral RNA encapsidation and reverse transcription, with these m5C mainly deposited by the cellular methyltransferase NSUN2. Loss of m5C from HBV RNA due to NSUN2 depletion resulted in a partial decrease in viral core protein (HBc) production, accompanied by a near-complete loss of the reverse transcribed viral DNA. Similarly, mutations introduced to remove the methylated cytidines resulted in a loss of HBc production and reverse transcription. Furthermore, pharmacological disruption of m5C deposition led to a significant decrease in HBV replication. Thus, our data indicate m5C methylations as a critical mediator of the epsilon elements' function in HBV virion production and reverse transcription, suggesting the therapeutic potential of targeting the m5C methyltransfer process on HBV epsilon as an antiviral strategy.

Rare and common variants in ROM1 and PRPH2 genes trans-modify Stargardt/ABCA4 disease.

Over 1,500 variants in the ABCA4 locus cause phenotypes ranging from severe, early-onset retinal degeneration to very late-onset maculopathies. The resulting ABCA4/Stargardt disease is the most prevalent Mendelian eye disorder, although its underlying clinical heterogeneity, including penetrance of many alleles, are not well-understood. We hypothesized that a share of this complexity is explained by trans-modifiers, i.e., variants in unlinked loci, which are currently unknown. We sought to identify these by performing exome sequencing in a large cohort for a rare disease of 622 cases and compared variation in seven genes known to clinically phenocopy ABCA4 disease to cohorts of ethnically matched controls. We identified a significant enrichment of variants in 2 out of the 7 genes. Moderately rare, likely functional, variants, at the minor allele frequency (MAF) <0.005 and CADD>25, were enriched in ROM1, where 1.3% of 622 patients harbored a ROM1 variant compared to 0.3% of 10,865 controls (p = 2.41E04; OR 3.81 95% CI [1.77; 8.22]). More importantly, analysis of common variants (MAF>0.1) identified a frequent haplotype in PRPH2, tagged by the p.Asp338 variant with MAF = 0.21 in the matched general population that was significantly increased in the patient cohort, MAF 0.25, p = 0.0014. Significant differences were also observed between ABCA4 disease subgroups. In the late-onset subgroup, defined by the hypomorphic p.Asn1868Ile variant and including c.4253+43G>A, the allele frequency for the PRPH2 p.Asp338 variant was 0.15 vs 0.27 in the remaining cohort, p = 0.00057. Known functional data allowed suggesting a mechanism by which the PRPH2 haplotype influences the ABCA4 disease penetrance. These associations were replicated in an independent cohort of 408 patients. The association was highly statistically significant in the combined cohorts of 1,030 cases, p = 4.00E-05 for all patients and p = 0.00014 for the hypomorph subgroup, suggesting a substantial trans-modifying role in ABCA4 disease for both rare and common variants in two unlinked loci.

Single Cell Analysis of Proteoforms.

We introduce single cell Proteoform imaging Mass Spectrometry (scPiMS), which realizes the benefit of direct solvent extraction and MS detection of intact proteins from single cells dropcast onto glass slides. Sampling and detection of whole proteoforms by individual ion mass spectrometry enable a scalable approach to single cell proteomics. This new scPiMS platform addresses the throughput bottleneck in single cell proteomics and boosts the cell processing rate by several fold while accessing protein composition with higher coverage.

Volatile Compounds Emitted by Plant Growth-Promoting Fungus Tolypocladium inflatum GT22 Alleviate Copper and Pathogen Stress.

Previous studies on the intricate interactions between plants and microorganisms have revealed that fungal volatile compounds (VCs) can affect plant growth and development. However, the precise mechanisms underlying these actions remain to be delineated. In this study, we discovered that VCs from the soilborne fungus Tolypocladium inflatum GT22 enhance the growth of Arabidopsis. Remarkably, priming Arabidopsis with GT22 VCs caused the plant to display an enhanced immune response and mitigated the detrimental effects of both pathogenic infections and copper stress. Transcriptomic analyses of Arabidopsis seedlings treated with GT22 VCs for 3, 24 and 48 h revealed that 90, 83 and 137 genes were differentially expressed, respectively. The responsive genes are known to be involved in growth, hormone regulation, defense mechanisms and signaling pathways. Furthermore, we observed the induction of genes related to innate immunity, hypoxia, salicylic acid biosynthesis and camalexin biosynthesis by GT22 VCs. Among the VCs emitted by GT22, exposure of Arabidopsis seedlings to limonene promoted plant growth and attenuated copper stress. Thus, limonene appears to be a key mediator of the interaction between GT22 and plants. Overall, our findings provide evidence that fungal VCs can promote plant growth and enhance both biotic and abiotic tolerance. As such, our study suggests that exposure of seedlings to T. inflatum GT22 VCs may be a means of improving crop productivity. This study describes a beneficial interaction between T. inflatun GT22 and Arabidopsis. Our investigation of microorganism function in terms of VC activities allowed us to overcome the limitations of traditional microbial application methods. The importance of this study lies in the discovery of T. inflatun GT22 as a beneficial microorganism. This soilborne fungus emits VCs with plant growth-promoting effects and the ability to alleviate both copper and pathogenic stress. Furthermore, our study offers a valuable approach to tracking the activities of fungal VC components via transcriptomic analysis and sheds light on the mechanisms through which VCs promote plant growth and induce resistance. This research significantly advances our knowledge of VC applications and provides an example for further investigations within this field.

Impaired cholesterol efflux in retinal pigment epithelium of individuals with juvenile macular degeneration.

Macular degeneration (MD) is characterized by the progressive deterioration of the macula and represents one of the most prevalent causes of blindness worldwide. Abnormal intracellular accumulation of lipid droplets and pericellular deposits of lipid-rich material in the retinal pigment epithelium (RPE) called drusen are clinical hallmarks of different forms of MD including Doyne honeycomb retinal dystrophy (DHRD) and age-related MD (AMD). However, the appropriate molecular therapeutic target underlying these disorder phenotypes remains elusive. Here, we address this knowledge gap by comparing the proteomic profiles of induced pluripotent stem cell (iPSC)-derived RPEs (iRPE) from individuals with DHRD and their isogenic controls. Our analysis and follow-up studies elucidated the mechanism of lipid accumulation in DHRD iRPE cells. Specifically, we detected significant downregulation of carboxylesterase 1 (CES1), an enzyme that converts cholesteryl ester to free cholesterol, an indispensable process in cholesterol export. CES1 knockdown or overexpression of EFEMP1R345W, a variant of EGF-containing fibulin extracellular matrix protein 1 that is associated with DHRD and attenuated cholesterol efflux and led to lipid droplet accumulation. In iRPE cells, we also found that EFEMP1R345W has a hyper-inhibitory effect on epidermal growth factor receptor (EGFR) signaling when compared to EFEMP1WT and may suppress CES1 expression via the downregulation of transcription factor SP1. Taken together, these results highlight the homeostatic role of cholesterol efflux in iRPE cells and identify CES1 as a mediator of cholesterol efflux in MD.

Cobalt Nitride Nanoparticles Encapsulated in N-Doped Carbon Nanotubes Modified Separator of Li-S Battery Achieving the Synergistic Effect of Restriction-Adsorption-Catalysis of Polysulfides.

Although lithium-sulfur (Li-S) batteries have broad market prospects due to their high theoretical energy density and potential cost-effectiveness, the practical applications still face serious shuttle effects of polysulfides (LiPSs) and slow redox reactions. Therefore, in this paper, cobalt nitride nanoparticles encapsulated in nitrogen-doped carbon nanotube (CoN@NCNT) are prepared as a functional layer for the separator of high-performance Li-S batteries. Carbon nanotubes with large specific surface areas not only promote the transport of ions and electrons but also weaken the migration of LiPSs and confine the dissolution of LiPSs in electrolytes. The lithiophilic heteroatom N adsorbs LiPSs by strong chemical adsorption, and the CoN particles with high catalytic activity greatly improve the kinetics of the conversion between LiPSs and Li2S2/Li2S during the charge-discharge process. Due to these advantages, the battery with CoN@NCNT modified separator has superior rate performance (initial discharge capacity of 834.7 mAh g-1 after activation at 1 C) and excellent cycle performance (capacity remains 729.7 mAh g-1 after 200 cycles at 0.2 C). This work proposes a strategy that can give the separator a strong ability to confinement-adsorption-catalysis of LiPSs in order to provide more possibilities for the development of Li-S batteries.

Function analysis and characterisation of a novel chitinase, MdCht9, in Musca domestica.

Insect chitinases have been proposed as potential targets for pest control. In this work, a novel group IV chitinase gene, MdCht9, from Musca domestica was found to have multiple functions in the physiological activity, including chitin regulation, development and antifungal immunity. The MdCht9 gene was cloned and sequenced, its phylogeny was analysed and its expression was determined in normal and 20E treated larvae. Subsequently, RNA interference (RNAi)-mediated MdCht9 knockdown was performed, followed by biochemical assays, morphological observations and transcriptome analysis. Finally, the recombinant protein MdCht9 (rMdCht9) was purified and tested for anti-microbial activity and enzyme characteristics. The results showed that MdCht9 consists of three domains, highly expressed in a larval salivary gland. RNAi silencing of MdCht9 resulted in significant down-regulation of chitin content and expression of 15 chitin-binding protein (CBP) genes, implying a new insight that MdCht9 might regulate chitin content by influencing the expression of CBPs. In addition, more than half of the lethality and partial wing deformity appeared due to the dsMdCht9 treatment. In addition, the rMdCht9 exhibited anti-microbial activity towards Candida albicans (fungus) but not towards Escherichia coli (G-) or Staphylococcus aureus (G+). Our work expands on previous studies of chitinase while providing a potential target for pest management.

Membranome-based identification of amino acid substitution in Haemophilus influenzae multidrug efflux pump HmrM for reduced chloramphenicol susceptibility.

A decreased chloramphenicol susceptibility in Haemophilus influenzae is commonly caused by the activity of chloramphenicol acetyltransferases (CATs). However, the involvement of membrane proteins in chloramphenicol susceptibility in H. influenzae remains unclear. In this study, chloramphenicol susceptibility testing, whole-genome sequencing, and analyses of membrane-related genes were performed in 51 H. influenzae isolates. Functional complementation assays and structure-based protein analyses were conducted to assess the effect of proteins with sequence substitutions on the minimum inhibitory concentration (MIC) of chloramphenicol in CAT-negative H. influenzae isolates. Six isolates were resistant to chloramphenicol and positive for type A-2 CATs. Of these isolates, A3256 had a similar level of CAT activity but a higher chloramphenicol MIC relative to the other resistant isolates; it also had 163 specific variations in 58 membrane genes. Regarding the CAT-negative isolates, logistic regression and receiver operator characteristic curve analyses revealed that 48T > G (Asn16Lys), 85 C > T (Leu29Phe), and 88 C > A (Leu30Ile) in HI_0898 (emrA), and 86T > G (Phe29Cys) and 141T > A (Ser47Arg) in HI_1177 (artM) were associated with enhanced chloramphenicol susceptibility, whereas 997G > A (Val333Ile) in HI_1612 (hmrM) was associated with reduced chloramphenicol susceptibility. Furthermore, the chloramphenicol MIC was lower in the CAT-negative isolates with EmrA-Leu29Phe/Leu30Ile or ArtM-Ser47Arg substitution and higher in those with HmrM-Val333Ile substitution, relative to their counterparts. The Val333Ile substitution was associated with enhanced HmrM protein stability and flexibility and increased chloramphenicol MICs in CAT-negative H. influenzae isolates. In conclusion, the substitution in H. influenzae multidrug efflux pump HmrM associated with reduced chloramphenicol susceptibility was characterised.

Antimicrobial efficacy and amino acid substitutions associated with susceptibility to the tellurium compound AS101 against Haemophilus influenzae and Haemophilus parainfluenzae.

The tellurite toxicity in Haemophilus influenzae and H. parainfluenzae remains unclear. To understand the potential of tellurite as a therapeutic option for these bacteria, we investigated the antimicrobial efficacy of AS101, a tellurium compound, against H. influenzae and H. parainfluenzae and the molecular basis of their differences in AS101 susceptibility. Through broth microdilution, we examined the minimum inhibitory concentration (MIC) of AS101 in 51 H. influenzae and 28 H. parainfluenzae isolates. Whole-genome sequencing was performed on the H. influenzae isolates to identify genetic variations associated with AS101 susceptibility. The MICs of AS101 were ≦ 4, 16-32, and ≧ 64 µg/mL in 9 (17.6%), 12 (23.5%), and 30 (58.8%) H. influenzae isolates, respectively, whereas ≦ 0.5 µg/mL in all H. parainfluenzae isolates, including multidrug-resistant isolates. Time-killing kinetic assay and scanning electron microscopy revealed the in vitro bactericidal activity of AS101 against H. parainfluenzae. Forty variations in nine tellurite resistance-related genes were associated with AS101 susceptibility. Logistic regression, receiver operator characteristic curve analysis, Venn diagram, and protein sequence alignment indicated that Val195Ile substitution in TerC, Ser93Gly in Gor (glutathione reductase), Pro44Ala/Ala50Pro in NapB (nitrate reductase), Val307Leu in TehA (tellurite resistance protein), Cys105Arg in CysK (cysteine synthase), and Thr364Ser in Csd (Cysteine desulfurase) were strongly associated with reduced AS101 susceptibility, whereas Ser155Pro in TehA with increased AS101 susceptibility. In conclusions, the antimicrobial efficacy of AS101 is high against H. parainfluenzae but low against H. influenzae. Genetic variations and corresponding protein changes relevant to AS101 non-susceptibility in H. influenzae were identified.

Robust inference for causal mediation analysis of recurrent event data.

Recurrent events, including cardiovascular events, are commonly observed in biomedical studies. Understanding the effects of various treatments on recurrent events and investigating the underlying mediation mechanisms by which treatments may reduce the frequency of recurrent events are crucial tasks for researchers. Although causal inference methods for recurrent event data have been proposed, they cannot be used to assess mediation. This study proposed a novel methodology of causal mediation analysis that accommodates recurrent outcomes of interest in a given individual. A formal definition of causal estimands (direct and indirect effects) within a counterfactual framework is given, and empirical expressions for these effects are identified. To estimate these effects, a semiparametric estimator with triple robustness against model misspecification was developed. The proposed methodology was demonstrated in a real-world application. The method was applied to measure the effects of two diabetes drugs on the recurrence of cardiovascular disease and to examine the mediating role of kidney function in this process.

Synthesis of a Triangle-Fused Six-Pointed Star and Its Electrocatalytic CO2 Reduction Activity.

As electrocatalysts, molecular catalysts with large aromatic systems (such as terpyridine, porphyrin, or phthalocyanine) have been widely applied in the CO2 reduction reaction (CO2RR). However, these monomeric catalysts tend to aggregate due to strong π-π interactions, resulting in limited accessibility of the active site. In light of these challenges, we present a novel strategy of active site isolation for enhancing the CO2RR. Six Ru(Tpy)2 were integrated into the skeleton of a metallo-organic supramolecule by stepwise self-assembly in order to form a rhombus-fused six-pointed star R1 with active site isolation. The turnover frequency (TOF) of R1 was as high as 10.73 s-1 at -0.6 V versus reversible hydrogen electrode (vs RHE), which is the best reported value so far at the same potential to our knowledge. Furthermore, by increasing the connector density on R1's skeleton, a more stable triangle-fused six-pointed star T1 was successfully synthesized. T1 exhibits exceptional stability up to 126 h at -0.4 V vs RHE and excellent TOF values of CO. The strategy of active site isolation and connector density increment significantly enhanced the catalytic activity by increasing the exposure of the active site. This work provides a starting point for the design of molecular catalysts and facilitates the development of a new generation of catalysts with a high catalytic performance.

Dynamic changes in quality of life in older patients with chronic obstructive pulmonary disease: a 7-year follow up.

PURPOSES: Chronic obstructive pulmonary disease (COPD) is a major cause of the rapid decline of health-related quality of life (HRQoL), associated with accelerated frailty in older populations. This study aimed to analyse the long-term dynamic changes of HRQoL and the predictive factors for the rapid decline of HRQoL in older patients with COPD. METHODS: Overall 244 patients with COPD, aged ≧ 65 years from one medical centre were enrolled between March 2012 and July 2020. Further, we prospectively assessed HRQoL scores with utility values, using EuroQol Five-Dimension (EQ-5D) questionnaires. Additionally, long-term dynamic changes in HRQoL were analysed using the Kernel smoothing method and examined the factors contributing to the deterioration of HRQoL using a linear mixed effects model. RESULTS: Older patients with COPD with forced expiration volume (FEV1) < 50% of prediction entered the phase of rapid and continuous decline of HRQoL ~ 2 years after enrolment, but patients with FEV1 ≥ 50% of prediction without rapidly declined HRQoL during 7 years follow up. Therefore, FEV1 < 50% of prediction is a novel predictor for the rapid decline of HRQoL. The course of rapidly declining HRQoL occurred, initially in the usual activities and pain/discomfort domains, followed by the morbidity, self-care, and depression/anxiety domains ~ 2 and 4 years after enrolment, respectively. The mixed effects model indicated that both FEV1 < 50% of prediction and a history of severe acute exacerbation (SAE) requiring hospitalisation were contributing factors for deterioration in HRQoL . CONCLUSIONS: Both FEV1 < 50% of prediction and exacerbations requiring hospitalisation were contributing factors for the deterioration of HRQoL in long-term follow up. Additionally, FEV1 < 50% of prediction was a novel predictor for patients entering the phase of rapid decline of HRQoL.

The multinomial mixed-effect regression model for predicting PCOC phases in hospice patients.

PURPOSE: The Palliative Care Outcomes Collaboration (PCOC) aims to enhance patient outcomes systematically. However, identifying crucial items and accurately determining PCOC phases remain challenging. This study aims to identify essential PCOC data items and construct a prediction model to accurately classify PCOC phases in terminal patients. METHODS: A retrospective cohort study assessed PCOC data items across four PCOC phases: stable, unstable, deteriorating, and terminal. From July 2020 to March 2023, terminal patients were enrolled. A multinomial mixed-effect regression model was used for the analysis of multivariate PCOC repeated measurement data. RESULTS: The dataset comprised 1933 terminally ill patients from 4 different hospice service settings. A total of 13,219 phases of care were analyzed. There were significant differences in the symptom assessment scale, palliative care problem severity score, Australia-modified Karnofsky performance status, and resource utilization groups-activities of daily living among the four PCOC phases of care. Clinical needs, including pain and other symptoms, declined from unstable to terminal phases, while psychological/spiritual and functional status for bed mobility, eating, and transfers increased. A robust prediction model achieved areas under the curves (AUCs) of 0.94, 0.94, 0.920, and 0.96 for stable, unstable, deteriorating, and terminal phases, respectively. CONCLUSIONS: Critical PCOC items distinguishing between PCOC phases were identified, enabling the development of an accurate prediction model. This model enhances hospice care quality by facilitating timely interventions and adjustments based on patients' PCOC phases.

Risk factors of chronic kidney disease in cisplatin-based hyperthermia intraperitoneal chemotherapy.

PURPOSE: Cisplatin is commonly prescribed in hyperthermic intraperitoneal chemotherapy (HIPEC) for peritoneal malignancy. Acute kidney injury (AKI) is regarded as a common complication after HIPEC combined with cytoreductive surgery (CRS). However, post-HIPEC chronic kidney disease (CKD) is scarce and less investigated. This study aims to investigate the incidence of CKD following cisplatin-based HIPEC and to analyse the associated risk factors. MATERIALS AND METHODS: From January 2016 to August 2021, a total of 55 patients treated with CRS and cisplatin-based HIPEC for peritoneal carcinomatosis were categorized retrospectively into groups, with and without CKD. Demographics, comorbidity, surgery, postoperative management, and complications were collected to evaluate risk factors for cisplatin-based HIPEC-related CKD. Univariate and multivariate analyses were conducted to confirm the correlation between different variables and CKD occurrence. RESULTS: Of the 55 patients, 24 (43.6%) patients developed AKI and 17 (70.8%) patients of these AKI patients progressed to CKD. Multivariate regression analysis identified intraoperative use of parecoxib (Odds Ratio (OR) = 4.39) and intraoperative maximum temperature > 38.5°C (OR = 6.40) as major risk factors for cisplatin-based HIPEC-related CKD occurrence. Though type II diabetes mellitus and intraoperative complications were the independent risk factors of AKI following cisplatin-based HIPEC, but they were not shown in CKD analysis. CONCLUSION: Intraoperative use of parecoxib during cisplatin-based HIPEC emerged as a significant risk factor for postoperative CKD. Clinicians should exercise caution in prescribing parecoxib during HIPEC procedures. Additionally, maintaining intraoperative body temperature below 38.5°C might be crucial to mitigate the risk of CKD development. This study underscores the importance of identifying and preventing specific risk factors to improve long-term renal outcomes in patients undergoing cisplatin-based HIPEC.

Implementation difficulties and solutions for a smart-clothes assisted home nursing care program for older adults with dementia or recovering from hip fracture.

BACKGROUND: Wearable devices have the advantage of always being with individuals, enabling easy detection of their movements. Smart clothing can provide feedback to family caregivers of older adults with disabilities who require in-home care. METHODS: This study describes the process of setting up a smart technology-assisted (STA) home-nursing care program, the difficulties encountered, and strategies applied to improve the program. The STA program utilized a smart-vest, designed specifically for older persons with dementia or recovering from hip-fracture surgery. The smart-vest facilitated nurses' and family caregivers' detection of a care receiver's movements via a remote-monitoring system. Movements included getting up at night, time spent in the bathroom, duration of daytime immobility, leaving the house, and daily activity. Twelve caregivers of older adults and their care receiver participated; care receivers included persons recovering from hip fracture (n = 5) and persons living with dementia (n = 7). Data about installation of the individual STA in-home systems, monitoring, and technical difficulties encountered were obtained from researchers' reports. Qualitative data about the caregivers' and care receivers' use of the system were obtained from homecare nurses' reports, which were explored with thematic analysis. RESULTS: Compiled reports from the research team identified three areas of difficulty with the system: incompatibility with the home environment, which caused extra hours of manpower and added to the cost of set-up and maintenance; interruptions in data transmissions, due to system malfunctions; and inaccuracies in data transmissions, due to sensors on the smart-vest. These difficulties contributed to frustration experienced by caregivers and care receivers. CONCLUSIONS: The difficulties encountered impeded implementation of the STA home nursing care. Each of these difficulties had their own unique problems and strategies to resolve them. Our findings can provide a reference for future implementation of similar smart-home systems, which could facilitate ease-of-use for family caregivers.

Kindergarten Visual-Perceptual and Motor Skills and Behavioral Traits Predict First-Grade Chinese Handwriting Legibility and Speed.

IMPORTANCE: Clarifying the relationship between kindergarteners' characteristics and their future handwriting performance is beneficial for the early detection of children at risk of handwriting difficulties. OBJECTIVE: To determine which visual-perceptual and motor skills and behavioral traits significantly predict kindergartners' Chinese handwriting legibility and speed in the first grade. DESIGN: One-year longitudinal, observational design. SETTING: Kindergarten and elementary schools. PARTICIPANTS: One hundred six kindergarten children (53 boys and 53 girls; ages 5 or 6 yr) were recruited. OUTCOMES AND MEASURES: The participants completed two subtests of the Bruininks-Oseretsky Test of Motor Proficiency-Second Edition, Test of Visual Perceptual Skills-Third Edition, Beery-Buktenica Developmental Test of Visual-Motor Integration (Beery™ VMI), and the Attention-Deficit/Hyperactivity Disorder Test-Chinese Version in kindergarten. Their handwriting legibility (character accuracy and construction) and speed were assessed by investigator-developed Chinese handwriting tests in the first grade. RESULTS: Multivariate regression analyses indicated the independent predictive power of spatial relationships (p = .042) and inattention (p = .004) for character accuracy. Visual-motor integration (VMI; p = .008) and inattention (p = .002) were the key predictors of character construction. Manual dexterity (p = .001) was the only significant predictor of writing speed. CONCLUSIONS AND RELEVANCE: Kindergarteners who perform poorly in spatial relationships, VMI, manual dexterity, and attention are likely to have less legible Chinese handwriting and slow writing speed in first grade. Plain-Language Summary: Children's visual-perceptual and motor skills and behavioral traits in kindergarten can predict their Chinese handwriting legibility and speed in first grade. This study found that kindergarteners who performed poorly in spatial relationships, VMI, manual dexterity, and attention were likely to have less legible Chinese handwriting and slow writing speed in the first grade.

The involvement of AtMKK1 and AtMKK3 in plant-deleterious microbial volatile compounds-induced defense responses.

KEY MESSAGE: Plant-deleterious microbial volatiles activate the transactivation of hypoxia, MAMPs and wound responsive genes in Arabidopsis thaliana. AtMKK1 and AtMKK3 are involved in the plant-deleterious microbial volatiles-induced defense responses. Microbial volatile compounds (mVCs) are a collection of volatile metabolites from microorganisms with biological effects on all living organisms. mVCs function as gaseous modulators of plant growth and plant health. In this study, the defense events induced by plant-deleterious mVCs were investigated. Enterobacter aerogenes VCs lead to growth inhibition and immune responses in Arabidopsis thaliana. E. aerogenes VCs negatively regulate auxin response and transport gene expression in the root tip, as evidenced by decreased expression of DR5::GFP, PIN3::PIN3-GFP and PIN4::PIN4-GFP. Data from transcriptional analysis suggests that E. aerogenes VCs trigger hypoxia response, innate immune responses and metabolic processes. In addition, the transcript levels of the genes involved in the synthetic pathways of antimicrobial metabolites camalexin and coumarin are increased after the E. aerogenes VCs exposure. Moreover, we demonstrate that MKK1 serves as a regulator of camalexin biosynthesis gene expression in response to E. aerogenes VCs, while MKK3 is the regulator of coumarin biosynthesis gene expression. Additionally, MKK1 and MKK3 mediate the E. aerogenes VCs-induced callose deposition. Collectively, these studies provide molecular insights into immune responses by plant-deleterious mVCs.

Uncovering molecular mechanisms involved in microbial volatile compounds-induced stomatal closure in Arabidopsis thaliana.

Microbial volatile compounds (mVCs) may cause stomatal closure to limit pathogen invasion as part of plant innate immune response. However, the mechanisms of mVC-induced stomatal closure remain unclear. In this study, we co-cultured Enterobacter aerogenes with Arabidopsis (Arabidopsis thaliana) seedlings without direct contact to initiate stomatal closure. Experiments using the reactive oxygen species (ROS)-sensitive fluorescent dye, H2DCF-DA, showed that mVCs from E. aerogenes enhanced ROS production in guard cells of wild-type plants. The involvement of ROS in stomatal closure was then demonstrated in an ROS production mutant (rbohD). In addition, we identified two stages of signal transduction during E. aerogenes VC-induced stomatal closure by comparing the response of wild-type Arabidopsis with a panel of mutants. In the early stage (3 h exposure), E. aerogenes VCs induced stomatal closure in wild-type and receptor-like kinase THESEUS1 mutant (the1-1) but not in rbohD, plant hormone-related mutants (nced3, erf4, jar1-1), or MAPK kinase mutants (mkk1 and mkk3). However, in the late stage (24 h exposure), E. aerogenes VCs induced stomatal closure in wild-type and rbohD but not in nced3, erf4, jar1-1, the1-1, mkk1 or mkk3. Taken together, our results suggest that E. aerogenes mVC-induced plant immune responses modulate stomatal closure in Arabidopsis by a multi-phase mechanism.

Cis-acting modifiers in the ABCA4 locus contribute to the penetrance of the major disease-causing variant in Stargardt disease.

Over 1200 variants in the ABCA4 gene cause a wide variety of retinal disease phenotypes, the best known of which is autosomal recessive Stargardt disease (STGD1). Disease-causing variation encompasses all mutation categories, from large copy number variants to very mild, hypomorphic missense variants. The most prevalent disease-causing ABCA4 variant, present in ~ 20% of cases of European descent, c.5882G > A p.(Gly1961Glu), has been a subject of controversy since its minor allele frequency (MAF) is as high as ~ 0.1 in certain populations, questioning its pathogenicity, especially in homozygous individuals. We sequenced the entire ~140Kb ABCA4 genomic locus in an extensive cohort of 644 bi-allelic, i.e. genetically confirmed, patients with ABCA4 disease and analyzed all variants in 140 compound heterozygous and 10 homozygous cases for the p.(Gly1961Glu) variant. A total of 23 patients in this cohort additionally harbored the deep intronic c.769-784C > T variant on the p.(Gly1961Glu) allele, which appears on a specific haplotype in ~ 15% of p.(Gly1961Glu) alleles. This haplotype was present in 5/7 of homozygous cases, where the p.(Gly1961Glu) was the only known pathogenic variant. Three cases had an exonic variant on the same allele with the p.(Gly1961Glu). Patients with the c.[769-784C > T;5882G > A] complex allele exhibit a more severe clinical phenotype, as seen in compound heterozygotes with some more frequent ABCA4 mutations, e.g. p.(Pro1380Leu). Our findings indicate that the c.769-784C > T variant is major cis-acting modifier of the p.(Gly1961Glu) allele. The absence of such additional allelic variation on most p.(Gly1961Glu) alleles largely explains the observed paucity of affected homozygotes in the population.