Agalactosyl IgG induces liver fibrogenesis via Fc gamma receptor 3a on human hepatic stellate cells.

The relevance of aberrant serum IgG N-glycosylation in liver fibrosis has been identified; however, its causal effect remains unclear. Because hepatic stellate cells (HSCs) contribute substantially to liver fibrosis, we investigated whether and through which mechanisms IgG N-glycosylation affects the fibrogenic properties of HSCs. Analysis of serum IgG1 N-glycome from 151 patients with chronic hepatitis B or liver cirrhosis revealed a positive correlation between Ishak fibrosis grading and IgG1 with agalactosyl N-glycoforms on the crystallizable fragment (Fc). Fc gamma receptor (FcγR) IIIa was observed in cultured human HSCs and HSCs in human liver tissues, and levels of FcγRIIIa in HSCs correlated with the severity of liver fibrosis. Additionally, agalactosyl IgG treatment caused HSCs to have a fibroblast-like morphology, enhanced migration and invasion capabilities, and enhanced expression of the FcγRIIIa downstream tyrosine-protein kinase SYK. Furthermore, agalactosyl IgG treatment increased fibrogenic factors in HSCs, including transforming growth factor (TGF)-ß1, total collagen, platelet-derived growth factor subunit B and its receptors, pro-collagen I-α1, α-smooth muscle actin, and matrix metalloproteinase 9. These effects were more pronounced in HSCs that stably expressed FCGR3A and were reduced in FCGR3A knockout cells. Agalactosyl IgG and TGF-ß1 each increased FCGR3A in HSCs. Furthermore, serum TGF-ß1 concentrations in patients were positively correlated with agalactosyl IgG1 levels and liver fibrosis severity, indicating a positive feedback loop involving agalactosyl IgG, HSC-FcγRIIIa, and TGF-ß1. In conclusion, agalactosyl IgG promotes fibrogenic characteristics in HSCs through FcγRIIIa. © 2024 The Pathological Society of Great Britain and Ireland.

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.

CHL1 functions as a nitrate sensor in plants.

Ions serve as essential nutrients in higher plants and can also act as signaling molecules. Little is known about how plants sense changes in soil nutrient concentrations. Previous studies showed that T101-phosphorylated CHL1 is a high-affinity nitrate transporter, whereas T101-dephosphorylated CHL1 is a low-affinity transporter. In this study, analysis of an uptake- and sensing-decoupled mutant showed that the nitrate transporter CHL1 functions as a nitrate sensor. Primary nitrate responses in CHL1T101D and CHLT101A transgenic plants showed that phosphorylated and dephosphorylated CHL1 lead to a low- and high-level response, respectively. In vitro and in vivo studies showed that, in response to low nitrate concentrations, protein kinase CIPK23 can phosphorylate T101 of CHL1 to maintain a low-level primary response. Thus, CHL1 uses dual-affinity binding and a phosphorylation switch to sense a wide range of nitrate concentrations in the soil, thereby functioning as an ion sensor in higher plants. For a video summary of this article, see the PaperFlick file with the Supplemental Data available online.

Molecular characterization of cefepime and aztreonam nonsusceptibility in Haemophilus influenzae.

BACKGROUND: Cefepime and aztreonam are highly efficacious against H. influenzae, and resistant strains are rare. In this study, we isolated cefepime- and aztreonam-nonsusceptible H. influenzae strains and addressed the molecular basis of their resistance to cefepime and aztreonam. METHODS: Two hundred and 28 specimens containing H. influenzae were screened, of which 32 isolates were enrolled and applied to antimicrobial susceptibility testing and whole-genome sequencing. Genetic variations that were detected in all nonsusceptible isolates with statistical significance by Fisher's exact tests were identified as cefepime or aztreonam nonsusceptibility related. Functional complementation assays were conducted to assess the in vitro effects of proteins with sequence substitutions on drug susceptibility. RESULTS: Three H. influenzae isolates were nonsusceptible to cefepime, one of which was also nonsusceptible to aztreonam. Genes encoding TEM, SHV and CTX-M extended-spectrum ß-lactamases were not detected in the cefepime- and aztreonam-nonsusceptible isolates. Five genetic variations in four genes and 10 genetic variations in five genes were associated with cefepime and aztreonam nonsusceptibility, respectively. Phylogenetic analyses revealed that changes in FtsI were correlated strongly with the MIC of cefepime and moderately with aztreonam. FtsI Thr532Ser-Tyr557His cosubstitution linked to cefepime nonsusceptibility and Asn305Lys-Ser385Asn-Glu416Asp cosubstitution to aztreonam nonsusceptibility. Functional complementation assays revealed that these cosubstitutions increased MICs of cefepime and aztreonam in susceptible H. influenzae isolates, respectively. CONCLUSIONS: Genetic variations relevant to resistant phenotypes of cefepime and aztreonam nonsusceptibility in H. influenzae were identified. Moreover, the effects of FtsI cosubstitutions on increasing MICs of cefepime and aztreonam in H. influenzae were demonstrated.

Molecular characterization of multidrug-resistant non-typeable Haemophilus influenzae with high-level resistance to cefuroxime, levofloxacin, and trimethoprim-sulfamethoxazole.

BACKGROUND: Non-typeable Haemophilus influenzae (NTHi) has become the major cause of invasive H. influenzae diseases in the post-H. influenzae type b vaccine era. The emergence of multidrug-resistant (MDR) NTHi is a growing public health problem. Herein, we investigated the molecular basis of MDR in NTHi. The isolated NTHi were subjected to antimicrobial susceptibility testing for 12 agents. Whole genome and plasmid sequencing were conducted and analyzed to identify significant genetic variations and plasmid-encoded genes conferred antibiotic resistance. RESULTS: Thirteen (50%) MDR NTHi isolates were obtained; of these, 92.3% were non-susceptible to ampicillin, 30.8% to amoxicillin-clavulanate, 61.5% to cefuroxime, 61.5% to ciprofloxacin/levofloxacin, 92.3% to trimethoprim-sulfamethoxazole, 30.8% to tetracycline, and 7.7% to azithromycin. Eight ampicillin-resistant isolates were ß-lactamase positive; of these, 6 carried blaTEM-1 and 2 carried blaROB-1, whereas 4 were ß-lactamase negative. Genetic variations in mrdA, mepA, and pbpG were correlated with amoxicillin-clavulanate non-susceptibility, whereas variations in ftsI and lpoA conferred cefuroxime resistance. Five variations in gyrA, 2 in gyrB, 3 in parC, 1 in parE, and 1 in the parC-parE intergenic region were associated with levofloxacin/ciprofloxacin non-susceptibility. Among these genes, 8 variations were linked to high-level levofloxacin resistance. Six variations in folA were associated with trimethoprim-sulfamethoxazole resistance. Plasmid-bearing tet(B) and mef(A) genes were responsible for tetracycline and azithromycin resistance in 4 and 1 MDR isolates, respectively. CONCLUSIONS: This study clarified the molecular epidemiology of MDR in NTHi. This can benefit the monitoring of drug resistance trends in NTHi and the adequate medical management of patients with NTHi infection.

Amino acid substitutions in the region between RpoB clusters II and III on rifampin susceptibility in Haemophilus influenzae.

BACKGROUND: Rifampin is a potent chemoprophylactic antibiotic for Haemophilus influenzae infection, and the resistance rate in H. influenzae is low. In this study, we assessed rifampin resistance-related genetic variations in H. influenzae. METHODS: Rifampin susceptibility testing and whole-genome sequencing were performed in 51 H. influenzae isolates. Variations associated with rifampin resistance were identified using Fisher's exact tests. Functional assays were performed to evaluate the effect of RpoB substitutions on rifampin susceptibility. RESULTS: Using the genome of the Rd KW20 H. influenzae strain as the reference, we detected 40 genetic variations in rpoB, which resulted in 39 deduced amino acid substitutions among the isolates. Isolate A0586 was resistant to rifampin, with a minimum inhibitory concentration (MIC) = 8 µg/mL. Phylogenetic analyses revealed that the RpoB sequence of isolate A0586 was distinct from other isolates. Five substitutions, including H526N located in cluster I and L623F, R628C, L645F, and L672F in the region between clusters II and III, were unique to isolate A0586. In two rifampin-susceptible H. influenzae isolates, RpoB-H526N alone and in combination with RpoB-L672F increased the MICs of rifampin to 4 and 8 µg/mL, respectively. RpoB-L672F did not affect cell growth and transcription in H. influenzae isolates. No amino acid substitutions in the AcrAB-TolC efflux pump or outer membrane proteins were found to be associated with rifampin resistance in H. influenzae. CONCLUSIONS: Our findings indicate that L672F substitution in the region between RpoB clusters II and III has an aggravating effect on rifampin resistance in H. influenzae.

Designs, applications, and limitations of genetically encoded fluorescent sensors to explore plant biology.

The understanding of signaling and metabolic processes in multicellular organisms requires knowledge of the spatial dynamics of small molecules and the activities of enzymes, transporters, and other proteins in vivo, as well as biophysical parameters inside cells and across tissues. The cellular distribution of receptors, ligands, and activation state must be integrated with information about the cellular distribution of metabolites in relation to metabolic fluxes and signaling dynamics in order to achieve the promise of in vivo biochemistry. Genetically encoded sensors are engineered fluorescent proteins that have been developed for a wide range of small molecules, such as ions and metabolites, or to report biophysical processes, such as transmembrane voltage or tension. First steps have been taken to monitor the activity of transporters in vivo. Advancements in imaging technologies and specimen handling and stimulation have enabled researchers in plant sciences to implement sensor technologies in intact plants. Here, we provide a brief history of the development of genetically encoded sensors and an overview of the types of sensors available for quantifying and visualizing ion and metabolite distribution and dynamics. We further discuss the pros and cons of specific sensor designs, imaging systems, and sample manipulations, provide advice on the choice of technology, and give an outlook into future developments.

Concept of Fluorescent Transport Activity Biosensor for the Characterization of the Arabidopsis NPF1.3 Activity of Nitrate.

The NRT1/PTR FAMILY (NPF) in Arabidopsis (Arabidopsis thaliana) plays a major role as a nitrate transporter. The first nitrate transporter activity biosensor NiTrac1 converted the dual-affinity nitrate transceptor NPF6.3 into fluorescence activity sensors. To test whether this approach is transferable to other members of this family, screening for genetically encoded fluorescence transport activity sensor was performed with the member of the NPF family in Arabidopsis. In this study, NPF1.3, an uncharacterized member of NPF in Arabidopsis, was converted into a transporter activity biosensor NiTrac-NPF1.3 that responds specifically to nitrate. The emission ratio change of NiTrac-NPF1.3 triggered by the addition of nitrate reveals the important function of NPF1.3 in nitrate transport in Arabidopsis. A functional analysis of Xenopus laevis oocytes confirmed that NPF1.3 plays a role as a nitrate transporter. This new technology is applicable in plant and medical research.

Feedback inhibition of AMT1 NH4+-transporters mediated by CIPK15 kinase.

BACKGROUND: Ammonium (NH4+), a key nitrogen form, becomes toxic when it accumulates to high levels. Ammonium transporters (AMTs) are the key transporters responsible for NH4+ uptake. AMT activity is under allosteric feedback control, mediated by phosphorylation of a threonine in the cytosolic C-terminus (CCT). However, the kinases responsible for the NH4+-triggered phosphorylation remain unknown. RESULTS: In this study, a functional screen identified protein kinase CBL-Interacting Protein Kinase15 (CIPK15) as a negative regulator of AMT1;1 activity. CIPK15 was able to interact with several AMT1 paralogs at the plasma membrane. Analysis of AmTryoshka, an NH4+ transporter activity sensor for AMT1;3 in yeast, and a two-electrode-voltage-clamp (TEVC) of AMT1;1 in Xenopus oocytes showed that CIPK15 inhibits AMT activity. CIPK15 transcript levels increased when seedlings were exposed to elevated NH4+ levels. Notably, cipk15 knockout mutants showed higher 15NH4+ uptake and accumulated higher amounts of NH4+ compared to the wild-type. Consistently, cipk15 was hypersensitive to both NH4+ and methylammonium but not nitrate (NO3-). CONCLUSION: Taken together, our data indicate that feedback inhibition of AMT1 activity is mediated by the protein kinase CIPK15 via phosphorylation of residues in the CCT to reduce NH4+-accumulation.

Extensively drug-resistant Haemophilus influenzae - emergence, epidemiology, risk factors, and regimen.

BACKGROUND: Concern about Haemophilus influenzae infection has been increasing over recent decades. Given the emergence of H. influenzae with severe drug resistance, we assessed the prevalence of as well as risk factors and potential therapies for extensively drug-resistant (XDR) H. influenzae infection in Taiwan. RESULTS: In total, 2091 H. influenzae isolates with disk diffusion-based antibiotic susceptibility testing from 2007 to 2018 were enrolled. H. influenzae strains resistant to ampicillin, chloramphenicol, levofloxacin, and trimethoprim-sulfamethoxazole tended to be isolated from patient wards (≧41%), whereas those resistant to amoxicillin-clavulanate, cefotaxime, and cefuroxime were more likely to be isolated from intensive care units (approximately 50%). XDR H. influenzae was first identified in 2007, and its incidence did not significantly change thereafter. Overall prevalence of single, multiple, and extensively drug-resistant H. influenzae over 2007-2018 was 21.5% (n = 450), 26.6% (n = 557), and 2.5% (n = 52), respectively. A stepwise logistic regression analysis revealed that blood culture (odds ratio: 4.069, 95% confidence intervals: 1.339-12.365, P = 0.013) was an independent risk factor for XDR H. influenzae infection. No nosocomial transmission of XDR H. influenzae observed. Antibiotic susceptibility testing results demonstrated that cefotaxime was effective against 78.8% (n = 41) of the XDR strains. CONCLUSIONS: The presence of XDR H. influenzae strains was identified in Taiwan, and cefotaxime was efficacious against most of these strains.

Hepatocellular carcinoma-associated single-nucleotide variants and deletions identified by the use of genome-wide high-throughput analysis of hepatitis B virus.

This study investigated hepatitis B virus (HBV) single-nucleotide variants (SNVs) and deletion mutations linked with hepatocellular carcinoma (HCC). Ninety-three HCC patients and 108 non-HCC patients were enrolled for HBV genome-wide next-generation sequencing (NGS) analysis. A systematic literature review and a meta-analysis were performed to validate NGS-defined HCC-associated SNVs and deletions. The experimental results identified 60 NGS-defined HCC-associated SNVs, including 41 novel SNVs, and their pathogenic frequencies. Each SNV was specific for either genotype B (n = 24) or genotype C (n = 34), except for nt53C, which was present in both genotypes. The pathogenic frequencies of these HCC-associated SNVs showed a distinct U-shaped distribution pattern. According to the meta-analysis and literature review, 167 HBV variants from 109 publications were categorized into four levels (A-D) of supporting evidence that they are associated with HCC. The proportion of NGS-defined HCC-associated SNVs among these HBV variants declined significantly from 75% of 12 HCC-associated variants by meta-analysis (Level A) to 0% of 10 HCC-unassociated variants by meta-analysis (Level D) (P < 0.0001). PreS deletions were significantly associated with HCC, in terms of deletion index, for both genotypes B (P = 0.030) and C (P = 0.049). For genotype C, preS deletions involving a specific fragment (nt2977-3013) were significantly associated with HCC (HCC versus non-HCC, 6/34 versus 0/32, P = 0.025). Meta-analysis of preS deletions showed significant association with HCC (summary odds ratio 3.0; 95% confidence interval 2.3-3.9). Transfection of Huh7 cells showed that all of the five novel NGS-defined HCC-associated SNVs in the small surface region influenced hepatocarcinogenesis pathways, including endoplasmic reticulum-stress and DNA repair systems, as shown by microarray, real-time polymerase chain reaction and western blot analysis. Their carcinogenic mechanisms are worthy of further research. Copyright © 2017 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Organisation-based self-esteem mediates the effects of social support and job satisfaction on intention to stay in nurses.

AIM: This study examined the effect of underlying contextual factors on the intention to stay in nurses. BACKGROUND: Workplace promotion, social support, work stress, job satisfaction and organisation-based self-esteem (OBSE) are psychosocial factors influencing the intention to stay in nurses. However, few studies have analysed the relationships among these factors. METHOD: A cross-sectional survey was conducted and a sample was recruited in convenience sampling method from a medical centre in Taiwan. A total of 791 nurses completed a self-report questionnaire over a 3-month period in 2013. RESULTS: Social support, work stress, job satisfaction and OBSE significantly affected the intention to stay in nurses. Furthermore, social support and job satisfaction showed a positive direct effect on the intention to stay and an indirect effect on the intention to stay; the indirect effect was mediated by OBSE. CONCLUSION: Organisation-based self-esteem mediates the effects of social support and job satisfaction on the intention to stay in nurses. IMPLICATIONS FOR NURSING MANAGEMENT: The results showing the influence of OBSE on the intention to stay in nurses can serve as insight for hospital managers to make decisions when encouraging and managing employees.

Aberrant serum immunoglobulin G glycosylation in chronic hepatitis B is associated with histological liver damage and reversible by antiviral therapy.

BACKGROUND: Aberrant serum immunoglobulin G (IgG) glycosylation and its immunomodulatory effect are rarely addressed in chronic hepatitis B virus (HBV) infection. METHODS: Serum IgG-Fc glycosylation profiles in 76 patients with HBV-related liver cirrhosis and 115 patients with chronic hepatitis B (CHB) before and after 48 weeks of anti-HBV nucleos(t)ide analogue treatment were analyzed using high-throughput liquid chromatography-mass spectrometry and were compared to profiles in 108 healthy controls. RESULTS: The level of aberrant serum IgG-Fc glycosylation ,: particularly galactose deficiency, was higher in patients with CHB and those with cirrhosis (P < .001 for both) than in healthy controls. IgG galactose deficiency was correlated with the severity of liver necroinflammation and fibrosis in CHB. Multivariate logistic regression analyses showed that the IgG-Fc glycoform with fucosylation and fully galactosylation was an independent factor for a total Knodell necroinflammation score of ≥ 7 (odds ratio, 0.74; 95% confidence interval, .56-.97) and an Ishak fibrosis score of ≥ 3 (odds ratio, 0.69; 95% confidence interval, .49-.97). Administration of antiviral therapy for 48 weeks reversed aberrant IgG-Fc glycosylation in patients with CHB from week 12 onward but did not reverse glycosylation in patients with cirrhosis. Attenuated IgG opsonization in patients with CHB, which was correlated with aberrant Fc-glycosylation, was reversed after treatment as well. CONCLUSIONS: Aberrant serum IgG-Fc glycosylation in CHB, which is highly associated with histological liver damage, affects IgG opsonizing activity and can be reversed by antiviral therapy.

Construct validity and reliability of the Chinese version of the Disaster Preparedness Evaluation Tool in Taiwan.

AIMS AND OBJECTIVES: To develop a Chinese version of the Disaster Preparedness Evaluation Tool and validate its psychometric properties. BACKGROUND: An adequate disaster preparation programme for nurses has not been developed in Taiwan. To develop an exhaustive and effective educational programme on disaster preparation for nurses, a multidimensional instrument is required for assessing the disaster preparation level of nurses. DESIGN: A cross-sectional study was conducted. METHODS: In total, 1550 of 2226 public health nurses in 15 counties completed the self-administrated questionnaire. We randomly selected 805 samples to examine the factor structure and factor model by using exploratory factor analysis and confirmatory factor analysis. The convergent validity was measured using the average variance extracted and composite reliability. RESULTS: Five factors, namely postdisaster management, skills, knowledge of self-preparation in a disaster, knowledge to respond in the community, and knowledge to respond in the workplace, were extracted, and explained 65·13% of the total variance. An acceptable model fit was identified using confirmatory factor analysis. The Cronbach's α coefficient of the Chinese version of the Disaster Preparedness Evaluation Tool was 0·97. Significant values of the average variance extracted greater than 0·5 indicated convergent validity. CONCLUSION: The Chinese version of the Disaster Preparedness Evaluation Tool is a reliable and valid instrument for measuring disaster preparation. RELEVANCE TO CLINICAL PRACTICE: The Chinese version of the Disaster Preparedness Evaluation Tool provides reliable and valid measures that can be used to evaluate the disaster preparedness of nurses. The items in the instrument can be used to identify the dimension of disaster management in all stages, and can form the essential foundation of an education and training programme for public health nurses to reduce the harm of disasters and promote community resilience.

Fluorescent Biosensor Imaging of Nitrate in Arabidopsis thaliana.

Nitrate (NO3-) is an essential element and nutrient for plants and animals. Despite extensive studies on the regulation of nitrate uptake and downstream responses in various cells, our knowledge of the distribution of nitrogen forms in different root cell types and their cellular compartments is still limited. Previous physiological models have relied on in vitro biochemistry and metabolite level analysis, which limits the ability to differentiate between cell types and compartments. Here, to address this, we report a nuclear-localized, genetically encoded fluorescent biosensor, which we named nlsNitraMeter3.0, for the quantitative visualization of nitrate concentration and distribution at the cellular level in Arabidopsis thaliana. This biosensor was specifically designed for nitrate measurements, not nitrite. Through genetic engineering to create and select sensors using yeast, Xenopus oocyte, and Arabidopsis expression systems, we developed a reversible and highly specific nitrate sensor. This method, combined with fluorescence imaging systems such as confocal microscopy, allows for the understanding and monitoring of nitrate transporter activity in plant root cells in a minimally invasive manner. Furthermore, this approach enables the functional analysis of nitrate transporters and the measurement of nitrate distribution in plants, providing a valuable tool for plant biology research. In summary, we provide a protocol for sensor development and a biosensor that can be used to monitor nitrate levels in plants. Key features This protocol builds upon the concept of FRET biosensors for in vivo visualization of spatiotemporal nitrate levels at a cellular resolution. Nitrate levels can be quantified utilizing the biosensor in conjunction with either a plate reader or a fluorescence microscope.

CIPK15-mediated inhibition of NH4+ transport protects Arabidopsis from submergence.

Ammonium (NH4+) serves as a vital nitrogen source for plants, but it can turn toxic when it accumulates in excessive amounts. Toxicity is aggravated under hypoxic/anaerobic conditions, e.g., during flooding or submergence, due to a lower assimilation capacity. AMT1; 1 mediates NH4+ uptake into roots. Under conditions of oxygen-deficiency, i.e., submergence, the CBL-interacting protein kinase OsCIPK15 has been shown to trigger SnRK1A signaling, promoting starch mobilization, thereby the increasing availability of ATP, reduction equivalents and acceptors for NH4+ assimilation in rice. Our previous study in Arabidopsis demonstrates that AtCIPK15 phosphorylates AMT1; 1 whose activity is under allosteric feedback control by phosphorylation of T460 in the cytosolic C-terminus. Here we show that submergence cause higher NH4+ accumulation in wild-type, plant but not of nitrate, nor in a quadruple amt knock-out mutant. In addition, submergence triggers rapid accumulation of AtAMT1;1 and AtCIPK15 transcripts as well as AMT1 phosphorylation. Significantly, cipk15 knock-out mutants do not exhibit an increase in AMT1 phosphorylation; however, they do display heightened sensitivity to submergence. These findings suggest that CIPK15 suppresses AMT activity, resulting in decreased NH4+ accumulation during submergence, a period when NH4+ assimilation capacity may be impaired.

A Novel Phytogenic Formulation, EUBIO-BPSG, as a Promising One Health Approach to Replace Antibiotics and Promote Reproduction Performance in Laying Hens.

Gut microbiota play a key role in health maintenance and disease pathogenesis in animals. Dietary phytochemicals are crucial factors shaping gut bacteria. Here, we investigated the function and mechanism of a phytogenic formulation, EUBIO-BPSG (BP), in laying hens. We found that BP dose-dependently improved health and egg production in 54-week-old hens. Furthermore, BP was correlated with increased fecal Lactobacillus, decreased Escherichia coli and Salmonella enterica, and reduced antibiotic resistance (AR) and antibiotic resistance genes (ARG) in chicken stools. The 16S rDNA data showed that BP increased seven genera of probiotics and reduced 13 genera of pathogens in chicken feces. In vitro co-culture experiments showed that BP at 4 µg/mL and above promoted growth of L. reuteri while large 100- and 200-fold higher doses suppressed growth of E. coli and S. enterica, respectively. Mechanistic studies indicated that L. reuteri and its supernatants antagonized growth of E. coli and S. enterica but not vice-versa. Five short-chain fatty acids and derivatives (SCFA) produced from L. reuteri directly killed both pathogens via membrane destruction. Furthermore, BP inhibited conjugation and recombination of ARG via interference with conjugation machinery and integrase activity in E. coli. Collectively, this work suggests that BP promotes host health and reproductive performance in laying hens through regulation of gut microbiota through increasing probiotics and decreasing pathogens and spreading ARG.

Factors influencing consumer adoption of USB-based Personal Health Records in Taiwan.

BACKGROUND: Usually patients receive healthcare services from multiple hospitals, and consequently their healthcare data are dispersed over many facilities' paper and electronic-based record systems. Therefore, many countries have encouraged the research on data interoperability, access, and patient authorization. This study is an important part of a national project to build an information exchange environment for cross-hospital digital medical records carried out by the Department of Health (DOH) of Taiwan in May 2008. The key objective of the core project is to set up a portable data exchange environment in order to enable people to maintain and own their essential health information.This study is aimed at exploring the factors influencing behavior and adoption of USB-based Personal Health Records (PHR) in Taiwan. METHODS: Quota sampling was used, and structured questionnaires were distributed to the outpatient department at ten medical centers which participated in the DOH project to establish the information exchange environment across hospitals. A total of 3000 questionnaires were distributed and 1549 responses were collected, out of those 1465 were valid, accumulating the response rate to 48.83%. RESULTS: 1025 out of 1465 respondents had expressed their willingness to apply for the USB-PHR. Detailed analysis of the data reflected that there was a remarkable difference in the "usage intention" between the PHR adopters and non-adopters (χ2 =182.4, p < 0.001). From the result of multivariate logistic regression analyses, we found the key factors affecting patients' adoption pattern were Usage Intention (OR, 9.43, 95%C.I., 5.87-15.16), Perceived Usefulness (OR, 1.60; 95%C.I., 1.11-2.29) and Subjective Norm (OR, 1.47; 95%C.I., 1.21-1.78). CONCLUSIONS: Higher Usage Intentions, Perceived Usefulness and Subjective Norm of patients were found to be the key factors influencing PHR adoption. Thus, we suggest that government and hospitals should promote the potential usefulness of PHR, and physicians should encourage patients' to adopt the PHR.

In vivo visualization of nitrate dynamics using a genetically encoded fluorescent biosensor.

Nitrate (NO3-) uptake and distribution are critical to plant life. Although the upstream regulation of NO3- uptake and downstream responses to NO3- in a variety of cells have been well studied, it is still not possible to directly visualize the spatial and temporal distribution of NO3- with high resolution at the cellular level. Here, we report a nuclear-localized, genetically encoded fluorescent biosensor, which we named NitraMeter3.0, for the quantitative visualization of NO3- distribution in Arabidopsis thaliana. This biosensor tracked the spatiotemporal distribution of NO3- along the primary root axis and disruptions by genetic mutation of transport (low NO3- uptake) and assimilation (high NO3- accumulation). The developed biosensor effectively monitors NO3- concentrations at the cellular level in real time and spatiotemporal changes during the plant life cycle.

Plasma proteome atlas for differentiating tumor stage and post-surgical prognosis of hepatocellular carcinoma and cholangiocarcinoma.

Although mass spectrometry-based plasma proteomics enables sensitive and large-scale discovery and validation of biomarkers for various diseases, its integrative application to hepatocellular carcinoma (HCC) and cholangiocarcinoma (CCA) is not well investigated. Therefore, we analyzed albumin- and immunoglobulin G-depleted plasma samples from 148 and 60 patients with HCC and CCA, respectively, using liquid chromatography-tandem mass spectrometry. The algorithm used to measure the content of each protein was the percentage of exponentially modified protein abundance index. From 5320 proteins assayed in plasma, 53 and 25 biomarker candidates were identified for HCC and CCA, respectively. The abundance of six and two HCC markers particularly protruded in stage II and III, respectively, whereas plasma serine protease inhibitor was the sole marker the level of which steadily decreased with CCA progression. From a prognostic facet, we showed candidate markers and their cutoff levels for evaluating probability of tumor recurrence and patient survival period. Combination Kaplan-Meier models showed that HCC stage III or IV and both the content of alpha-2-HS-glycoprotein and apolipoprotein CIII <0.2% exhibited the poorest post-surgical recurrence-free and overall survivals. Furthermore, the content of afamin ≥0.2% played a significant role on the poor prognosis in patients with CCA. Our findings, taken together, characterized novel plasma biomarker signatures in dissecting tumor stages and post-surgical outcomes of HCC and CCA.