A Putative Apoplastic Effector of Clavibacter capsici, ChpGCc as Hypersensitive Response and Virulence (Hrv) Protein in Plants.

Clavibacter bacteria use secreted apoplastic effectors, such as putative serine proteases, for virulence in host plants and for hypersensitive response (HR) induction in nonhost plants. Previously, we have shown that Clavibacter capsici ChpGCc is important for the necrosis development in pepper (Capsicum annuum) leaves. Here, we determine the function of ChpGCc, along with three paralogous proteins, for HR induction in the apoplastic space of a nonhost plant, Nicotiana tabacum. The full-length and signal peptide-deleted (ΔSP) mature forms of all proteins fused with the tobacco PR1b signal sequence were generated. The full-length and ΔSP forms of ChpGCc and only the ΔSP forms of ChpECc and Pat-1Cc, but none of the ChpCCc, triggered HR. Based on the predicted protein structures, ChpGCc carries amino acids for a catalytic triad and a disulfide bridge in positions like Pat-1Cm. Substituting these amino acids of ChpGCc with alanine abolished or reduced HR-inducing activity. To determine whether these residues are important for necrosis development in pepper, alanine-substituted chpGCc genes were transformed into the C. capsici PF008ΔpCM1 strain, which lacks the intact chpGCc gene. The strain with any variants failed to restore the necrosis-causing ability. These results suggest that ChpGCc has a dual function as a virulence factor in host plants and an HR elicitor in nonhost plants. Based on our findings and previous results, we propose Clavibacter apoplastic effectors, such as ChpGCc, Pat-1Cm, Chp-7Cs, and ChpGCm, as hypersensitive response and virulence (Hrv) proteins that display phenotypic similarities to the hypersensitive response and pathogenicity (Hrp) proteins found in gram-negative bacteria. [Formula: see text] Copyright © 2024 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Bacteriophage cocktail for biocontrol of soft rot disease caused by Pectobacterium species in Chinese cabbage.

Pectobacterium spp. are necrotrophic plant pathogens that cause the soft rot disease in Chinese cabbage, resulting in severe yield loss. The use of conventional antimicrobial agents, copper-based bactericides, and antibiotics has encountered several limitations, such as bioaccumulation on plants and microbial resistance. Bacteriophages (phages) are considered promising alternative antimicrobial agents against diverse phytopathogens. In this study, we isolated and characterized two virulent phages (phiPccP-2 and phiPccP-3) to develop a phage cocktail. Morphological and genomic analyses revealed that two phages belonged to the Tevenvirinae and Mccorquodalevirinae subfamilies, respectively. The phiPccP-2 and phiPccP-3 phages, which have a broad host range, were stable at various environmental conditions, such as various pHs and temperatures and exposure to ultraviolet light. The phage cocktail developed using these two lytic phages inhibited the emergence of phage-resistant bacteria compared to single-phage treatments in in vitro challenge assays. The phage cocktail treatment effectively prevented the development of soft rot symptom in matured Chinese cabbage leaves. Additionally, the phage cocktail comprising three phages (phiPccP-1, phiPccP-2, and phiPccP-3) showed superior biocontrol efficacy against the mixture of Pectobacterium strains in Chinese cabbage seedlings. These results suggest that developing phage cocktails is an effective approach for biocontrol of soft rot disease caused by Pectobacterium strains in crops compared to single-phage treatments. KEY POINTS: •Two newly isolated Pectobacterium phages, phiPccP-2 and phiPccP-3, infected diverse Pectobacterium species and effectively inhibited the emergence of phage-resistant bacteria. •Genomic and physiological analyses suggested that both phiPccP-2 and phiPccP-3 are lytic phages and that their lytic activities are stable in the environmental conditions under which Chinese cabbage grows. •Treatment using a phage cocktail comprising phiPccP-2 and phiPccP-3 efficiently suppressed soft rot disease in detached mature leaves and seedlings of Chinese cabbage, indicating the applicability of the phage cocktail as an alternative antimicrobial agent.

Functional Characterization of Two Cellulase Genes in the Gram-Positive Pathogenic Bacterium Clavibacter michiganensis for Wilting in Tomato.

Diverse plant pathogens secrete cellulases to degrade plant cell walls. Previously, the plasmid-borne cellulase gene celA was shown to be important for the virulence of the gram-positive bacterium Clavibacter michiganensis in tomato. However, details of the contribution of cellulases to the development of wilting in tomato have not been well-determined. To better understand the contribution of cellulases to the virulence of C. michiganensis in tomato, a mutant lacking cellulase activity was generated and complemented with truncated forms of certain cellulase genes, and virulence of those strain was examined. A celA mutant of the C. michiganensis type strain LMG7333 lost its cellulase activity and almost all its ability to cause wilting in tomato. The cellulase catalytic domain and cellulose-binding domain of CelA together were sufficient for both cellulase activity and the development of wilting in tomato. However, the expansin domain did not affect virulence or cellulase activity. The celA ortholog of Clavibacter sepedonicus restored the full virulence of the celA mutant of C. michiganensis. Another cellulase gene, celB, located in the chromosome, carries a single-base deletion in most C. michiganensis strains but does not carry a functional signal peptide in its N terminus. Nevertheless, an experimentally modified CelB protein with a CelA signal peptide was secreted and able to cause wilting in tomato. These results indicate that cellulases are major virulence factors of C. michiganensis that causes wilting in tomato. Furthermore, there are natural variations among cellulase genes directly affecting their function.

Multiple plasmid-borne virulence genes of Clavibacter michiganensis ssp. capsici critical for disease development in pepper.

Clavibacter michiganensis ssp. capsici is a Gram-positive plant-pathogenic bacterium causing bacterial canker disease in pepper. Virulence genes and mechanisms of C. michiganensis ssp. capsici in pepper have not yet been studied. To identify virulence genes of C. michiganensis ssp. capsici, comparative genome analyses with C. michiganensis ssp. capsici and its related C. michiganensis subspecies, and functional analysis of its putative virulence genes during infection were performed. The C. michiganensis ssp. capsici type strain PF008 carries one chromosome (3.056 Mb) and two plasmids (39 kb pCM1Cmc and 145 kb pCM2Cmc ). The genome analyses showed that this bacterium lacks a chromosomal pathogenicity island and celA gene that are important for disease development by C. michiganensis ssp. michiganensis in tomato, but carries most putative virulence genes in both plasmids. Virulence of pCM1Cmc -cured C. michiganensis ssp. capsici was greatly reduced compared with the wild-type strain in pepper. The complementation analysis with pCM1Cmc -located putative virulence genes showed that at least five genes, chpE, chpG, ppaA1, ppaB1 and pelA1, encoding serine proteases or pectate lyase contribute to disease development in pepper. In conclusion, C. michiganensis ssp. capsici has a unique genome structure, and its multiple plasmid-borne genes play critical roles in virulence in pepper, either separately or together.

Identification of a molecular marker tightly linked to bacterial wilt resistance in tomato by genome-wide SNP analysis.

KEY MESSAGE: Genotyping of disease resistance to bacterial wilt in tomato by a genome-wide SNP analysis Bacterial wilt caused by Ralstonia pseudosolanacearum is one of the destructive diseases in tomato. The previous studies have identified Bwr-6 (chromosome 6) and Bwr-12 (chromosome 12) loci as the major quantitative trait loci (QTLs) contributing to resistance against bacterial wilt in tomato cultivar 'Hawaii7996'. However, the genetic identities of two QTLs have not been uncovered yet. In this study, using whole-genome resequencing, we analyzed genome-wide single-nucleotide polymorphisms (SNPs) that can distinguish a resistant group, including seven tomato varieties resistant to bacterial wilt, from a susceptible group, including two susceptible to the same disease. In total, 5259 non-synonymous SNPs were found between the two groups. Among them, only 265 SNPs were located in the coding DNA sequences, and the majority of these SNPs were located on chromosomes 6 and 12. The genes that both carry SNP(s) and are near Bwr-6 and Bwr-12 were selected. In particular, four genes in chromosome 12 encode putative leucine-rich repeat (LRR) receptor-like proteins. SNPs within these four genes were used to develop SNP markers, and each SNP marker was validated by a high-resolution melting method. Consequently, one SNP marker, including a functional SNP in a gene, Solyc12g009690.1, could efficiently distinguish tomato varieties resistant to bacterial wilt from susceptible varieties. These results indicate that Solyc12g009690.1, the gene encoding a putative LRR receptor-like protein, might be tightly linked to Bwr-12, and the SNP marker developed in this study will be useful for selection of tomato cultivars resistant to bacterial wilt.

Functional roles of the pepper leucine-rich repeat protein and its interactions with pathogenesis-related and hypersensitive-induced proteins in plant cell death and immunity.

MAIN CONCLUSION: Pepper leucine-rich repeat protein (CaLRR1) interacts with defense response proteins to regulate plant cell death and immunity. This review highlights the current understanding of the molecular functions of CaLRR1 and its interactor proteins. Plant cell death and immune responses to microbial pathogens are controlled by complex and tightly regulated molecular signaling networks. Xanthomonas campestris pv. vesicatoria (Xcv)-inducible pepper (Capsicum annuum) leucine-rich repeat protein 1 (CaLRR1) serves as a molecular marker for plant cell death and immunity signaling. In this review, we discuss recent advances in elucidating the functional roles of CaLRR1 and its interacting plant proteins, and understanding how they are involved in the cell death and defense responses. CaLRR1 physically interacts with pepper pathogenesis-related proteins (CaPR10 and CaPR4b) and hypersensitive-induced reaction protein (CaHIR1) to regulate plant cell death and defense responses. CaLRR1 is produced in the cytoplasm and trafficked to the extracellular matrix. CaLRR1 binds to CaPR10 in the cytoplasm and CaPR4b and CaHIR1 at the plasma membrane. CaLRR1 synergistically accelerates CaPR10-triggered hypersensitive cell death, but negatively regulates CaPR4b- and CaHIR1-triggered cell death. CaHIR1 interacts with Xcv filamentous hemagglutinin (Fha1) to trigger disease-associated cell death. The subcellular localization and cellular function of these CaLRR1 interactors during plant cell death and defense responses were elucidated by Agrobacterium-mediated transient expression, virus-induced gene silencing, and transgenic overexpression studies. CaPR10, CaPR4b, and CaHIR1 positively regulate defense signaling mediated by salicylic acid and reactive oxygen species, thereby activating hypersensitive cell death and disease resistance. A comprehensive understanding of the molecular functions of CaLRR1 and its interacting protein partners in cell death and defense responses will provide valuable information for the molecular genetics of plant disease resistance, which could be exploited as a sustainable disease management strategy.

Clavibacter michiganensis subsp. capsici subsp. nov., causing bacterial canker disease in pepper.

Clavibacter michiganensis is a Gram-stain-positive bacterium with eight subspecies. One of these subspecies is C. michiganensis subsp. michiganensis, which causes bacterial canker disease in tomato. Bacterial strains showing very similar canker disease symptoms to those of a strain originally classified as C. michiganensis have been isolated from pepper. In this paper, we reclassified strains isolated from pepper. On the basis of phylogenetic analysis with 16S rRNA gene sequences, the strains isolated from pepper were grouped in a separate clade from other subspecies of C. michiganensis. Biochemical, physiological and genetic characteristics of strain PF008T, which is the representative strain of the isolates from pepper, were examined in this study. Based on multi-locus sequence typing and other biochemical and physiological features including colony color, utilization of carbon sources and enzyme activities, strain PF008T was categorically differentiated from eight subspecies of C. michiganensis. Moreover, genome analysis showed that the DNA G+C content of strain PF008T is 73.2 %. These results indicate that PF008T is distinct from other known subspecies of C. michiganensis. Therefore, we propose a novel subspecies, C. michiganensis subsp. capsici, causing bacterial canker disease in pepper, with a type strain of PF008T (=KACC 18448T=LMG 29047T).

Pathogenesis-related protein 4b interacts with leucine-rich repeat protein 1 to suppress PR4b-triggered cell death and defense response in pepper.

To control defense and cell-death signaling, plants contain an abundance of pathogen recognition receptors such as leucine-rich repeat (LRR) proteins. Here we show that pepper (Capsicum annuum) LRR1 interacts with the pepper pathogenesis-related (PR) protein 4b, PR4b, in yeast and in planta. PR4b is synthesized in the endoplasmic reticulum, interacts with LRR1 in the plasma membrane, and is secreted to the apoplast via the plasma membrane. Binding of PR4b to LRR1 requires the chitin-binding domain of PR4b. Purified PR4b protein inhibits spore germination and mycelial growth of plant fungal pathogens. Transient expression of PR4b triggers hypersensitive cell death. This cell death is compromised by co-expression of LRR1 as a negative regulator in Nicotiana benthamiana leaves. LRR1/PR4b silencing in pepper and PR4b over-expression in Arabidopsis thaliana demonstrated that LRR1 and PR4b are necessary for defense responses to Pseudomonas syringae pv. tomato and Hyaloperonospora arabidopsidis (Hpa) infection. The mutant of the PR4b Arabidopsis ortholog, pr4, showed enhanced susceptibility to Hpa infection. Together, our results suggest that PR4b functions as a positive modulator of plant cell death and defense responses. However, the activity of PR4b is suppressed by interaction with LRR1.

Two nuclear export signals of Cdc6 are differentially associated with CDK-mediated phosphorylation residues for cytoplasmic translocation.

Cdc6 is cleaved at residues 442 and 290 by caspase-3 during apoptosis producing p49-tCdc6 and p32-tCdc6, respectively. While p32-tCdc6 is unable to translocate into the cytoplasm, p49-tCdc6 retains cytoplasmic translocation activity, but it has a lower efficiency than wild-type Cdc6. We hypothesized that a novel nuclear export signal (NES) sequence exists between amino acids 290 and 442. Cdc6 contains a novel NES in the region of amino acids 300-315 (NES2) that shares sequence similarity with NES1 at residues 462-476. In mutant versions of Cdc6, we replaced leucine with alanine in NES1 and NES2 and co-expressed the mutant constructs with cyclin A. We observed that the cytoplasmic translocation of these mutants was reduced in comparison to wild-type Cdc6. Moreover, the cytoplasmic translocation of a mutant in which all four leucine residues were mutated to alanine was significantly inhibited in comparison to the translocation of wild-type Cdc6. The Crm1 binding activities of Cdc6 NES mutants were consistent with the efficiency of its cytoplasmic translocation. Further studies have revealed that L468 and L470 of NES1 are required for cytoplasmic translocation of Cdc6 phosphorylated at S74, while L311 and L313 of NES2 accelerate the cytoplasmic translocation of Cdc6 phosphorylated at S54. These results suggest that the two NESs of Cdc6 work cooperatively and distinctly for the cytoplasmic translocation of Cdc6 phosphorylated at S74 and S54 by cyclin A/Cdk2.

The Pepper Lipoxygenase CaLOX1 Plays a Role in Osmotic, Drought and High Salinity Stress Response.

In plants, lipoxygenases (LOXs) are involved in various physiological processes, including defense responses to biotic and abiotic stresses. Our previous study had shown that the pepper 9-LOX gene, CaLOX1, plays a crucial role in cell death due to pathogen infection. Here, the function of CaLOX1 in response to osmotic, drought and high salinity stress was examined using CaLOX1-overexpressing (CaLOX1-OX) Arabidopsis plants. Changes in the temporal expression pattern of the CaLOX1 gene were observed when pepper leaves were treated with drought and high salinity, but not when treated with ABA, the primary hormone in response to drought stress. During seed germination and seedling development, CaLOX1-OX plants were more tolerant to ABA, mannitol and high salinity than wild-type plants. In contrast, expression of the ABA-responsive marker genes RAB18 and RD29B was higher in CaLOX1-OX Arabidopsis plants than in wild-type plants. In response to high salinity, CaLOX1-OX plants exhibited enhanced tolerance, compared with the wild type, which was accompanied by decreased accumulation of H2O2 and high levels of RD20, RD29A, RD29B and P5CS gene expression. Similarly, CaLOX1-OX plants were also more tolerant than wild-type plants to severe drought stress. H2O2 production and the relative increase in lipid peroxidation were lower, and the expression of COR15A, DREB2A, RD20, RD29A and RD29B was higher in CaLOX1-OX plants, relative to wild-type plants. Taken together, our results indicate that CaLOX1 plays a crucial role in plant stress responses by modulating the expression of ABA- and stress-responsive marker genes, lipid peroxidation and H2O2 production.

Requirement of the cytosolic interaction between PATHOGENESIS-RELATED PROTEIN10 and LEUCINE-RICH REPEAT PROTEIN1 for cell death and defense signaling in pepper.

Plants recruit innate immune receptors such as leucine-rich repeat (LRR) proteins to recognize pathogen attack and activate defense genes. Here, we identified the pepper (Capsicum annuum) pathogenesis-related protein10 (PR10) as a leucine-rich repeat protein1 (LRR1)-interacting partner. Bimolecular fluorescence complementation and coimmunoprecipitation assays confirmed the specific interaction between LRR1 and PR10 in planta. Avirulent Xanthomonas campestris pv vesicatoria infection induces PR10 expression associated with the hypersensitive cell death response. Transient expression of PR10 triggers hypersensitive cell death in pepper and Nicotiana benthamiana leaves, which is amplified by LRR1 coexpression as a positive regulator. LRR1 promotes the ribonuclease activity and phosphorylation of PR10, leading to enhanced cell death signaling. The LRR1-PR10 complex is formed in the cytoplasm, resulting in its secretion into the apoplastic space. Engineered nuclear confinement of both proteins revealed that the cytoplasmic localization of the PR10-LRR1 complex is essential for cell death-mediated defense signaling. PR10/LRR1 silencing in pepper compromises resistance to avirulent X. campestris pv vesicatoria infection. By contrast, PR10/LRR1 overexpression in Arabidopsis thaliana confers enhanced resistance to Pseudomonas syringae pv tomato and Hyaloperonospora arabidopsidis. Together, these results suggest that the cytosolic LRR-PR10 complex is responsible for cell death-mediated defense signaling.

Antioxidant treatment during manipulation procedures prevents mitochondrial and DNA damage and enhances nuclear reprogramming of bovine somatic cell nuclear transfer embryos.

We tried to prevent the mitochondrial and DNA damage caused by mechanical stress-associated reactive oxygen species (ROS), and to improve the reprogramming of bovine somatic cell nuclear transfer (SCNT) embryos by antioxidant treatment during the manipulation procedures of SCNT. Bovine recipient oocytes and reconstituted oocytes were treated with antioxidants during manipulation procedures. The H2O2 level, mitochondrial morphology, membrane potential and apoptosis at the one-cell stage, and in vitro development and DNA methylation status of blastocysts were evaluated. Antioxidant treatment during manipulation procedures reduced the H2O2 level of SCNT embryos. Antioxidant-treated SCNT embryos normally formed mitochondrial clumps, similar to IVF embryos, and showed higher mitochondrial membrane potential versus the SCNT control (P<0.05). Apoptosis and DNA fragmentation were reduced by antioxidant treatment. The development rate to the blastocyst stage was higher (P<0.05) in the antioxidant treatment groups (30.5±2.5 to 30.6±1.6%) versus the control (23.0±1.9%). The DNA methylation status of blastocysts in the antioxidant treatment groups was lower (P<0.05) than that of the control and similar to that of IVF embryos. These results indicate that antioxidant treatment during manipulation procedures can prevent cellular damage that may be caused by mechanical stress-associated ROS, and improve nuclear reprogramming.

The pepper cysteine/histidine-rich DC1 domain protein CaDC1 binds both RNA and DNA and is required for plant cell death and defense response.

Plant defense against microbial pathogens is coordinated by a complex regulatory network. Cysteine/histidine-rich DC1 domain proteins mediate a variety of cellular processes involved in plant growth, development and stress responses. We identified a pepper (Capsicum annuum) cysteine/histidine-rich DC1 domain protein gene, CaDC1, which positively regulates plant defense during microbial infection, based on gene silencing and transient expression in pepper, as well as ectopic expression in Arabidopsis. Induction of CaDC1 by avirulent Xanthomonas campestris pv vesicatoria (Xcv) infection was pronounced at both transcriptional and translational levels in pepper leaves. Purified CaDC1 protein bound to both DNA and RNA in vitro, especially in the presence of Zn(2+). CaDC1 was localized to both the nucleus and the cytoplasm, which was required for plant cell death signaling. The nuclear localization of CaDC1 was dependent on the divergent C1 (DC1) domain. CaDC1 silencing in pepper conferred increased susceptibility to Xcv infection, which was accompanied by reduced salicylic acid accumulation and defense-related gene expression. Ectopic expression of CaDC1 in Arabidopsis enhanced resistance to Hyaloperonospora arabidopsidis. CaDC1 binds both RNA and DNA and functions as a positive regulator of plant cell death and SA-dependent defense responses.

A novel virulence gene, cviA1 of Clavibacter michiganensis for necrosis development in the Nicotiana benthamiana plant.

Clavibacter michiganensis is a Gram-positive bacterium that causes diverse disease symptoms in tomatoes and Nicotiana benthamiana, a surrogate host plant, including canker, blister lesions, and wilting. Previously, we reported that C. michiganensis also causes necrosis in N. benthamiana leaves. Here, to identify novel virulence genes of C. michiganensis required for necrosis development in N. benthamiana leaves, we screened 1,862 transposon-inserted mutants and identified a mutant strain that exhibited weak and delayed necrosis, whereas there was no discernible difference in blister lesions, canker, or wilting symptoms. Notably, this mutant caused canker similar to that of the wild-type strain, but caused mild wilting in tomato. This mutant carried a transposon in a chromosomal gene, called Clavibactervirulence gene A1 (cviA1). CviA1 encodes a 180-amino acid protein with a signal peptide (SP) at the N-terminus and two putative transmembrane domains (TMs) at the C-terminus. Interestingly, deletion of the SP or the C-terminus, including the two putative TMs, in CviA1 failed to restore full necrosis in the mutant, highlighting the importance of protein secretion and the putative TMs for necrosis. A paralog of cviA1, cviA2 is located on the large plasmid pCM2 of C. michiganensis. Despite its high similarity to cviA1, the introduction of cviA2 into the cviA1 mutant strain did not restore virulence. Similarly, the introduction of cviA1 into the Clavibacter capsici type strain PF008, which initially lacks cviA1, did not enhance necrosis symptoms. These results reveals that the chromosomal cviA1 gene in C. michiganensis plays an important role in necrosis development in N. benthamiana leaves.

Label-Free Surface-Enhanced Raman Scattering Detection of Fire Blight Pathogen Using a Pathogen-Specific Bacteriophage.

Fire blight is one of the most devastating plant diseases, causing severe social and economic problems. Herein, we report a novel method based on label-free surface-enhanced Raman scattering (SERS) combined with an Erwinia amylovora-specific bacteriophage that allows detecting efficiently fire blight bacteria E. amylovora for the first time. To achieve the highest SERS signals for E. amylovora, we synthesized and compared plasmonic nanoparticles (PNPs) with different sizes, i.e., bimetallic gold core-silver shell nanoparticles (Au@AgNPs) and monometallic gold nanoparticles (AuNPs) and utilized the coffee-ring effect for the self-assembly of PNPs and enrichment of fire blight bacteria. Furthermore, we investigated the changes in the SERS spectra of E. amylovora after incubation with an E. amylovora-specific bacteriophage, and we found considerable differences in the SERS signals as a function of the bacteriophage incubation time. The results indicate that our bacteriophage-based label-free SERS analysis can specifically detect E. amylovora without the need for peak assignment on the SERS spectra but simply by monitoring the changes in the SERS signals over time. Therefore, our facile method holds great potential for the label-free detection of pathogenic bacteria and the investigation of viral-bacterial interactions.

Visceral obesity as a risk factor of incisional hernia after single-port laparoscopic gynecologic surgery.

OBJECTIVE: We aimed to evaluate associations between abdominal fat distribution (AFD) parameters and incisional hernia (IH) in patients who underwent transumbilical single-port laparoscopic surgery (SPLS) for gynecological disease. METHODS: Medical records of 2116 patients who underwent SPLS for gynecological disease at Daejeon St. Mary's Hospital between March 2014 and February 2021 were reviewed. Among 21 (1.0%) patients who developed IH requiring surgical treatment after SPLS, 18 had preoperative abdominopelvic computed tomography (CT) images. As a control group, we randomly selected 72 patients who did not develop IH and who had undergone preoperative abdominopelvic CT scan, matched to test patients by type of surgery. Total fat area (TFA), visceral fat area (VFA), subcutaneous fat area (SFA), visceral-to-subcutaneous fat ratio (VSR), and waist circumference (WC) were measured at the level of the third lumbar vertebral body on the preoperative abdominopelvic CT images, using National Institutes of Health (NIH) ImageJ version 1.53 k. RESULTS: Receiver operating curve analysis showed that VFA has the highest predictive value for IH among AFD parameters (AUC = 0.749, 95% CI 0.630-0.869, p < 0.001). Univariate analysis showed that age, BMI, hypertension, dyslipidemia, TFA, VFA, VSR and WC were significant factors for IH. In multivariate analysis, only high VFA was identified as an independent risk factor for IH (HR 6.18, 95% CI 1.13-33.87, p = 0.04), whereas BMI, TFA, SFA, VSR, and WC failed to show statistical significance. CONCLUSION: We could find high VFA as an independent risk factor of IH in patients who underwent SPLS for gynecologic disease.

On-site applicable diagnostic fluorescent probe for fire blight bacteria.

Fire blight is a representative plant infection that contaminates edible plants and causes socio-economic problems in agricultural and livestock industries globally. It is caused by the pathogen Erwinia amylovora (E. amylovora) creates lethal plant necrosis and spreads rapidly across plant organs. We newly disclose the fluorogenic probe B-1 for real-time on-site detection of fire blight bacteria for the first time. B-1 exhibited no emission signals but manifested bright emission properties in the presence of fire blight bacteria. Based on these features, fluorescence imaging of the fire blight bacteria and its real-time detection from the infected host plant tissues were conducted. The detection limit against E. amylovora was 102 CFU/mL, which had excellent sensitivity. The fluorogenic probe-based on-site diagnostic technology was supplemented by introducing a new portable UV device. This work holds enormous potential to be a new advanced tool for detecting fire blight in agricultural and livestock industries.

Developing Core Elements and Checklist Items for Implementing Antimicrobial Stewardship Programs in Korean General Hospitals: A Modified Delphi Survey.

BACKGROUND: Antimicrobial stewardship programs (ASPs) aim to optimize antimicrobial use by minimizing the spread of antimicrobial resistance. The core elements for implementing ASPs in healthcare facilities have been developed by the World Health Organization, international research group and government agencies of various countries. However, to date, there is no documented core elements for implementation of ASP in Korea. This survey aimed to establish a national consensus on a set of core elements and their related checklist items for the implementation of ASPs in Korean general hospitals. MATERIALS AND METHODS: The survey was conducted from July 2022 to August 2022 by the Korean Society for Antimicrobial Therapy with support from the Korea Disease Control and Prevention Agency. A literature review was conducted by searching Medline and relevant websites to retrieve a list of core elements and checklist items. These core elements and checklist items were evaluated by a multidisciplinary panel of experts using a structured modified Delphi consensus procedure, using two-step survey included online in-depth questionnaires and in-person meeting. RESULTS: The literature review identified 6 core elements (Leadership commitment, Operating system, Action, Tracking, Reporting, and Education) and 37 related checklist items. Fifteen experts participated in the consensus procedures. Ultimately, all 6 core elements were retained, and 28 checklist items were proposed, all with ≥80% agreement; in addition 9 items were merged into 2 items, 2 items were deleted, and 15 items were rephrased. CONCLUSION: This Delphi survey provides useful indicators for the implementation of ASP in Korea and suggests national policy improvement about the barriers (e.g., shortage of staffing and financial support) existing in Korea for optimal implementation of ASPs.

Pepper asparagine synthetase 1 (CaAS1) is required for plant nitrogen assimilation and defense responses to microbial pathogens.

Asparagine synthetase is a key enzyme in the production of the nitrogen-rich amino acid asparagine, which is crucial to primary nitrogen metabolism. Despite its importance physiologically, the roles that asparagine synthetase plays during plant defense responses remain unknown. Here, we determined that pepper (Capsicum annuum) asparagine synthetase 1 (CaAS1) is essential for plant defense to microbial pathogens. Infection with Xanthomonas campestris pv. vesicatoria (Xcv) induced early and strong CaAS1 expression in pepper leaves and silencing of this gene resulted in enhanced susceptibility to Xcv infection. Transgenic Arabidopsis (Arabidopsis thaliana) plants that overexpressed CaAS1 exhibited enhanced resistance to Pseudomonas syringae pv. tomato DC3000 and Hyaloperonospora arabidopsidis. Increased CaAS1 expression influenced early defense responses in diseased leaves, including increased electrolyte leakage, reactive oxygen species and nitric oxide bursts. In plants, increased conversion of aspartate to asparagine appears to be associated with enhanced resistance to bacterial and oomycete pathogens. In CaAS1-silenced pepper and/or CaAS1-overexpressing Arabidopsis, CaAS1-dependent changes in asparagine levels correlated with increased susceptibility or defense responses to microbial pathogens, respectively. Linking transcriptional and targeted metabolite studies, our results suggest that CaAS1 is required for asparagine synthesis and disease resistance in plants.

Overexpression of Xanthomonas campestris pv. vesicatoria effector AvrBsT in Arabidopsis triggers plant cell death, disease and defense responses.

Recognition of bacterial effector proteins by plant cells is crucial for plant disease and defense response signaling. The Xanthomonas campestris pv. vesicatoria (Xcv) type III effector protein, AvrBsT, is secreted into plant cells from Xcv strain Bv5-4a. Here, we demonstrate that dexamethasone (DEX): avrBsT overexpression triggers cell death signaling in healthy transgenic Arabidopsis plants. AvrBsT overexpression in Arabidopsis also reduced susceptibility to infection with the obligate biotrophic oomycete Hyaloperonospora arabidopsidis. Overexpression of avrBsT significantly induced some defense-related genes in Arabidopsis leaves. A high-throughput in planta proteomics screen identified TCP-1 chaperonin, SEC7-like guanine nucleotide exchange protein and calmodulin-like protein, which were differentially expressed in DEX:avrBsT-overexpression (OX) Arabidopsis plants during Hp. arabidopsidis infection. Treatment with purified GST-tagged AvrBsT proteins distinctly inhibited the growth and sporulation of Hp. arabidopsidis on Arabdiopsis cotyledons. In contrast, DEX:avrBsT-OX plants exhibited enhanced susceptibility to Pseudomonas syringae pv. tomato (Pst) DC3000 infection. Notably, susceptible cell death and enhanced electrolyte leakage were significantly induced in the Pst-infected leaves of DEX:avrBsT-OX plants. Together, these results suggest that Xcv effector AvrBsT overexpression triggers plant cell death, disease and defense signaling leading to both disease and defense responses to microbial pathogens of different lifestyles.