Arabidopsis hydathodes are sites of auxin accumulation and nutrient scavenging.

Hydathodes are small organs found on the leaf margins of vascular plants which release excess xylem sap through a process called guttation. While previous studies have hinted at additional functions of hydathode in metabolite transport or auxin metabolism, experimental support is limited. We conducted comprehensive transcriptomic, metabolomic and physiological analyses of mature Arabidopsis hydathodes. This study identified 1460 genes differentially expressed in hydathodes compared to leaf blades, indicating higher expression of most genes associated with auxin metabolism, metabolite transport, stress response, DNA, RNA or microRNA processes, plant cell wall dynamics and wax metabolism. Notably, we observed differential expression of genes encoding auxin-related transcriptional regulators, biosynthetic processes, transport and vacuolar storage supported by the measured accumulation of free and conjugated auxin in hydathodes. We also showed that 78% of the total content of 52 xylem metabolites was removed from guttation fluid at hydathodes. We demonstrate that NRT2.1 and PHT1;4 transporters capture nitrate and inorganic phosphate in guttation fluid, respectively, thus limiting the loss of nutrients during this process. Our transcriptomic and metabolomic analyses unveil an organ with its specific physiological and biological identity.

Comparative transcriptomics reveals a highly polymorphic Xanthomonas HrpG virulence regulon.

BACKGROUND: Bacteria of the genus Xanthomonas cause economically significant diseases in various crops. Their virulence is dependent on the translocation of type III effectors (T3Es) into plant cells by the type III secretion system (T3SS), a process regulated by the master response regulator HrpG. Although HrpG has been studied for over two decades, its regulon across diverse Xanthomonas species, particularly beyond type III secretion, remains understudied. RESULTS: In this study, we conducted transcriptome sequencing to explore the HrpG regulons of 17 Xanthomonas strains, encompassing six species and nine pathovars, each exhibiting distinct host and tissue specificities. We employed constitutive expression of plasmid-borne hrpG*, which encodes a constitutively active form of HrpG, to induce the regulon. Our findings reveal substantial inter- and intra-specific diversity in the HrpG* regulons across the strains. Besides 21 genes directly involved in the biosynthesis of the T3SS, the core HrpG* regulon is limited to only five additional genes encoding the transcriptional activator HrpX, the two T3E proteins XopR and XopL, a major facility superfamily (MFS) transporter, and the phosphatase PhoC. Interestingly, genes involved in chemotaxis and genes encoding enzymes with carbohydrate-active and proteolytic activities are variably regulated by HrpG*. CONCLUSIONS: The diversity in the HrpG* regulon suggests that HrpG-dependent virulence in Xanthomonas might be achieved through several distinct strain-specific strategies, potentially reflecting adaptation to diverse ecological niches. These findings enhance our understanding of the complex role of HrpG in regulating various virulence and adaptive pathways, extending beyond T3Es and the T3SS.

A fully sequenced collection of homozygous EMS mutants for forward and reverse genetic screens in Arabidopsis thaliana.

Genetic screens are powerful tools for biological research and are one of the reasons for the success of the thale cress Arabidopsis thaliana as a research model. Here, we describe the whole-genome sequencing of 871 Arabidopsis lines from the Homozygous EMS Mutant (HEM) collection as a novel resource for forward and reverse genetics. With an average 576 high-confidence mutations per HEM line, over three independent mutations altering protein sequences are found on average per gene in the collection. Pilot reverse genetics experiments on reproductive, developmental, immune and physiological traits confirmed the efficacy of the tool for identifying both null, knockdown and gain-of-function alleles. The possibility of conducting subtle repeated phenotyping and the immediate availability of the mutations will empower forward genetic approaches. The sequence resource is searchable with the ATHEM web interface (https://lipm-browsers.toulouse.inra.fr/pub/ATHEM/), and the biological material is distributed by the Versailles Arabidopsis Stock Center.

CSF1R inhibition depletes brain macrophages and reduces brain virus burden in SIV-infected macaques.

Perivascular macrophages (PVMs) and, to a lesser degree, microglia are targets and reservoirs of HIV and simian immunodeficiency virus (SIV) in the brain. Previously, we demonstrated that colony-stimulating factor 1 receptor (CSF1R) in PVMs was upregulated and activated in chronically SIV-infected rhesus macaques with encephalitis, correlating with SIV infection of PVMs. Herein, we investigated the role of CSF1R in the brain during acute SIV infection using BLZ945, a brain-penetrant CSF1R kinase inhibitor. Apart from three uninfected historic controls, nine Indian rhesus macaques were infected acutely with SIVmac251 and divided into three groups (n = 3 each): an untreated control and two groups treated for 20-30 days with low- (10 mg/kg/day) or high- (30 mg/kg/day) dose BLZ945. With the high-dose BLZ945 treatment, there was a significant reduction in cells expressing CD163 and CD206 across all four brain areas examined, compared with the low-dose treatment and control groups. In 9 of 11 tested regions, tissue viral DNA (vDNA) loads were reduced by 95%-99% following at least one of the two doses, and even to undetectable levels in some instances. Decreased numbers of CD163+ and CD206+ cells correlated significantly with lower levels of vDNA in all four corresponding brain areas. In contrast, BLZ945 treatment did not significantly affect the number of microglia. Our results indicate that doses as low as 10 mg/kg/day of BLZ945 are sufficient to reduce the tissue vDNA loads in the brain with no apparent adverse effect. This study provides evidence that infected PVMs are highly sensitive to CSF1R inhibition, opening new possibilities to achieve viral clearance.

Bacterial host adaptation through sequence and structural variations of a single type III effector gene.

Molecular mechanisms underlying quantitative variations of pathogenicity remain elusive. Here, we identified the Xanthomonas campestris XopJ6 effector that triggers disease resistance in cauliflower and Arabidopsis thaliana. XopJ6 is a close homolog of the Ralstoniapseudosolanacearum PopP2 YopJ family acetyltransferase. XopJ6 is recognized by the RRS1-R/RPS4 NLR pair that integrates a WRKY decoy domain mimicking effector targets. We identified a XopJ6 natural variant carrying a single residue substitution in XopJ6 WRKY-binding site that disrupts interaction with WRKY proteins. This mutation allows XopJ6 to evade immune perception while retaining some XopJ6 virulence functions. Interestingly, xopJ6 resides in a Tn3-family transposon likely contributing to xopJ6 copy number variation (CNV). Using synthetic biology, we demonstrate that xopJ6 CNV tunes pathogen virulence on Arabidopsis through gene dosage-mediated modulation of xopJ6 expression. Together, our findings highlight how sequence and structural genetic variations restricted at a particular effector gene contribute to bacterial host adaptation.

Prognostic value of B-score for predicting joint replacement in the context of osteoarthritis phenotypes: Data from the osteoarthritis initiative.

Objective: Developing new therapies for knee osteoarthritis (KOA) requires improved prediction of disease progression. This study evaluated the prognostic value of clinical clusters and machine-learning derived quantitative 3D bone shape B-score for predicting total and partial knee replacement (KR). Design: This retrospective study used longitudinal data from the Osteoarthritis Initiative. A previous study used patients' clinical profiles to delineate phenotypic clusters. For these clusters, the distribution of B-scores was assessed (employing Tukey's method). The value of both cluster allocation and B-score for KR-prediction was then evaluated using multivariable Cox regression models and Kaplan-Meier curves for time-to-event analyses. The impact of using B-score vs. cluster was evaluated using a likelihood ratio test for the multivariable Cox model; global performances were assessed by concordance statistics (Harrell's C-index) and time dependent receiver operating characteristic (ROC) curves. Results: B-score differed significantly for the individual clinical clusters (p â€‹< â€‹0.001). Overall, 9.4% of participants had a KR over 9 years, with a shorter time to event in clusters with high B-score at baseline. Those clusters were characterized clinically by a high rate of comorbidities and potential signs of inflammation. Both phenotype and B-score independently predicted KR, with better prediction if combined (P â€‹< â€‹0.001). B-score added predictive value in groups with less pain and radiographic severity but limited physical activity. Conclusions: B-scores correlated with phenotypes based on clinical patient profiles. B-score and phenotype independently predicted KR surgery, with higher predictive value if combined. This can be used for patient stratification in drug development and potentially risk prediction in clinical practice.

Attitudes and Behaviors towards Food and Weight in Late Pregnancy: A Comparative Approach between Individuals with and without Previous Bariatric Surgery.

The aims of this study were to compare, between pregnant individuals with and without bariatric surgery: (1) eating behaviors, (2) intuitive eating components and, (3) attitudes towards weight gain. This retrospective study included data collected in healthy pregnant individuals with and without previous bariatric surgery who were recruited at the Centre Hospitalier Universitaire (CHU) de Québec-Université Laval. Pregnant individuals who underwent bariatric surgery (biliopancreatic bypass with duodenal switch [n = 14] or sleeve gastrectomy [n = 5]) were individually matched, for age (±0.4 years) and body mass index (BMI) (±0.3 kg/m2), with pregnant individuals who have not received bariatric surgery. In the second trimester, participants completed the Three Factor Eating Questionnaire (TFEQ) and the Intuitive Eating Scale 2 (IES-2). In the third trimester, participants completed the French version of the Pregnancy Weight Gain Attitude Scale assessing attitudes towards weight gain. Pregnant individuals who have had bariatric surgery had a higher score for flexible restraint and a lower score for situational susceptibility to disinhibition compared to individuals who have not had undergone bariatric surgery (2.89 ± 1.15 vs. 1.95 ± 1.31; p = 0.04 and 1.11 ± 1.29 vs. 2.79 ± 1.44, respectively; p < 0.001). Regarding intuitive eating, pregnant individuals who experienced bariatric surgery had a higher score for reliance on internal hunger and satiety cues and a lower one for unconditional permission to eat compared with those who had not experienced bariatric surgery (3.99 ± 0.81 vs. 3.30 ± 1.03; p = 0.02 and 3.28 ± 0.54 vs. 3.61 ± 0.68, respectively; p = 0.03). No difference in attitudes towards weight gain was observed between groups. Overall, pregnant individuals who had undergone bariatric surgery had different eating behaviors and intuitive eating components compared to pregnant individuals without bariatric surgery. These results need to be confirmed in further studies with larger sample sizes.

Grouping Boundary Proposals for Fast Interactive Image Segmentation.

Geodesic models are known as an efficient tool for solving various image segmentation problems. Most of existing approaches only exploit local pointwise image features to track geodesic paths for delineating the objective boundaries. However, such a segmentation strategy cannot take into account the connectivity of the image edge features, increasing the risk of shortcut problem, especially in the case of complicated scenario. In this work, we introduce a new image segmentation model based on the minimal geodesic framework in conjunction with an adaptive cut-based circular optimal path computation scheme and a graph-based boundary proposals grouping scheme. Specifically, the adaptive cut can disconnect the image domain such that the target contours are imposed to pass through this cut only once. The boundary proposals are comprised of precomputed image edge segments, providing the connectivity information for our segmentation model. These boundary proposals are then incorporated into the proposed image segmentation model, such that the target segmentation contours are made up of a set of selected boundary proposals and the corresponding geodesic paths linking them. Experimental results show that the proposed model indeed outperforms state-of-the-art minimal paths-based image segmentation approaches.

Computing geodesic paths encoding a curvature prior for curvilinear structure tracking.

In this paper, we introduce an efficient method for computing curves minimizing a variant of the Euler-Mumford elastica energy, with fixed endpoints and tangents at these endpoints, where the bending energy is enhanced with a user-defined and data-driven scalar-valued term referred to as the curvature prior. In order to guarantee that the globally optimal curve is extracted, the proposed method involves the numerical computation of the viscosity solution to a specific static Hamilton-Jacobi-Bellman (HJB) partial differential equation (PDE). For that purpose, we derive the explicit Hamiltonian associated with this variant model equipped with a curvature prior, discretize the resulting HJB PDE using an adaptive finite difference scheme, and solve it in a single pass using a generalized fast-marching method. In addition, we also present a practical method for estimating the curvature prior values from image data, designed for the task of accurately tracking curvilinear structure centerlines. Numerical experiments on synthetic and real-image data illustrate the advantages of the considered variant of the elastica model with a prior curvature enhancement in complex scenarios where challenging geometric structures appear.

Arabidopsis thaliana Early Foliar Proteome Response to Root Exposure to the Rhizobacterium Pseudomonas simiae WCS417.

Pseudomonas simiae WCS417 is a plant growth-promoting rhizobacterium that improves plant health and development. In this study, we investigate the early leaf responses of Arabidopsis thaliana to WCS417 exposure and the possible involvement of formate dehydrogenase (FDH) in such responses. In vitro-grown A. thaliana seedlings expressing an FDH::GUS reporter show a significant increase in FDH promoter activity in their roots and shoots after 7 days of indirect exposure (without contact) to WCS417. After root exposure to WCS417, the leaves of FDH::GUS plants grown in the soil also show an increased FDH promoter activity in hydathodes. To elucidate early foliar responses to WCS417 as well as FDH involvement, the roots of A. thaliana wild-type Col and atfdh1-5 knock-out mutant plants grown in soil were exposed to WCS417, and proteins from rosette leaves were subjected to proteomic analysis. The results reveal that chloroplasts, in particular several components of the photosystems PSI and PSII, as well as members of the glutathione S-transferase family, are among the early targets of the metabolic changes induced by WCS417. Taken together, the alterations in the foliar proteome, as observed in the atfdh1-5 mutant, especially after exposure to WCS417 and involving stress-responsive genes, suggest that FDH is a node in the early events triggered by the interactions between A. thaliana and the rhizobacterium WCS417. [Formula: see text] Copyright © 2023 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Gap Balancing Throughout the Arc of Motion With Navigated TKA and a Novel Force-Controlled Distractor: A Review of the First 273 Cases.

BACKGROUND: This study evaluated the ability to achieve the targeted soft-tissue balance in terms of medio-lateral (ML) laxity and gap values when using a computer-assisted orthopedic surgery (CAOS) system featuring an intra-articular force-controlled distractor and assessed learning curves associated with the adoption of this technology. METHODS: The first 273 cases using this technology were reported without exclusions comparing 1) final ML laxity and 2) final average gap to their predefined targets. For both parameters, the signed and unsigned differentials were reported. The linear mixed model was used to evaluate laxity curve differences between surgeons. A cumulative sum control chart (CUSUM) was applied to assess surgeon learning curves regarding surgical time. RESULTS: Both the average signed ML laxity and gap differentials were neutral throughout the full arc of motion. Both the average unsigned ML laxity and gap differentials were linear. Signature of ML laxity and gap differential curves tended to be surgeon-specific. The CUSUM analyses of surgical times demonstrated either a short learning curve or the absence of a discernible learning pattern for surgeons. CONCLUSION: Data from all users involved with the pilot release of the balancing device were considered to capture variability in familiarity with the technique and learning curve cases were included. A high ability to achieve targeted gap balance throughout the arc of motion using the proposed method was observed.

CRISPRi in Xanthomonas demonstrates functional convergence of transcription activator-like effectors in two divergent pathogens.

Functional analysis of large gene families in plant pathogens can be cumbersome using classical insertional mutagenesis. Additionally, Cas9 toxicity has limited the application of CRISPR-Cas9 for directed mutagenesis in bacteria. Here, we successfully applied a CRISPR interference strategy to investigate the cryptic role of the transcription activator-like effector (tale) multigene family in several plant-pathogenic Xanthomonas bacterial species, owing to their contribution to pathogen virulence. Single guide RNAs (sgRNAs) designed against Xanthomonas phaseoli pv manihotis tale conserved gene sequences efficiently silenced expression of all tales, with concomitant decrease in virulence and TALE-induced host gene expression. The system is readily translatable to other Xanthomonas species infecting rice, citrus, Brassica, and cassava, silencing up to 16 tales in a given strain using a single sgRNA. Complementation with plasmid-borne designer tales lacking the sgRNA-targeted sequence restored molecular and virulence phenotypes in all pathosystems. Our results evidenced that X. campestris pv campestris CN08 tales are relevant for symptom development in cauliflower. They also show that the MeSWEET10a sugar transporter is surprisingly targeted by the nonvascular cassava pathogen X. cassavae, highlighting a new example of TALE functional convergence between phylogenetically distant Xanthomonas. Overall, this novel technology provides a platform for discovery and rapid functional understanding of highly conserved gene families.

The eINTACT system dissects bacterial exploitation of plant osmosignalling to enhance virulence.

Bacteria inject effector proteins into host cells to manipulate cellular processes that promote disease. Since bacteria deliver minuscule amounts of effectors only into targeted host cells, it is technically challenging to capture effector-dependent cellular changes from bulk-infected host tissues. Here, we report a new technique called effector-inducible isolation of nuclei tagged in specific cell types (eINTACT), which facilitates affinity-based purification of nuclei from Arabidopsis plant cells that have received Xanthomonas bacterial effectors. Analysis of purified nuclei reveals that the Xanthomonas effector XopD manipulates the expression of Arabidopsis abscisic acid signalling-related genes and activates OSCA1.1, a gene encoding a calcium-permeable channel required for stomatal closure in response to osmotic stress. The loss of OSCA1.1 causes leaf wilting and reduced bacterial growth in infected leaves, suggesting that OSCA1.1 promotes host susceptibility. eINTACT allows us to uncover that XopD exploits host OSCA1.1/abscisic acid osmosignalling-mediated stomatal closure to create a humid habitat that favours bacterial growth and opens up a new avenue for accurately elucidating functions of effectors from numerous gram-negative plant bacteria in native infection contexts.

Geodesic Models With Convexity Shape Prior.

The minimal geodesic models established upon the eikonal equation framework are capable of finding suitable solutions in various image segmentation scenarios. Existing geodesic-based segmentation approaches usually exploit image features in conjunction with geometric regularization terms, such as euclidean curve length or curvature-penalized length, for computing geodesic curves. In this paper, we take into account a more complicated problem: finding curvature-penalized geodesic paths with a convexity shape prior. We establish new geodesic models relying on the strategy of orientation-lifting, by which a planar curve can be mapped to an high-dimensional orientation-dependent space. The convexity shape prior serves as a constraint for the construction of local geodesic metrics encoding a particular curvature constraint. Then the geodesic distances and the corresponding closed geodesic paths in the orientation-lifted space can be efficiently computed through state-of-the-art Hamiltonian fast marching method. In addition, we apply the proposed geodesic models to the active contours, leading to efficient interactive image segmentation algorithms that preserve the advantages of convexity shape prior and curvature penalization.

Treatment of Dissecting Vertebral Pseudoaneurysms: A Single-Center Experience.

Background: Dissecting vertebral artery pseudoaneurysms represent a unique clinical challenge with careful appreciation for location of the posterior inferior cerebellar artery. Limited data is available in terms of outcomes regarding the various treatment modalities. Methods: 11 patients with dissecting pseudoaneurysms were identified from 2013-2021. Pseudoaneurysm size and morphology, clinical presentation, and treatment approach was collected. Success of treatment was recorded based on post-operative imaging as well as documented overall patient outcomes. Three primary treatment modalities emerged: coil embolization, stent assisted coiling, and flow diversion. Results: Of the 11 patients, 5 were female and 6 were male with an age from 36 to 69.7. 7 had ruptured pseudoaneurysms at time of treatment. Size of pseudoaneurysm ranged from 3 to 6 mm. 8 were on the right and 3 were on the left vertebral artery. 8 were proximal to PICA and 3 were distal. Co-dominance of vertebral filling was seen in 5 patients, 5 with dominance through right vertebral artery, and 1 with dominance through left vertebral artery. Variability existed in treatment approaches with 4 patients undergoing coil occlusion, 5 patients undergoing flow diversion stenting, and 2 patients undergoing flow diversion stenting with jailed coiling. 1 patient had enlargement of pseudoaneurysm while inpatient and required a second flow diversion device. 1 patient had two flow diversion devices placed initially at time of treatment due to morphology of PA. 6 patients had repeat angiograms between 6 to 9 months with complete occlusion. 3 had CTA or MRA with complete occlusion for those that had flow diversion, they were transitioned from aspirin and clopidogrel to aspirin monotherapy after first repeat angiogram. 6 patients required shunt placement for hydrocephalus. 1 patient died prior to discharge due to sepsis. 2 patients died post discharge: 1 with myocardial infarction and the 2nd due to urosepsis.Dissecting vertebral pseudoaneurysm has high morbidity and mortality if rupture occurs. Location of PICA origin influences treatment approach. Patients with poor Hunt/Hess scores upon arrival had increased risk for systemic infection and mortality.

Venous Outflow for Brain Arteriovenous Malformations: Overview and Treatment Implications.

Introduction: Recent evidence has demonstrated a close relationship between the cerebral venous and lymphatic systems. Venous congestion has been implicated in a host of neurologic disorders, with relevance for vascular etiologies. Objective: The authors aim to review the literature as it pertains to brain arteriovenous malformations' (BAVMs) venous hemodynamics and glymphatic pathways, as well as the implications of BAVM treatment. Results: BAVMs offer a unique challenge, with sudden alteration in flow dynamics leading to increased hemorrhage risk and difficult challenges post-treatment. Conclusion: Recent progress in the understanding of CNS fluid dynamics and glymphatic pathways have revealed important implications for BAVM pathology and treatment. As imaging techniques and treatment modalities advance, there is a need to further investigate this relationship as it relates to therapeutic options and post-treatment sequalae.

A ß-glucuronidase (GUS) Based Bacterial Competition Assay to Assess Fine Differencesin Fitness during Plant Infection.

Competition assays are a simple phenotyping strategy that confront two bacterial strains to evaluate their relative fitness. Because they are more accurate than single-strain growth assays, competition assays can be used to highlight slight differences that would not otherwise be detectable. In the frame of host-pathogens interactions, they can be very useful to study the contribution of individual bacterial genes to bacterial fitness and lead to the identification of new adaptive traits. Here, we describe how to perform such competition assays by taking the example of the model phytopathogenic bacterium Xanthomonas campestris pv. campestris during infection of the mesophyll of its cauliflower host. This phenotypic assay is based on the use of a Competitive Index (CI) that compares the relative abundance of co-inoculated strains before and after inoculation. Since multiplication is a direct proxy for bacterial fitness, the evolution of the ratio between both strains in the mixed population is a direct way to assess differences in fitness in a given environment. In this protocol, we exploit the blue staining of GUS-expressing bacteria to count blue vs. white colonies on plates and estimate the competitiveness of the strains of interest in plant mesophyll.

Hydathodes.

Bellenot et al. introduce hydathodes, an oft-overlooked plant organ that acts as a pressure valve to expel excess guttation sap at the leaf margin, typically visible at dawn.

Genome Sequences of 17 Strains from Eight Races of Xanthomonas campestris pv. campestris.

Xanthomonas campestris pv. campestris is a group of phytopathogenic bacteria causing black rot disease on Brassicaceae crops. Here, we report on draft genome sequences of 17 strains representing eight of nine known races of this pathogen, including the pathotype strain CFBP 6865.

Genome-wide identification of fitness determinants in the Xanthomonas campestris bacterial pathogen during early stages of plant infection.

Plant diseases are an important threat to food production. While major pathogenicity determinants required for disease have been extensively studied, less is known on how pathogens thrive during host colonization, especially at early infection stages. Here, we used randomly barcoded-transposon insertion site sequencing (RB-TnSeq) to perform a genome-wide screen and identify key bacterial fitness determinants of the vascular pathogen Xanthomonas campestris pv campestris (Xcc) during infection of the cauliflower host plant (Brassica oleracea). This high-throughput analysis was conducted in hydathodes, the natural entry site of Xcc, in xylem sap and in synthetic media. Xcc did not face a strong bottleneck during hydathode infection. In total, 181 genes important for fitness were identified in plant-associated environments with functional enrichment in genes involved in metabolism but only few genes previously known to be involved in virulence. The biological relevance of 12 genes was independently confirmed by phenotyping single mutants. Notably, we show that XC_3388, a protein with no known function (DUF1631), plays a key role in the adaptation and virulence of Xcc possibly through c-di-GMP-mediated regulation. This study revealed yet unsuspected social behaviors adopted by Xcc individuals when confined inside hydathodes at early infection stages.