An NLR integrated domain toolkit to identify plant pathogen effector targets.

Plant immune receptors, known as NOD-like receptors (NLRs), possess unique integrated decoy domains that enable plants to attract pathogen effectors and initiate a specific immune response. The present study aimed to create a library of these integrated domains (IDs) and screen them with pathogen effectors to identify targets for effector virulence and NLR-effector interactions. This works compiles IDs found in NLRs from seven different plant species and produced a library of 78 plasmid clones containing a total of 104 IDs, representing 43 distinct InterPro domains. A yeast two-hybrid assay was conducted, followed by an in planta interaction test, using 32 conserved effectors from Ralstonia pseudosolanacearum type III. Through these screenings, three interactions involving different IDs (kinase, DUF3542, WRKY) were discovered interacting with two unrelated type III effectors (RipAE and PopP2). Of particular interest was the interaction between PopP2 and ID#85, an atypical WRKY domain integrated into a soybean NLR gene (GmNLR-ID#85). Using a Förster resonance energy transfer-fluorescence lifetime imaging microscopy technique to detect protein-protein interactions in living plant cells, PopP2 was demonstrated to physically associate with ID#85 in the nucleus. However, unlike the known WRKY-containing Arabidopsis RRS1-R NLR receptor, GmNLR-ID#85 could not be acetylated by PopP2 and failed to activate RPS4-dependent immunity when introduced into the RRS1-R immune receptor. The generated library of 78 plasmid clones, encompassing these screenable IDs, is publicly available through Addgene. This resource is expected to be valuable for the scientific community with respect to discovering targets for effectors and potentially engineering plant immune receptors.

SWI/SNF chromatin remodeling subunit Smarca4/BRG1 is essential for female fertility†.

Mammalian folliculogenesis is a complex process that involves the regulation of chromatin structure for gene expression and oocyte meiotic resumption. The SWI/SNF complex is a chromatin remodeler using either Brahma-regulated gene 1 (BRG1) or BRM (encoded by Smarca4 and Smarca2, respectively) as its catalytic subunit. SMARCA4 loss of expression is associated with a rare type of ovarian cancer; however, its function during folliculogenesis remains poorly understood. In this study, we describe the phenotype of BRG1 mutant mice to better understand its role in female fertility. Although no tumor emerged from BRG1 mutant mice, conditional depletion of Brg1 in the granulosa cells (GCs) of Brg1fl/fl;Amhr2-Cre mice caused sterility, whereas conditional depletion of Brg1 in the oocytes of Brg1fl/fl;Gdf9-Cre mice resulted in subfertility. Recovery of cumulus-oocyte complexes after natural mating or superovulation showed no significant difference in the Brg1fl/fl;Amhr2-Cre mutant mice and significantly fewer oocytes in the Brg1fl/fl;Gdf9-Cre mutant mice compared with controls, which may account for the subfertility. Interestingly, the evaluation of oocyte developmental competence by in vitro culture of retrieved two-cell embryos indicated that oocytes originating from the Brg1fl/fl;Amhr2-Cre mice did not reach the blastocyst stage and had higher rates of mitotic defects, including micronuclei. Together, these results indicate that BRG1 plays an important role in female fertility by regulating granulosa and oocyte functions during follicle growth and is needed for the acquisition of oocyte developmental competence.

Patient and Provider Satisfaction With Telehealth Preanesthesia Examinations.

PURPOSE: Telehealth has become commonplace in many healthcare systems across the United States. Due to the COVID-19 pandemic, a large academic medical center in the southeast USA has increased the use of telehealth in the anesthesia department to complete preanesthetic assessments before the patient's surgery. This has allowed high-risk patient populations to limit potential exposure to COVID-19 and limit the financial burden of traveling from neighboring counties to complete their preanesthetic assessment in person. The purpose of this quality improvement project was to evaluate the effectiveness of a preanesthetic assessment via telehealth among patients undergoing anesthesia. DESIGN: A quality improvement project. METHODS: Two separate surveys assessing patient and provider satisfaction with telehealth preanesthesia assessments were developed. Patients undergoing anesthesia at the medical center completed a telehealth preanesthesia assessment and received a satisfaction survey via email post-procedure. Certified registered nurse anesthetists (CRNAs) who cared for patients that completed a telehealth preanesthesia assessment also received an emailed satisfaction survey. FINDINGS: Results demonstrate high satisfaction scores for both patients and providers. CONCLUSIONS: Current telehealth preanesthetic assessments prompt high satisfaction scores, supporting continued, and expanded use.

Convergent Rewiring of the Virulence Regulatory Network Promotes Adaptation of Ralstonia solanacearum on Resistant Tomato.

The evolutionary and adaptive potential of a pathogen is a key determinant for successful host colonization and proliferation but remains poorly known for most of the pathogens. Here, we used experimental evolution combined with phenotyping, genomics, and transcriptomics to estimate the adaptive potential of the bacterial plant pathogen Ralstonia solanacearum to overcome the quantitative resistance of the tomato cultivar Hawaii 7996. After serial passaging over 300 generations, we observed pathogen adaptation to within-plant environment of the resistant cultivar but no plant resistance breakdown. Genomic sequence analysis of the adapted clones revealed few genetic alterations, but we provide evidence that all but one were gain of function mutations. Transcriptomic analyses revealed that even if different adaptive events occurred in independently evolved clones, there is convergence toward a global rewiring of the virulence regulatory network as evidenced by largely overlapping gene expression profiles. A subset of four transcription regulators, including HrpB, the activator of the type 3 secretion system regulon and EfpR, a global regulator of virulence and metabolic functions, emerged as key nodes of this regulatory network that are frequently targeted to redirect the pathogen's physiology and improve its fitness in adverse conditions. Significant transcriptomic variations were also detected in evolved clones showing no genomic polymorphism, suggesting that epigenetic modifications regulate expression of some of the virulence network components and play a major role in adaptation as well.

Diagnostic Neuroradiology Subspecialty Training: 1 Versus 2 Years; the Canadian Perspective.

BACKGROUND: Canada began a national reform of its post-graduate medical education training programs to a Competence By Design (CBD) model. Trends from accredited neuroradiology programs from the past 10 years were investigated to inform educators and stakeholders for this process. METHODS: A 13-question electronic survey was sent to program directors of all 8 accredited neuroradiology training programs in Canada. Data was requested for each year on the 2008-2019 graduating classes. Questions pertained to program enrolment; program completion; post-training employment; and the sufficiency of 1-year training programs. RESULTS: Response rate was 100%. Over the timeframe studied, the 2-year programs increased in size (P = 0.007), while the 1-year programs remained steady (P = 0.27). 12.2% of trainees enrolled in the 2-year program dropped out after 1 year, and were considered 1-year trainees thereafter. A higher proportion of 2-year trainees obtain positions within academic institutions (89.5 vs 67.2%, P = 0.0007), whereas a higher proportion of 1-year trainees obtain positions within non-academic institutions (29.3 vs 8.1%, P = 0.0007). A higher proportion of those with Canadian board certification in diagnostic radiology who completed a 2-year program obtained a position within a Canadian academic institution compared to non-certified 2-year trainees (P < 0.001). 71.4% of program directors agreed that a 1-year program was sufficient for non-academic staff positions. CONCLUSION: The length of the training program has significant impact on employment in academic vs non-academic institutions. This information can be used to guide the upcoming CBD initiative for neuroradiology programs.

LY75 Suppression in Mesenchymal Epithelial Ovarian Cancer Cells Generates a Stable Hybrid EOC Cellular Phenotype, Associated with Enhanced Tumor Initiation, Spreading and Resistance to Treatment in Orthotopic Xenograft Mouse Model.

The implications of the epithelial-mesenchymal transition (EMT) mechanisms in the initiation and progression of epithelial ovarian cancer (EOC) remain poorly understood. We have previously shown that suppression of the antigen receptor LY75 directs mesenchymal-epithelial transition (MET) in EOC cell lines with the mesenchymal phenotype, associated with the loss of Wnt/ß-catenin signaling activity. In the present study, we used the LY75-mediated modulation of EMT in EOC cells as a model in order to investigate in vivo the specific role of EOC cells, with an epithelial (E), mesenchymal (M) or mixed epithelial plus mesenchymal (E+M) phenotype, in EOC initiation, dissemination and treatment response, following intra-bursal (IB) injections of SKOV3-M (control), SKOV3-E (Ly75KD) and a mixed population of SKOV3-E+M cells, into severe combined immunodeficiency (SCID) mice. We found that the IB-injected SKOV3-E cells displayed considerably higher metastatic potential and resistance to treatment as compared to the SKOV3-M cells, due to the acquisition of a Ly75KD-mediated hybrid phenotype and stemness characteristics. We also confirmed in vivo that the LY75 depletion directs suppression of the Wnt/ß-catenin pathway in EOC cells, suggestive of a protective role of this pathway in EOC etiology. Moreover, our data raise concerns regarding the use of LY75-targeted vaccines for dendritic-cell EOC immunotherapy, due to the possible occurrence of undesirable side effects.

Cervical Internal Carotid Occlusion versus Pseudo-occlusion at CT Angiography in the Context of Acute Stroke: An Accuracy, Interobserver, and Intraobserver Agreement Study.

Purpose To evaluate the diagnostic accuracy and reliability of computed tomographic (CT) angiography to distinguish true cervical internal carotid artery (ICA) occlusion from pseudo-occlusion (defined as an isolated intracranial thrombus that impedes ascending blood flow) in the context of acute stroke. Materials and Methods This was a retrospective study of patients who underwent thrombectomy with preprocedural CT angiography that helps to demonstrate a lack of attenuation in the cervical ICA on the symptomatic side (24 men and 13 women; mean age, 63 years; age range, 30-86 years). Seven readers, including five neuroradiologists and two interventional neuroradiology fellows, independently reviewed the CT angiography images to assess whether there was true cervical ICA occlusion. Their results were compared with digital subtraction angiography (DSA) as the reference standard. Sensitivity and specificity for detecting true occlusion as well as accuracy and diagnostic odds ratio were calculated, with inter- and intraobserver κ statistics. Results Cervical ICA pseudo-occlusion occurred in 12 of 37 patients (32.4%) with nonattenuation of the cervical ICA on the symptomatic side. Interobserver agreement coefficients did not reach the substantial value of 0.61 for either pairs or groups of readers. The cohort's average sensitivity and specificity was 68% (95% confidence interval [CI]: 59%, 76%) and 75% (95% CI: 71%, 80%), respectively, with a diagnostic odds ratio of 8 (95% CI: 3, 18) and only fair interobserver agreement (κ = 0.32; 95% CI: 0.16, 0.47). Conclusion In the context of acute ischemic stroke with ipsilateral ICA nonattenuation at single-phase CT angiography, even specialized radiologists may not reliably distinguish true cervical occlusion from pseudo-occlusion. © RSNA, 2017 Online supplemental material is available for this article.

Follicle capacitation: a meta-analysis to investigate the transcriptome dynamics following follicle-stimulating hormone decline in bovine granulosa cells.

In recent years, exciting progress was made to improve the embryo outcome after ovarian stimulation in domestic animals. The practice of follicle-stimulating hormone (FSH) withdrawal, which is defined as the period of time between the last injection of FSH and oocyte retrieval, resulted in embryo yields significantly superior. Since then, specific changes in the transcriptome of granulosa cells were associated with the increase and also the decline in oocyte developmental competence following the FSH decline. In this study, we integrated large datasets from a public depository using a meta-analysis in order to elucidate the molecular changes occurring in granulosa cells following FSH decline in association with oocyte developmental competence. The meta-analysis revealed that the gene expression patterns observed during this period resulted from the downregulation of proliferative signals, and the upregulation of differentiation signals and early apoptotic signals. Additionally, FSH decline induced cellular hypoxia and triggered the expression of proinflammatory molecules which resulted in early atresia and mimicked the luteinizing hormone (LH) surge signaling to ovulation. To characterize this unique differentiation period, we suggest using the term "follicle capacitation" to refer to the functional changes occurring within the follicle in order to prepare the molecular machinery for the LH surge and ovulation following FSH decline. During this period, the follicle confers the oocyte with developmental competence to become a viable embryo. However, if this period is not rapidly followed by an LH surge, apoptosis signals are increased to generate follicular atresia and decrease oocyte quality.

Expression of atresia biomarkers in granulosa cells after ovarian stimulation in heifers.

The use of younger gamete donors in dairy cattle genetic selection programs significantly accelerates genetic gains by decreasing the interval between generations. Ovarian stimulation (OS) and the practice of follicle-stimulating hormone (FSH) withdrawal, also known as coasting, are intensively used in pre-pubertal heifers without detrimental effects on subsequent reproductive performance but generally with lower embryo yields. However, recent data from embryo transfer programs showed similar embryo yields in younger and sexually mature animals but with a significant difference in the coasting period. The aim of the present study was to identify a set of granulosa cell biomarkers capable of distinguishing optimal follicle differentiation from late differentiation and atresia in order to assess the differences in coasting dynamics between pre- and post-pubertal donors. We integrated transcriptomic data sets from a public depository and used vote counting meta-analysis in order to elucidate the molecular changes occurring in granulosa cells during late follicle differentiation and atresia. The meta-analysis revealed the gene expression associated with follicle demise, and most importantly, identified potential biomarkers of that status in bovine granulosa cells. The comparison of the expression of six biomarkers between pre- and post-pubertal donors revealed that younger donors had more signs of atresia after the same period of coasting. We found different follicular dynamics following coasting in younger donors. It is possible that younger donors are less capable to sustain follicular survival most likely due to insufficient luteinizing hormone signaling. In summary, the pre-pubertal status influences follicular dynamics and reduces the oocyte developmental competence curve following OS and FSH withdrawal in heifers.

Effect of heifer age on the granulosa cell transcriptome after ovarian stimulation.

Genomic selection is accelerating genetic gain in dairy cattle. Decreasing generation time by using younger gamete donors would further accelerate breed improvement programs. Although ovarian stimulation of peripubertal animals is possible and embryos produced in vitro from the resulting oocytes are viable, developmental competence is lower than when sexually mature cows are used. The aim of the present study was to shed light on how oocyte developmental competence is acquired as a heifer ages. Ten peripubertal Bos taurus Holstein heifers underwent ovarian stimulation cycles at the ages of 8, 11 (mean 10.8) and 14 (mean 13.7) months. Collected oocytes were fertilised in vitro with spermatozoa from the same adult male. Each heifer served as its own control. The transcriptomes of granulosa cells recovered with the oocytes were analysed using microarrays. Differential expression of certain genes was measured using polymerase chain reaction. Principal component analysis of microarray data revealed that the younger the animal, the more distinctive the gene expression pattern. Using ingenuity pathway analysis (IPA) and NetworkAnalyst (www.networkanalyst.ca), the main biological functions affected in younger donors were identified. The results suggest that cell differentiation, inflammation and apoptosis signalling are less apparent in peripubertal donors. Such physiological traits have been associated with a lower basal concentration of LH.

Steroidogenic genes expressions are repressed by high levels of leptin and the JAK/STAT signaling pathway in MA-10 Leydig cells.

The adipose tissue is an important endocrine organ secreting numerous peptide hormones, including leptin. Increased circulating levels of leptin, as a result of hormonal resistance in obese individuals, may contribute to lower androgen production in obese males. However, the molecular mechanisms involved need to be better defined. Androgens are mainly produced by Leydig cells within the testis. In male rodents, activation of the leptin receptor modulates a cascade of intracellular signal transduction pathways which may lead to regulation of transcription factors having influences on steroidogenesis in Leydig cells. Thus, as a result of high leptin levels interacting with its receptor and modulating the activity of the JAK/STAT signaling pathway, the activity of transcription factors important for steroidogenic genes expressions may be inhibited in Leydig cells. Here we show that Lepr is increasingly expressed within Leydig cells according to postnatal development. Although high levels of leptin (corresponding to obesity condition) alone had no effect on Leydig cells' steroidogenic genes expression, it downregulated cAMP-dependent activations of the cholesterol transporter Star and of the rate-limiting steroidogenic enzyme Cyp11a1. Our results suggest that STAT transcriptional activity is downregulated by high levels of leptin, leading to reduced cAMP-dependent steroidogenic genes (Star and Cyp11a1) expressions in MA-10 Leydig cells. However, other transcription factors such as members of the SMAD and NFAT families may be involved and need further investigation to better define how leptin regulates their activities and their relevance for Leydig cells function.

Comparative analysis of granulosa cell gene expression in association with oocyte competence in FSH-stimulated Holstein cows.

Ovarian stimulation with exogenous FSH followed by FSH withdrawal or 'coasting' is an effective means of increasing the number of oocytes obtainable for the in vitro production of cattle embryos. However, the quality of the oocytes thus obtained varies considerably from one cow to the next. The aim of the present study was to gain a better understanding of the follicular conditions associated with low oocyte developmental competence. Granulosa cells from 94 Holstein cows in a commercial embryo production facility were collected following ovarian stimulation and coasting. Microarray analysis showed 120 genes expressed with a differential of at least 1.5 when comparing donors of mostly competent with donors of mostly incompetent oocytes. Using ingenuity pathway analysis, we revealed the main biological functions and potential upstream regulators that distinguish donors of mostly incompetent oocytes. These are involved in cell proliferation, apoptosis, lipid metabolism, retinol availability and insulin signalling. In summary, we demonstrated that differences in follicle maturity at collection could explain differences in oocyte competence associated with individual animals. We also revealed deficiencies in lipid metabolism and retinol signalling in granulosa cells from donors of mostly incompetent oocytes.

Frailty in end-stage renal disease: comparing patient, caregiver, and clinician perspectives.

BACKGROUND: Frailty is associated with poor outcomes for patients on dialysis and is traditionally measured using tools that assess physical impairment. Alternate measurement tools highlight cognitive and functional domains, requiring clinician, patient, and/or caregiver input. In this study, we compared frailty measures for incident dialysis patients that incorporate patient, clinician, and caregiver perspectives with an aim to contrast the measured prevalence of frailty using tools derived from different conceptual frameworks. METHODS: A prospective cohort study of incident dialysis patients was conducted between February 2014 and June 2015. Frailty was assessed at dialysis onset using: 1) modified definition of Fried Phenotype (Dialysis Morbidity Mortality Study definition, DMMS); 2) Clinical Frailty Scale (CFS); 3) Frailty Assessment Care Planning Tool (provides CFS grading, FACT-CFS); and 4) Frailty Index (FI). Measures were compared via correlation and sensitivity/specificity analyses. RESULTS: A total of 98 patients participated (mean age of 61 ± 14 years). Participants were primarily Caucasian (91%), male (58%), and the majority started on hemodialysis (83%). The median score for both the CFS and FACT-CFS was 4 (interquartile range of 3-5). The mean FI score was 0.31 (standard deviation ± 0.16). The DMMS identified 78% of patients as frail. The FACT-CFS demonstrated highest correlation (r = 0.71) with the FI, while the DMMS was most sensitive (97%, 100%) and a CFS ≥ 5 most specific (100%, 77%) at corresponding FI cutoff values (>0.21, >0.45). CONCLUSIONS: Frailty assessments of incident dialysis patients that include clinician, caregiver and patient perspectives have moderate to strong correlation with the FI. At specified FI cutoff values, the FACT-CFS and DMMS are highly sensitive measures of frailty. The CFS and FACT-CFS may represent viable alternative screening tools in dialysis patients.

Transcriptome meta-analysis of three follicular compartments and its correlation with ovarian follicle maturity and oocyte developmental competence in cows.

Oocyte developmental competence in superstimulated cows is dependent in part on the duration of the FSH coasting. FSH coasting refers to superstimulation with FSH (2 days of endogenous FSH following follicle ablation and 3 days of FSH injections) followed by no FSH for a specific duration. The optimal duration varies among individuals. FSH coasting appears to modulate the transcriptome of different follicular compartments, which cooperate as a single functional unit. However, the integrative effects of FSH coasting on different follicular compartments remain ambiguous. Meta-analysis of three independent transcriptome studies, each focused on a single cell type (granulosa, cumulus, and oocyte) during FSH coasting, allowed the identification of 12 gene clusters with similar time-course expression patterns in all three compartments. Network analysis identified HNF4A (involved in metabolic functions) and ELAVL1 (an RNA-binding protein) as hub genes regulated respectively upward and downward in the clusters enriched at the optimal coasting time, and APP (involved in mitochondrial functions) and COPS5 (a member of the COP9 signalosome) as hub genes regulated respectively upwards and downwards in the clusters enriched progressively throughout the coasting period. We confirmed the effects on HNF4A downstream targets (TTR, PPL) and other hub genes (ELAVL1, APP, MYC, and PGR) in 30 cows with RT-quantitative PCR. The correlation of hub gene expression levels with FSH coasting indicated that a combination of these genes could predict oocyte competence with 83% sensitivity, suggesting that they are potential biomarkers of follicle differentiation. These findings could be used to optimize FSH coasting on an individual basis.

Human-impacted areas of France are environmental reservoirs of the Pseudallescheria boydii/Scedosporium apiospermum species complex.

Species of the Pseudallescheria boydii/Scedosporium apiospermum complex (PSC) are emerging fungal pathogens able to chronically colonize the airways of patients with cystic fibrosis (CF). As P. boydii was found more frequently colonizing the lungs of CF patients in France than in other European countries in a previous report, the present study was conducted in order to clarify distribution of PSC species in France and to characterize their natural habitat. The highest densities of PSC isolates were found in human-impacted areas, i.e. agricultural areas, fluids obtained from wastewater treatment plants, playgrounds and industrial areas. PSC was not detected from soil samples collected in forests. Most PSC culture-positive soil samples exhibited a pH range of 6-8. Scedosporium dehoogii, the most abundant species, was detected in all human-impacted area types except vineyards, whereas Scedosporium aurantiacum was mostly found in agricultural areas. Pseudallescheria boydii and S. apiospermum were predominantly isolated from seashores and playgrounds respectively. Pseudallescheria minutispora was found only once from a playground. This study highlights potential sources of contamination of the patients, especially in the CF context.

DRESS syndrome: cerebral vasculitic-like presentation.

INTRODUCTION: DRESS (drug reaction with eosinophilia and systemic symptoms) syndrome is a severe adverse drug-induced reaction. It manifests with pyrexia, eosinophilia, and lymphadenopathy, with multiple organ involvement, mainly the skin, liver, and kidneys. The purpose of this article is to demonstrate that DRESS syndrome can be associated with cerebral manifestations, a concept not well known in the neuroradiological literature. METHODS: We describe three cases of DRESS syndrome associated with cerebral vasculitic-like lesions and realize a review of the literature to demonstrate that this association represents a very rare entity. RESULTS: Acute ischemic lesions were found among two patients. In all cases, perivascular enhancement was present. Magnetic resonance angiography (MRA) sequence was normal. Although no cerebral biopsy was performed, this enhancement pattern is strongly suggestive of a vasculitic process associated with DRESS syndrome. CONCLUSION: Diagnosis of cerebral vasculitic-like associated lesions must be considered in patients with DRESS syndrome since it can be reversed completely by withdrawing the causal medication and instigating corticosteroid treatment in a timely fashion.

Dilatation of the bile duct in patients after cholecystectomy: a retrospective study.

PURPOSE: Retrospective assessment of impact of cholecystectomy, age, and sex on bile duct (BD) diameter. MATERIALS AND METHODS: We retrospectively reviewed abdominal contrast-enhanced multidetector computed tomography and laboratory reports of 290 consecutive patients (119 men; mean age, 55.9 years) who presented without cholestasis to the emergency department of our institution between June 2009 and August 2010. BD diameters were measured in 3 locations, by 2 independent observers, twice, at 1-month intervals. Reproducibility and agreement were evaluated by intraclass correlation coefficients and Bland-Altman analyses. The effects of cholecystectomy, age, and sex on BD diameter were analysed with linear mixed models. RESULTS: BD diameter inter-reader reproducibility and agreement were excellent at the level of the right hepatic artery (intraclass correlation coefficient, 0.94). Sixty-one patients (21.0%) had a history of cholecystectomy. Among them, the 95th percentile of BD diameters at hepatic artery level was 7.9 mm (<50 years) and 12.3 mm (≥50 years). Among those without cholecystectomy, BD diameter was 6.2 mm (<50 years) and 7.7 mm (≥50 years). Cholecystectomy was associated with significantly larger BD diameters in both age groups (P < .001). Older age was associated with larger BD diameters (P = .004). Sex had no impact on BD diameter (P = .842). CONCLUSION: Patients after cholecystectomy may present with an enlarged BD unrelated to cholestasis. The BD diameter increases with age. Clinicians should rely on cholecystectomy status, age, and laboratory results to determine needs of further investigation.

A receptor required for chitin perception facilitates arbuscular mycorrhizal associations and distinguishes root symbiosis from immunity.

Plants establish symbiotic associations with arbuscular mycorrhizal fungi (AMF) to facilitate nutrient uptake, particularly in nutrient-limited conditions. This partnership is rooted in the plant's ability to recognize fungal signaling molecules, such as chitooligosaccharides (chitin) and lipo-chitooligosaccharides. In the legume Medicago truncatula, chitooligosaccharides trigger both symbiotic and immune responses via the same lysin-motif-receptor-like kinases (LysM-RLKs), notably CERK1 and LYR4. The nature of plant-fungal engagement is opposite according to the outcomes of immunity or symbiosis signaling, and as such, discrimination is necessary, which is challenged by the dual roles of CERK1/LYR4 in both processes. Here, we describe a LysM-RLK, LYK8, that is functionally redundant with CERK1 for mycorrhizal colonization but is not involved in chitooligosaccharides-induced immunity. Genetic mutation of both LYK8 and CERK1 blocks chitooligosaccharides-triggered symbiosis signaling, as well as mycorrhizal colonization, but shows no further impact on immunity signaling triggered by chitooligosaccharides, compared with the mutation of CERK1 alone. LYK8 interacts with CERK1 and forms a receptor complex that appears essential for chitooligosaccharides activation of symbiosis signaling, with the lyk8/cerk1 double mutant recapitulating the impact of mutations in the symbiosis signaling pathway. We conclude that this novel receptor complex allows chitooligosaccharides activation specifically of symbiosis signaling and helps the plant to differentiate between activation of these opposing signaling processes.

The grapevine LysM receptor-like kinase VvLYK5-1 recognizes chitin oligomers through its association with VvLYK1-1.

The establishment of defense reactions to protect plants against pathogens requires the recognition of invasion patterns (IPs), mainly detected by plasma membrane-bound pattern recognition receptors (PRRs). Some IPs, also termed elicitors, are used in several biocontrol products that are gradually being developed to reduce the use of chemicals in agriculture. Chitin, the major component of fungal cell walls, as well as its deacetylated derivative, chitosan, are two elicitors known to activate plant defense responses. However, recognition of chitooligosaccharides (COS) in Vitis vinifera is still poorly understood, hampering the improvement and generalization of protection tools for this important crop. In contrast, COS perception in the model plant Arabidopsis thaliana is well described and mainly relies on a tripartite complex formed by the cell surface lysin motif receptor-like kinases (LysM-RLKs) AtLYK1/CERK1, AtLYK4 and AtLYK5, the latter having the strongest affinity for COS. In grapevine, COS perception has for the moment only been demonstrated to rely on two PRRs VvLYK1-1 and VvLYK1-2. Here, we investigated additional players by overexpressing in Arabidopsis the two putative AtLYK5 orthologs from grapevine, VvLYK5-1 and VvLYK5-2. Expression of VvLYK5-1 in the atlyk4/5 double mutant background restored COS sensitivity, such as chitin-induced MAPK activation, defense gene expression, callose deposition and conferred non-host resistance to grapevine downy mildew (Erysiphe necator). Protein-protein interaction studies conducted in planta revealed a chitin oligomer-triggered interaction between VvLYK5-1 and VvLYK1-1. Interestingly, our results also indicate that VvLYK5-1 mediates the perception of chitin but not chitosan oligomers showing a part of its specificity.

Incidence and predictive factors of retropharyngeal lymph node metastases in patients with oropharyngeal cancer undergoing multimodality treatment planning imaging.

BACKGROUND: We investigated the incidence and predictive factors of retropharyngeal lymph node (RPLN) metastases in patients with oropharyngeal cancer (OPC) undergoing multimodality treatment planning imaging before radiotherapy. METHODS: Consecutive patients with OPC treated with curative-intent radiotherapy from 2017 to 2019 were retrospectively analyzed. Treatment planning comprised contrast-enhanced computed tomography (CT), magnetic resonance imaging (MRI), and fluorodeoxyglucose-positron emission tomography (FDG-PET) unless contraindicated. RESULTS: Of 300 patients, 66 (22%) had radiological evidence of RPLN involvement on planning images, compared to 17 (6%) on diagnostic CT alone. On multivariate analysis, RPLN involvement was statistically (p < 0.05) associated with tonsil, soft palate, and posterior pharyngeal wall primaries, and with disease extension to the soft palate or vallecula. CONCLUSIONS: Multimodality treatment planning imaging reveals a high rate of RPLN metastases from OPC compared to diagnostic CT alone. Patients with tonsil, soft palate, or posterior pharyngeal wall primaries or disease extending to the soft palate or vallecula appear at higher risk.