RESUMEN
The control of stem canker disease of Brassica napus (rapeseed), caused by the fungus Leptosphaeria maculans is based largely on plant genetic resistance: single-gene specific resistance (Rlm genes) or quantitative, polygenic, adult-stage resistance. Our working hypothesis was that quantitative resistance partly obeys the gene-for-gene model, with resistance genes 'recognizing' fungal effectors expressed during late systemic colonization. Five LmSTEE (stem-expressed effector) genes were selected and placed under the control of the AvrLm4-7 promoter, an effector gene highly expressed at the cotyledon stage of infection, for miniaturized cotyledon inoculation test screening of a gene pool of 204 rapeseed genotypes. We identified a rapeseed genotype, 'Yudal', expressing hypersensitive response to LmSTEE98. The LmSTEE98-RlmSTEE98 interaction was further validated by inactivation of the LmSTEE98 gene with a CRISPR-Cas9 approach. Isolates with mutated versions of LmSTEE98 induced more severe stem symptoms than the wild-type isolate in 'Yudal'. This single-gene resistance was mapped in a 0.6 cM interval of the 'Darmor_bzh' × 'Yudal' genetic map. One typical gene-for-gene interaction contributes partly to quantitative resistance when L. maculans colonizes the stems of rapeseed. With numerous other effectors specific to stem colonization, our study provides a new route for resistance gene discovery, elucidation of quantitative resistance mechanisms and selection for durable resistance.
Asunto(s)
Ascomicetos , Brassica napus , Resistencia a la Enfermedad , Enfermedades de las Plantas , Ascomicetos/genética , Ascomicetos/patogenicidad , Brassica napus/genética , Brassica napus/microbiología , Cotiledón , Resistencia a la Enfermedad/genética , Genes de Plantas , Enfermedades de las Plantas/genética , Enfermedades de las Plantas/microbiologíaRESUMEN
In phytopathogenic fungi, the expression of hundreds of small secreted protein (SSP)-encoding genes is induced upon primary infection of plants while no or a low level of expression is observed during vegetative growth. In some species such as Leptosphaeria maculans, this coordinated in-planta upregulation of SSP-encoding genes expression relies on an epigenetic control but the signals triggering gene expression in-planta are unknown. In the present study, biotic and abiotic factors that may relieve suppression of SSP-encoding gene expression during axenic growth of L. maculans were investigated. Some abiotic factors (temperature, pH) could have a limited effect on SSP gene expression. In contrast, two types of cellular stresses induced by antibiotics (cycloheximide, phleomycin) activated strongly the transcription of SSP genes. A transcriptomic analysis to cycloheximide exposure revealed that biological processes such as ribosome biosynthesis and rRNA processing were induced whereas important metabolic pathways such as glycogen and nitrogen metabolism, glycolysis and tricarboxylic acid cycle activity were down-regulated. A quantitatively different expression of SSP-encoding genes compared to plant infection was also detected. Interestingly, the same physico-chemical parameters as those identified here for L. maculans effectors were identified to regulate positively or negatively the expression of bacterial effectors. This suggests that apoplastic phytopathogens may react to similar physiological parameters for regulation of their effector genes.
Asunto(s)
Ascomicetos/genética , Proteínas Fúngicas/biosíntesis , Enfermedades de las Plantas/microbiología , Estrés Fisiológico/genética , Ascomicetos/crecimiento & desarrollo , Proteínas Fúngicas/genética , Perfilación de la Expresión Génica , Regulación Fúngica de la Expresión Génica , Hojas de la Planta/microbiologíaRESUMEN
Project Orbis was initiated in May 2019 by the Oncology Center of Excellence to facilitate faster patient access to innovative cancer therapies by providing a framework for concurrent submissions and review of oncology products among international partners. Since its inception, Australia's Therapeutic Goods Administration (TGA), Canada's Health Canada (HC), Singapore's Health Sciences Authority (HSA), Switzerland's Swissmedic (SMC), Brazil's National Health Surveillance Agency (ANVISA), United Kingdom's Medicines and Healthcare Products Regulatory Agency (MHRA), and most recently Israel's Ministry of Health (IMoH) Medical Technologies, Health Information, Innovation and Research (MTIIR) Directorate, have joined Project Orbis. While each country has its own expedited review pathways to bring promising therapies to patients, there are some similarities and differences in pathways and timelines. FDA's fast-track designation and MHRA's marketing authorization under exceptional circumstances (MAEC) allow non-clinical and limited clinical evidence to support approval under these programs. HC's Extraordinary Use New Drug (EUND) pathway allows granting exceptional use authorization with limited clinical evidence. ANVISA, HSA, MTIIR, and TGA do not have standard pathways that allow non-clinical evidence and limited clinical evidence. While there is no definite regulatory pathway for HSA, the current framework for approval does allow flexibility in the type of data (non-clinical or clinical) required to demonstrate the benefit-risk profile of a product. HSA may register a product if the agency is satisfied that the overall benefit outweighs the risk. All Project Orbis Partner (POP) countries have similar programs to the FDA accelerated approval program except ANVISA. Although HSA and MTIIR do not have defined pathways for accelerated approval programs, there are opportunities to request accelerated approval per these agencies. All POP countries have pathways like the FDA priority review except MHRA. Priority review timelines for new drugs range from 120 to 264 calendar days (cd). Standard review timelines for new drugs range from 180 to 365 cd.
Asunto(s)
Medicina , Neoplasias , Estados Unidos , Humanos , Aprobación de Drogas , United States Food and Drug Administration , CanadáRESUMEN
Osteoarthritis, the leading cause of chronic joint pain, is studied through different animal models, but none of them is ideal in terms of reliability and translational value. In this pilot study of female rats, 3 surgical models of osteoarthritic pain, i.e., destabilization of the medial meniscus (DMM), cranial cruciate ligament transection (CCLT), and the combination of both surgical models (COMBO) and 1 chemical model [intra-articular injection of monosodium iodoacetate (MIA)] were compared for their impact on functional pain outcomes [static weight-bearing (SWB) and punctate tactile paw withdrawal threshold (PWT)] and spinal neuropeptides [substance P (SP), calcitonin gene-related peptide (CGRP), bradykinin (BK), and somatostatin (SST)]. Six rats were assigned to each model group and a sham group. Both the chemical model (MIA) and surgical COMBO model induced functional alterations in SWB and PWT, with the changes being more persistent in the surgical combination group. Both models also produced an increase in levels of pro-nociceptive and anti-nociceptive neuropeptides at different timepoints. Pain comparison with the MIA model showed the advantage of a surgical model, especially the combination of the DMM and CCLT models, whereas each surgical model alone only led to temporary functional alterations and no change in neuropeptidomics.
L'arthrose, la principale cause de douleur chronique articulaire, est étudiée à travers différents modèles animaux, mais aucun d'eux n'est idéal en termes de fiabilité et de valeur translationnelle. Trois modèles chirurgicaux de douleur arthrosique, c'est-à-dire, la déstabilisation du ménisque médial, la transsection du ligament croisé crânial et la combinaison des deux, ainsi qu'un modèle chimique (injection intraarticulaire de mono-iodoacétate de sodium) ont été comparés dans cette étude pilote chez des rattes quant à leurs impacts sur les évaluations fonctionnelles de la douleur (distribution pondérale statique, évaluation ponctuelle de l'allodynie tactile) et les neuropeptides spinaux (substance P, peptide relié au gène de la calcitonine, bradykinine et somatostatine). Six rats ont été assignés à chacun des modèles et un groupe Sham. Autant le modèle du mono-iodoacétate de sodium que celui de la combinaison chirurgicale ont tous les deux induits des altérations fonctionnelles de la distribution pondérale statique et du seuil de retrait de la patte suite à une stimulation ponctuelle tactile, mais avec des changements plus persistants dans le groupe de la combinaison chirurgicale. Ces deux modèles ont également engendré une augmentation des niveaux en neuropeptides pro-nociceptifs et anti-nociceptifs à différents moments. Un intérêt du modèle chirurgical a été démontré suite à la comparaison de la douleur avec le modèle du mono-iodoacétate de sodium, en particulier la déstabilisation du ménisque médial combinée à la transsection du ligament croisé crânial, tandis que les inductions chirurgicales unique entraînaient des altérations fonctionnelles temporaires avec aucun changement neuropeptidomique.(Traduit par les auteurs).
Asunto(s)
Modelos Animales de Enfermedad , Regulación de la Expresión Génica/fisiología , Neuropéptidos/metabolismo , Osteoartritis/etiología , Dolor/metabolismo , Animales , Femenino , Inyecciones Intraarticulares , Dimensión del Dolor , Proyectos Piloto , Distribución Aleatoria , Ratas , Ratas Sprague-Dawley , Soporte de PesoRESUMEN
Leptosphaeria maculans, the causal agent of stem canker disease, colonizes oilseed rape (Brassica napus) in two stages: a short and early colonization stage corresponding to cotyledon or leaf colonization, and a late colonization stage during which the fungus colonizes systemically and symptomlessly the plant during several months before stem canker appears. To date, the determinants of the late colonization stage are poorly understood; L. maculans may either successfully escape plant defences, leading to stem canker development, or the plant may develop an 'adult-stage' resistance reducing canker incidence. To obtain an insight into these determinants, we performed an RNA-sequencing (RNA-seq) pilot project comparing fungal gene expression in infected cotyledons and in symptomless or necrotic stems. Despite the low fraction of fungal material in infected stems, sufficient fungal transcripts were detected and a large number of fungal genes were expressed, thus validating the feasibility of the approach. Our analysis showed that all avirulence genes previously identified are under-expressed during stem colonization compared with cotyledon colonization. A validation RNA-seq experiment was then performed to investigate the expression of candidate effector genes during systemic colonization. Three hundred and seven 'late' effector candidates, under-expressed in the early colonization stage and over-expressed in the infected stems, were identified. Finally, our analysis revealed a link between the regulation of expression of effectors and their genomic location: the 'late' effector candidates, putatively involved in systemic colonization, are located in gene-rich genomic regions, whereas the 'early' effector genes, over-expressed in the early colonization stage, are located in gene-poor regions of the genome.
Asunto(s)
Ascomicetos/genética , Brassica napus/microbiología , Cotiledón/microbiología , Tallos de la Planta/microbiología , Recuento de Colonia Microbiana , Regulación hacia Abajo/genética , Perfilación de la Expresión Génica , Regulación Fúngica de la Expresión Génica , Ontología de Genes , Genes Fúngicos , ARN Mensajero/genética , ARN Mensajero/metabolismo , Reproducibilidad de los Resultados , Análisis de Secuencia de ARN , Regulación hacia Arriba/genéticaRESUMEN
OBJECTIVES: To assess the potential of a new single-session surgical procedure, the overlapping pubic and ischiatic osteotomy (OPIO) for modification of bilateral hip conformation. We hypothesized that OPIO would be simple to perform with currently available surgical equipment, through a single surgical approach, with minimal potential morbidity, and that it would allow adequate simultaneous bilateral improvement of coxofemoral joint conformation in patients at risk of canine hip dysplasia. METHODS: The OPIO procedure was performed in the pelves of five large breed canine cadavers. Computed tomography images of each cadaver were compared by measurement of the dorsal acetabular rim angle (DARA), acetabular angle (AA), dorso-ventral sacroiliac ratio (SR), and pubic inlet area before and after OPIO. RESULTS: Coxofemoral joint conformation was improved after OPIO. Postoperative DARA was significantly decreased (mean: -5.09°) and AA was significantly increased (mean: 3.54°) after OPIO. The SR was not significantly different after OPIO, indicating minimal impact on the sacro-illiac joints by the procedure. Pubic inlet dimensions and area were significantly decreased after OPIO, but the overall effect on pelvic inlet area was clinically insignificant. CLINICAL SIGNIFICANCE: An OPIO allows some improvement of coxofemoral joint conformation in canine cadavers.
Asunto(s)
Acetábulo/cirugía , Enfermedades de los Perros/cirugía , Luxación de la Cadera/veterinaria , Osteotomía/veterinaria , Animales , Perros , Luxación de la Cadera/cirugía , Osteotomía/métodos , Hueso Púbico/cirugíaRESUMEN
BACKGROUND: Lack of validity in osteoarthritis pain models and assessment methods is suspected. Our goal was to 1) assess the repeatability and reproducibility of measurement and the influence of environment, and acclimatization, to different pain assessment outcomes in normal rats, and 2) test the concurrent validity of the most reliable methods in relation to the expression of different spinal neuropeptides in a chemical model of osteoarthritic pain. METHODS: Repeatability and inter-rater reliability of reflexive nociceptive mechanical thresholds, spontaneous static weight-bearing, treadmill, rotarod, and operant place escape/avoidance paradigm (PEAP) were assessed by the intraclass correlation coefficient (ICC). The most reliable acclimatization protocol was determined by comparing coefficients of variation. In a pilot comparative study, the sensitivity and responsiveness to treatment of the most reliable methods were tested in the monosodium iodoacetate (MIA) model over 21 days. Two MIA (2 mg) groups (including one lidocaine treatment group) and one sham group (0.9 % saline) received an intra-articular (50 µL) injection. RESULTS: No effect of environment (observer, inverted circadian cycle, or exercise) was observed; all tested methods except mechanical sensitivity (ICC <0.3), offered good repeatability (ICC ≥0.7). The most reliable acclimatization protocol included five assessments over two weeks. MIA-related osteoarthritic change in pain was demonstrated with static weight-bearing, punctate tactile allodynia evaluation, treadmill exercise and operant PEAP, the latter being the most responsive to analgesic intra-articular lidocaine. Substance P and calcitonin gene-related peptide were higher in MIA groups compared to naive (adjusted P (adj-P) = 0.016) or sham-treated (adj-P = 0.029) rats. Repeated post-MIA lidocaine injection resulted in 34 times lower downregulation for spinal substance P compared to MIA alone (adj-P = 0.029), with a concomitant increase of 17 % in time spent on the PEAP dark side (indicative of increased comfort). CONCLUSION: This study of normal rats and rats with pain established the most reliable and sensitive pain assessment methods and an optimized acclimatization protocol. Operant PEAP testing was more responsive to lidocaine analgesia than other tests used, while neuropeptide spinal concentration is an objective quantification method attractive to support and validate different centralized pain functional assessment methods.
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Artritis Experimental , Neuropéptidos/análisis , Osteoartritis , Dimensión del Dolor/métodos , Proteómica/métodos , Aclimatación/fisiología , Animales , Artritis Experimental/complicaciones , Artritis Experimental/metabolismo , Cromatografía Líquida de Alta Presión , Condicionamiento Operante , Femenino , Ácido Yodoacético/toxicidad , Osteoartritis/complicaciones , Osteoartritis/metabolismo , Dolor/etiología , Umbral del Dolor , Ratas , Ratas Sprague-Dawley , Reproducibilidad de los Resultados , Espectrometría de Masas en TándemRESUMEN
PURPOSE: This paper presents the results of a qualitative study on the values underlying the decision-making process of an interdisciplinary team working in a work rehabilitation facility of a Québec teaching hospital. METHODS: In order to document the values underlying the decision-making process, a single case observational study was conducted. Interdisciplinary team weekly discussions on ongoing cases of 22 workers absent from work due to musculoskeletal disorders were videotaped. All discourses were transcribed and analyzed following an inductive and iterative approach. The values identified were validated by feedback from team members. RESULTS: Ten common decision values emerged from the data: (1) team unity and credibility, (2) collaboration with stakeholders, (3) worker's internal motivation, (4) worker's adherence to the program, (5) worker's reactivation, (6) single message, (7) reassurance, (8) graded intervention, (9) pain management and (10) return to work as a therapy. The analysis of these values led to the design of a model describing interrelations between them. CONCLUSIONS: This study throws light on some mechanisms underlying the decisions made by the team and determining its action. This improves understanding of the actions taken by an interdisciplinary team in work rehabilitation and may facilitate knowledge transfer in the training of other teams.
Asunto(s)
Toma de Decisiones , Enfermedades Musculoesqueléticas/rehabilitación , Enfermedades Profesionales/rehabilitación , Planificación de Atención al Paciente , Grupo de Atención al Paciente , Femenino , Humanos , Masculino , Quebec , Valores Sociales , Grabación de Cinta de VideoRESUMEN
INTRODUCTION: Various obstacles to and facilitators of collaboration between an interdisciplinary work rehabilitation team and the stakeholders (workers, insurers, physicians, and employers) exist, but are not well characterized. METHODS: An observational study was conducted, using videotapes of interdisciplinary team discussions of ongoing cases involving 22 workers absent from work due to musculoskeletal disorders. The actions taken and strategies adopted by the team in an effort to overcome the obstacles to collaboration were studied. RESULTS: Various factors influence collaboration between the rehabilitation team and the stakeholders. In general, stakeholder endorsement of the team's therapeutic principles and confidence in their approach emerged as particularly important factors. Diverse strategies, most often, education and awareness-raising, were used by the team to foster collaboration among the parties. CONCLUSIONS: This study provides greater insight into the factors affecting collaboration among a rehabilitation team, an injured worker and other stakeholders. The results may improve understanding of the actions taken by rehabilitation teams and help to optimize their practices.