The SARS-CoV-2 viral load in COVID-19 patients is lower on face mask filters than on nasopharyngeal swabs.

Face masks and personal respirators are used to curb the transmission of SARS-CoV-2 in respiratory droplets; filters embedded in some personal protective equipment could be used as a non-invasive sample source for applications, including at-home testing, but information is needed about whether filters are suited to capture viral particles for SARS-CoV-2 detection. In this study, we generated inactivated virus-laden aerosols of 0.3-2 microns in diameter (0.9 µm mean diameter by mass) and dispersed the aerosolized viral particles onto electrostatic face mask filters. The limit of detection for inactivated coronaviruses SARS-CoV-2 and HCoV-NL63 extracted from filters was between 10 to 100 copies/filter for both viruses. Testing for SARS-CoV-2, using face mask filters and nasopharyngeal swabs collected from hospitalized COVID-19-patients, showed that filter samples offered reduced sensitivity (8.5% compared to nasopharyngeal swabs). The low concordance of SARS-CoV-2 detection between filters and nasopharyngeal swabs indicated that number of viral particles collected on the face mask filter was below the limit of detection for all patients but those with the highest viral loads. This indicated face masks are unsuitable to replace diagnostic nasopharyngeal swabs in COVID-19 diagnosis. The ability to detect nucleic acids on face mask filters may, however, find other uses worth future investigation

REPONSES ADAPTATIVES D'ALTERNARIA BRASSICICOLA AU STRESS OXYDATIF LORS DE L'INTERACTION AVEC LES BRASSICACEES. Rôle du métabolisme du mannitol et des Glutathion-S-transférases

The induction of oxidative stress during infection plays a crucial role in host defense mechanisms. , The plant family Brassicaceae includes many important crop plants and the model species Arabidospsis thaliana. Within this plant family, the oxidative stress can be generated either by reactive oxygen species (ROS) released around the infected area during the oxidative burst, or by plant secondary metabolites, such as isothiocyanates (ITC). The necrotrophic fungus Alternaria brassicicola is the causative agent of black spot disease of Bassicaceae. Despite this host defense system, this fungus is still able to complete its infectious disease cycle, indicating the existence of strategies to cope with this oxidative. In this study, we showed that oxidative stress tolerance of A. brassicicola was partly dependent on the mannitol metabolism, which was involved in conidial resistance during the in planta oxidative burst, and in protection against intracellular damages caused by ITC during the early step of infection. Our results also supported the potential involvement of Glutathion-S-transférase enzymes in ITC detoxification mechanisms. Alteration of mannitol metabolism or inactivation of selected GST resulted in reduced virulence indicating that their targeting would be an attractive option for antifungal strategies.L'induction par un hôte d'un stress oxydatif chez un parasite représente l'une des stratégies de défense chimique les plus répandues. Chez les Brassicacées, famille de plantes comprenant des espèces à fort intérêt agronomique comme le chou ou le colza, ou d'intérêt scientifique comme Arabidopsis thaliana, le stress oxydatif peut être généré soit par des formes actives d'oxygène libérées au niveau de la zone d'agression lors du burst oxydatif soit par des molécules issues du métabolisme secondaire de la plante telles que les isothiocyanates (ITC). Bien que fortement exposé à un tel stress lors de l'infection, Alternaria brassicicola, champignon pathogène nécrotrophe inféodé aux Brassicacées, est capable d'accomplir son cycle parasitaire et donc de s'y adapter. Cette thèse met en évidence que la tolérance du champignon aux stress oxydatif est partiellement dépendante du métabolisme du mannitol qui participe à la résistance des conidies lors du burst oxydatif in planta ou à la protection contre les dommages intracellulaires des FAO générées par les ITC lors des phases précoces de l'interaction. Les travaux réalisés montrent également que la charge fongitoxique des dérivés des glucosinolates peut être inhibée par leur conjugaison au glutathion via des Glutathion-Stransférases (GST) spécifiques. L'altération du métabolisme du mannitol ou l'inactivation de ces GST diminue significativement le pouvoir pathogène d'A. brassicicola et la régulation de ces voies métaboliques est donc à considérer dans l'optique de développer de nouvelles stratégies de protection des cultures

Réponses adaptatives d'Alternaria Brassicicola au stress oxydatif lors de l'interaction avec les brassicacees (rôle du métabolisme du mannitol et des Glutathion-S-transférases)

L'induction par un hôte d'un stress oxydatif chez un parasite représente l'une des stratégies de défense chimique les plus répandues. Chez les Brassicacées, famille de plantes comprenant des espèces à fort intérêt agronomique comme le chou ou le colza, ou d'intérêt scientifique comme Arabidopsis thaliana, le stress oxydatif peut être généré soit par des formes actives d'oxygène libérées au niveau de la zone d'agression lors du burst oxydatif soit par des molécules issues du métabolisme secondaire de la plante telles que les isothiocyanates (ITC). Bien que fortement exposé à un tel stress lors de l'infection, Alternaria brassicicola, champignon pathogène nécrotrophe inféodé aux Brassicacées, est capable d'accomplir son cycle parasitaire et donc de s'y adapter. Cette thèse met en évidence que la tolérance du champignon aux stress oxydatif est partiellement dépendante du métabolisme du mannitol qui participe à la résistance des conidies lors du burst oxydatif in planta ou à la protection contre les dommages intracellulaires des FAO générées par les ITC lors des phases précoces de l'interaction. Les travaux réalisés montrent également que la charge fongitoxique des dérivés des glucosinolates peut être inhibée par leur conjugaison au glutathion via des Glutathion-Stransférases (GST) spécifiques. L'altération du métabolisme du mannitol ou l'inactivation de ces GST diminue significativement le pouvoir pathogène d'A. brassicicola et la régulation de ces voies métaboliques est donc à considérer dans l'optique de développer de nouvelles stratégies de protection des cultures.The induction of oxidative stress during infection plays a crucial role in host defense mechanisms. , The plant family Brassicaceae includes many important crop plants and the model species Arabidospsis thaliana. Within this plant family, the oxidative stress can be generated either by reactive oxygen species (ROS) released around the infected area during the oxidative burst, or by plant secondary metabolites, such as isothiocyanates (ITC). The necrotrophic fungus Alternaria brassicicola is the causative agent of black spot disease of Bassicaceae. Despite this host defense system, this fungus is still able to complete its infectious disease cycle, indicating the existence of strategies to cope with this oxidative. In this study, we showed that oxidative stress tolerance of A. brassicicola was partly dependent on the mannitol metabolism, which was involved in conidial resistance during the in planta oxidative burst, and in protection against intracellular damages caused by ITC during the early step of infection. Our results also supported the potential involvement of Glutathion-S-transférase enzymes in ITC detoxification mechanisms. Alteration of mannitol metabolism or inactivation of selected GST resulted in reduced virulence indicating that their targeting would be an attractive option for antifungal strategies.ANGERS-BU Lettres et Sciences (490072106) / SudocSudocFranceF

Impact of the UPR on the virulence of the plant fungal pathogen [i]A. brassicicola[/i]

The fungal genus [i]Alternaria[/i] contains many destructive plant pathogens, including [i]Alternaria brassicicola[/i], which causes black spot disease on a wide range of Brassicaceae plants and which is routinely used as a model necrotrophic pathogen in studies with [i]Arabidopsis thaliana[/i]. During host infection, many fungal proteins that are critical for disease progression are processed in the endoplasmic reticulum (ER)/Golgi system and secreted in planta. The unfolded protein response (UPR) is an essential part of ER protein quality control that ensures efficient maturation of secreted and membrane-bound proteins in eukaryotes. This review highlights the importance of the UPR signaling pathway with respect to the ability of [i]A. brassicicola[/i] to efficiently accomplish key steps of its pathogenic life cycle. Understanding the pathogenicity mechanisms that fungi uses during infection is crucial for the development of new antifungal therapies. Therefore the UPR pathway has emerged as a promising drug target for plant disease control

Data from: Characterization of glutathione transferases involved in the pathogenicity of Alternaria brassicicola

Background: Glutathione transferases (GSTs) represent an extended family of multifunctional proteins involved in detoxification processes and tolerance to oxidative stress. We thus anticipated that some GSTs could play an essential role in the protection of fungal necrotrophs against plant-derived toxic metabolites and reactive oxygen species that accumulate at the host-pathogen interface during infection. Results: Mining the genome of the necrotrophic Brassica pathogen Alternaria brassicicola for glutathione transferase revealed 23 sequences, 17 of which could be clustered into the main classes previously defined for fungal GSTs and six were ‘orphans’. Five isothiocyanate-inducible GSTs from five different classes were more thoroughly investigated. Analysis of their catalytic properties revealed that two GSTs, belonging to the GSTFuA and GTT1 classes, exhibited GSH transferase activity with isothiocyanates (ITC) and peroxidase activity with cumene hydroperoxide, respectively. Mutant deficient for these two GSTs were however neither more susceptible to ITC nor less aggressive than the wild-type parental strain. By contrast mutants deficient for two other GSTs, belonging to the Ure2pB and GSTO classes, were distinguished by their hyper-susceptibility to ITC and low aggressiveness against Brassica oleracea. In particular AbGSTO1 could participate in cell tolerance to ITC due to its glutathione-dependent thioltransferase activity. The fifth ITC-inducible GST belonged to the MAPEG class and although it was not possible to produce the soluble active form of this protein in a bacterial expression system, the corresponding deficient mutant failed to develop normal symptoms on host plant tissues. Conclusions: Among the five ITC-inducible GSTs analyzed in this study, three were found essential for full aggressiveness of A. brassicicola on host plant. This, to our knowledge is the first evidence that GSTs might be essential virulence factors for fungal necrotrophs

Characterization of glutathione transferases involved in the pathogenicity of Alternaria brassicicola

Background Glutathione transferases (GSTs) represent an extended family of multifunctional proteins involved in detoxification processes and tolerance to oxidative stress. We thus anticipated that some GSTs could play an essential role in the protection of fungal necrotrophs against plant-derived toxic metabolites and reactive oxygen species that accumulate at the host-pathogen interface during infection. Results Mining the genome of the necrotrophic Brassica pathogen Alternaria brassicicola for glutathione transferase revealed 23 sequences, 17 of which could be clustered into the main classes previously defined for fungal GSTs and six were ‘orphans’. Five isothiocyanate-inducible GSTs from five different classes were more thoroughly investigated. Analysis of their catalytic properties revealed that two GSTs, belonging to the GSTFuA and GTT1 classes, exhibited GSH transferase activity with isothiocyanates (ITC) and peroxidase activity with cumene hydroperoxide, respectively. Mutant deficient for these two GSTs were however neither more susceptible to ITC nor less aggressive than the wild-type parental strain. By contrast mutants deficient for two other GSTs, belonging to the Ure2pB and GSTO classes, were distinguished by their hyper-susceptibility to ITC and low aggressiveness against Brassica oleracea. In particular AbGSTO1 could participate in cell tolerance to ITC due to its glutathione-dependent thioltransferase activity. The fifth ITC-inducible GST belonged to the MAPEG class and although it was not possible to produce the soluble active form of this protein in a bacterial expression system, the corresponding deficient mutant failed to develop normal symptoms on host plant tissues. Conclusions Among the five ITC-inducible GSTs analyzed in this study, three were found essential for full aggressiveness of A. brassicicola on host plant. This, to our knowledge is the first evidence that GSTs might be essential virulence factors for fungal necrotrophs

Role of mannitol metabolism in the pathogenicity of the necrotrophic fungus Alternaria brassicicola

In this study, the physiological functions of fungal mannitol metabolism in the pathogenicity and protection against environmental stresses were investigated in the necrotrophic fungus Alternada brassicicola. Mannitol metabolism was examined during infection of Brass/ca oleracea leaves by sequential HPLC quantification of the major soluble carbohydrates and expression analysis of genes encoding two proteins of mannitol metabolism, i.e., a mannitol dehydrogenase (AbMdh), and a mannito1-1-phosphate dehydrogenase (AbMpd). Knockout mutants deficient for AbMdf7 or AbMpd and a double mutant lacking both enzyme activities were constructed. Their capacity to cope with various oxidative and drought stresses and their pathogenic behavior were evaluated. Metabolic and gene expression profiling indicated an increase in mannitol production during plant infection. Depending on the mutants, distinct pathogenic processes, such as leaf and silique colonization, sporulation, survival on seeds, were impaired by comparison to the wild-type. This pathogenic alteration could be partly explained by the differential susceptibilities of mutants to oxidative and drought stresses. These results highlight the importance of mannitol metabolism with respect to the ability of A. brassicrcola to efficiently accomplish key steps of its pathogenic life cycle

supp data sequences

fasta file of the sequences used for the phylogenetic analysi

Coupling evolutionary dynamics of Venturia inaequalis effectors and functional genomic to decipher mechanisms of virulence and to identify durable resistance genes in apple.

During infection, pathogens secrete small secreted proteins (SSPs), called effectors, that promote disease. Plant receptors encoded by resistance R genes might recognize such effectors (also called avirulence factors AVRs), resulting in plant immunity. Pathogens evade recognition thanks to the emergence of virulent alleles present in populations. It has been demonstrated that avirulent effectors are crucial for the pathogen infection cycle and that their loss-of-function may induce a substantial fitness cost. This kind of effector is expected to be under purifying selective pressure. Here, we aim at identifying the effector repertoire of Venturia inaequalis, the agent of apple scab, assessing its evolutionary dynamics and studying the role of candidate effectors in virulence. We sequenced de novo 90 strains, collected on apple and on their wild relatives and differing in their host range or virulence to study allelic polymorphism at 880 putative effector loci. The top-20 hits for highly conserved sequences were selected as candidates for further functional analyses. In planta gene expression showed a significant induction of these conserved SSP at the early stage of plant infection. Their functions were investigated using targeted deletion mutants. Remarkably, loss of two conserved SSPs resulted in reduced aggressiveness without any alteration in growth in vitro. GFP-tagged protein and heterologous expression were used to assess their sub-cellular localization in infected apple leaves. Involvement of theses SSP in the modulation of host defence was also investigated using an apple full-transcript microarray. Highly conserved effectors will be used to screen for novel R genes in Malus genotypes characterized for their high resistance to scab. This combined knowledge should enable us to understand strategies used by the pathogen to overcome defences in apple and consequently to build more durable resistance towards apple scab