Category Archives: Academic productions

Article: Forest pathology

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Do higher summer temperatures restrict the dissemination of Hymenoscyphus fraxineus in France? M Grosdidier, R Ioos, B Marçais. Forest Pathology

Summary

Hymenoscyphus fraxineus is an invasive pathogen that severely affects European ashes, jeopardizing the use of this species in forestry. However, even aggressive invasive pathogens require environmental conditions conducive to disease development. Hence, H. fraxineus survival at temperatures above 35°C is limited, which could reduce its impact in southern Europe. This study examined the effect of environmental conditions, mainly summer temperatures, on ash dieback development in southeast France between 2015 and 2016. Fourteen sites were selected according to altitudinal and longitudinal gradients covering a wide range of mean summer temperatures. Disease severity, density of infected and healthy rachises in the litter and quantity of apothecia produced on infected rachises were measured. Results point to limited disease severity in this area because of the occurrence of high summer temperatures that are unfavourable to the pathogen. In a context of global warming, increases in temperatures could have a positive impact on ashes by limiting ash dieback development.

Article: Applied and Environmental Microbiology

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Assessing airborne fungal communities by high-throughput sequencing using passive traps J Aguayo, C Fourrier-Jeandel, C Husson, R Ioos Applied and Environmental Microbiology, AEM. 02637-17

ABSTRACT

Techniques based on the high-throughput sequencing (HTS) of environmental DNA have provided a new way of studying fungal diversity. However, these techniques suffer from a number of methodological biases which may appear at any of the steps involved in a metabarcoding study. Air is one of the most important environments where fungi can be found because it is the primary medium of dispersal for many species. Looking ahead to future developments, it was decided to test 20 protocols including different passive spore traps, spore recovery procedures, DNA extraction kits and barcode loci. HTS was performed with the Illumina MiSeq platform targeting two sub loci of the fungal internal transcribed spacer. Multivariate analysis and generalized linear models showed that the type of passive spore trap, the spore recovery procedure and the barcode all impact the description of fungal communities in terms of richness and diversity when assessed by HTS metabarcoding. In contrast, DNA extraction kits did not significantly impact these results. Although passive traps may be used to describe airborne fungal communities, a study using specific real-time PCR and a mock community showed that these kinds of trap are affected by environmental conditions that may induce losses of biological material, impacting diversity and community composition results.

Importance The advent of high throughput sequencing (HTS) methods such as those offered by next-generation sequencing (NGS) techniques has opened a new era in the study of fungal diversity in different environmental substrates. In this study we show that the assessment of the diversity of airborne fungal communities can reliably be achieved by the use of simple and robust passive spore traps. However, comparison of sample processing protocols showed that several methodological biases may impact results of fungal diversity when assessed by metabarcoding. Our data suggest that identifying these biases is of paramount importance to enable a correct identification and relative quantification of community members.

Article: Annals of Forest Science

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Comparison of teak wood properties according to forest management: short versus long rotation DE Rizanti, W Darmawan, B George, A Merlin, S Dumarcay, H Chapuis, … Annals of Forest Science 75 (2), 39

Context

In Indonesia, teak wood has been supplied from the state forests (Perhutani) for long rotation teak and from community teak plantations for short rotation teak. Short rotation teak has been harvested at 7–10 years and long rotation teak at 40–60 years.

Aims

This paper discusses the characterization of technical properties of short and long rotation teak wood based on the chemical, anatomical, physical, and mechanical properties.

Methods

The properties of short rotation and long rotation teak woods were characterized by measuring their density, extractive contents, chemical composition, swelling, wettability, water sorption isotherm, decay resistance, anatomical properties, bending strength (modulus of rupture (MOR), modulus of elasticity (MOE)), and hardness.

Results

The results indicate that short rotation teak was not particularly different in swelling, MOE and MOR, and Brinell hardness compared to long rotation teak, although it was less dense and less durable due to lower heartwood and extractive contents. Therefore, careful attention should be given to the use of short rotation teak in some wood-processing technologies.

Conclusion

Lower wood density and durability of the short rotation compared to the long rotation teak will restrict its utilization to some extent for both indoor and outdoor applications. Fast-growing teak from community cannot be used as usual heartwood teak from Perhutani because of the very low proportion of useful heartwood in the stem.

Article: FEBS letters

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Structural snapshots along the reaction mechanism of the atypical poplar thioredoxin‐like2. 1 K Chibani, F Saul, C Didierjean, N Rouhier, A Haouz FEBS letters

Abstract

Plastidial thioredoxin (TRX)‐like2.1 proteins are atypical thioredoxins possessing a WCRKC active site signature and using glutathione for recycling. To obtain structural information supporting the peculiar catalytic mechanisms and target proteins of these TRXs, we solved the crystal structures of poplar TRX‐like2.1 in oxidized and reduced states and of mutated variants. These structures share similar folding with TRXs exhibiting the canonical WCGPC signature. Moreover, the overall conformation is not altered by reduction of the catalytic disulfide bond or in a C45S/C67S variant that formed a disulfide‐bridged dimer possibly mimicking reaction intermediates with target proteins. Modeling of the interaction of TRX‐like2.1 with both NADPH‐ and ferredoxin‐thioredoxin reductases (FTR) indicates that the presence of Arg43 and Lys44 residues likely precludes reduction by the plastidial FTR.

Article: FEMS Microbiology Ecology

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Tracking the invasion: dispersal of Hymenoscyphus fraxineus airborne inoculum at different scales. M Grosdidier, R Ioos, C Husson, O Cael, T Scordia… FEMS Microbiology Ecology.

Summary

Ash dieback is caused by an invasive pathogen, Hymenoscyphus fraxineus, which emerged in Europe in the 1990s and jeopardizes the management of ash stands. Although the biological cycle of the pathogen is well understood, its dispersal patterns via airborne spores remain poorly described. We investigated the seasonal and spatial patterns of dispersal in France using both a passive spore-trapping method coupled with a real-time PCR assay and reports of ash dieback based on symptom observations. Spores detection varies from year to year, with a detection ability of 30 to 47%, depending on meteorological conditions, which affect both production of inoculum and efficiency of the trapping. Nevertheless, our results are consistent and we showed that sporulation peak occurred from June to August and that spores were detected up to 50-100 km ahead of the disease front, proving the presence of the pathogen before any observation of symptoms. The spore dispersal gradient was steep, most of inoculum remaining within 50 m of infected ashes. Two dispersal kernels were fitted using Bayesian methods to estimate the mean dispersal distance of H. fraxineus from inoculum sources. The estimated mean distances of dispersal, either local or regional scale, were 1.4 km and 2.6 km, respectively, the best fitting kernel being the inverse power-law. This information may help to design disease management strategies.

Article: Current opinion in microbiology

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Show me the way: rust effector targets in heterologous plant systems. C Lorrain, B Petre, S Duplessis. Current opinion in microbiology 46, 19-25

Abstract

For years, the study of rust fungal effectors has been impeded by the lack of molecular genetic tools in rust pathosystems. The recent use of heterologous plants to perform effector screens (effectoromics)-including effector localisation (cellular targets) and protein interactors (molecular targets) in plant cells-has changed the game. These screens revealed that many candidate effectors from various rust fungi target specific plant cell compartments, including chloroplasts, and associate with specific plant protein complexes. Such information represents unparalleled opportunities to understand how effectors sustain extreme parasitic interactions and obligate biotrophy. Despite their limitations, we here portray how the use of heterologous expression systems has been essential for gaining new insight into rust effectors.

Article: FEMS Microbiology Reviews

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Bacterial-Fungal Interactions: ecology, mechanisms and challenges A Deveau, G Bonito, J Uehling, M Paoletti, M Becker, S Bindschedler, … FEMS microbiology reviews

Abstract

Fungi and bacteria are found living together in a wide variety of environments. Their interactions are significant drivers of many ecosystem functions and are important for the health of plants and animals. A large number of fungal and bacterial families are engaged in complex interactions that lead to critical behavioural shifts of the microorganisms ranging from mutualism to pathogenicity. The importance of bacterial-fungal interactions (BFI) in environmental science, medicine and biotechnology has led to the emergence of a dynamic and multidisciplinary research field that combines highly diverse approaches including molecular biology, genomics, geochemistry, chemical and microbial ecology, biophysics and ecological modelling. In this review, we discuss most recent advances that underscore the roles of BFI across relevant habitats and ecosystems. A particular focus is placed on the understanding of BFI within complex microbial communities and in regards of the metaorganism concept. We also discuss recent discoveries that clarify the (molecular) mechanisms involved in bacterial-fungal relationships, and the contribution of new technologies to decipher generic principles of BFI in terms of physical associations and molecular dialogues. Finally, we discuss future directions for researches in order to catalyse a synergy within the BFI research area and to resolve outstanding questions.

Article: Applied and Environmental Microbiology

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Enzyme activities of two recombinant heme-including peroxidases TvDyP1 and TvVP2 identified from the secretome of Trametes versicolor S Amara, T Perrot, D Navarro, A Deroy, A Benkhelfallah, A Chalak, … Applied and Environmental Microbiology, AEM. 02826-17

ABSTRACT

Trametes versicolor is a wood inhabiting Agaricomycete known for its ability to cause strong white rot decay on hardwood and for its high tolerance toward phenolic compounds. The goal of the present work was to give insights on the molecular biology and biochemistry of heme-including class-II and dye-decolorizing peroxidases secreted from this fungus. Proteomic analysis of the secretome of T. versicolor BRFM1218 grown on oak wood revealed a set of 200 secreted proteins among which were a dye-decolorizing peroxidase TvDyP1 and a versatile peroxidase TvVP2. Both peroxidases were heterologously produced in E. coli, and were biochemically characterized and tested for their capacity to oxidize complex substrates. Both peroxidases were found to be active against several substrates in acidic conditions, and TvDyP1 was very stable in a relatively large range of pH (pH 2.0 to 6.0) while TvVP2 was more stable at pH 5.0-6.0 only. Thermostability of both enzymes was also tested and TvDyP1 was globally found to be more stable than TvVP2. After 180 min of incubation at T°C ranging from 30°C to 50°C, activities of TvVP2 drastically decreased retaining 10% to 30% of the its initial activity. In the same conditions, TvDyP1 retained 20% to 80% of enzyme activity. The two proteins were catalytically characterized and TvVP2 was shown to accept a wider range of reducing substrates than TvDyP1. Furthermore, both enzymes were found to be active against two flavonoids, quercetin and catechin, found in oak wood, TvVP2 displaying a more rapid oxidation of the two compounds. They were tested for their potential interest in dye decolourization of five industrial dyes and TvVP2 presented a wider oxidation and decolourization capacity towards the dye substrates than TvDyP1.

IMPORTANCE Trametes versicolor is a wood inhabiting Agaricomycete known for its ability to cause strong white rot decay on hardwood and for its high tolerance toward phenolic compounds. Among white-rot fungi, the basidiomycete T. versicolor has been extensively studied for its efficiency to degrade wood, and specifically lignin, thanks to an extracellular oxidative enzymatic system. The corresponding oxidative system was previously studied in several works for classical lignin and manganese peroxidases, and in this study, two new components of the oxidative system of T. versicolor, one dye-decoloririzing peroxidase and one versatile peroxidase were biochemically characterized in depth and compare to other fungal peroxidases.

Article: Frontiers in Microbiology

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Genome-Wide Analysis of Corynespora cassiicola Leaf Fall Disease Putative Effectors. D Lopez, S Ribeiro, P Label, B Fumanal, JS Venisse, A Kohler, … Frontiers in Microbiology 9, 276

Abstract

Corynespora cassiicola is an Ascomycetes fungus with a broad host range and diverse life styles. Mostly known as a necrotrophic plant pathogen, it has also been associated with rare cases of human infection. In the rubber tree, this fungus causes the Corynespora leaf fall (CLF) disease, which increasingly affects natural rubber production in Asia and Africa. It has also been found as an endophyte in South American rubber plantations where no CLF outbreak has yet occurred. The C. cassiicola species is genetically highly diverse, but no clear relationship has been evidenced between phylogenetic lineage and pathogenicity. Cassiicolin, a small glycosylated secreted protein effector, is thought to be involved in the necrotrophic interaction with the rubber tree but some virulent C. cassiicola isolates do not have a cassiicolin gene. This study set out to identify other putative effectors involved in CLF. The genome of a highly virulent C. cassiicola isolate from the rubber tree (CCP) was sequenced and assembled. In silico prediction revealed 2870 putative effectors, comprising CAZymes, lipases, peptidases, secreted proteins and enzymes associated with secondary metabolism. Comparison with the genomes of 44 other fungal species, focusing on effector content, revealed a striking proximity with phylogenetically unrelated species (Colletotrichum acutatum, Colletotrichum gloesporioides, Fusarium oxysporum, nectria hematococca and Botrosphaeria dothidea) sharing life style plasticity and broad host range. Candidate effectors involved in the compatible interaction with the rubber tree were identified by transcriptomic analysis. Differentially expressed genes included 92 putative effectors, among which cassiicolin and two other secreted singleton proteins. Finally, the genomes of 35 C. cassiicola isolates representing the genetic diversity of the species were sequenced and assembled, and putative effectors identified. At the intraspecific level, effector-based classification was found to be highly consistent with the phylogenomic trees. Identification of lineage-specific effectors is a key step toward understanding C. cassiicola virulence and host specialization mechanisms.

Article: Journal of Biological Chemistry

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Function and maturation of the Fe–S center in dihydroxyacid dehydratase from Arabidopsis H Gao, T Azam, S Randeniya, J Couturier, N Rouhier, MK Johnson. Journal of Biological Chemistry, jbc. RA117. 001592

Abstract

Dihydroxyacid dehydratase (DHAD) is the third enzyme required for branched-chain amino acid biosynthesis in bacteria, fungi, and plants. DHAD enzymes contain two distinct types of active-site Fe–S clusters. The best characterized examples are Escherichia coli DHAD, which contains an oxygen-labile [Fe4S4] cluster, and spinach DHAD, which contains an oxygen-resistant [Fe2S2] cluster. Although the Fe–S cluster is crucial for DHAD function, little is known about the cluster-coordination environment or the mechanism of catalysis and cluster biogenesis. Here, using the combination of UV-visible absorption and circular dichroism, resonance Raman and electron paramagnetic resonance, we spectroscopically characterized the Fe–S center in DHAD from Arabidopsis thaliana (At). Our results indicated that AtDHAD can accommodate [Fe2S2] and [Fe4S4] clusters. However, only the [Fe2S2] cluster–bound form is catalytically active. We found that the [Fe2S2] cluster is coordinated by at least one non-cysteinyl ligand, which can be replaced by the thiol group(s) of dithiothreitol.In vitro cluster transfer and reconstitution reactions revealed that [Fe2S2] cluster–containing NFU2 protein is likely the physiological cluster donor for in vivo maturation of AtDHAD. In summary, AtDHAD binds either one [Fe4S4] or one [Fe2S2] cluster, with only the latter being catalytically competent and capable of substrate and product binding, and NFU2 appears to be the physiological [Fe2S2] cluster donor for DHAD maturation. This work represents the first in vitro characterization of recombinant AtDHAD, providing new insights into the properties, biogenesis, and catalytic role of the active-site Fe–S center in a plant DHAD.