Transcriptomic markers of fungal growth, respiration and carbon-use efficiency

Fungal metabolic carbon acquisition and its subsequent partitioning between biomass production and respiration, i.e. the carbon-use efficiency (CUE), are central parameters in biogeochemical modeling. However, current available techniques for estimating these parameters are all associated with practical and theoretical shortcomings, making assessments unreliable. Gene expression analyses hold the prospect of phenotype prediction by indirect means, providing new opportunities to obtain information about metabolic priorities. We cultured four different fungal isolates (Chalara longipes, Laccaria bicolor, Serpula lacrymans and Trichoderma harzianum) in liquid media with contrasting nitrogen availability and measured growth rates and respiration to calculate CUE. By relating gene expression markers to measured carbon fluxes, we identified genes coding for 1,3-β-glucan synthase and 2-oxoglutarate dehydrogenase as suitable markers for growth and respiration, respectively, capturing both intraspecific variation as well as within-strain variation dependent on growth medium. A transcript index based on these markers correlated significantly with differences in CUE between the fungal isolates. Our study paves the way for the use of these markers to assess differences in growth, respiration and CUE in natural fungal communities, using metatranscriptomic or the RT-qPCR approach

Where does all the phosphorus go? Mass balance modelling of phosphorus in the Swedish long-term soil fertility experiments

To gain insights into phosphorus (P) dynamics in soils and the ability to predict soil responses to varying fertilizer inputs, mass balance models prove to be valuable tools. In this study, a new dynamic mass balance model, PBalD8, was used to describe the change in extracted P in the A horizon of soils subjected to diverse fertilizer treatments over a period of 50 to 60 years in five soil fertility experiments. The model employed a Freundlich equation to describe soil-solution partitioning of P and assumed that acid-lactate-extractable P represented a labile pool of P in instant equilibrium with soil solution P. Additionally, oxalate-extractable inorganic P was presumed to comprise the sum of the labile and stable pools of P, with mass flux to and from the latter described by Fick's first law. The model was evaluated using results from extractions and P K-edge XANES spectroscopy. Notably, organic P, as revealed by P K-edge XANES, did not substantially contribute to long-term changes in soil P content and was therefore excluded from consideration. In general, the model offered reasonable fits to the extracted P concentrations. However, for the P-depleted treatments, a prerequisite was that the P removal through harvest was lower compared to measurements. Conversely, in three of the soils, the modelled fertilizer inputs needed to be reduced to 70 % to 85 % of the known additions. These discrepancies may be attributed to the involvement of deeper soil horizons, including deep crop uptake and mixing with lower soil layers, although other factors such as lateral dispersion and inaccuracies in estimating applied fertilizers cannot be discounted. These results underscore the necessity of gaining a more comprehensive understanding of how deeper soil horizons influence P mass balances in agricultural soils. In one of the soils, Fja center dot rdingslo center dot v, P K-edge XANES results demonstrated the formation of calcium phosphate over time in the highest fertilization treatment, consistent with the model. Additionally, in two soils, Kungsa center dot ngen and the P-depleted Vreta Kloster soil, the model predicted a significant contribution from mineral weathering. However, the PBalD8 model also projected higher P leaching rates than those observed, suggesting that the model may not fully capture this P output term

Fungal ecological strategies reflected in gene transcription - a case study of two litter decomposers.

Microbial communities interplay with their environment through their functional traits that can be a response or an effect on the environment. Here, we explore how a functional trait-the decomposition of organic matter, can be addressed based on genetic markers and how the expression of these markers reflect ecological strategies of two fungal litter decomposer Gymnopus androsaceus and Chalara longipes. We sequenced the genomes of these two fungi, as well as their transcriptomes at different steps of Pinus sylvestris needles decomposition in microcosms. Our results highlighted that if the gene content of the two species could indicate similar potential decomposition abilities, the expression levels of specific gene families belonging to the glycoside hydrolase category reflected contrasting ecological strategies. Actually, C. longipes, the weaker decomposer in this experiment, turned out to have a high content of genes involved in cell wall polysaccharides decomposition but low expression levels, reflecting a versatile ecology compare to the more competitive G. androsaceus with high expression levels of keystone functional genes. Thus, we established that sequential expression of genes coding for different components of the decomposer machinery indicated adaptation to chemical changes in the substrate as decomposition progressed

Retrospective evaluation of whole exome and genome mutation calls in 746 cancer samples

Funder: NCI U24CA211006Abstract: The Cancer Genome Atlas (TCGA) and International Cancer Genome Consortium (ICGC) curated consensus somatic mutation calls using whole exome sequencing (WES) and whole genome sequencing (WGS), respectively. Here, as part of the ICGC/TCGA Pan-Cancer Analysis of Whole Genomes (PCAWG) Consortium, which aggregated whole genome sequencing data from 2,658 cancers across 38 tumour types, we compare WES and WGS side-by-side from 746 TCGA samples, finding that ~80% of mutations overlap in covered exonic regions. We estimate that low variant allele fraction (VAF < 15%) and clonal heterogeneity contribute up to 68% of private WGS mutations and 71% of private WES mutations. We observe that ~30% of private WGS mutations trace to mutations identified by a single variant caller in WES consensus efforts. WGS captures both ~50% more variation in exonic regions and un-observed mutations in loci with variable GC-content. Together, our analysis highlights technological divergences between two reproducible somatic variant detection efforts

Impact de l’essence forestière sur les processus de dégradation et d’assimilation des polysaccharides végétaux par la communauté fongique des sols forestiers

The degradation of plant biomass is an essential process for the proper functioning of forest soils and terrestrial carbon cycling. Mechanisms involved in these processes are strongly controlled by saprotrophic fungi which secrete several hydrolytic enzymes to access at their primary nutrient sources found under the form of polysaccharides (cellulose and hemicelluloses). Enzymatic hydrolysis of plant polymers releases a high diversity of low molecular weight compounds (mono- and oligosaccharides). These molecules enter in fungal cell using transmembrane transporter systems. Consequently, the presence/absence and the substrate specificity of these transporters might contribute to the metabolic versatility of soil fungi. Several studies have demonstrated that tree species strongly affect diversity and composition of fungal communities. In this context, we hypothesized that the fungal communities selected by the different tree species expressed specific lignocellulolytic enzymes and sugar transporters; and thereby each fungal community was specifically adapted to the nature of litter produced by the tree species considered. We assessed, by the high-throughput sequencing of gene-fragments amplified from soil cDNA, the impact of tree species (Beech vs Spruce) on the diversity of genes encoding either lignocellulolytic enzymes or sugar porters expressed by soil fungi in two mono-specific forests. Our results revealed that most detected genes, encoding either lignocellulolytic enzymes or sugar transporters, have an unknown origin and are specifically found (for more than 80% of them) in one of the two forest soils. This work showed a significant “tree species effect” on the composition of functional genes expressed by soil fungi and suggests that beyond the species level, functional diversity of fungal communities must be addressed to better understand ecosystem functioning. Moreover, by using a functional metatranscriptomic approach, we identified functional transporter sequences differing with respect to their substrate specificities. From a spruce cDNA library, and for the first time, we identified high affinity or mannose specific transporters. Coincidently, as opposed to beech, spruce is indeed a tree species with a large proportion of mannose in its hemicellulosesEn milieu forestier, la décomposition de la matière organique d’origine végétale et son assimilation par les microorganismes sont des processus essentiels au bon fonctionnement des sols et du cycle du carbone. Les mécanismes impliqués dans ces phénomènes de dégradation sont fortement contrôlés par les champignons saprotrophes qui sécrètent de nombreuses enzymes hydrolytiques afin d’accéder à leur principale source de nutriments qui se trouve sous la forme de polysaccharides (cellulose et hémicelluloses). L’hydrolyse enzymatique de ces polymères végétaux engendre une grande diversité de composés de faible poids moléculaire (mono- et oligosaccharides). Ces molécules pénètrent dans la cellule fongique via des systèmes de transporteurs membranaires dont la présence/absence et la spécificité de substrat contribuent à la versatilité métabolique de ces microorganismes du sol. La nature de l’essence forestière affecte fortement la diversité et la composition des communautés fongiques. Dans ce contexte, nous avons émis l’hypothèse que les communautés fongiques sélectionnées par les différentes essences forestières diffèrent entre elles par la diversité des mécanismes qu’elles mettent en place lors du processus de décomposition de la matière organique d'origine végétale et lors du transport des produits de dégradation ; et que de ce fait chaque communauté fongique est spécifiquement adaptée à la nature de la litière produite par l’espèce végétale considérée. Par des approches de séquençage haut-débit d’amplicons générés à partir d’ADNc environnementaux, nous avons analysé la diversité des transcrits codant des protéines fongiques impliquées à la fois dans la dégradation des polysaccharides végétaux et dans le transport des sucres issus de cette hydrolyse enzymatique au sein de sols localisés sous des forêts de hêtres et d’épicéas. L’analyse des données obtenues a mis en évidence des transcrits jusqu’alors inconnus et spécifiquement retrouvés pour plus 80% d’entre eux sous l’un des deux couverts végétaux conduisant ainsi à un effet significatif de l’essence forestière sur la composition des gènes exprimés par les communautés fongiques. Des transporteurs potentiellement spécifiques du mannose ont été détectés, pour la première fois, sous la forêt d’épicéas par une approche de crible fonctionnel dans la levure de banques d’ADNc environnementaux. Or, cette essence forestière est connue pour posséder d’importantes quantités de mannose au niveau de ses hémicelluloses. Les résultats obtenus au cours de cette thèse soulignent l’importance d’étudier la diversité fonctionnelle des communautés fongiques pour comprendre l’impact du couvert végétal sur leur composition et le fonctionnement des écosystèmes forestier

Impact of tree species on the mechanisms developed by fungal communities to degrade and assimilate plant organic matter in forest soils

En milieu forestier, la décomposition de la matière organique d’origine végétale et son assimilation par les microorganismes sont des processus essentiels au bon fonctionnement des sols et du cycle du carbone. Les mécanismes impliqués dans ces phénomènes de dégradation sont fortement contrôlés par les champignons saprotrophes qui sécrètent de nombreuses enzymes hydrolytiques afin d’accéder à leur principale source de nutriments qui se trouve sous la forme de polysaccharides (cellulose et hémicelluloses). L’hydrolyse enzymatique de ces polymères végétaux engendre une grande diversité de composés de faible poids moléculaire (mono- et oligosaccharides). Ces molécules pénètrent dans la cellule fongique via des systèmes de transporteurs membranaires dont la présence/absence et la spécificité de substrat contribuent à la versatilité métabolique de ces microorganismes du sol. La nature de l’essence forestière affecte fortement la diversité et la composition des communautés fongiques. Dans ce contexte, nous avons émis l’hypothèse que les communautés fongiques sélectionnées par les différentes essences forestières diffèrent entre elles par la diversité des mécanismes qu’elles mettent en place lors du processus de décomposition de la matière organique d'origine végétale et lors du transport des produits de dégradation ; et que de ce fait chaque communauté fongique est spécifiquement adaptée à la nature de la litière produite par l’espèce végétale considérée. Par des approches de séquençage haut-débit d’amplicons générés à partir d’ADNc environnementaux, nous avons analysé la diversité des transcrits codant des protéines fongiques impliquées à la fois dans la dégradation des polysaccharides végétaux et dans le transport des sucres issus de cette hydrolyse enzymatique au sein de sols localisés sous des forêts de hêtres et d’épicéas. L’analyse des données obtenues a mis en évidence des transcrits jusqu’alors inconnus et spécifiquement retrouvés pour plus 80% d’entre eux sous l’un des deux couverts végétaux conduisant ainsi à un effet significatif de l’essence forestière sur la composition des gènes exprimés par les communautés fongiques. Des transporteurs potentiellement spécifiques du mannose ont été détectés, pour la première fois, sous la forêt d’épicéas par une approche de crible fonctionnel dans la levure de banques d’ADNc environnementaux. Or, cette essence forestière est connue pour posséder d’importantes quantités de mannose au niveau de ses hémicelluloses. Les résultats obtenus au cours de cette thèse soulignent l’importance d’étudier la diversité fonctionnelle des communautés fongiques pour comprendre l’impact du couvert végétal sur leur composition et le fonctionnement des écosystèmes forestiersThe degradation of plant biomass is an essential process for the proper functioning of forest soils and terrestrial carbon cycling. Mechanisms involved in these processes are strongly controlled by saprotrophic fungi which secrete several hydrolytic enzymes to access at their primary nutrient sources found under the form of polysaccharides (cellulose and hemicelluloses). Enzymatic hydrolysis of plant polymers releases a high diversity of low molecular weight compounds (mono- and oligosaccharides). These molecules enter in fungal cell using transmembrane transporter systems. Consequently, the presence/absence and the substrate specificity of these transporters might contribute to the metabolic versatility of soil fungi. Several studies have demonstrated that tree species strongly affect diversity and composition of fungal communities. In this context, we hypothesized that the fungal communities selected by the different tree species expressed specific lignocellulolytic enzymes and sugar transporters; and thereby each fungal community was specifically adapted to the nature of litter produced by the tree species considered. We assessed, by the high-throughput sequencing of gene-fragments amplified from soil cDNA, the impact of tree species (Beech vs Spruce) on the diversity of genes encoding either lignocellulolytic enzymes or sugar porters expressed by soil fungi in two mono-specific forests. Our results revealed that most detected genes, encoding either lignocellulolytic enzymes or sugar transporters, have an unknown origin and are specifically found (for more than 80% of them) in one of the two forest soils. This work showed a significant “tree species effect” on the composition of functional genes expressed by soil fungi and suggests that beyond the species level, functional diversity of fungal communities must be addressed to better understand ecosystem functioning. Moreover, by using a functional metatranscriptomic approach, we identified functional transporter sequences differing with respect to their substrate specificities. From a spruce cDNA library, and for the first time, we identified high affinity or mannose specific transporters. Coincidently, as opposed to beech, spruce is indeed a tree species with a large proportion of mannose in its hemicellulose

Transcriptomic markers of fungal growth, respiration and carbon-use efficiency

Fungal metabolic carbon acquisition and its subsequent partitioning between biomass production and respiration, i.e. the carbon-use efficiency (CUE), are central parameters in biogeochemical modeling. However, current available techniques for estimating these parameters are all associated with practical and theoretical shortcomings, making assessments unreliable. Gene expression analyses hold the prospect of phenotype prediction by indirect means, providing new opportunities to obtain information about metabolic priorities. We cultured four different fungal isolates (Chalara longipes, Laccaria bicolor, Serpula lacrymans and Trichoderma harzianum) in liquid media with contrasting nitrogen availability and measured growth rates and respiration to calculate CUE. By relating gene expression markers to measured carbon fluxes, we identified genes coding for 1,3-beta-glucan synthase and 2-oxoglutarate dehydrogenase as suitable markers for growth and respiration, respectively, capturing both intraspecific variation as well as within-strain variation dependent on growth medium. A transcript index based on these markers correlated significantly with differences in CUE between the fungal isolates. Our study paves the way for the use of these markers to assess differences in growth, respiration and CUE in natural fungal communities, using metatranscriptomic or the RT-qPCR approach

Where does all the phosphorus go? Mass balance modelling of phosphorus in the Swedish long-term soil fertility experiments

To gain insights into phosphorus (P) dynamics in soils and the ability to predict soil responses to varying fertilizer inputs, mass balance models prove to be valuable tools. In this study, a new dynamic mass balance model, PBalD8, was used to describe the change in extracted P in the A horizon of soils subjected to diverse fertilizer treatments over a period of 50 to 60 years in five soil fertility experiments. The model employed a Freundlich equation to describe soil-solution partitioning of P and assumed that acid-lactate-extractable P represented a labile pool of P in instant equilibrium with soil solution P. Additionally, oxalate-extractable inorganic P was presumed to comprise the sum of the labile and stable pools of P, with mass flux to and from the latter described by Fick’s first law. The model was evaluated using results from extractions and P K-edge XANES spectroscopy. Notably, organic P, as revealed by P K-edge XANES, did not substantially contribute to long-term changes in soil P content and was therefore excluded from consideration. In general, the model offered reasonable fits to the extracted P concentrations. However, for the P-depleted treatments, a prerequisite was that the P removal through harvest was lower compared to measurements. Conversely, in three of the soils, the modelled fertilizer inputs needed to be reduced to 70 % to 85 % of the known additions. These discrepancies may be attributed to the involvement of deeper soil horizons, including deep crop uptake and mixing with lower soil layers, although other factors such as lateral dispersion and inaccuracies in estimating applied fertilizers cannot be discounted. These results underscore the necessity of gaining a more comprehensive understanding of how deeper soil horizons influence P mass balances in agricultural soils. In one of the soils, Fjärdingslöv, P K-edge XANES results demonstrated the formation of calcium phosphate over time in the highest fertilization treatment, consistent with the model. Additionally, in two soils, Kungsängen and the P-depleted Vreta Kloster soil, the model predicted a significant contribution from mineral weathering. However, the PBalD8 model also projected higher P leaching rates than those observed, suggesting that the model may not fully capture this P output term