Organic-matter decomposition as a bioassessment tool of stream functioning: A comparison of eight decomposition-based indicators exposed to different environmental changes.

Organic-matter decomposition has long been proposed as a tool to assess stream functional integrity, but this indicator largely depends on organic-matter selection. We assessed eight decomposition-based indicators along two well-known environmental gradients, a nutrient-enrichment gradient (0.2-1.4 mg DIN/L) in central Portugal and an acidification gradient (pH: 4.69-7.33) in north-eastern France to identify the most effective organic-matter indicator for assessing stream functional integrity. Functional indicators included natural leaf litter (alder and oak) in 10-mm and 0.5-mm mesh bags, commercial tea (Lipton green and rooibos teas in 0.25-mm mesh bags), wood sticks (wood tongue depressors) and cotton strips. Biotic indices based on benthic macroinvertebrates (IPtIN for Portugal and IBGN for France) were calculated to compare the effectiveness of structural and functional indicators in detecting stream impairment and to assess the relationship between both types of indicators. The effectiveness of organic-matter decomposition rates as a functional indicator depended on the stressor considered and the substrate used. Decomposition rates generally identified nutrient enrichment and acidification in the most acidic streams. Decomposition rates of alder and oak leaves in coarse-mesh bags, green and rooibos teas and wood sticks were positively related with pH. Only decomposition rates of rooibos tea and wood sticks were related with DIN concentration; decomposition rates along the nutrient-enrichment gradient were confounded by differences in shredder abundance and temperature among streams. Stream structural integrity was good to excellent across streams; the IPtIN index was unrelated to DIN concentration, while the IBGN index was positively related with pH. The relationships between decomposition rates and biotic indices were loose in most cases, and only decomposition rates of alder leaves in coarse-mesh bags and green tea were positively related with the IBGN. Commercial substrates may be a good alternative to leaf litter to assess stream functional integrity, especially in the case of nutrient enrichment.

Inter- and intraspecific functional variability of aquatic fungal decomposers and freshwater ecosystem processes.

Although considerable intraspecific trait variation is common, research dedicated to ecosystem functioning has focused mainly on species diversity. Organic matter breakdown, a key ecosystem-level process in woodland streams is mainly driven by aquatic hyphomycetes. These aquatic fungal decomposers constitute a critical link between plant litter and invertebrate detritivores in detritus-based food webs in streams. In this study, we evaluated the functional variability across a set of ten isolates each belonging to five widespread aquatic hyphomycete species, namely Articulospora tetracladia, Anguillospora crassa, Lemonniera terrestris, Neonectria lugdunensis and Tetracladium marchalianum. All the isolates originated from undisturbed streams. We estimated inter- and intraspecific variability on growth rates, litter decomposition and sporulation rates of the isolates. In addition, we also assessed the invertebrate consumption rates on leaves colonized by different isolates. Significant differences were observed within the fungal species in growth rates (A. crassa, L. terrestris, N. lugdunensis and T. marchalianum), leaf litter decomposition (A. tetracladia, L. terrestris and N. lugdunensis) and sporulation rates (A. crassa, A. tetracladia, L. terrestris and N. lugdunensis). The relative consumption rates of the shredder Schizopelex festiva significantly differed when fed with leaves colonized by isolates of L. terrestris and N. lugdunensis, however differences were not seen when fed with leaves conditioned by different species. Overall, results indicate substantial intraspecific functional variability among the isolates of aquatic hyphomycetes. Besides, our study also provides a novel insight that intraspecific functional variability is a natural phenomenon exhibited by isolates not only from polluted but also from undisturbed streams. Here the isolates demonstrated marked inter- and intraspecific functional variability, calling for a greater understanding of the functional role of aquatic hyphomycetes and its ability to influence higher trophic levels.

Co-translational assembly of proteasome subunits in NOT1-containing assemblysomes.

The assembly of large multimeric complexes in the crowded cytoplasm is challenging. Here we reveal a mechanism that ensures accurate production of the yeast proteasome, involving ribosome pausing and co-translational assembly of Rpt1 and Rpt2. Interaction of nascent Rpt1 and Rpt2 then lifts ribosome pausing. We show that the N-terminal disordered domain of Rpt1 is required to ensure efficient ribosome pausing and association of nascent Rpt1 protein complexes into heavy particles, wherein the nascent protein complexes escape ribosome quality control. Immunofluorescence and in situ hybridization studies indicate that Rpt1- and Rpt2-encoding messenger RNAs co-localize in these particles that contain, and are dependent on, Not1, the scaffold of the Ccr4-Not complex. We refer to these particles as Not1-containing assemblysomes, as they are smaller than and distinct from other RNA granules such as stress granules and GW- or P-bodies. Synthesis of Rpt1 with ribosome pausing and Not1-containing assemblysome induction is conserved from yeast to human cells.

Global patterns and drivers of ecosystem functioning in rivers and riparian zones.

River ecosystems receive and process vast quantities of terrestrial organic carbon, the fate of which depends strongly on microbial activity. Variation in and controls of processing rates, however, are poorly characterized at the global scale. In response, we used a peer-sourced research network and a highly standardized carbon processing assay to conduct a global-scale field experiment in greater than 1000 river and riparian sites. We found that Earth's biomes have distinct carbon processing signatures. Slow processing is evident across latitudes, whereas rapid rates are restricted to lower latitudes. Both the mean rate and variability decline with latitude, suggesting temperature constraints toward the poles and greater roles for other environmental drivers (e.g., nutrient loading) toward the equator. These results and data set the stage for unprecedented "next-generation biomonitoring" by establishing baselines to help quantify environmental impacts to the functioning of ecosystems at a global scale.

Rapid characterization of aquatic hyphomycetes by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

Protein fingerprinting using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI--TOF MS) is a rapid, reliable, and economical method to characterize isolates of terrestrial fungi and other microorganisms. The objective of our study was to evaluate the suitability of MALDI-TOF MS for the identification of aquatic hyphomycetes, a polyphyletic group of fungi that play crucial roles in stream ecosystems. To this end, we used 34 isolates of 21 aquatic hyphomycete species whose identity was confirmed by spore morphology and internal transcribed spacer (ITS1-5.8S-ITS2 = ITS) nuc rDNA sequencing. We tested the efficiency of three protein extraction methods, including chemical and mechanical treatments using 13 different protocols, with the objective of producing high-quality MALDI-TOF mass spectra. In addition to extraction protocols, mycelium age was identified as a key parameter affecting protein extraction efficiency. The dendrogram based on mass-spectrum similarity indicated good and relevant taxonomic discrimination; the tree structure was comparable to that of the phylogram based on ITS sequences. Consequently, MALDI-TOF MS could reliably identify the isolates studied and provided greater taxonomic accuracy than classical morphological methods. MALDI-TOF MS seems suited for rapid characterization and identification of aquatic hyphomycete species.

Interactive effects of dissolved nitrogen, phosphorus and litter chemistry on stream fungal decomposers.

The enrichment of ecosystems by nutrients such as nitrogen (N) and phosphorus (P) has important ecological consequences. These include effects on plant litter decomposition in forest soils and forested headwater streams, where fungi play a pivotal role. However, our understanding of nutrient relationships on fungal communities associated with decomposing litter remains surprisingly incomplete. We conducted a fully factorial microcosm experiment with known communities of fungal decomposers from streams to assess the importance of dissolved N and P supply, as well as the atomic nutrient ratio (N:P), on fungal community succession, diversity, biomass and reproduction on three leaf-litter species differing in nutrient and lignin concentrations. Fungal biomass accrual and spore production were strongly controlled by external N supply, whereas P supply was much less important. The magnitude of these effects was mediated by litter quality, with stronger effects of dissolved N and P on lignin-poor and high N:P litter. N supply also influenced fungal diversity and species composition, acting as a pacemaker of community succession. Collectively, our data indicate that N was in much greater demand than predicted by standard stoichiometric models. The most parsimonious explanation for this deviation relates to the need of litter fungi to invest large amounts of N into degradative exoenzymes.

Quantitative real-time PCR as a promising tool for the detection and quantification of leaf-associated fungal species - A proof-of-concept using Alatospora pulchella.

Traditional methods to identify aquatic hyphomycetes rely on the morphology of released conidia, which can lead to misidentifications or underestimates of species richness due to convergent morphological evolution and the presence of non-sporulating mycelia. Molecular methods allow fungal identification irrespective of the presence of conidia or their morphology. As a proof-of-concept, we established a quantitative real-time polymerase chain reaction (qPCR) assay to accurately quantify the amount of DNA as a proxy for the biomass of an aquatic hyphomycete species (Alatospora pulchella). Our study showed discrimination even among genetically closely-related species, with a high sensitivity and a reliable quantification down to 9.9 fg DNA (3 PCR forming units; LoD) and 155.0 fg DNA (47 PCR forming units; LoQ), respectively. The assay's specificity was validated for environmental samples that harboured diverse microbial communities and likely contained PCR-inhibiting substances. This makes qPCR a promising tool to gain deeper insights into the ecological roles of aquatic hyphomycetes and other microorganisms.

Harmful effects of silver nanoparticles on a complex detrital model system.

The rapid proliferation of silver nanoparticles (AgNP) in industry and the environment requires realistic toxicity assessments based on approaches that consider the biological complexity of ecosystems. Here we assessed the acute toxicity of carbonate-coated AgNP and, for comparison, AgNO3 (Ag(+)) by using a model system consisting of decomposing plant litter and the associated fungal and bacterial decomposers as central players in the functioning of stream ecosystems. Little variation in size and surface charge during the experiment indicated that the AgNP used were essentially stable. AgNP disrupted bacterial growth (≤83% reduction in protein biosynthesis, EC50 = 0.3 µM), clearly affected fungal growth (≤61% reduction in ergosterol synthesis, EC50 = 47 µM) with both endpoints more sensitive to AgNP than to Ag(+). Fungal reproduction, in contrast, was stimulated by AgNP, but not Ag(+), at concentrations up to 25 µM. Both AgNP and Ag(+ )also stimulated extracellular alkaline phosphatase but reduced leucine aminopeptidase, whereas ß-glucosidase was stimulated by AgNP and reduced by Ag(+). Importantly, the provision of cysteine, a chelating ligand that complexes free Ag(+), failed to alleviate AgNP toxicity to microbial growth, clearly demonstrating particle-mediated toxicity independent of the presence of ionic silver. This contrasts with the observed inhibition of leucine aminopeptidase by Ag(+), which accounted for 2-6% of the total silver in treatments receiving AgNP. These results show that although outcomes of AgNP and Ag(+ )exposure assessed by different functional endpoints vary widely, AgNP strongly interferes with bacterial growth and a range of other microbial processes, resulting in severe consequences for natural microbial communities and ecosystem functioning.

Litter identity mediates predator impacts on the functioning of an aquatic detritus-based food web.

During past decades, several mechanisms such as resource quality and habitat complexity have been proposed to explain variations in the strength of trophic cascades across ecosystems. In detritus-based headwater streams, litter accumulations constitute both a habitat and a resource for detritivorous macroinvertebrates. Because litter edibility (which promotes trophic cascades) is usually inversely correlated with its structural complexity (which weakens trophic cascades), there is a great scope for stronger trophic cascades in litter accumulations that are dominated by easily degradable litter species. However, it remains unclear how mixing contrasting litter species (conferring both habitat complexity and high quality resource) may influence top-down controls on communities and processes. In enclosures exposed in a second-order stream, we manipulated litter species composition by using two contrasting litter (alder and oak), and the presence-absence of a macroinvertebrate predator (Cordulegaster boltonii larvae), enabling it to effectively exert predation pressure, or not, on detritivores (consumptive versus non-consumptive predation effects). Leaf mass loss, detritivore biomass and community structure were mostly controlled independently by litter identity and mixing and by predator consumption. However, the strength of predator control was mediated by litter quality (stronger on alder), and to a lesser extent by litter mixing (weaker on mixed litter). Refractory litter such as oak leaves may contribute to the structural complexity of the habitat for stream macroinvertebrates, allowing the maintenance of detritivore communities even when strong predation pressure occurs. We suggest that considering the interaction between top-down and bottom-up factors is important when investigating their influence on natural communities and ecosystem processes in detritus-based ecosystems.

Aquatic hyphomycete species are screened by the hyporheic zone of woodland streams.

Aquatic hyphomycetes strongly contribute to organic matter dynamics in streams, but their abilities to colonize leaf litter buried in streambed sediments remain unexplored. Here, we conducted field and laboratory experiments (slow-filtration columns and stream-simulating microcosms) to test the following hypotheses: (i) that the hyporheic habitat acting as a physical sieve for spores filters out unsuccessful strategists from a potential species pool, (ii) that decreased pore size in sediments reduces species dispersal efficiency in the interstitial water, and (iii) that the physicochemical conditions prevailing in the hyporheic habitat will influence fungal community structure. Our field study showed that spore abundance and species diversity were consistently reduced in the interstitial water compared with surface water within three differing streams. Significant differences occurred among aquatic hyphomycetes, with dispersal efficiency of filiform-spore species being much higher than those with compact or branched/tetraradiate spores. This pattern was remarkably consistent with those found in laboratory experiments that tested the influence of sediment pore size on spore dispersal in microcosms. Furthermore, leaves inoculated in a stream and incubated in slow-filtration columns exhibited a fungal assemblage dominated by only two species, while five species were codominant on leaves from the stream-simulating microcosms. Results of this study highlight that the hyporheic zone exerts two types of selection pressure on the aquatic hyphomycete community, a physiological stress and a physical screening of the benthic spore pool, both leading to drastic changes in the structure of fungal community.

Leaf-associated fungal diversity in acidified streams: insights from combining traditional and molecular approaches.

We combined microscopic and molecular methods to investigate fungal assemblages on alder leaf litter exposed in the benthic and hyporheic zones of five streams across a gradient of increasing acidification for 4 weeks. The results showed that acidification and elevated Al concentrations strongly depressed sporulating aquatic hyphomycetes diversity in both zones of streams, while fungal diversity assessed by denaturing gradient gel electrophoresis (DGGE) appeared unaffected. Clone library analyses revealed that fungal communities on leaves were dominated by members of Ascomycetes and to a lesser extent by Basidiomycetes and Chytridiomycetes. An important contribution of terrestrial fungi was observed in both zones of the most acidified stream and in the hyporheic zone of the reference circumneutral stream. The highest leaf breakdown rate was observed in the circumneutral stream and occurred in the presence of both the highest diversity of sporulating aquatic hyphomycetes and the highest contribution to clone libraries of sequences affiliated with aquatic hyphomycetes. Both methods underline the major role played by aquatic hyphomycetes in leaf decomposition process. Our findings also bring out new highlights on the identity of leaf-associated fungal communities and their responses to anthropogenic alteration of running water ecosystems.

Benthic algae stimulate leaf litter decomposition in detritus-based headwater streams: a case of aquatic priming effect?

In detritus-based ecosystems, autochthonous primary production contributes very little to the detritus pool. Yet primary producers may still influence the functioning of these ecosystems through complex interactions with decomposers and detritivores. Recent studies have suggested that, in aquatic systems, small amounts of labile carbon (C) (e.g., producer exudates), could increase the mineralization of more recalcitrant organic-matter pools (e.g., leaf litter). This process, called priming effect, should be exacerbated under low-nutrient conditions and may alter the nature of interactions among microbial groups, from competition under low-nutrient conditions to indirect mutualism under high-nutrient conditions. Theoretical models further predict that primary producers may be competitively excluded when allochthonous C sources enter an ecosystem. In this study, the effects of a benthic diatom on aquatic hyphomycetes, bacteria, and leaf litter decomposition were investigated under two nutrient levels in a factorial microcosm experiment simulating detritus-based, headwater stream ecosystems. Contrary to theoretical expectations, diatoms and decomposers were able to coexist under both nutrient conditions. Under low-nutrient conditions, diatoms increased leaf litter decomposition rate by 20% compared to treatments where they were absent. No effect was observed under high-nutrient conditions. The increase in leaf litter mineralization rate induced a positive feedback on diatom densities. We attribute these results to the priming effect of labile C exudates from primary producers. The presence of diatoms in combination with fungal decomposers also promoted decomposer diversity and, under low-nutrient conditions, led to a significant decrease in leaf litter C:P ratio that could improve secondary production. Results from our microcosm experiment suggest new mechanisms by which primary producers may influence organic matter dynamics even in ecosystems where autochthonous primary production is low.

Effect of acidification on leaf litter decomposition in benthic and hyporheic zones of woodland streams.

Anthropogenic acidification has deleterious effects on both structure and functioning of surface water ecosystems. This study examined how it may affect the leaf decomposition rate and the community structure and activity of decomposers in both benthic and hyporheic zones of five headwater streams along an acidification gradient from highly acidic (pH 4.6) to circumneutral (pH 7.4). Overall, responses to acidification in hyporheic zones were less pronounced, but followed the same pattern as in their benthic counterparts. Leaf decomposition was much faster in the circumneutral stream, both in the hyporheic and benthic zones (k = 0.0068 and 0.0534 d(-1), respectively), than in the most acidic one (k = 0.0016 and 0.0055 d(-1), respectively), and correlated well with the acidic gradient in both compartments. Interestingly, leaf litter decomposition was less affected by acidification in hyporheic compared to benthic compartments, likely due to the relatively low sensitivity of fungi, which were the main decomposers of buried coarse particulate organic matter. These results argue in favour of conserving hyporheic habitats in acidified streams as they can maintain matter and species fluxes that are essential to the ecosystem.

Leaf litter recycling in benthic and hyporheic layers in agricultural streams with different types of land use.

Changes in land use and intensification of agricultural pressure have greatly accelerated the alteration of the landscape in most developed countries. These changes may greatly disturb the adjacent ecosystems, particularly streams, where the effects of pollution are amplified. In this study, we used the leaf litter breakdown rate to assess the functional integrity of stream ecosystems and river sediments along a gradient of either traditional extensive farming or a gradient of vineyard area. In the benthic layer, the total litter breakdown process integrates the temporal variability of the anthropogenic disturbances and is strongly influenced by land use changes in the catchment even though a low concentration of toxics was measured during the study period. This study also confirmed the essential role played by amphipods in the litter breakdown process. In contrast, microbial processes may have integrated the variations in available nutrients and dissolved oxygen concentrations, but failed to respond to the disturbances induced by vineyard production (the increase in pesticides and metal concentrations) during the study period. The response of microbes may not be sensitive enough for assessing the global effect of seasonal agricultural practices. Finally, the leaf litter breakdown measured in the hyporheic zone seemed mainly driven by microbial activities and was hence more affected by vertical exchanges with surface water than by land use practices. However, the breakdown rate of leaf litter in the hyporheic zone may constitute a relevant way to evaluate the impact on river functioning of any human activities that induce massive soil erosion and sediment clogging.