Non-interactive effects drive multiple stressor impacts on the taxonomic and functional diversity of atlantic stream macroinvertebrates.

Freshwaters are considered among the most endangered ecosystems globally due to multiple stressors, which coincide in time and space. These local stressors typically result from land-use intensification or hydroclimatic alterations, among others. Despite recent advances on multiple stressor effects, current knowledge is still limited to manipulative approaches minimizing biological and abiotic variability. Thus, the assessment of multiple stressor effects in real-world ecosystems is required. Using an extensive survey of 50 stream reaches across North Portugal, we evaluated taxonomic and functional macroinvertebrate responses to multiple stressors, including marked gradients of nutrient enrichment, flow reduction, riparian vegetation structure, thermal stress and dissolved oxygen depletion. We analyzed multiple stressor effects on two taxonomic (taxon richness, Shannon-diversity) and two trait-based diversity indices (functional richness, functional dispersion), as well as changes in trait composition. We found that multiple stressors had additive effects on all diversity metrics, with nutrient enrichment identified as the most important stressor in three out of four metrics, followed by dissolved oxygen depletion and thermal stress. Taxon richness, Shannon-diversity and functional richness responded similarly, whereas functional dispersion was driven by changes in flow velocity and thermal stress. Functional trait composition changed along a major stress gradient determined by nutrient enrichment and oxygen depletion, which was positively correlated with organisms possessing fast-living strategies, aerial respiration, adult phases, and gathering-collector feeding habits. Overall, our results reinforce the need to consider complementary facets of biodiversity to better identify assembly processes in response to multiple stressors. Our data suggest that stressor interactions may be less frequent in real-word streams than predicted by manipulative experiments, which can facilitate mitigation strategies. By combining an extensive field survey with an integrative consideration of multiple biodiversity facets, our study provides new insights that can help to better assess and manage rivers in a global change context.

Antioxidant and anti-inflammatory activity of curcumin transdermal gel in an IL-10 knockout mouse model of inflammatory bowel disease.

Inflammatory bowel diseases are a group of inflammatory disorders of the gastrointestinal tract. Their prevalence is still low in Brazil, but the incidence is increasing annually. A variety of compounds present in Curcuma longa L., particularly curcumin, have been shown to reduce oxidative stress and aid in the prevention of associated diseases. This study aimed to assess the effect of curcumin transdermal gel on oxidative stress and intestinal inflammation in IL-10 knockout mice. Female mice were divided into four groups: a control group (C0) treated with vehicle and three experimental groups treated with transdermal gel containing 50 (C50), 75 (C75), and 100 (C100) mg curcumin kg-1 body weight. Colon malondialdehyde concentrations were lower in C50 and C75 groups. C100 treatment led to reduced catalase activity in the small intestine, whereas C50, C75, and C100 treatments resulted in decreased catalase activity in the colon. In contrast, superoxide dismutase activity increased in the small intestine of C50 and C75 mice and decreased in the colon of C50, C75, and C100 mice. Glutathione S-transferase activity increased in the small intestine and decreased in the colon of C75 animals. These findings suggest that curcumin transdermal gel exerts a protective effect against oxidative stress.

TGF-ß acts as a dual regulator of COX-2/PGE2 tumor promotion depending of its cross-interaction with H-Ras and Wnt/ß-catenin pathways in colorectal cancer cells.

Transforming growth factor-ß (TGF-ß) plays a dual role acting as tumor promoter or suppressor. Along with cyclooxygenase-2 (COX-2) and oncogenic Ras, this multifunctional cytokine is deregulated in colorectal cancer. Despite their individual abilities to promote tumor growth and invasion, the mechanisms of cross regulation between these pathways is still unclear. Here, we investigate the effects of TGF-ß, Ras oncogene and COX-2 in the colorectal cancer context. We used colon adenocarcinoma cell line HT-29 and Ras-transformed IEC-6 cells, both treated with prostaglandin E2 (PGE2 ), TGF-ß or a combined treatment with these agents. We demonstrated that PGE2 alters the subcellular localization of E-cadherin and ß-catenin and enhanced the tumorigenic potential in HT-29 cells. This effect was inhibited by TGF-ß, indicating a tumor suppressor role. Conversely, in Ras-transformed IEC-6 cells, TGF-ß induced COX-2 expression and increased invasiveness, acting as a tumor promoter. In IEC-6 Ras-transformed cells, TGF-ß increased nuclear ß-catenin and Wnt/ß-catenin activation, opposite to what was seen in the PGE2 and TGF-ß joint treatment in HT-29 cells. Together, our findings show that TGF-ß increases COX-2 levels and induces invasiveness cooperating with Ras in a Wnt/ß-catenin activation-dependent manner. This shows TGF-ß dual regulation over COX-2/PGE2 tumor promotion depending on the H-Ras and Wnt/ß-catenin pathways activation status in intestinal cancer cells.

Water Quality Assessment of the Pardo River Basin, Brazil: A Multivariate Approach Using Limnological Parameters, Metal Concentrations and Indicator Bacteria.

Located in the southeast of Brazil, the Pardo River Basin has a large population and an economy focused on agriculture, with a strong predominance of the sugarcane agro-industry. The purpose of the study was to assess the water quality of the Pardo River Basin under a multivariate approach using limnological parameters, metal concentrations, and indicator bacteria. Nine sampling campaigns were performed during both the dry and rainy seasons. Element concentrations were determined by inductively coupled plasma spectrometry (ICP-MS, Perkin Elmer Elan 6000). A battery of test to determine limnological parameters was performed (in situ). Total coliforms and Escherichia coli were detected and quantified using Defined Substrate Technology Colilert® and multiple tube dilutions. Principal component analysis and hierarchical cluster analysis were used as multivariate exploratory analysis. In general, the results suggest the influence of rain, possible sewage discharges into the watercourse, and the input of organic matter in some sampling points in both seasons, besides the absence of riparian vegetation in much of the Pardo River. The likely influence of industrial activities that do not have great prominence in the region was supported by temporal/spatial assessment of Cr and V. The water quality monitoring of Pardo River is an important tool for environmental management, and its continuity is indicated to obtain a consistent series of systematic data and thereby support concretely the actions of planning and controlling the use of water from the Pardo River and soil around them.

Copper and zinc affect the activity of plasma membrane H+-ATPase and thiol content in aquatic fungi.

Aquatic hyphomycetes are the major microbial decomposers of plant litter in streams. We selected three aquatic hyphomycete species with different abilities to tolerate, adsorb and accumulate copper and zinc, and we investigated the effects of these metals on H+-ATPase activity as well as on the levels of thiol (SH)-containing compounds. Before metal exposure, the species isolated from a metal-polluted stream (Heliscus submersus and Flagellospora curta) had higher levels of thiol compounds than the species isolated from a clean stream (Varicosporium elodeae). However, V. elodeae rapidly increased the levels of thiols after metal exposure, emphasizing the importance of these compounds in fungal survival under metal stress. The highest amounts of metals adsorbed to fungal mycelia were found in the most tolerant species to each metal, i.e. in H. submersus exposed to copper and in V. elodeae exposed to zinc. Short-term (10 min) exposure to copper completely inhibited the activity of H+-ATPase of H. submersus and V. elodeae, whilst zinc only led to a similar effect on H. submersus. However, at longer exposure times (8 days) the most metal-tolerant species exhibited increased H+-ATPase activities, suggesting that the plasma membrane proton pump may be involved in the acclimation of aquatic hyphomycetes to metals.

Integrating three tools for the environmental assessment of the Pardo River, Brazil.

There is a growing need for strategic assessment of environmental conditions in river basins around the world. In spite of the considerable water resources, Brazil has been suffering from water quality decrease in recent years. Pardo River runs through Minas Gerais and São Paulo, two of the most economically important states in Brazil, and is being currently promoted as a future drinking water source. This study aimed at integrating three different tools to conduct a hydromorphological assessment focused on the spatial complexity, connectivity, and dynamism of the Pardo River, Brazil. Twelve sampling stretches were evaluated in four sampling campaigns, in dry and rainy seasons. In each stretch, permanent preservation areas (PPAs), hydromorphological integrity by rapid assessment protocol (RAP), and physicochemical parameters were qualified. The kappa coefficient was used to assess statistical agreement among monitoring tools. The PPA analysis showed that in all stretches, the vegetation was modified. RAP results revealed environmental deterioration in stretches located near human activities and less variability of substrates available for aquatic fauna and sediment deposition as well. Low values for dissolved oxygen in the river mouth were noted in the rainy season. Electrical conductivity was higher in stretches near sugarcane crops. The poor agreement (k<0.35) between the RAP and physicochemical parameters indicates that the tools generate different and complementary information, while they are not replaceable. Potential changes of the hydromorphological characteristics and variations in physicochemical indicators must be related to extensive PPA modification.

Spatial accumulation of flood-driven riverside litter in two Northern Atlantic Rivers.

The escalation of litter accumulation in aquatic environments is recognized as an emerging global concern. Although rivers represent the main conduits for land-based waste into the oceans, the spatial dynamics of litter accumulation in these systems remain poorly investigated, especially after hydro-climatic extreme events. Floods have been identified as major drivers of litter mobilization, including macroplastics, within rivers. However, predicting flood-induced litter accumulation along riverbanks is complex due to the cumulative interplay of multiple environmental (geomorphological and riparian) and anthropogenic factors. Using empirical data collected from 14 stream reaches in two Northern Atlantic rivers in Portugal, our study evaluates which factors, among geomorphological, riparian, and anthropogenic descriptors, best drive riverside litter accumulation after floods. Taking into account the longitudinal gradient and the spatial heterogeneity of the studied reaches, our study enhances how the accumulation and characteristics (type, size) of riverside litter vary across a rural-urban continuum. Our model reveals that the combination of the human population density and the stream slope at river reach showed the highest explanatory power for the accumulation of riverside litter. Our findings indicate that litter tends to be retained close to the source, even under flood conditions. We also found that the structure of riparian vegetation showed low explanatory power for litter accumulation. However, riparian trapping could be influenced by litter input (density and type) which varies with anthropogenic activities. This work highlights the importance of gathering field data to identify critical areas of riverside litter accumulation within river basins. Our findings can further support environmental managers in designing and implementing effective cleanup campaigns and implementing plastic recovery strategies at specific areas. Nevertheless, it is crucial to enhance coordinated efforts across the entire value chain to reduce plastic pollution, promote innovative approaches for plastic litter valorization, and establish effective prevention pathways.

Assemblage and diversity of fungi on wood and seaweed litter of seven northwest portuguese beaches.

Three hundred and fifty woody litter and one hundred and forty seaweed litter sampled from seven beaches of Northwest Portugal were assessed for the filamentous fungal assemblage and diversity. The woody litter was screened for fungi up to 42 months using damp chamber incubation. They consisted of 36 taxa (ascomycetes, 21; basidiomycetes, 3; anamorphic taxa, 12) comprising 10 core group taxa (≥10%) (ascomycetes, 8; basidiomycete, 1; anamorphic taxa, 1). The total fungal isolates ranged between 150 and 243, while the number of fungal taxa per wood ranged between 3 and 4.9. The seaweed litter was screened up to four months in damp chamber incubation. They encompassed 29 taxa (ascomycetes, 16; basidiomycetes, 2; anamorphic taxa, 11) comprising 15 core group taxa (ascomycetes, 9; basidiomycete, 1; anamorphic taxa, 5). Total fungal isolates ranged between 56 and 120, while the number of fungal taxa per seaweed segment ranged between 4.8 and 6.3. Fifteen taxa of ascomycetes, two of basidiomycetes, and four anamorphic taxa were common to wood and seaweed litter. On both the substrates, two arenicolous fungi Arenariomyces trifurcates and Corollospora maritima were the predominant fungi (72.6-85.9%). The species abundance curves showed higher frequency of occurrence of fungal taxa in seaweed than woody litter. Our study revealed rich assemblage and diversity of marine fungi on wood and seaweed litter of Northwest Portugal beaches. The fungal composition and diversity of this survey have been compared with earlier investigations on marine fungi of Portugal coast.

Remote sensing depicts riparian vegetation responses to water stress in a humid Atlantic region.

Riparian areas in the Cantabrian Atlantic ecoregion (northwest Portugal) play a key role in soil formation and conservation, regulation of nutrient and water cycle, creation of landscape aesthetic value and the preservation of biodiversity. The maintenance of their ecological integrity is crucial given the ever increase in multiple anthropogenic (water demand and agriculture) and climatic pressures (droughts and extreme events). We developed a transferable remote sensing approach, taking advantage of the latest freely available technologies (Sentinel-2 and Copernicus Land products), to detect intra-annual and inter-annual changes in riparian vegetation productivity at the river basin scale related to water stress. This study has used the normalized difference vegetation index (NDVI) to investigate riparian vegetation productivity dynamics on three different vegetation types (coniferous, broadleaved and grassland) over the past 5 years (2015-2019). Our results indicated that inter-annual seasonality differed between drier (2017) and wetter (2016) years. We found that intra-annual dynamics of NDVI were influenced by the longitudinal river zonation. Our model ranked first (r2m = 0.73) showed that the productivity of riparian vegetation during the dry season was positively influenced by annual rainfall and by the type of riparian vegetation. The emergent long lags between climatic variation and riparian plant productivity provides opportunities to forecast early warnings of climatically-driven impacts. In addition, the different average productivity levels among vegetation types should be considered when assessing climatic impacts on riparian vegetation. Future applications of Sentinel 2 products could seek to distinguish riparian areas that are likely to be more vulnerable to changes in the annual water balance from those that are more resistant under longer-term changes in climate.

Quantitative analysis of proton movements associated with the uptake of weak carboxylic acids. The yeast Candida utilis as a model.

A quantitative analysis of the proton movements associated with the initial uptake rate of weak short-chain carboxylic acids was developed in order to estimate proton/carboxylate symports stoichiometries. The yeast Candida utilis was used as a biological model and the deduced equations were applied on the elucidation of the proton/carboxylate symports stoichiometries of lactate, succinate and citrate in a strain of that yeast species at different pH values. At pH 5.0, the proton/lactate and the proton/succinate symport stoichiometry was 1:1. In the cases of the proton/lactate and proton/citrate symports it appears that the stoichiometry ratio increased with increasing extracellular pH.

Utilization and transport of acetic acid in Dekkera anomala and their implications on the survival of the yeast in acidic environments.

The yeast Dekkera anomala IGC 5153 exhibited a restricted ability to use weak acids as the only carbon and energy sources. Of the monocarboxylic, dicarboxylic, and tricarboxylic acids tested, only acetic acid was used in such a way. The cells were able to grow at acetic acid concentrations of 0.1 to 3% (vol/vol) over a pH range of 3.5 to 5.5, and the specific growth rates decreased exponentially with the increase of the undissociated acetic acid concentration in the culture medium. Transport assays carried out in cells that exhibited higher specific growth rates showed the presence of an acetate-proton symport associated with a simple diffusion component of the undissociated acetic acid, the weight of the latter increasing with the undissociated acid concentration in the culture media. The acetate carrier was shared by propionic, formic, and sorbic acids and was inducible and repressed by glucose and concentrations of undissociated acetic acid in the culture medium above 0.3% (vol/vol). In undissociated acetic acid repression conditions, the lowest values for the yeast specific growth rates were obtained, and the simple diffusion of the undissociated acid was the only mechanism involved in the acetic acid uptake by the cells. The results will be discussed in terms of the high tolerance of D. anomala to the acidic stress conditions present in wine.

Impacts of warming on aquatic decomposers along a gradient of cadmium stress.

We evaluated the effects of cadmium and temperature on plant-litter decomposition by examining diversity and activity of aquatic fungi and leaf consumption by Limnephilus sp., a typical invertebrate shredder of Iberian streams. Freshly fallen leaves were immersed in a stream to allow microbial colonization, and were exposed in microcosms to a gradient of cadmium (≤11 levels, ≤35 mg L(-1)). Microcosms were kept at 15 °C, a temperature typically found in Iberian streams in autumn, and at 21 °C to simulate a warming scenario. The increase in temperature stimulated leaf decomposition by microbes, fungal reproduction and leaf consumption by the shredder. Conversely, increased cadmium concentrations inhibited fungal reproduction and diversity, and leaf consumption by the invertebrate. Cadmium concentration inhibiting 50% of fungal reproduction, microbial decomposition and leaf consumption by the shredder was higher at 15 °C than at 21 °C, suggesting that higher temperatures can lead to increased metal toxicity to aquatic decomposers.

Low- and high-affinity transport systems for citric acid in the yeast Candida utilis.

Citric acid-grown cells of the yeast Candida utilis induced two transport systems for citric acid, presumably a proton symport and a facilitated diffusion system for the charged and the undissociated forms of the acid, respectively. Both systems could be observed simultaneously when the transport was measured at 25 degrees C with labelled citric acid at pH 3.5 with the following kinetic parameters: for the low-affinity system, Vmax, 1.14 nmol of undissociated citric acid s-1 mg (dry weight) of cells-1, and Km, 0.59 mM undissociated acid; for the high-affinity system, Vmax, 0.38 nmol of citrate s-1 mg (dry weight) of cells-1, and Km, 0.056 mM citrate. At high pH values (above 5.0), the low-affinity system was absent or not measurable. The two transport systems exhibited different substrate specificities. Isocitric acid was a competitive inhibitor of citric acid for the high-affinity system, suggesting that these tricarboxylic acids used the same transport system, while aconitic, tricarballylic, trimesic, and hemimellitic acids were not competitive inhibitors. With respect to the low-affinity system, isocitric acid, L-lactic acid, and L-malic acid were competitive inhibitors, suggesting that all of these mono-, di-, and tricarboxylic acids used the same low-affinity transport system. The two transport systems were repressed by glucose, and as a consequence diauxic growth was observed. Both systems were inducible, and not only citric acid but also lactic acid and malic acid may induce those transport systems. The induction of both systems was not dependent on the relative concentration of the anionic form(s) and of undissociated citric acid in the culture medium.(ABSTRACT TRUNCATED AT 250 WORDS)

A comparative study on the transport of L(-)malic acid and other short-chain carboxylic acids in the yeast Candida utilis: evidence for a general organic acid permease.

Cells of the yeast Candida utilis grown in medium with short-chain mono-, di- or tricarboxylic acids transported L(-)malic acid by two transport systems at pH 3.0. Results indicate that probably a proton symport for the ionized form of the acid and a facilitated diffusion for the undissociated form were present. Dicarboxylic acids such as succinic, fumaric, oxaloacetic and alpha-ketoglutaric acids were competitive inhibitors of the malic acid for the high-affinity system, suggesting that these acids used the same transport system. In turn, competitive inhibition uptake studies of labelled carboxylic acid in the low-affinity range indicated that this system was non-specific and able to accept not only carboxylic (mono-, di- or tri-) acids but also some amino acids. Additionally, under the same growth conditions, C. utilis produced two mediated transport systems for lactic acid: a proton symport for the anionic form which appeared to be a common monocarboxylate carrier and a facilitated diffusion system for the undissociated acid displaying a substrate specificity similar to that observed for the low-affinity dicarboxylic acid transport. The mediated carboxylic acid transport systems were inducible and subjected to repression by glucose. In glucose-grown cells the undissociated dicarboxylic acids entered the cells slowly by simple diffusion. Repressed glucose-grown cells were only able to produce both transport systems if an inducer, at low concentration (0.5%, w/v), was present during starvation in buffer. This process was inhibited by the presence of cycloheximide indicating that induction requires de novo protein synthesis. If a higher acid concentration was used, only the low-affinity transport system was detectable, showing that the high-affinity system was also repressed by high concentrations of the inducer.

Reconstitution of lactate proton symport activity in plasma membrane vesicles from the yeast Candida utilis.

Lactic acid transport was studied in plasma membrane vesicles from the yeast Candida utilis IGC 3092 which were fused with liposomes containing cytochrome c oxidase. After the addition of an electron donor system, these hybrid membrane vesicles were able to generate a proton-motive force of about--150 mV, inside alkaline and negative. In vesicles prepared from lactic acid-grown cells, the uptake of labelled lactic acid, at pH 6.2, under energized conditions, was expressed by a kinetics consistent with the involvement of a mediated transport system. This carrier exhibited a substrate specificity pattern identical to the one found for the lactate-proton symport in intact cells. The transport of labelled lactic acid was accumulative and strongly sensitive to the effects of the protonophore carbonyl cyanide p-(trifluoromethoxy)phenylhydrazone, consistent with the involvement of the proton-motive force in acid uptake, hence with the presence of a proton symport for lactate. Dissipation of the transmembrane electric potential by valinomycin did not have a significant effect on lactate accumulation, whereas abolishing the transmembrane pH gradient (delta pH) by nigericin prevented the accumulation and led to a rapid efflux of the accumulated acid. The data support that the delta pH is the main component of the proton-motive force involved in the transport of the acid and its accumulation. The lactate-proton symport stoichiometry was 1:1, being independent of the pH. Vesicles prepared from glucose-grown cells did not display the capacity to transport and accumulate lactate. However, activity for the carrier was also reconstituted in vesicles obtained from glucose-grown cells after incubation in buffer containing lactic acid. These results were consistent with those obtained in intact cells, which demonstrated that the lactate-proton symport of the yeast C. utilis is inducible.

Effects of zinc on leaf decomposition by fungi in streams: studies in microcosms.

The effect of zinc on leaf decomposition by aquatic fungi was studied in microcosms. Alder leaf disks were precolonized for 15 days at the source of the Este River and exposed to different zinc concentrations during 25 days. Leaf mass loss, fungal biomass (based on ergosterol concentration), fungal production (rates of [1-14C]acetate incorporation into ergosterol), sporulation rates, and species richness of aquatic hyphomycetes were determined. At the source of the Este River decomposition of alder leaves was fast and 50% of the initial mass was lost in 25 days. A total of 18 aquatic hyphomycete species were recorded during 42 days of leaf immersion. Articulospora tetracladia was the dominant species, followed by Lunulospora curvula and two unidentified species with sigmoid conidia. Cluster analysis suggested that zinc concentration and exposure time affected the structure of aquatic hyphomycete assemblages, even though richness had not been severely affected. Both zinc concentration and exposure time significantly affected leaf mass loss, fungal production and sporulation, but not fungal biomass. Zinc exposure reduced leaf mass loss, inhibited fungal production and affected fungal reproduction by either stimulating or inhibiting sporulation rates. The results of this work suggested zinc pollution might depress leaf decomposition in streams due to changes in the structure and activity of aquatic fungi.

Transport of lactate and other short-chain monocarboxylates in the yeast Saccharomyces cerevisiae.

Saccharomyces cerevisiae IGC4072 grown in lactic acid medium transported lactate by an accumulative electroneutral proton-lactate symport with a proton-lactate stoichiometry of 1:1. The accumulation ratio measured with propionate increased with decreasing pH from ca. 24-fold at pH 6.0 to ca. 1,400-fold at pH 3.0. The symport accepted the following monocarboxylates (Km values at 25 degrees C and pH 5.5): D-lactate (0.13 mM), L-lactate (0.13 mM), pyruvate (0.34 mM), propionate (0.09 mM), and acetate (0.05 mM), whereas apparently a different proton symport accepted formate (0.13 mM). The lactate system was inducible and was subject to glucose repression. Undissociated lactic acid entered the cells by simple diffusion. The permeability of the plasma membrane for undissociated lactic acid increased exponentially with pH, and the diffusion constant increased 40-fold when the pH was increased from 3.0 to 6.0.