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The plastisphere, a unique microbial biofilm community colonizing plastic debris and microplastics (MPs) in aquatic environments, has attracted increasing attention owing to its ecological and public health implications. This review consolidates current state of knowledge on freshwater plastisphere, focussing on its biodiversity, community assembly, and interactions with environmental factors. Current biomolecular approaches revealed a variety of prokaryotic and eukaryotic taxa associated with plastic surfaces. Despite their ecological importance, the presence of potentially pathogenic bacteria and mobile genetic elements (i.e., antibiotic resistance genes) raises concerns for ecosystem and human health. However, the extent of these risks and their implications remain unclear. Advanced sequencing technologies are promising for elucidating the functions of plastisphere, particularly in plastic biodegradation processes. Overall, this review emphasizes the need for comprehensive studies to understand plastisphere dynamics in freshwater and to support effective management strategies to mitigate the impact of plastic pollution on freshwater resources.
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In last 10 years, the interest about the presence of microplastics (MPs) in the environment has strongly grown. Wastewaters function as a carrier for MPs contamination from source to the aquatic environment, so the knowledge of the fate of this emerging contaminant in wastewater treatment plants (WWTPs) is a priority. This work aims to review the presence of MPs in the influent wastewater (WW) and the effectiveness of the treatments of conventional WWTPs. Moreover, the negative impacts of MPs on the management of the processes have been also discussed. The work also focuses on the possible approaches to tackle MPs contamination enhancing the effectiveness of the WWTPs. Based on literature results, despite WWTPs are not designed for MPs removal from WW, they can effectively remove the MPs (up to 99 % in some references). Nevertheless, they normally act as "hotspots" of MPs contamination considering the remaining concentration of MPs in WWTPs' effluents can be several orders of magnitude higher than receiving waters. Moreover, MPs removed from WW are concentrated in sewage sludge (potentially >65 % of MPs entering the WWTP) posing a concern in case of the potential reuse as a soil improver. This work aims to present a paradigm shift intending WWTPs as key barriers for environmental protection. Approaches for increasing effectiveness against MPs have been discussed in order to define the optimal point(s) of the WWTP in which these technologies should be located. The need of a future legislation about MPs in water and sludge is discussed.
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The monitoring of plastics in freshwater ecosystems has witnessed a significant increase in recent years, driven by the awareness that approximately 80 % of marine plastic litter originates from terrestrial sources transported to the seas through lakes and rivers. Consequently, it is imperative to develop monitoring plans that offer a comprehensive understanding of plastic contamination in these aquatic environments, given their seasonal variations in hydrochemical characteristics and anthropogenic sources. Historically, most global lake monitoring campaigns have been limited to one-time or, at most, seasonal sampling. In this context, the primary objective of the present study was to assess the quantitative and qualitative monthly variations of floating plastics in Lake Maggiore, a large European lake with high ecological and economic significance. Twelve transverse transects were conducted from January to December 2022 using a Manta-net with a 100 µm mesh. Characterization of each plastic particle was performed using a µ-Fourier Transform Infrared Spectroscope (µFT-IR). The results revealed relatively low levels of contamination in Lake Maggiore when compared with other lakes worldwide exclusively from a secondary origin. However, a considerable heterogeneity was observed, both quantitatively and qualitatively. Notably, we identified a 13-fold difference between the minimum (0.02 plastics/m3 in September) and maximum (0.29 plastics/m3 in December) concentrations of plastics, accompanied by significant variations in polymer composition. Our monitoring underscored the necessity of also considering the temporal variation as a potential factor influencing plastic contamination in a lake. Moreover, frequent sampling emerged as a crucial requirement to accurately gauge the extent of plastic pollution, yielding robust and valuable data essential for effective environmental management.
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Lagos , Contaminantes Químicos del Agua , Plásticos , Ecosistema , Contaminantes Químicos del Agua/análisis , Monitoreo del Ambiente , ItaliaRESUMEN
Microplastic particles are persistent micropollutants that provide a substrate for the growth of bacterial biofilms, posing a threat to the environment. This study explores the changes in commercially available food containers made of conventional (polypropylene PP, polyethylene terephthalate PET), innovative biodegradable (Mater-Bi) and natural (wood and cellulose) materials, when introduced in the surface waters of Lake Maggiore for 43 days. Spectral changes revealed by FT-IR spectroscopy in PET and Mater-Bi, and changes in thermal properties of all human-made material tested indicated a degradation process occurred during environmental exposure. Despite similar bacterial richness, biofilms on PET, PP, and Mater-Bi differed from natural material biofilms and the planktonic community. Human-made material communities showed a higher proportion of potential pathogens, with PET and PP also exhibiting increased abundances of antibiotic resistance genes. Overall, these findings stress the need for dedicated strategies to curb the spread of human-made polymers in freshwaters, including innovative materials that, due to their biodegradable properties, might be perceived less hazardous for the environment.
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Plásticos Biodegradables , Humanos , Plásticos , Plancton , Espectroscopía Infrarroja por Transformada de Fourier , Polipropilenos , Microplásticos , BacteriasRESUMEN
Plastic debris is thought to be widespread in freshwater ecosystems globally1. However, a lack of comprehensive and comparable data makes rigorous assessment of its distribution challenging2,3. Here we present a standardized cross-national survey that assesses the abundance and type of plastic debris (>250 µm) in freshwater ecosystems. We sample surface waters of 38 lakes and reservoirs, distributed across gradients of geographical position and limnological attributes, with the aim to identify factors associated with an increased observation of plastics. We find plastic debris in all studied lakes and reservoirs, suggesting that these ecosystems play a key role in the plastic-pollution cycle. Our results indicate that two types of lakes are particularly vulnerable to plastic contamination: lakes and reservoirs in densely populated and urbanized areas and large lakes and reservoirs with elevated deposition areas, long water-retention times and high levels of anthropogenic influence. Plastic concentrations vary widely among lakes; in the most polluted, concentrations reach or even exceed those reported in the subtropical oceanic gyres, marine areas collecting large amounts of debris4. Our findings highlight the importance of including lakes and reservoirs when addressing plastic pollution, in the context of pollution management and for the continued provision of lake ecosystem services.
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Lagos , Plásticos , Contaminación del Agua , Abastecimiento de Agua , Ecosistema , Lagos/química , Plásticos/análisis , Plásticos/clasificación , Contaminación del Agua/análisis , Contaminación del Agua/estadística & datos numéricos , Encuestas y Cuestionarios , Urbanización , Actividades HumanasRESUMEN
Salmonid species are main actors in the Italian socio-ecological landscape of inland fisheries. We present novel data on the size-age structure of one of the remnant Italian populations of the critically endangered marble trout Salmo marmoratus, which co-occurs with other stocked non-native salmonids in a large glacial river of the Lake Maggiore basin (Northern Italy-Southern Switzerland). Like other Italian native trout populations, the Toce River marble trout population is affected by anthropogenic introgression with the non-native brown trout S. trutta. Our sample includes 579 individuals, mainly collected in the Toce River main channel. We estimated the length-weight relationship, described the population size-age structure, estimated the age-specific growth trajectories, and fit an exponential mortality model. A subset of the sample was also used to measure numerical and biomass density. The estimated asymptotic maximum length is ~105 cm total length (TL). Mean length at first maturity is ~55 cm TL, and mean length at maximum yield per recruit is ~68 cm TL. Approximately 45-70% of the population are estimated to die annually, along with a fishing annual mortality of ~37%, with an exploitation ratio of ~0.5. The frequency distribution of length classes in a sample collected by angling shows that ~80% of the individuals that could be retained according to the current recreational fishing regulations likely never reproduced, and large fish disproportionally contributing to recruitment are fished and retained. We identify possible overfishing risks posed by present regulations, and propose updated harvest-slot length limits to mitigate such risks. More detailed and long-term datasets on this system are needed to more specifically inform the fishery management and monitor the effects of any change in the management strategy on the size-age structure of the marble trout population of the Toce River.
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Salmo salar , Trucha , Animales , Conservación de los Recursos Naturales , Explotaciones Pesqueras , ItaliaRESUMEN
Stochastic or deterministic processes control the bacterial community assembly in waters and their understanding is a fundamental question to correctly manage aquatic environments exposed to the release of antibiotics from anthropogenic sources. It has been suggested that microdiversity (i.e. the rare biosphere) convers freshwater communities with stability, meaning that previously rare taxa bloom when the community is disturbed. Since there might be a seed bank of similar, but not abundant, bacterial taxa in different waters, we tested whether a disturbance by an antibiotic cocktail would increase similarity in bacterial communities from different freshwater systems (a wastewater effluent and two lakes). In a continuous culture set-up in chemostats, we show that disturbance with antibiotics causes communities from different environments to become more similar. Once the antibiotic pressure is released the communities tend to become more dissimilar again. This shows that there is a similar shift in community composition even in waters from very different origins when they are disturbed by antibiotics, even at low concentrations. Antibiotics impact the bacterial communities at the cell and the community level, independently by the original degree of anthropogenic stress they are adapted to, altering the original phenotypes, genotypes, and the relations between bacteria.
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Antibacterianos , Microbiota , Antibacterianos/toxicidad , Bacterias/genética , Lagos/microbiología , Aguas ResidualesRESUMEN
The risk for human health posed by polluted aquatic environments, and especially those carrying antibiotic resistance genes (ARGs) of clinical interest, is still debated. This is because of our limited knowledge of the dynamics of antimicrobial resistance in the environment, the selection mechanisms underlying the spread of ARGs, and the ecological factors potentially favoring their return to humans. The Class 1 integron is one of the most effective platforms for the dissemination of ARGs. In this study we investigated a freshwater system consisting of a lake-river-lake continuum, determining the abundance of class 1 integrons and their associated ARGs by a modulated metagenomic approach. Bacterial abundance and community composition were used to identify the potential carriers of class 1 integrons and their associated ARGs over a period of six months. Class 1 integrons and their ARG cargoes were significantly more abundant in riverine sampling sites receiving treated wastewater. Further, class 1 integrons carried ARGs ranked at the highest risk for human health (e.g., catB genes), in particular, genes encoding resistance to aminoglycosides. Genera of potential pathogens, such as Pseudomonas and Escherichia-Shigella, were correlated with class 1 integrons. The lake-river-lake system demonstrated a clear relationship between the integrase gene of class 1 integrons (intI1) and anthropogenic impact, but also a strong environmental filtering that favored the elimination of intI1 once the human derived stressors were reduced. Overall, the results of this study underline the role class 1 integrons as proxy of anthropogenic pollution and suggest this genetic platform as an important driver of aminoglycoside resistance genes, including high risk ARGs, of potential concern for human health.
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Antibacterianos , Integrones , Humanos , Integrones/genética , Antibacterianos/farmacología , Efectos Antropogénicos , Farmacorresistencia Bacteriana/genética , Genes Bacterianos , LagosRESUMEN
Research on microplastics (MPs) in the terrestrial environment is currently at a still embryonal stage. The current knowledge concerning poorly known diffuse sources of MPs pollution in terrestrial ecosystems have been considered in this work. In addition, a particular focus on the presence, mechanism of absorption and effects of MPs in plants has also been provided. Research concerning microplastics in urban areas and their intake by Tyre and Road Wear Particulates (TWRP) demonstrated a high contribution of this plastic debris to microplastic pollution, although quantification of these inputs is challenging to assess because studies are still very few. Around 50% of particles are expected to remain in the roadside soil, while the rest is transported away by the runoff with high concentrations of TRWP with a size ranging between 0.02 and 0.1 mm. Natural and anthropic environments like temporary ponds, stormwater retention ponds and small waterbodies were considered sensitive connecting ecosystems rich in biodiversity between terrestrial and aquatic environments. Even if studies are not yet exhaustive and just eight studies were currently published concerning these ecosystems, considerable values of MPs were already observed both in the sediment and water phase of ponds. Although still poorly explored, agricultural environments were already demonstrated to hide a significant number of microplastics linked mainly to agricultural activities and practices (e.g. mulch, sewage and compost fertilisation). However, the microplastics transportation processes into the soil are still understudied, and a few works are available. Microplastics and primarily nanoplastics presence was also observed in common edible plants (fruit and vegetables) with alarming Estimated Daily Intakes ranging from 2.96 × 1004 to 4.62 × 1005 (p kg-1 day-1) for adults depending on species. In addition, adverse effects on plants growth, photosynthetic activity, antioxidant system and nutritional values of several common fruits and vegetables were also demonstrated by several studies.
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Microplásticos , Plásticos , Ecosistema , Contaminación Ambiental , SueloRESUMEN
Wastewater treatment plants (WWTPs) efficiently retain microplastic particles (MPs) generated within urban areas. Among the wastewater treatment steps, disinfection has not been characterized for its potential MPs retention activity, although it has been reported that processes used to abate the bacterial load could also affect MPs concentration. For this reason, we evaluated the MPs concentration across the overall wastewater treatment process and before and after the disinfection step in four small/medium WWTPs located in the north of Italy. Most of the MPs found in the samples were fibers or fragments, smaller than 500 µm, mainly composed of polyethylene, polypropylene, or polyethylene terephthalate. The retention efficiency at the outlets was higher than 94% in all the plants analyzed. More interestingly, the disinfection treatments adopted by the different WWTPs reduced MPs concentration from a minimum of 9.1% (UV treatment) to a maximum of 67.6% (chlorination), promoting a further increase of the overall retention efficiency of the WWTPs from 0.4 to 0.7%. Quantitatively, the disinfection contributes to the MPs reduction in the outlets by retaining 0.5-6.7 million MPs per day, in WWTPs that discharge 2.7-12 million MPs per day. The results of the present work underline the importance of a careful choice of the steps that constitute the wastewater treatment, including disinfection, in order to minimize MPs discharge into the natural ecosystems.
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Contaminantes Químicos del Agua , Purificación del Agua , Desinfección , Ecosistema , Monitoreo del Ambiente , Microplásticos , Plásticos , Eliminación de Residuos Líquidos , Aguas Residuales , Contaminantes Químicos del Agua/análisisRESUMEN
Microplastic particles (MPs) contamination of aquatic environments has raised a growing concern in recent decades because of their numerous potential toxicological effects. Although fish are among the most studied aquatic organisms, reports on MPs ingestion in freshwater environments are still scarce. Thus, there is still much to study to understand the uptake mechanisms, their potential accumulation among the food webs and their ecotoxicological effects. Here, MPs presence in the digestive system of one of the most widespread and commercially exploited freshwater fish, the perch (Perca fluviatilis, Linnaeus 1758), was investigated in four different south-alpine lakes, to assess the extent of ingestion and evaluate its relation to the body health condition. A total of 80 perch specimen have been sampled from the Italian lakes Como, Garda, Maggiore and Orta. Microplastic particles occurred in 86% of the analysed specimens, with average values ranging from 1.24 ± 1.04 MPs fish-1 in L. Como to 5.59 ± 2.61 MPs fish-1 in L. Garda. The isolated particles were mainly fragments, except in L. Como where films were more abundant. The most common polymers were polyethylene, polyethylene terephthalate, polyamide, and polycarbonate, although a high degree of degradation was found in 43% of synthetic particles, not allowing their recognition up to a single polymer. Despite the high number of ingested MPs, fish health (evaluated by means of Fulton's body condition and hepatosomatic index) was not affected. Instead, fullness index showed an inverse linear relationship with the number of ingested particles, which suggests that also in perch MPs presence could interfere with feeding activity, as already described for other taxa.
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Percas , Contaminantes Químicos del Agua , Animales , Monitoreo del Ambiente , Italia , Lagos , Microplásticos , Plásticos , Contaminantes Químicos del Agua/análisisRESUMEN
Microplastic Particles (MPs) are ubiquitous pollutants widely found in aquatic ecosystems. Although MPs are mostly retained in wastewater treatment plants (WWTPs), a high number of MPs reaches the open waters potentially contributing to the spread of pathogenic bacteria and antibiotic resistance genes in the environment. Nowadays, a limited number of studies have focused on the role of MPs as carriers of potentially pathogenic and antibiotic resistant bacteria in WWTPs. Thus, an investigation on the community composition (by 16S rRNA gene amplicon sequencing) and the abundance of antibiotic and metal resistance genes (by qPCR) of the biofilm on MPs (the plastisphere) and of planktonic bacteria in treated (pre- and post-disinfection) wastewaters was performed. MPs resulted to be very similar in terms of type, color, size, and chemical composition, before and after the disinfection. The bacterial community on MPs differed from the planktonic community in terms of richness, composition, and structure of the community network. Potentially pathogenic bacteria generally showed higher abundances in treated wastewater than in the biofilm on MPs. Furthermore, among the tested resistance genes, only sul2 (a common resistance gene against sulfonamides) resulted to be more abundant in the plastisphere than in the planktonic bacterial community. Our results suggest that the wastewater plastisphere could promote the spread of pathogenic bacteria and resistance genes in aquatic environment although with a relatively lower contribution than the wastewater planktonic bacterial community.
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Microplásticos , Aguas Residuales , Antibacterianos , Bacterias/genética , Ecosistema , Genes Bacterianos , Plásticos , ARN Ribosómico 16S/genéticaRESUMEN
BACKGROUND: Oleaginous yeasts are able to accumulate very high levels of neutral lipids especially under condition of excess of carbon and nitrogen limitation (medium with high C/N ratio). This makes necessary the use of two-steps processes in order to achieve high level of biomass and lipid. To simplify the process, the decoupling of lipid synthesis from nitrogen starvation, by establishing a cytosolic acetyl-CoA formation pathway alternative to the one catalysed by ATP-citrate lyase, can be useful. RESULTS: In this work, we introduced a new cytoplasmic route for acetyl-CoA (AcCoA) formation in Rhodosporidium azoricum by overexpressing genes encoding for homologous phosphoketolase (Xfpk) and heterologous phosphotransacetylase (Pta). The engineered strain PTAPK4 exhibits higher lipid content and produces higher lipid concentration than the wild type strain when it was cultivated in media containing different C/N ratios. In a bioreactor process performed on glucose/xylose mixture, to simulate an industrial process for lipid production from lignocellulosic materials, we obtained an increase of 89% in final lipid concentration by the engineered strain in comparison to the wild type. This indicates that the transformed strain can produce higher cellular biomass with a high lipid content than the wild type. The transformed strain furthermore evidenced the advantage over the wild type in performing this process, being the lipid yields 0.13 and 0.05, respectively. CONCLUSION: Our results show that the overexpression of homologous Xfpk and heterologous Pta activities in R. azoricum creates a new cytosolic AcCoA supply that decouples lipid production from nitrogen starvation. This metabolic modification allows improving lipid production in cultural conditions that can be suitable for the development of industrial bioprocesses using lignocellulosic hydrolysates.
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Basidiomycota/metabolismo , Lignina/metabolismo , Lípidos/biosíntesis , Ingeniería Metabólica/métodos , Acetilcoenzima A/metabolismo , Aldehído-Liasas/genética , Aldehído-Liasas/metabolismo , Bacillus subtilis/genética , Biomasa , Citoplasma/metabolismo , Proteínas Fúngicas/genética , Genes Bacterianos , Genes Fúngicos , Ingeniería Genética , Recombinación Homóloga , Metabolismo de los Lípidos/genética , Nitrógeno/metabolismo , Fosfato Acetiltransferasa/genética , Fosfato Acetiltransferasa/metabolismo , Proteínas Recombinantes , TransfecciónRESUMEN
Plastic debris are at present recognized as an emerging potential threat for natural environments, wildlife and humans. In the past years an increasing attention has been addressed to investigate the presence and concentration of plastic debris in the ecosystems, including surface waters. Scientific literature extensively describes the ingestion by aquatic fauna, the transfer into food webs and the potential action as a vector for toxic compounds or alien microorganisms. Although the scientific community addresses this issue with considerable effort, many questions remain open. In particular, new sources of microplastics have been recently recognized, possibly representing major environmental inputs compared to those previously considered. In addition to the already renowned sources such as the embrittlement of plastic litter and microbeads released from personal care products, microplastic can be released also by washing of synthetic clothes, abrasion of vehicles tyres and from the weathering of different kind of paints. This review tries to exhaustively enumerate all the possible sources of plastic litter that have been identified so far and to report quantitative assessments of their inputs on microplastics pollution to natural environments reported in scientific and grey literature, with an emphasis on surface waters.
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Treated wastewater discharged into the environment acts as a disturbance of the natural microbial communities in terms of taxonomic composition and of functional gene pool, including antibiotic resistance genes. We tested whether stochastic and heterogeneous site-specific trajectories or generalities, potentially driven by deterministic processes, control the fate of allochthonous bacteria from anthropogenic sources and the persistence of their functional traits in freshwater. Finding generalities would allow the identification of wastewater treatments that could be effective in abating determinants of antibiotic resistance. We analysed the short-term response of native bacterial communities in waters exposed to the disturbance of wastewater at different dilutions, using a metagenomic approach that revealed both microbial community composition and the scope and abundance of the resistome that can pose indirect risks to human health. We found that the taxonomic composition of the communities after the disturbance was driven by case-specific stochastic processes, whereas the resistome had a deterministic trajectory, rapidly stabilising its functional traits with higher proportions of wastewater effluents, regardless of differences in taxonomic composition, richness of antibiotic resistance genes and of bacterial taxa, phenotypic features of the bacterial communities, and type of wastewater treatment. The observed deterministic proliferation of resistomes in freshwater bodies receiving wastewater effluents, suggests that this process may contribute to the global propagation of antibiotic resistance, and thus calls for new legislations promoting alternative tertiary treatments for the wastewater reuse, and targeting bacterial functional traits and not only bacterial abundances.
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Antibacterianos , Aguas Residuales , Farmacorresistencia Microbiana , Agua Dulce , Genes Bacterianos , HumanosRESUMEN
The influence of cultural conditions on lipid production was investigated in two species, Trichosporon oleaginosus and Rhodosporidium azoricum. We showed that nitrogen limitation is not the main factor triggering the mechanism of lipid accumulation in T. oleaginosus. Moreover, a scarce availability of oxygen negatively affected lipid synthesis to a lesser extent in T. oleaginosus than in R. azoricum. This highlights how the importance of controlling fermentation parameters is strictly linked to the yeast species employed. We showed that these parameters affect the activity of important enzymes, influencing the metabolic fluxes into different pathways, in particular pentose phosphate pathway and cytoplasmic pyruvate bypass. Furthermore, T. oleaginosus exhibited wider substrate flexibility, faster growth and higher lipid accumulation in fed-batch cultivation. Microbial oils obtained from both yeasts proved a valuable feedstock, alternative to vegetable oils, for advanced diesel biofuel production.
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Basidiomycota , Lípidos , Trichosporon , Aceites , LevadurasRESUMEN
The Crabtree positive yeasts, such as Saccharomyces cerevisiae, prefer fermentation to respiration, even under fully aerobic conditions. The selective pressures that drove the evolution of this trait remain controversial because of the low ATP yield of fermentation compared to respiration. Here we propagate experimental populations of the weak-Crabtree yeast Lachancea kluyveri, in competitive co-culture with bacteria. We find that L. kluyveri adapts by producing quantities of ethanol lethal to bacteria and evolves several of the defining characteristics of Crabtree positive yeasts. We use precise quantitative analysis to show that the rate advantage of fermentation over aerobic respiration is insufficient to provide an overall growth advantage. Thus, the rapid consumption of glucose and the utilization of ethanol are essential for the success of the aerobic fermentation strategy. These results corroborate that selection derived from competition with bacteria could have provided the impetus for the evolution of the Crabtree positive trait.
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Bacterias/crecimiento & desarrollo , Evolución Molecular , Saccharomycetales/crecimiento & desarrollo , Aerobiosis , Técnicas de Cultivo Celular por Lotes , Técnicas de Cocultivo , Etanol/metabolismo , Glucólisis , Mutación INDEL , Polimorfismo de Nucleótido Simple , ARN de Hongos/química , ARN de Hongos/aislamiento & purificación , ARN de Hongos/metabolismo , Saccharomycetales/genética , Saccharomycetales/metabolismo , Análisis de Secuencia de ARN , TranscriptomaRESUMEN
Vinylphenol reductase of Dekkera bruxellensis, the characteristic enzyme liable for "Brett" sensory modification of wine, has been recently recognized to belong to the short chain dehydrogenases/reductases family. Indeed, a preliminary biochemical characterisation has conferred to the purified protein a dual significance acting as superoxide dismutase and as a NADH-dependent reductase. The present study aimed for providing a certain identification of the enzyme by cloning the VPR gene in S. cerevisiae, a species not producing ethyl phenols. Transformed clones of S. cerevisiae resulted capable of expressing a biologically active form of the heterologous protein, proving its role in the conversion of 4-vinyl guaiacol to 4-ethyl guaiacol. A VPR specific protein activity of 9 ± 0.6 mU/mg was found in crude extracts of S. cerevisiae recombinant strain. This result was confirmed in activity trials carried out with the protein purified from transformant cells of S. cerevisiae by a his-tag purification approach; in particular, VPR-enriched fractions showed a specific activity of 1.83 ± 0.03 U/mg at pH 6.0. Furthermore, in agreement with literature, the purified protein behaves like a SOD, with a calculated specific activity of approximatively 3.41 U/mg. The comparative genetic analysis of the partial VPR gene sequences from 17 different D. bruxellesis strains suggested that the observed polymorphism (2.3%) and the allelic heterozygosity state of the gene do not justify the well described strain-dependent character in producing volatile phenols of this species. Actually, no correlation exists between genotype membership of the analysed strains and their capability to release off-flavours. This work adds valuable knowledge to the study of D. bruxellensis wine spoilage and prepare the ground for interesting future industrial applications.
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Dekkera/genética , Oxidorreductasas/genética , Saccharomyces cerevisiae/genética , Clonación Molecular , Dekkera/enzimología , Fermentación , Microbiología de Alimentos , Genotipo , Oxidorreductasas/química , Oxidorreductasas/metabolismo , Fenoles/metabolismo , Polimorfismo Genético , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Saccharomyces cerevisiae/metabolismo , Vino/análisisRESUMEN
Dekkera bruxellensis and Saccharomyces cerevisiae are considered two phylogenetically distant relatives, but they share several industrial relevant traits such as the ability to produce ethanol under aerobic conditions (Crabtree effect), high tolerance towards ethanol and acids, and ability to grow without oxygen. Beside a huge adaptability, D. bruxellensis exhibits a broader spectrum in utilization of carbon and nitrogen sources in comparison to S. cerevisiae. With the aim to better characterize its carbon source metabolism and regulation, the usage of galactose and the role that glucose plays on sugar metabolism were investigated in D. bruxellensis CBS 2499. The results indicate that in this yeast galactose is a non-fermentable carbon source, in contrast to S. cerevisiae that can ferment it. In particular, its metabolism is affected by the nitrogen source. Interestingly, D. bruxellensis CBS 2499 exhibits the 'short-term Crabtree effect', and the expression of genes involved in galactose utilization and in respiratory metabolism is repressed by glucose, similarly to what occurs in S. cerevisiae.
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Brettanomyces/genética , Brettanomyces/metabolismo , Galactosa/metabolismo , Redes y Vías Metabólicas/genética , Ácido Acético/metabolismo , Carbono/metabolismo , Etanol/metabolismo , Regulación Fúngica de la Expresión Génica , Glucosa/metabolismo , Nitrógeno/metabolismoRESUMEN
Dekkera bruxellensis is a yeast known to affect the quality of wine and beer. This species, due to its high ethanol and acid tolerance, has been reported also to compete with Saccharomyces cerevisiae in distilleries producing fuel ethanol. In order to understand how this species responds when exposed to low temperatures, some mechanisms like synthesis and accumulation of intracellular metabolites, changes in lipid composition and activation of the HOG-MAPK pathway were investigated in the genome sequenced strain CBS 2499. We show that cold stress caused intracellular accumulation of glycogen, but did not induce accumulation of trehalose and glycerol. The cellular fatty acid composition changed after the temperature downshift, and a significant increase of palmitoleic acid was observed. RT-PCR analysis revealed that OLE1 encoding for Δ9-fatty acid desaturase was up-regulated, whereas TPS1 and INO1 didn't show changes in their expression. In D. bruxellensis Hog1p was activated by phosphorylation, as described in S. cerevisiae, highlighting a conserved role of the HOG-MAP kinase signaling pathway in cold stress response.