Aproximación metaproteómica para evaluar el efecto de metales pesados en un diseño experimental de microcosmos

Premio extraordinario de Trabajo Fin de Máster curso 2020/2021. Máster en BiotecnologíaLa contaminación es uno de los grandes problemas a los que se enfrenta la sociedad actual, que afecta al bienestar de los ecosistemas. Destacan los ecosistemas acuáticos por su relevancia económica y ecológica. La liberación de metales pesados en las aguas se ha potenciado como consecuencia de distintas actividades antropogénicas. Los cambios en la estructura y funcionalidad de las poblaciones microbianas se han propuesto como marcadores biológicos de respuesta a la contaminación ambiental. En este trabajo, se ha estudiado el efecto de la combinación de los metales plata y cadmio a distintas concentraciones (0, 5 y 50 μg/L) sobre el microbioma de sedimentos en un sistema de microcosmos acuático artificial. Si evaluamos la distribución de las familias a las que pertenecen las proteínas identificadas, la exposición aguda a la mezcla de metales dio lugar a una pérdida de la diversidad y a una disminución en el número de identificaciones en numerosas familias de microorganismos, destacando Rhizobiaceae, Rhodobacteraceae y Rhodospirillaceae. Además, la exposición a estos contaminantes supuso una alteración en el patrón de expresión de las proteínas identificadas. A la concentración más baja de la mezcla de metales se produjo un aumento en la diversidad y número de funciones, destacando los procesos biológicos de transporte y unión a metales, que podrían relacionarse con la activación de mecanismos de resistencia frente a estos contaminantes. No obstante, muchas funciones como la respuesta a estrés o la unión a metales desaparecieron a la mayor concentración de la mezcla, lo cual podría indicar que la capacidad de defensa de las bacterias se ve comprometida como consecuencia de la exposición aguda. El análisis de clústeres de las proteínas con cambios significativos en las distintas condiciones permitió distinguir 4 patrones diferentes. Estos cambios, asociados a las funciones biológicas de las proteínas identificadas, podrían emplearse como biomarcadores específicos de este tipo de contaminación. Los estudios metaproteómicos constituyen, por tanto, una buena aproximación para estudiar los efectos biológicos de los contaminantes y sus posibles consecuencias toxicológicas.Pollution is one of the major problems faced by today’s society, which affects the well-being of ecosystems. Aquatic ecosystems stand out for their economic and ecological relevance. The release of heavy metals into water bodies has been enhanced because of different anthropogenic activities. Changes in the structure and functionality of microbial populations have been proposed as biological markers of response to environmental pollution. In this study, the effect of the combination of heavy metals cadmium and silver, at different concentrations (0, 5 and 50 μg/L) on the microbiome of the artificial aquatic microcosms’ sediments, has been investigated. If we evaluate the distribution of the microbial families whose proteins have been identified, the acute exposure to the mixture of metals led to a loss of diversity and a decrease in the number of identifications in numerous families of microorganisms, highlighting Rhizobiaceae, Rhodobacteraceae and Rhodospirillaceae. Furthermore, the exposure to these pollutants altered the expression patterns of the identified proteins. At the lowest concentration of the metal mixture, there was an increase in the diversity and number of functions, highlighting biological processes as transport and metal-binding, which could be related to the activation of resistance mechanisms against metals. However, many functions such as stress response or metal-binding, disappeared at the acute concentration of the mixture, which could indicate that the defence capacity of bacteria is compromised due to this exposure. Cluster analysis of proteins with significant changes under the different conditions allowed us to distinguish 4 different patterns. These changes, associated with the biological functions of the identified proteins, could be used as specific biomarkers of this type of pollution. Metaproteomics studies are, therefore, a good approach to study the biological effects of pollutants and their possible toxicological consequences

Biomethanisation of sewage sludge: Sonication pretreatment and monitoring of microbial communities

The improvement of mesophilic biomethanisation of recalcitrant sewage sludge derived from urban wastewater treatment through the application of a sonication pretreatment was evaluated in parallel in two pilot-scale anaerobic digesters (two biological replicates: reactors RA and RB). The valorisation process was monitored through a novel and holistic approach that related the biomethanisation yield, and its main batch operational parameters, with the abundance of archaeal and bacterial communities in the anaerobic inocula. Sonication allowed achieving a methane yield coefficient derived from sewage sludge of 240 ± 20 mLSTPCH4/g VS (volatile solids) at the load range of 0.8–4.0 g VS/L in both reactors. The process was more stable in reactor B, with a wider range of loads being allowed (up to 5.29 g VS/L). Monitoring the presence of Archaea in the mixing liquor revealed a variation in their abundance throughout the process which was directly related to the availability of organic matter and pH. Advanced metagenomic analysis showed the phylogenetic and functional diversity of the complex microbiome involved. While Bacteria were widely distributed in 35 phyla, Archaea fitted in only two. Euryarchaeota was the majoritarian archaeal phylum (99.5 %) and its more abundant families are linked to methanogenic metabolism. Functional analysis revealed several relevant metabolic pathways that followed similar trends in both reactors. “Methane metabolism” clearly diminished at the end of the process in concordance with the exhaust of methane generation, while “ABC transporters” or “two-component systems”, involved in bacterial survival to changing environments, followed the opposite pattern. This integrated approach could help to increase the methanogenic valorisation of sewage sludge