Origen del microbioma de placas solares

[ES] Durante los últimos años, los avances en las técnicas metagenómicas y bioinformáticas han permitido explorar la biodiversidad microbiana presente en distintos tipos de ambientes. Entre todos ellos, los ambientes extremos y artificiales han despertado un especial interés en la comunidad científica debido a su alto valor ecológico y su enorme potencial como fuente de microorganismos con aplicaciones industriales. Las placas solares son una de las estructuras hechas por el hombre más extendidas y se caracterizan por combinar estas dos propiedades: son ambientes artificiales sometidos a condiciones fisicoquímicas extremas. En un trabajo anterior, nuestro grupo caracterizó este ecosistema utilizando una aproximación metagenómica en placas solares de la ciudad de Valencia, y determinó que la comunidad microbiana presente tenía más relación con ambientes naturales extremos, como desiertos, que con otro tipo de ambientes relacionados con los humanos o las áreas urbanas. El objetivo de este trabajo es discernir el origen de esta comunidad microbiana. Para ello, se partió de la hipótesis de que el microbioma de placas solares dependía en gran medida de las migraciones microbianas desde los desiertos a través de las tormentas de polvo. Se analizaron diferentes perfiles taxonómicos extraídos de la bibliografía y correspondientes a ecosistemas susceptibles de ser el germen de la biocenosis presente en los paneles fotovoltaicos (aire sin polvo, aire con polvo desértico y suelo desértico), y se compararon con muestras de placas solares de Valencia y Berkeley secuenciadas por nuestro grupo. Los resultados demostraron que la comunidad microbiana presente en los paneles solares no es fruto de una simple deposición de microorganismos aéreos, sino que se trata de una comunidad compleja en la que se seleccionan aquellos organismos mejor adaptados. Se encontró que tanto el aire como los desiertos ejercían una importante influencia en la formación del microbioma de placas solares. Además, las comunidades microbianas que habitan estas estructuras presentaban un elevado grado de similitud entre las dos localizaciones estudiadas, y un reducido núcleo de esta población se encontraba significativamente infrarrepresentado en los otros ecosistemas analizados. Este core se compone de bacterias resistentes a la desecación y la radiación que se caracterizan por su capacidad de sintetizar carotenoides.[EN] During the last years, new advances in metagenomic and bioinformatic techniques have allowed the exploration of the biodiversity present in different types of environments. Among them, extreme and artificial environments have been found of special interest by the scientific community due to their high ecological value and their potential as a source of microorganisms with industrial applications. Solar panels are one of most widespread man-made structures that combine these two characteristics: they are artificial environments that are subjected to extreme physicochemical conditions. In a previous work, our group characterized this ecosystem by means of a metagenomic approach performed on solar panels in the city of Valencia. The microbial community present in the surface of these structures proved more similar to that associated to other extreme environments, such as deserts, than to other human-associated or urban-associated areas. The objective of this work is to explore the origin of this microbial community, under the hypothesis that the microbiome present on solar panels is largely dependent on microbial migrations coming from deserts through dust storms. Different taxonomic profiles that corresponded to the ecosystems susceptible to be the germ of the solar panels biocenosis (air without dust, air with dust, and desert soil) were retrieved from the literature and compared with those of solar panels sampled in Valencia and Berkeley that had been previously sequenced by our group. Our results showed that the microbial community present in the surface of solar panels was not a simple deposition of air microorganisms, but a complex community in which resistant microorganisms are selected. Air and deserts proved to have an important influence on the formation of the solar panels microbiome. Furthermore, the microbial communities inhabiting solar panels were found to be strikingly similar in the two locations analyzed, and a reduced core of those communities was found to be significantly underrepresented in the other ecosystems studied. This core is composed of desiccationand radiation-resistant bacteria, characterized by the ability to synthesize carotenoids.Latorre Pérez, A. (2017). Origen del microbioma de placas solares. http://hdl.handle.net/10251/86418TFG

Optimizing Nanopore-based microbiome sequencing for characterizing biotechnologically-relevant ecosystems

La secuenciación de siguiente generación (NGS) ha cambiado completamente la manera en la que estudiamos el mundo microbiano. A lo largo de los últimos años, las plataformas Illumina han sido las más usadas para secuenciar ADN. No obstante, la secuenciación por nanoporos, un nuevo método impulsado por Oxford Nanopore Technologies (ONT), está incrementando gradualmente su popularidad dadas sus múltiples ventajas sobre las plataformas NGS. Estas ventajas incluyen la portabilidad y la capacidad para generar secuencias largas en tiempo real. A pesar de estos avances, la adopción de la secuenciación ONT se ha visto limitada por la elevada tasa de error asociada a esta tecnología en comparación con Illumina. El objetivo de la presente tesis fue optimizar la secuenciación por nanoporos para analizar comunidades microbianas de interés industrial y/o biotecnológico. Específicamente, se evaluó el uso de esta técnica de secuenciación para tres aplicaciones diferentes: la monitorización industrial, la bioprospección microbiana y el ensamblaje metagenómico. En el primer caso, se estudiaron las comunidades microbianas implicadas en la producción de biogás, un combustible obtenido por digestión anaerobia (DA), mediante secuenciación metataxonómica. Este trabajo permitió medir el efecto de diferentes parámetros sobre los microbiomas responsables de la DA y detectar marcadores microbianos relacionados con una producción optimizada de biogás. En segundo lugar, se usó la secuenciación ONT in situ para caracterizar distintas muestras durante una expedición de bioprospección al Desierto de Tabernas (Almería, España). Los análisis bioinformáticos se centraron en detectar bacterias resistentes a la radiación y a la desecación, que eran el objetivo de la bioprospección. Los datos obtenidos mediante secuenciación fueron consistentes con los resultados de las técnicas de cultivo, demostrando que la secuenciación portátil puede ser usada para predecir el potencial biotecnológico de las muestras in situ. Finalmente, se investigó el impacto de la secuenciación ONT en el ensamblaje metagenómico usando datos generados a partir de comunidades microbianas artificiales. Después de evaluar varias herramientas de ensamblaje, metaFlye mostró el mejor rendimiento, permitiendo recuperar varios genomas extremadamente contiguos directamente desde el metagenoma. En términos generales, esta tesis evidencia que las plataformas ONT pueden ser aplicadas con éxito para caracterizar microbiomas de relevancia biotecnológica y analiza críticamente las direcciones futuras de la secuenciación por nanoporos, teniendo en cuenta las limitaciones y ventajas de esta tecnología.Next-generation sequencing (NGS) has completely changed the way the microbial world is studied. Illumina platforms have been the most widely used DNA sequencers during the last 8-10 years. However, Nanopore sequencing, a third-generation sequencing method powered by Oxford Nanopore Technologies (ONT), is becoming increasingly popular due to its multiple advantages over NGS, including portability and the ability to read long DNA molecules in real time. Despite these improvements, the adoption of ONT platforms has been hampered by the higher error rate of this technology compared to Illumina. This thesis aimed at optimizing Nanopore sequencing to analyze microbial communities of industrial and/or biotechnological interest. Specifically, this sequencing technique was evaluated for its use in three different applications: industrial monitoring, microbial bioprospecting and metagenome assembly. In the first case, metataxonomic sequencing was applied to characterize the archaeal and bacterial communities involved in the production of biogas, which is an industrially relevant biofuel obtained by anaerobic digestion (AD). This work enabled to measure the effect of different operating parameters on the AD microbiome and to detect microbial markers associated with an improved production of biogas. Secondly, in situ Nanopore sequencing was used to study several samples during a bioprospecting expedition to the Tabernas Desert (Almeria, Spain). Microbiome analyses were focused on the detection of radiation- and desiccation-resistant bacteria, which were the target of the bioprospecting activities. Nanopore data was consistent with the results obtained by culture methods, thus demonstrating that portable sequencing can be used to predict the biotechnological potential of the samples in situ. Finally, the impact of Nanopore sequencing on metagenome assembly was investigated by using data generated from mock communities with different levels of complexity. After benchmarking various assembly tools, metaFlye showed the best overall performance, retrieving highly contiguous genomes directly from metagenomic data. Altogether, this thesis proves that ONT platforms can be efficiently applied to characterize biotechnologically-relevant ecosystems. Moreover, future directions of Nanopore sequencing are critically discussed considering all the advantages and limitations of this technology

A lab in the field: applications of real-time, in situ metagenomic sequencing

High-throughput metagenomic sequencing is considered one of the main technologies fostering the development of microbial ecology. Widely used second-generation sequencers have enabled the analysis of extremely diverse microbial communities, the discovery of novel gene functions, and the comprehension of the metabolic interconnections established among microbial consortia. However, the high cost of the sequencers and the complexity of library preparation and sequencing protocols still hamper the application of metagenomic sequencing in a vast range of real-life applications. In this context, the emergence of portable, third-generation sequencers is becoming a popular alternative for the rapid analysis of microbial communities in particular scenarios, due to their low cost, simplicity of operation, and rapid yield of results. This review discusses the main applications of real-time, in situ metagenomic sequencing developed to date, highlighting the relevance of this technology in current challenges (such as the management of global pathogen outbreaks) and in the next future of industry and clinical diagnosis.Adriel Latorre is a recipient of a Doctorado Industrial fellowship from the Ministerio de Ciencia, Innovacion y Universidades (reference DI-17-09613).Peer reviewe

Assembly methods for nanopore-based metagenomic sequencing: a comparative study

Abstract Metagenomic sequencing has allowed for the recovery of previously unexplored microbial genomes. Whereas short-read sequencing platforms often result in highly fragmented metagenomes, nanopore-based sequencers could lead to more contiguous assemblies due to their potential to generate long reads. Nevertheless, there is a lack of updated and systematic studies evaluating the performance of different assembly tools on nanopore data. In this study, we have benchmarked the ability of different assemblers to reconstruct two different commercially-available mock communities that have been sequenced using Oxford Nanopore Technologies platforms. Among the tested tools, only metaFlye, Raven, and Canu performed well in all the datasets. These tools retrieved highly contiguous genomes (or even complete genomes) directly from the metagenomic data. Despite the intrinsic high error of nanopore sequencing, final assemblies reached high accuracy (~ 99.5 to 99.8% of consensus accuracy). Polishing strategies demonstrated to be necessary for reducing the number of indels, and this had an impact on the prediction of biosynthetic gene clusters. Correction with high quality short reads did not always result in higher quality draft assemblies. Overall, nanopore metagenomic sequencing data-adapted to MinION’s current output-proved sufficient for assembling and characterizing low-complexity microbial communities

Thermoelectric heat exchange and growth regulation in a continuous yeast culture

We have designed a thermoelectric heat exchanger (TEHE) for microbial fermentations that is able to produce electric power from a microbial continuous culture using the intrinsic heat generated by microbial growth. While the TEHE was connected, the system proved able to stably self-maintain both the temperature and the optical density of the culture. This paves the way toward a more sustainable operation of microbial fermentations, in which energy could be saved by converting part of the metabolic heat into usable electric power.This work was funded by the “Valoritza i Transfereix” program (CPI-13-128) from the University of Valencia.Peer reviewe

Lessons from a survey on the public perception of synthetic biology and related disciplines

Financial support from the European CSA on biological standardization BIOROBOOST (EU grant number 820699, http://standardsinsynbio.eu) is acknowledged. EMM is funded with a Formación de Profesorado Universitario (FPU) grant from the Spanish Government (Ministerio de Ciencia, Innovación y Universidades), with reference FPU17/04184. ALP is a recipient of a Doctorado Industrial fellowship from the Ministerio de Ciencia, Innovación y Universidades (Spain), with reference DI‐17‐09613.Peer reviewe

Beyond archaea: The table salt bacteriome

Commercial table salt is a condiment with food preservative properties by decreasing water activity and increasing osmotic pressure. Salt is also a source of halophilic bacteria and archaea. In the present research, the diversity of halotolerant and halophilic microorganisms was studied in six commercial table salts by culture-dependent and culture-independent techniques. Three table salts were obtained from marine origins: Atlantic Ocean, Mediterranean (Ibiza Island), and Odiel marshes (supermarket marine salt). Other salts supplemented with mineral and nutritional ingredients were also used: Himalayan pink, Hawaiian black, and one with dried vegetables known as Viking salt. The results of 16S rRNA gene sequencing reveal that the salts from marine origins display a similar archaeal taxonomy, but with significant variations among genera. Archaeal taxa Halorubrum, Halobacterium, Hallobellus, Natronomonas, Haloplanus, Halonotius, Halomarina, and Haloarcula were prevalent in those three marine salts. Furthermore, the most abundant archaeal genera present in all salts were Natronomonas, Halolamina, Halonotius, Halapricum, Halobacterium, Haloarcula, and uncultured Halobacterales. Sulfitobacter sp. was the most frequent bacteria, represented almost in all salts. Other genera such as Bacillus, Enterococcus, and Flavobacterium were the most frequent taxa in the Viking, Himalayan pink, and black salts, respectively. Interestingly, the genus Salinibacter was detected only in marine-originated salts. A collection of 76 halotolerant and halophilic bacterial and haloarchaeal species was set by culturing on different media with a broad range of salinity and nutrient composition. Comparing the results of 16S rRNA gene metataxonomic and culturomics revealed that culturable bacteria Acinetobacter, Aquibacillus, Bacillus, Brevundimonas, Fictibacillus, Gracilibacillus, Halobacillus, Micrococcus, Oceanobacillus, Salibacterium, Salinibacter, Terribacillus, Thalassobacillus, and also Archaea Haloarcula, Halobacterium, and Halorubrum were identified at least in one sample by both methods. Our results show that salts from marine origins are dominated by Archaea, whereas salts from other sources or salt supplemented with ingredients are dominated by bacteria.This research was financially supported by the Spanish Government on SETH Project (reference: RTI2018-095584-B-C41-42-43-44, co-financed by FEDER funds and Ministerio de Ciencia, Innovación y Universidades), the Helios project (reference: BIO2015-66960-C3-1-R), and the European CSA on biological standardization BIOROBOOST (EU grant no. 820699). LS was funded by the European project BIOROBOOST. AL-P was funded by Doctorado Industrial fellowship from the Ministerio de Ciencia, Innovación y Universidades (reference: DI-17-09613).Peer reviewe

Chemically stressed bacterial communities in anaerobic digesters exhibit resilience and ecological flexibility

Anaerobic digestion is a technology known for its potential in terms of methane production. During the digestion process, multiple metabolites of high value are synthesized. However, recent works have demonstrated the high robustness and resilience of the involved microbiomes; these attributes make it difficult to manipulate them in such a way that a specific metabolite is predominantly produced. Therefore, an exact understanding of the manipulability of anaerobic microbiomes may open up a treasure box for bio-based industries. In the present work, the effect of nalidixic acid, γ-aminobutyric acid (GABA), and sodium phosphate on the microbiome of digested sewage sludge from a water treatment plant fed with glucose was investigated. Despite of the induced process perturbations, high stability was observed at the phylum level. However, strong variations were observed at the genus level, especially for the genera Trichococcus, Candidatus Caldatribacterium, and Phascolarctobacterium. Ecological interactions were analyzed based on the Lotka–Volterra model for Trichococcus, Rikenellaceae DMER64, Sedimentibacter, Candidatus Cloacimonas, Smithella, Cloacimonadaceae W5 and Longilinea. These genera dynamically shifted among positive, negative or no correlation, depending on the applied stressor, which indicates a surprisingly dynamic behavior. Globally, the presented work suggests a massive resilience and stability of the methanogenic communities coupled with a surprising flexibility of the particular microbial key players involved in the process.We are grateful for funding of the work by the German Ministry of Economic Affairs and Energy (grant numbers 16KN070128 and 16KN070126). Moreover, we thank the Spanish Ministry of Science, Innovation and Universities for funding the Ph.D. of Adriel Latorre-Pérrez (Doctorado Industrial Fellowship, reference DI-17-09613). Finally, we are grateful for open access funding by the publication fund of the TU Dresden and for funding by the European Union through the BioRoboost project, H2020-NMBP-TR-IND-2018-2020/BIOTEC-01-2018 (CSA), Project ID 210491758.Peer reviewe

The car tank lid bacteriome: a reservoir of bacteria with potential in bioremediation of fuel

Bioprospecting of microorganisms suitable for bioremediation of fuel or oil spills is often carried out in contaminated environments such as gas stations or polluted coastal areas. Using next-generation sequencing (NGS) we analyzed the microbiota thriving below the lids of the fuel deposits of diesel and gasoline cars. The microbiome colonizing the tank lids differed from the diversity found in other hydrocarbon-polluted environments, with Proteobacteria being the dominant phylum and without clear differences between gasoline or diesel-fueled vehicles. We observed differential growth when samples were inoculated in cultures with gasoline or diesel as the main carbon source, as well as an increase in the relative abundance of the genus Pseudomonas in diesel. A collection of culturable strains was established, mostly Pseudomonas, Stenotrophomonas, Staphylococcus, and Bacillus genera. Strains belonging to Bacillus, Pseudomonas, Achromobacter, and Isoptericola genera showed a clear diesel degradation pattern when analyzed by GC-MS, suggesting their potential use for bioremediation and a possible new species of Isoptericola was further characterized as hydrocarbon degrader.Financial support from the Spanish Government (Grant SETH, ref. RTI2018-095584-B-C41-42-43-44 co-financed by ERDF and AEI-Ministerio de Ciencia e Innovación MICINN) and European Union H2020 (BioRobooST project ID 210491758; Micro4Biogas project ID101000470; MIPLACE project ref. PCI2019-111845-2, 427 Programación Conjunta Internacional 2019, AEI) are acknowledged. À.V.-V. is a recipient of a Formación de Profesorado Universitario (FPU) grant from the Spanish Ministerio de Universidades, with reference FPU18/02578. A.L.-P. is funded with a Doctorado Industrial fellowship from Ministerio de Ciencia, Innovación y Universidades, with reference DI-17-09613.Peer reviewe

A round trip to the desert: In situ nanopore sequencing informs targeted bioprospecting

Bioprospecting expeditions are often performed in remote locations, in order to access previously unexplored samples. Nevertheless, the actual potential of those samples is only assessed once scientists are back in the laboratory, where a time-consuming screening must take place. This work evaluates the suitability of using Nanopore sequencing during a journey to the Tabernas Desert (Spain) for forecasting the potential of specific samples in terms of bacterial diversity and prevalence of radiation- and desiccation-resistant taxa, which were the target of the bioprospecting activities. Samples collected during the first day were analyzed through 16S rRNA gene sequencing using a mobile laboratory. Results enabled the identification of locations showing the greatest and the least potential, and a second, informed sampling was performed focusing on those sites. After finishing the expedition, a culture collection of 166 strains belonging to 50 different genera was established. Overall, Nanopore and culturing data correlated well, since samples holding a greater potential at the microbiome level also yielded a more interesting set of microbial isolates, whereas samples showing less biodiversity resulted in a reduced (and redundant) set of culturable bacteria. Thus, we anticipate that portable sequencers hold potential as key, easy-to-use tools for in situ-informed bioprospecting strategies.Financial support from the Spanish Government grant SETH (ref: RTI2018-095584-B-C41-42-43-44) is acknowledged. AL-P is a recipient of a Doctorado Industrial fellowship from the Spanish Ministerio de Ciencia, Innovación y Universidades (reference DI-17-09613).Peer reviewe