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[Abstract] The northwest of Spain has an abundance of non-volcanic hot springs that, until recently, had only been used for thermalism activities. One of such hot springs, Muiño da Veiga, has now been explored using metagenomics to study the microbial community that inhabits these high-temperature circumneutral continental waters. Sequencing of the metagenome allowed the characterization of its composition, diversity, metabolic connections and potential as a source for thermozymes, as well as its ability to assemble MAGs. A diverse microbial community dominated by Bacteria domain members was revealed, particularly from the early-branching Aquificales group. The most abundant genus was Sulfurihydrogenibium, known for its implication in sulfur cycling and for forming mats that enable novel niches. The variety of primary producers with autotrophic pathways (and specifically the sulfur oxidizing pathway) expands the range of available nutrients, and the increase in biomass forms thicker mats, resulting in more available niches and broader microbial diversity. Nonetheless, certain metabolic pathways were attributed to less abundant members of the microbial community, reinforcing the idea that the rare biosphere plays important roles in the network of interactions present in an ecosystem and acts as genetic reservoirs. In addition, three of the assembled MAGs represent novel microbial diversity found in this hot spring. Moreover, the presence of enzymes and microorganisms with possible biotechnological applications was confirmed, including proteases, lipases and cell-wall degrading enzymes, pointing to the potential for the hot spring as a source for thermozymes.

[Abstract] Cancer stem cells are a small subpopulation within the tumor with high capacity for self-renewal, differentiation and reconstitution of tumor heterogeneity. Cancer stem cells are major contributors of tumor initiation, metastasis and therapy resistance in cancer. Emerging evidence indicates that ubiquitination-mediated post-translational modification plays a fundamental role in the maintenance of cancer stem cell characteristics. In this review, we will discuss how protein degradation controlled by the E3 ubiquitin ligases plays a fundamental role in the self-renewal, maintenance and differentiation of cancer stem cells, highlighting the possibility to develop novel therapeutic strategies against E3 ubiquitin ligases targeting CSCs to fight cancer.

[Abstract] With their circumneutral pH and their moderate temperature (66 and 68°C, respectively), As Burgas and Muiño da Veiga are two important human-use hot springs, previously studied with traditional culture methods, but never explored with a metagenomic approach. In the present study, we have performed metagenomic sequence-based analyses to compare the taxonomic composition and functional potential of these hot springs. Proteobacteria, Deinococcus-Thermus, Firmicutes, Nitrospirae, and Aquificae are the dominant phyla in both geothermal springs, but there is a significant difference in the abundance of these phyla between As Burgas and Muiño da Veiga. Phylum Proteobacteria dominates As Burgas ecosystem while Aquificae is the most abundant phylum in Muiño da Veiga. Taxonomic and functional analyses reveal that the variability in water geochemistry might be shaping the differences in the microbial communities inhabiting these geothermal springs. The content in organic compounds of As Burgas water promotes the presence of heterotrophic populations of the genera Acidovorax and Thermus, whereas the sulfate-rich water of Muiño da Veiga favors the co-dominance of genera Sulfurihydrogenibium and Thermodesulfovibrio. Differences in ammonia concentration exert a selective pressure toward the growth of nitrogen-fixing bacteria such as Thermodesulfovibrio in Muiño da Veiga. Temperature and pH are two important factors shaping hot springs microbial communities as was determined by comparative analysis with other thermal springs.

[Abstract] Hot spring metagenomes are described as the DNA of whole microbial communities inhabiting high temperature inland water habitats. The focus of the present work was to search for thermozymes (that maintain their activity at high temperatures) encoded in such metagenomes, as they find biotechnological applications in all kinds of industries. Additional information like the metabolic interactions between community members have been studied as well. The first step of the metagenomic study consisted in the sampling of two hot springs: As Burgas and Muiño da Veiga. The metagenomic DNA from these samples was extracted and used to construct two metagenomic libraries. These libraries were screened to detect potentially relevant enzymatic activities using a wide range of substrates, resulting in the discovery of a novel thermozyme with lipolytic activity (LipB12_A11) which was biochemically characterized. The metagenomic DNA from one of the hot springs was also sequenced with a shotgun sequencing strategy. This led to the description of the microbial community from the hot spring, based on sequence reads abundance and taxonomical and functional assignment. Reads assembly into larger sequences allowed the prediction of several putative proteins with biotechnological interest. Of these, a novel endoglucanase (Cel776) was expressed and biochemically characterized., [Resumen] Los metagenomas de aguas termales consisten en el conjunto del ADN de comunidades microbianas procedentes de hábitats de aguas continentales a altas temperaturas. El objetivo de este trabajo es la búsqueda en estos metagenomas de termozimas (que mantienen su actividad a altas temperaturas) codificadas en ellos, ya que tienen aplicaciones biotecnológicas en todo tipo de industrias. Además, se han estudiado las interacciones metabólicas que existen entre los miembros de la comunidad microbiana. El primer paso en el estudio metagenómico ha sido el muestreo de dos fuentes termales: As Burgas y Muiño da Veiga. Se extrajo el ADN metagenómico de estas muestras y se construyeron dos metagenotecas. Se realizó una búsqueda funcional usando un abanico de sustratos en las metagenotecas para encontrar actividades enzimáticas potencialmente relevantes, y se caracterizó bioquímicamente una nueva termozima con actividad lipolítica (LipB12_A11). Además, se secuenció el ADN metagenómico de una de las fuentes termales con una estrategia shotgun. En base a la abundancia de las lecturas y su asignación taxonómica y funcional se describió la comunidad microbiana de la fuente termal. Las lecturas se ensamblaron en secuencias más largas para predecir proteínas putativas con interés biotecnológico. De ellas, se expresó y caracterizó una nueva endoglucanasa (Cel776)., [Resumo] Os metaxenomas de augas termais consisten no conxunto do ADN de comunidades microbianas de hábitats de augas continentais a alta temperatura. O obxectivo deste traballo é a procura nestes metaxenomas de termoencimas (que manteñen a súa actividade a altas temperaturas) que neles se codifican, xa que teñen aplicacións biotecnolóxicas en todo tipo de industrias. Ademáis, estudáronse as interaccións metabólicas que existen entre os membros da comunidade microbiana. O primeiro paso do estudo metaxenómico foi a toma de mostras de dúas fontes termais: As Burgas e Muiño da Veiga. Destas mostras foi extraído o ADN metaxenómico e construíronse dúas metaxenotecas. Realizouse unha busca funcional empregando unha serie de substratos nas metaxenotecas para atopar actividades encimáticas potencialmente relevantes e caracterizouse bioquímicamente unha nova termocima con actividade lipolítica (LipB12_A11). Ademais, o ADN metaxenómico dunha das augas termais secuenciouse mediante unha estratexia de shotgun. Baseado na abundancia de lecturas e na súa asignación taxonómica e funcional, describimos a comunidade microbiana da fonte termal. As lecturas reuníronse en secuencias máis longas para predicir proteínas con suposto interese biotecnolóxico. Destas, expresouse e caracterizouse unha nova endoglucanasa (Cel776).

[Abstract] The present work was carried out on 133 halophilic strains isolated on MGM (Modified Growth Medium) medium with 12 and 23% (w/v) of salt. A screening of the extracellular proteolytic activities, carried out on the same medium supplemented with casein or gelatin at 1% (w/v), allowed us to select 24 bacterial and 21 archaeal strains presenting a precipitate around the colonies for casein and/or a translucent halo (after addition of Frazier’s reagent) for gelatin. The enzymatic test was performed on liquid medium in microculture in a 2 mL Eppendorf tube. The assay of the proteolytic activity, using Azocasein as substrate, followed two protocols—the first with PBS and the second with Tris HCl, with positive and negative controls—and demonstrated interesting results for 10 strains among the 45 tested including five bacteria and five archaea. These underwent morphological, physiological and molecular characterizations based on amplification and sequencing of the 16S ribosomal RNA gene.

[Abstract] Cellulases are a heterogeneous group of enzymes that synergistically catalyze the hydrolysis of cellulose, the major component of plant biomass. Such reaction has biotechnological applications in a broad spectrum of industries, where they can provide a more sustainable model of production. As a prerequisite for their implementation, these enzymes need to be able to operate in the conditions the industrial process requires. Thus, cellulases retrieved from extremophiles, and more specifically those of thermophiles, are likely to be more appropriate for industrial needs in which high temperatures are involved. Metagenomics, the study of genes and gene products from the whole community genomic DNA present in an environmental sample, is a powerful tool for bioprospecting in search of novel enzymes. In this review, we describe the cellulolytic systems, we summarize their biotechnological applications, and we discuss the strategies adopted in the field of metagenomics for the discovery of new cellulases, focusing on those of thermophilic microorganisms.

[Abstract] Proteases have numerous biotechnological applications and the bioprospection for newly-thermostable proteases from the great biodiversity of thermophilic microorganisms inhabiting hot environments, such as geothermal sources, aims to discover more effective enzymes for processes at higher temperatures. We report in this paper the production and the characterization of a purified acid protease from strain OA30, a moderate thermophilic bacterium isolated from an Algerian hot spring. Phenotypic and genotypic study of strain OA30 was followed by the production of the extracellular protease in a physiologically-optimized medium. Strain OA30 showed multiple extracellular proteolytic enzymes and protease 32-F38 was purified by chromatographic methods and its biochemical characteristics were studied. Strain OA30 was affiliated with Brevibacillus thermoruber species. Protease 32-F38 had an estimated molecular weight of 64.6 kDa and was optimally active at 50 C. It showed a great thermostability after 240 min and its optimum pH was 6.0. Protease 32-F38 was highly stable in the presence of different detergents and solvents and was inhibited by metalloprotease inhibitors. The results of this work suggest that protease 32-F38 might have interesting biotechnological applications.