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The principle of proportionality is considered the main legal tool to control restrictive measures of rights, both in ordinary courts and at a constitutional level. In addition to its general limitations, new shortcomings of the principle have played a central role during the pandemic, questioning the principleʼs efficacy in situations of factual uncertainty, especially in technically or scientifically complex contexts. This article analyses this efficacy problem and exemplifies it with specific measures adopted to prevent COVID-19. It also analyses potential ways to counter those shortcomings, such as refining the principle itself, emphasising judicial deference to legislative and executive powers, or adopting prior decisions as to the information that must be taken into account in case of uncertainty. Finally, the article proposes some additional checks that could complement the culture of justification promoted by the principle and strengthen the control of public powers when restricting rights under conditions of uncertainty.

ABSTRACT The pks island is one of the most prevalent pathogenicity islands among the Escherichia coli strains that colonize the colon of colorectal carcinoma (CRC) patients. This pathogenic island encodes the production of a nonribosomal polyketide-peptide named colibactin, which induces double-strand breaks in DNA molecules. Detection or even depletion of this pks-producing bacteria could help to understand the role of these strains in the context of CRC. In this work, we performed a large-scale in silico screening of the pks cluster in more than 6,000 isolates of E. coli. The results obtained reveal that not all the pks-detected strains could produce a functional genotoxin and, using antibodies against pks-specific peptides from surface cell proteins, a methodology for detection and depletion of pks+ bacteria in gut microbiotas was proposed. With our method, we were able to deplete a human gut microbiota of this pks+ strains, opening the door to strain-directed microbiota modification and intervention studies that allow us to understand the relation between these genotoxic strains and some gastrointestinal diseases. IMPORTANCE The human gut microbiome has also been hypothesized to play a crucial role in the development and progression of colorectal carcinoma (CRC). Between the microorganisms of this community, the Escherichia coli strains carrying the pks genomic island were shown to be capable of promoting colon tumorigenesis in a colorectal cancer mouse model, and their presence seems to be directly related to a distinct mutational signature in patients suffering CRC. This work proposes a novel method for the detection and depletion of pks-carrying bacteria in human gut microbiotas. In contrast to methods based on probes, this methodology allows the depletion of low-abundance bacterial strains maintaining the viability of both targeted and non-targeted fractions of the microbiota, allowing the study of the contribution of these pks-carrying strains to different diseases, such as CRC, and their role in other physiological, metabolic or immune processes.

ABSTRACT Faecalibacterium represents one of the most abundant bacterial groups in the human intestinal microbiota of healthy adults and can represent more than 10% of the total bacterial population, Faecalibacterium prausnitzii being the only recognized species up to the past year. Reduction in the abundance of F. prausnitzii in the human gut has been linked to several human disorders, such as Crohn’s disease. In this study, we developed a strategy to modify the relative abundance of F. prausnitzii in fecal microbiotas as a means of evaluating its contribution to the immunomodulatory effect of intestinal microbiotas with different F. prausnitzii contents using a peripheral blood mononuclear cell (PBMC) model. We used a polyclonal antibody against the surface of F. prausnitzii M21 to capture the bacterium from synthetic and human fecal microbiotas using immunoseparation techniques. As a proof-of-principle study, the levels of immunomodulation exerted by microbiotas of healthy donors (HDs) with different relative abundances of F. prausnitzii, achieved with the above-mentioned immunoseparation technique, were evaluated in a PBMC model. For this purpose, PBMCs were cocultivated with the modified microbiotas or a pure culture of F. prausnitzii and, subsequently, the microbiota of Crohn’s donors was added to the coculture. The cytokine concentration was determined, showing that our experimental model supports the anti-inflammatory effects of this bacterium. IMPORTANCE There is increasing interest in deciphering the contribution of gut microbiota species to health and disease amelioration. The approach proposed herein provides a novel and affordable strategy to probe deeply into microbiota-host interactions by strategically modifying the relative abundance of specific gut microbes, hence facilitating the study of their contribution to a given trait of the microbiota.

La «gestión de riesgos» en España parece haber relegado al Parlamento a una posición secundaria, frente a un Poder Ejecutivo omnímodo y poco controlado. Esta tendencia, que ha podido observarse en el derecho de necesidad, pero también en las normas ordinarias relacionadas con el riesgo, viene provocada en gran medida por la interpretación y el fundamento otorgado hasta el momento al llamado «principio de precaución», una norma que inspira y vertebra el derecho ordinario y extraordinario frente a riesgos inciertos. Tras analizar brevemente el origen del principio, su recepción en España y la definición mayoritaria que le han otorgado la doctrina y los tribunales, analizamos diversas objeciones a la concepción actual del principio, de las que trae causa la impropia subordinación del Parlamento en este ámbito. A la vista de dichas objeciones y del erróneo fundamento atribuido a las normas de precaución hasta ahora, trataremos de reformular estas últimas para recuperar el protagonismo del Parlamento en la más amplia «canalización institucionalizada del riesgo» (ya no simple gestión) que la propia arquitectura constitucional le ha reservado. De ello extraeremos algunas consecuencias prácticas y mejoras para el Parlamento, relacionadas con la técnica legislativa a la hora de regular riesgos inciertos y la evaluación técnica y científica de los mismos dentro del procedimiento legislativo.

Abstract This work describes a new procedure that allows the targeted modification of the human gut microbiota by using antibodies raised against bacterial surface-associated proteins specific to the microorganism of interest. To this end, a polyclonal antibody recognising the surface-associated protein Surface Layer Protein A of Lactobacillus acidophilus DSM20079T was developed. By conjugating this antibody with fluorescent probes and magnetic particles, we were able to specifically identify this bacterium both in a synthetic, and in real gut microbiotas by means of a flow cytometry approach. Further, we demonstrated the applicability of this antibody to deplete complex human gut microbiotas from L. acidophilus in a single step. L. acidophilus was found to interact with other bacteria both in synthetic and in real microbiotas, as reflected by its concomitant depletion together with other species. Further optimization of the procedure including a trypsin step enabled to achieve the selective and complete isolation of this species. Depleting a single species from a gut microbiota, using antibodies recognizing specific cell surface elements of the target organism, will open up novel ways to tackle research on the specific immunomodulatory and metabolic contributions of a bacterium of interest in the context of a complex human gut microbiota, including the investigation into therapeutic applications by adding/depleting a key bacterium. This represents the first work in which an antibody/flow-cytometry based application enabled the targeted edition of human gut microbiotas, and represents the basis for the design of precision microbiome-based therapies.

Abstract In recent years the role of extracellular vesicles (EVs) of Gram-positive bacteria in host-microbe cross-talk has become increasingly appreciated, although the knowledge of their biogenesis, release and host-uptake is still limited. The aim of this study was to characterize the EVs released by the dairy isolate Lactiplantibacillus plantarum BGAN8 and to gain an insight into the putative mechanism of EVs uptake by intestinal epithelial cells. The cryo-TEM observation undoubtedly demonstrated the release of EVs (20 to 140 nm) from the surface of BGAN8, with exopolysaccharides seems to be part of EVs surface. The proteomic analysis revealed that the EVs are enriched in enzymes involved in central metabolic pathways, such as glycolysis, and in membrane components with the most abundant proteins belonging to amino acid/peptide ABC transporters. Putative internalization pathways were evaluated in time-course internalization experiments with non-polarized HT29 cells in the presence of inhibitors of endocytic pathways: chlorpromazine and dynasore (inhibitors of clathrin-mediated endocytosis—CME) and filipin III and nystatin (disrupting lipid rafts). For the first time, our results revealed that the internalization was specifically inhibited by dynasore and chlorpromazine but not by filipin III and nystatin implying that one of the entries of L. plantarum vesicles was through CME pathway.

Abstract Background The microbial populations of the human intestinal tract and their relationship to specific diseases have been extensively studied during the last decade. However, the characterization of the human bile microbiota as a whole has been hampered by difficulties in accessing biological samples and the lack of adequate methodologies to assess molecular studies. Although a few reports have described the biliary microbiota in some hepatobiliary diseases, the bile microbiota of healthy individuals has not been described. With this in mind, the goal of the present study was to generate fundamental knowledge on the composition and activity of the human bile microbiota, as well as establishing its potential relationship with human bile-related disorders. Results Human bile samples from the gallbladder of individuals from a control group, without any record of hepatobiliary disorder, were obtained from liver donors during liver transplantation surgery. A bile DNA extraction method was optimized together with a quantitative PCR (qPCR) assay for determining the bacterial load. This allows the selection of samples to perform functional metagenomic analysis. Bile samples from the gallbladder of individuals suffering from lithiasis were collected during gallbladder resection and the microbial profiles assessed, using a 16S rRNA gene-based sequencing analysis, and compared with those of the control group. Additionally, the metabolic profile of the samples was analyzed by nuclear magnetic resonance (NMR). We detected, for the first time, bacterial communities in gallbladder samples of individuals without any hepatobiliary pathology. In the biliary microecosystem, the main bacterial phyla were represented by Firmicutes, Bacteroidetes, Actinobacteria, and Proteobacteria. Significant differences in the relative abundance of different taxa of both groups were found. Sequences belonging to the family Propionibacteriaceae were more abundant in bile samples from control subjects; meanwhile, in patients with cholelithiasis members of the families Bacteroidaceae, Prevotellaceae, Porphyromonadaceae, and Veillonellaceae were more frequently detected. Furthermore, the metabolomics analysis showed that the two study groups have different metabolic profiles. Conclusions Our results indicate that the gallbladder of human individuals, without diagnosed hepatobiliary pathology, harbors a microbial ecosystem that is described for the first time in this study. Its bacterial representatives and metabolites are different from those detected in people suffering from cholelithiasis. In this regard, since liver donors have been subjected to the specific conditions of the hospital’s intensive care unit, including an antibiotic treatment, we must be cautious in stating that their bile samples contain a physiologically normal biliary microbiome. In any case, our results open up new possibilities to discover bacterial functions in a microbial ecosystem that has not previously been explored.

Peptides encrypted in the intestinal microbial-exoproteome mediate the host-microbiota crosstalk, which is disrupted in inflammatory bowel disease (IBD). Here, the MAHMI database was used for the identification of 20 novel intestinal bacterial peptides. Our results revealed that serum IgA levels directed towards the peptides, but not IgG, discriminated healthy controls from IBD patients. Indeed, they also differentiated patients with ulcerative colitis from Crohńs disease and, within them, patients with and without intestinal inflammation. All peptides were immunomodulatory as they changed the intestinal cytokine milieu following human lamina propria mononuclear cells culture (with/out LPS), revealing a Bifidobacterium longum subsp. longum peptide with the highest tolerogenic properties. Therefore, bacterial peptides encrypted in the human gut metaproteome may have utility as non-invasive biomarkers to aid on IBD diagnosis and monitoring. These peptides also display immunomodulatory effects on the intestinal mucosa revealing them as novel functional compounds for non-drug therapeutic strategies in IBD.

Bacteria-host interactions are mediated by different microbial associated molecular patterns which are most often surface structures such as, among others, exopolysaccharides (EPSs). In this work, the capability of two isogenic EPS-producing Bifidobacterium animalis subsp. lactis strains to modulate the gut microbiota of healthy mice, was assessed. Each strain produces a different type of polymer; the ropy strain S89L synthesized a rhamnose-rich, high-molecular weight EPS in highest abundance than the non-ropy DMS10140 one. BALB/c mice were orally fed for 10 days with milk-bifidobacterial suspensions and followed afterward for 7 post-intervention days (wash-out period). The colonic content of mice was collected in several sampling points to perform a metataxonomic analysis. In addition, the influence of specific microbial clades, apparently stimulated by the ropy and non-ropy strains, on mouse plasmatic cytokine levels was investigated through hierarchical association testing. Analysis of 16S rRNA gene sequences showed that the abundance of Firmicutes phylum significantly increased 7 days after cessing the treatment with both strains. The relative abundance of Alloprevotella genus also rose, but after shorter post-treatment times (3 days for both DMS10140 and S89L strains). Some bacterial clades were specifically modulated by one or another strain. As such, the non-ropy DMS10140 strain exerted a significant influence on Intestinomonas genus, which increased after 4 post-administration days. On the other hand, feeding with the ropy strain S89L led to an increase in sequences of Faecalibaculum genus at 4 post-treatment days, while the abundance of Erysipelotrichaceae and Lactobacillaceae families increased for prolonged times. Association testing revealed that several lactobacilli and bifidobacterial significantly stimulated by ropy S89L strain were positively associated with the levels of certain cytokines, including IL-5 and IL-27. These results highlight relevant changes in mice gut microbiota produced after administration of the ropy S89L strain that were associated to a potential immune modulation effect.

This work supports the massive presence of potential immunomodulatory peptides in the human gut metaproteome. These peptides were identified through the MAHMI database as potentially anti-inflammatory, and sixteen of them synthesized for characterize their mechanism of action. From them, peptide HM14 was encrypted in an extracellular protein produced by Bifidobacterium longum, a common member of the human microbiota, and displayed the highest anti-inflammatory capability. Molecular mechanism of action of HM14 pointed to a specific interaction between this immunomodulatory peptide and antigen presenting cells, which resulted in a higher formation of iTreg cells. Moreover, HM14 was effective in decreasing pro-inflammatory parameters in PBMCs isolated from a cohort of Crohn’s patients. Finally, non-targeted metabolomics confirmed the ability of HM14 to modulate the metabolic activity of PBMCs to fulfil its energy and biosynthetic requirements. Overall, our combined in silico/multiomics approach supports the human gut metaproteome as a source for immunomodulatory peptides.

Lactobacillus genus includes both probiotic and representative strains of the human gut microbiota. Independent studies have reported on the anti-inflammatory properties of different Lactobacillus strains, although we are far from understanding the underlying molecular interplay. In this work we show that a daily administration of Lactobacillus acidophilus DSM20079T (DSM20079) to healthy piglets resulted in plasmatic increases of the anti-inflammatory IL10, whilst IL12 and the pro-inflammatory ratio IL12+TNFα/IL10 decreased. The extracellular protein fraction of DSM20079 was identified as the responsible for the crosstalk interaction that elicited these tolerogenic effects. This strain was able to activate innate immune pathways in dendritic cells and to decrease the production of pro-inflammatory cytokines in both CD4+/CD8+ T cell subsets in healthy donors and in Crohn’s Disease patients. The tolerogenic effect exerted by the extracellular proteins of this strain suggests their potential use as coadjutant for therapeutic applications targeting chronic inflammatory illnesses.

Probiotics are used to improve human health due to their capability to induce beneficial effects in gastrointestinal diseases and modulate the immune response. However, scarce information is known about the bacterial active compounds and the metabolic pathways of interaction with immune cells. In the present study, we analyzed the immune response elicited by surface-associated proteins, the whole surface extract (WSE) and the genomic DNA of three strains from the main representative probiotic species for human consumption, L. acidophilus DSM20079T, L. rhamnosus GG, B. longum NCIMB 8809, using an in vitro model of human PBMCs from healthy donors. Eighteen cytokines were quantified using Luminex Technology to understand the immune pathways modulate by bacterial extracts. Overall, the bacterial fractions elicited a different immune response in PBMCs, whereas the isolated DNA was able to induce a higher Th17-like response that could lead to a protective effect in the epithelial barrier function.

Convergence of a stochastic process is an intrinsic property quite relevant for its successful practical for example for the function optimization problem. Lyapunov functions are widely used as tools to prove convergence of optimization procedures. However, identifying a Lyapunov function for a specific stochastic process is a difficult and creative task. This work aims to provide a geometric explanation to convergence results and to state and identify conditions for the convergence of not exclusively optimization methods but any stochastic process. Basically, we relate the expected directions set of a stochastic process with the half-space of a conservative vector field, concepts defined along the text. After some reasonable conditions, it is possible to assure convergence when the expected direction resembles enough to some vector field. We translate two existent and useful convergence results into convergence of processes that resemble to particular conservative vector fields. This geometric point of view could make it easier to identify Lyapunov functions for new stochastic processes which we would like to prove its convergence.

This work presents a novel in silico approach to the prediction and characterization of the glycolytic capacities of the beneficial intestinal bacterium Faecalibacterium prausnitzii. Available F. prausnitzii genomes were explored taking the glycolytic capacities of F. prausnitzii SL3/3 and F. prausnitzii L2-6 as reference. The comparison of the generated glycolytic profiles offered insights into the particular capabilities of F. prausnitzii SL3/3 and F. prausnitzii L2-6 as well as the potential of the rest of strains. Glycoside hydrolases were mostly detected in the pathways responsible for the starch and sucrose metabolism and the biosynthesis of secondary metabolites, but this analysis also identified some other potentially interesting, but still uncharacterized activities, such as several hexosyltransferases and some hydrolases. Gene neighborhood maps offered additional understanding of the genes coding for relevant glycoside hydrolases. Although information about the carbohydrate preferences of F. prausnitzii is scarce, the in silico metabolic predictions were consistent with previous knowledge about the impact of fermentable sugars on the growth promotion and metabolism of F. prausnitzii. So, while the predictions still need to be validated using culturing methods, the approach holds the potential to be reproduced and scaled to accommodate the analysis of other strains (or even families and genus) as well as other metabolic activities. This will allow the exploration of novel methodologies to design or obtain targeted probiotics for F. prausnitzii and other strains of interest.

Bile is a biological fluid synthesized in the liver, mainly constituted by bile acids and cholesterol, which functions as a biological detergent that emulsifies and solubilizes lipids, thereby playing an essential role in fat digestion. Besides, bile acids are important signaling molecules that regulate key functions at intestinal and systemic levels in the human body, affecting glucose and lipid metabolism, and immune homeostasis. Apart from this, due to their amphipathic nature, bile acids are toxic for bacterial cells and, thus, exert a strong selective pressure on the microbial populations inhabiting the human gut, decisively shaping the microbial profiles of our gut microbiota, which has been recognized as a metabolic organ playing a pivotal role in host health. Remarkably, bacteria in our gut also display a range of enzymatic activities capable of acting on bile acids and, to a lesser extent, cholesterol. These activities can have a direct impact on host physiology as they influence the composition of the intestinal and circulating bile acid pool in the host, affecting bile homeostasis. Given that bile acids are important signaling molecules in the human body, changes in the microbiota-residing bile biotransformation ability can significantly impact host physiology and health status. Elucidating ways to fine-tune microbiota-bile acids-host interplay are promising strategies to act on bile and cholesterol-related disorders. This manuscript summarizes the current knowledge on bile and cholesterol metabolism by intestinal bacteria, as well as its influence on host physiology, identifying knowledge gaps and opportunities to guide further advances in the field.

Dihydrochalcones, phlorizin (PZ) and its aglycone phloretin (PT), have evidenced immunomodulatory effects through several mechanisms. However, the differential metabolic signatures that lead to these properties are largely unknown. Since macrophages play an important role in the immune response, our study aimed to characterise human THP-1 macrophages under PZ and PT exposure. A multiplatform-based untargeted metabolomics approach was used to reveal metabolites associated with the anti-inflammatory mechanisms triggered by the dihydrochalcones in LPS-stimulated macrophages, for the first time. Results showed differential phenotypic response in macrophages for all treatments. Dihydrochalcone treatment in LPS-stimulated macrophages mimics the response under normal conditions, suggesting inhibition of LPS response. Antagonistic effects of dihydrochalcones against LPS was mainly observed in glycerophospholipid and sphingolipid metabolism besides promoting amino acid biosynthesis. Moreover, PT showed greater metabolic activity than PZ. Overall, the findings of this study yielded knowledge about the mechanisms of action PZ and PT at metabolic level in modulating inflammatory response in human cells.

Background: Oats provide important nutritional and pharmacological properties, although their safety in coeliac patients remains controversial. Previous studies have confirmed that the reactivity of the anti-33-mer monoclonal antibody with different oat varieties is proportional to the immune responses in terms of T-cell proliferation. Although the impact of these varieties on the adaptive response has been studied, the role of the dendritic cells (DC) is still poorly understood. The aim of this study is to characterize different oat fractions and to study their effect on DC from coeliac patients. Methods and results: Protein fractions were isolated from oat grains and analyzed by SDS–PAGE. Several proteins were characterized in the prolamin fraction using immunological and proteomic tools, and by Nano-LC-MS/MS. These proteins, analogous to α- and γ-gliadin-like, showed reactive sequences to anti-33-mer antibody suggesting their immunogenic potential. That was further confirmed as some of the newly identified oat peptides had a differential stimulatory capacity on circulating DC from coeliac patients compared with healthy controls. Conclusions: This is the first time, to our knowledge, where newly identified oat peptides have been shown to elicit a differential stimulatory capacity on circulating DC obtained from coeliac patients, potentially identifying immunogenic properties of these oat peptides.

Bifidobacteria are among the most abundant microorganisms inhabiting the intestine of humans and many animals. Within the genus Bifidobacterium, several beneficial effects have been attributed to strains belonging to the subspecies Bifidobacterium longum subsp. longum and Bifidobacterium longum subsp. infantis, which are often found in infants and adults. The increasing numbers of sequenced genomes belonging to these two subspecies, and the availability of novel computational tools focused on predicting glycolytic abilities, with the aim of understanding the capabilities of degrading specific carbohydrates, allowed us to depict the potential glycoside hydrolases (GH) of these bacteria, with a focus on those GH profiles that differ in the two subspecies. We performed an in silico examination of 188 sequenced B. longum genomes and depicted the commonly present and strain-specific GHs and GH families among representatives of this species. Additionally, GH profiling, genome-based and 16S rRNA-based clustering analyses showed that the subspecies assignment of some strains does not properly match with their genetic background. Furthermore, the analysis of the potential GH component allowed the distinction of clear GH patterns. Some of the GH activities, and their link with the two subspecies under study, are further discussed. Overall, our in silico analysis poses some questions about the suitability of considering the GH activities of B. longum subsp. longum and B. longum subsp. infantis to gain insight into the characterization and classification of these two subspecies with probiotic interest.

There are strong evidences that probiotics influence the immune status of the host, in a strain-specific manner, acting in the gastrointestinal tract. On the hypothesis that certain extracellular proteins and peptides from gut bacteria may mediate part of this immunomodulation and assuming they are able to diffuse through the mucus layer and interact with immune cells we have developed this work. Our study attempts to understand the immunomodulatory mechanisms of (i) Pext, the extracellular protein fraction of Lactobacillus acidophilus DSM20079T, (ii) HM14, a peptide encrypted in an extracellular glycoside hydrolase from Bifidobacterium longum NCIMB 8809 and (iii) Escherichia coli O111:B4 lipopolysaccharide (LPS), a well-known pro-inflammatory molecule, over human peripheral blood mononuclear cells (PBMCs). An untargeted LC-ESI-QTOF-MS metabolomics approach was applied to reveal intracellular changes in treated-PBMCs isolated from healthy donors. Differences in NADH arrest, NAD+ concentration reduction, as well as increases in palmitic acid and methanephrin were observed in HM14 and Pext treated-cells compared to those stimulated with LPS. This would support an anti-inflammatory molecular mechanism of action of such proteinaceous molecules. Moreover, this methodology has confirms the importance of metabolomics approaches to better understanding immune cell responses to gut bacterial-derived molecules.

BACKGROUND:The wide range of potential applications has made the Basic Local Alignment Search Tool (BLAST) a ubiquitous tool in the field of Molecular Biology. Within this context, it is increasingly appealing to embed BLAST services within larger Web applications. RESULTS:This work introduces BlasterJS viewer, a new JavaScript library for the lightweight development of Web-based applications supporting the visualisation of BLAST outputs. BlasterJS detaches from similar data viewers by focusing on the visual and interactive display of sequence similarity results and being completely independent of BLAST services. BlasterJS is compatible with the text outputs generated by the BLAST family of programs, namely BLASTp, BLASTn, BLASTx, tBLASTn, and tBLASTx, and works in all major Web browsers. Furthermore, BlasterJS is available through the EBI's BioJS registry 5, which extends its potential use to a wider scope of bioinformatics applications. CONCLUSIONS:BlasterJS is new Javascript library that enables easy and seamless integration of visual and interactive representations of BLAST outputs in Web-based applications supporting sequence similarity search. BlasterJS is free accessible at http://sing-group.org/blasterjs/.

Bifidobacterium represents a genus within the phylum Actinobacteria which is one of the major phyla in the healthy intestinal tract of humans. Bifidobacterium is one of the most abundant genera in adults, but its predominance is even more pronounced in infants, especially during lactation, when they can constitute the majority of the total bacterial population. They are one of the pioneering colonizers of the early gut microbiota, and they are known to play important roles in the metabolism of dietary components, otherwise indigestible in the upper parts of the intestine, and in the maturation of the immune system. Bifidobacteria have been shown to interact with human immune cells and to modulate specific pathways, involving innate and adaptive immune processes. In this mini-review, we provide an overview of the current knowledge on the immunomodulatory properties of bifidobacteria and the mechanisms and molecular players underlying these processes, focusing on the corresponding implications for human health. We deal with in vitro models suitable for studying strain-specific immunomodulatory activities. These include peripheral blood mononuclear cells and T cell-mediated immune responses, both effector and regulatory cell responses, as well as the modulation of the phenotype of dendritic cells, among others. Furthermore, preclinical studies, mainly germ-free, gnotobiotic, and conventional murine models, and human clinical trials, are also discussed. Finally, we highlight evidence supporting the immunomodulatory effects of bifidobacterial molecules (proteins and peptides, exopolysaccharides, metabolites, and DNA), as well as the role of bifidobacterial metabolism in maintaining immune homeostasis through cross-feeding mechanisms.

Torralba es un yacimiento del Pleistoceno medio, con dataciones numéricas que le sitúan en el MIS 7 (191/243 ka). Su industria, que procede de niveles fluviales y se encuentra fundamentalmente en posición secundaria, fue elaborada sobre materias primas introducidas desde distancias que pueden alcanzar varias decenas de kilómetros. El estudio tecno-económico de las series industriales obtenidas en las excavaciones realizadas por el marqués de Cerralbo (1909-1913) y Clark Howell (1961-1963) confirma la identidad achelense de la industria y la realización de procesos de producción, configuración y mantenimiento en el yacimiento, aunque al menos una parte del macroutillaje fue introducida ya configurada en el mismo. Se identifican aspectos tecnológicos singulares en el contexto del achelense europeo, como el uso frecuente del sílex en la elaboración de hendedores.

Scientific studies focused on the role of the human microbiome over human health have generated billions of gigabits of genetic information during the last decade. Nowadays integration of all this information in public databases and development of pipelines allowing us to biotechnologically exploit this information are urgently needed. Prediction of the potential bioactivity of the products encoded by the human gut microbiome, or metaproteome, is the first step for identifying proteins responsible for the molecular interaction between microorganisms and the immune system. We have recently published the Mechanism of Action of the Human Microbiome (MAHMI) database (http://www.mahmi.org), conceived as a resource compiling peptide sequences with a potential immunomodulatory activity. Fifteen out of the 300 hundred million peptides contained in the MAHMI database were synthesized. These peptides were identified as being encrypted in proteins produced by gut microbiota members, they do not contain cleavage points for the major intestinal endoproteases and displayed high probability to have immunomodulatory bioactivity. The bacterial peptides FR-16 and LR-17 encrypted in proteins from Bifidobacterium longum DJ010A and Bifidobacterium fragilis YCH46 respectively, showed the higher immune modulation capability over human peripheral blood mononuclear cells. Both peptides modulated the immune response toward increases in the Th17 and decreases in the Th1 cell response, together with an induction of IL-22 production. These results strongly suggest the combined use of bioinformatics and in vitro tools as a first stage in the screening of bioactive peptides encrypted in the human gut metaproteome.

Typical bacterial strain differentiation methods are often challenged by high genetic similarity between strains. To address this problem, we introduce a novel in silico peptide fingerprinting method based on conventional wet-lab protocols that enables the identification of potential strain-specific peptides. These can be further investigated using in vitro approaches, laying a foundation for the development of biomarker detection and application-specific methods. This novel method aims at reducing large amounts of comparative peptide data to binary matrices while maintaining a high phylogenetic resolution. The underlying case study concerns the Bacillus cereus group, namely the differentiation of Bacillus thuringiensis, Bacillus anthracis and Bacillus cereus strains. Results show that trees based on cytoplasmic and extracellular peptidomes are only marginally in conflict with those based on whole proteomes, as inferred by the established Genome-BLAST Distance Phylogeny (GBDP) method. Hence, these results indicate that the two approaches can most likely be used complementarily even in other organismal groups. The obtained results confirm previous reports about the misclassification of many strains within the B. cereus group. Moreover, our method was able to separate the B. anthracis strains with high resolution, similarly to the GBDP results as benchmarked via Bayesian inference and both Maximum Likelihood and Maximum Parsimony. In addition to the presented phylogenomic applications, whole-peptide fingerprinting might also become a valuable complementary technique to digital DNA-DNA hybridization, notably for bacterial classification at the species and subspecies level in the future.

Accumulated evidence suggests a relationship between specific allergic processes, such as atopic eczema in children, and an aberrant fecal microbiota. However, little is known about the complete microbiota profile of adult individuals suffering from asthma. We determined the fecal microbiota in 21 adult patients suffering allergic asthma (age 39.43 ± 10.98 years old) and compare it with the fecal microbiota of 22 healthy controls (age 39.29 ± 9.21 years old) using culture independent techniques. An Ion-Torrent 16S rRNA gene-based amplification and sequencing protocol was used to determine the fecal microbiota profile of the individuals. Sequence microbiota analysis showed that the microbial alpha-diversity was not significantly different between healthy and allergic individuals and no clear clustering of the samples was obtained using an unsupervised principal component analysis. However, the analysis of specific bacterial groups allowed us to detect significantly lower levels of bifidobacteria in patients with long-term asthma. Also, in allergic individuals the Bifidobacterium adolescentis species prevailed within the bifidobacterial population. The reduction in the levels on bifidobacteria in patients with long-term asthma suggests a new target in allergy research and opens possibilities for the therapeutic modulation of the gut microbiota in this group of patients.

ABSTRACT Systemic lupus erythematosus (SLE) is the prototypical systemic autoimmune disease in humans and is characterized by the presence of hyperactive immune cells and aberrant antibody responses to nuclear and cytoplasmic antigens, including characteristic anti–double-stranded DNA antibodies. We performed a cross-sectional study in order to determine if an SLE-associated gut dysbiosis exists in patients without active disease. A group of 20 SLE patients in remission, for which there was strict inclusion and exclusion criteria, was recruited, and we used an optimized Ion Torrent 16S rRNA gene-based analysis protocol to decipher the fecal microbial profiles of these patients and compare them with those of 20 age- and sex-matched healthy control subjects. We found diversity to be comparable based on Shannon’s index. However, we saw a significantly lower Firmicutes/Bacteroidetes ratio in SLE individuals (median ratio, 1.97) than in healthy subjects (median ratio, 4.86; P < 0.002). A lower Firmicutes/Bacteroidetes ratio in SLE individuals was corroborated by quantitative PCR analysis. Notably, a decrease of some Firmicutes families was also detected. This dysbiosis is reflected, based on in silico functional inference, in an overrepresentation of oxidative phosphorylation and glycan utilization pathways in SLE patient microbiota. IMPORTANCE Growing evidence suggests that the gut microbiota might impact symptoms and progression of some autoimmune diseases. However, how and why this microbial community influences SLE remains to be elucidated. This is the first report describing an SLE-associated intestinal dysbiosis, and it contributes to the understanding of the interplay between the intestinal microbiota and the host in autoimmune disorders.

Bifidobacteria are known as anaerobic/microaerophilic and fermentative microorganisms, which commonly inhabit the gastrointestinal tract of various animals and insects. Analysis of the 2,167,301 bp genome of Bifidobacterium asteroides PRL2011, a strain isolated from the hindgut of Apis mellifera var. ligustica, commonly known as the honey bee, revealed its predicted capability for respiratory metabolism. Conservation of the latter gene clusters in various B. asteroides strains enforces the notion that respiration is a common metabolic feature of this ancient bifidobacterial species, which has been lost in currently known mammal-derived Bifidobacterium species. In fact, phylogenomic based analyses suggested an ancient origin of B. asteroides and indicates it as an ancestor of the genus Bifidobacterium. Furthermore, the B. asteroides PRL2011 genome encodes various enzymes for coping with toxic products that arise as a result of oxygen-mediated respiration.

The human gastrointestinal tract is exposed to a huge variety of microorganisms, either commensal or pathogenic; at this site, a balance between immunity and immune tolerance is required. Intestinal dendritic cells (DCs) control the mechanisms of immune response/tolerance in the gut. In this paper we have identified a peptide (STp) secreted by Lactobacillus plantarum, characterized by the abundance of serine and threonine residues within its sequence. STp is encoded in one of the main extracellular proteins produced by such species, which includes some probiotic strains, and lacks cleavage sites for the major intestinal proteases. When studied in vitro, STp expanded the ongoing production of regulatory IL-10 in human intestinal DCs from healthy controls. STp-primed DC induced an immunoregulatory cytokine profile and skin-homing profile on stimulated T-cells. Our data suggest that some of the molecular dialogue between intestinal bacteria and DCs may be mediated by immunomodulatory peptides, encoded in larger extracellular proteins, secreted by commensal bacteria. These peptides may be used for the development of nutraceutical products for patients with IBD. In addition, this kind of peptides seem to be absent in the gut of inflammatory bowel disease patients, suggesting a potential role as biomarker of gut homeostasis.

Spain and three of its closest neighbours (Germany, Austria and Portugal) have recently undergone drastic legal changes regarding euthanasia and assisted suicide . Our article performs a Comparative Law analysis of the solutions adopted in each country. It seems especially relevant if we consider that, despite their apparent similarity at first sight, those solutions present significant differences among them. These differences also show some broader constitutional issues underlying the central problem, which are pointed out pending further discussion. [ABSTRACT FROM AUTHOR]