Notas briológicas de la provincia de Vizcaya. I. Estudio de las comunidades de esfagnos del Puerto de Urquiola

We have studied the sphagnum and the damp boggy comunities of the Puerto de Urquiola (Vizcaya). We have collected 10 species of Sphagnum,from these, 8 species are new records for Vizcaya. From the syntaxonomic study of the comunities, we describe for the first time Helodo-Sphagnetum Lemee 1937, Sphagnetosum crassicladisubas. nova.Hemos estudiado los esfagnos y las comunidades higro-turbosas del Puerto de Urquiola (Vizcaya). Se han recolectado 10 especies de Sphagnum, de ellas, 8 especies son nuevas citas para Vizcaya. Del estudio fitosociológico de las comunidades, se describe la subasociación Sphagnetosum crassicladi subas. nov

Data on recovery of 21 amino acids, 9 biogenic amines and ammonium ions after spiking four different beers with five concentrations of these analytes

A novel chromatographic method for the simultaneous analysis of nine biogenic amines, 21 amino acids and ammonium ions in beer has been recently described in “A UHPLC method for the simultaneous analysis of biogenic amines, amino acids and ammonium ions in beer” (Redruello et al., 2017) [1]. The present article provides recovery data of the 31 analytes after spiking four different beers with five concentrations of each analyte (15, 30, 60, 120 and 240 µM).This work was performed with the financial support of the Spanish Ministry of Economy and Competitiveness (AGL2013-45431-R) and the Plan for Science, Technology and Innovation 2013–2017 of the Principality of Asturias, which is co-funded by the European Regional Development Fund (GRUPIN14-137).Peer reviewe

Implementation of the agmatine-controlled expression system for inducible gene expression in Lactococcus lactis

[Background] Lactococcus lactis has been safely consumed in fermented foods for millennia. This Gram-positive bacterium has now become of industrial importance as an expression host for the overproduction of lipopolysaccharide-free recombinant proteins used as food ingredients, therapeutic proteins and biotechnological enzymes.[Results] This paper reports an agmatine-controlled expression (ACE) system for L. lactis, comprising the lactococcal agmatine-sensor/transcriptional activator AguR and its target promoter P aguB . The usefulness and efficiency of this system was checked via the reporter gene gfp and by producing PEP (Myxococcus xanthus prolyl-endopeptidase), an enzyme of biomedical interest able to degrade the immunotoxic peptides produced during the gastrointestinal breakdown of gluten.[Conclusion] The ACE system developed in this work was suitable for the efficient expression of the functional recombinant proteins GFP and PEP. The expression system was tightly regulated by the agmatine concentration and allowed high protein production without leakiness.This work was performed with the financial support of the Spanish Ministry of Economy and Competitiveness (AGL2013-45431-R), the Principality of Asturias Plan for Science, Technology and Innovation 2013-2017, and FEDER funds (GRUPIN14-137). We acknowledge support of the publication fee by the CSIC Open Access Publication Support Initiative through its Unit of Information Resources for Research (URICI). P.A.S. was the beneficiary of a fellowship from the FICYT, Principality of Asturias, Spain (BP09093). D.M.L. and B.d.R. were beneficiaries of JAE DOC contracts (CSIC).Peer Reviewe

Putrescine production by Lactococcus lactis subsp. cremoris CECT 8666 is reduced by NaCl via a decrease in bacterial growth and the repression of the genes involved in putrescine production

The reduction of NaCl in food is a public health priority; high NaCl intakes have been associated with serious health problems. However, it is reported that reducing the NaCl content of cheeses may lead to an increase in the content of biogenic amines (BAs). The present work examines the effect of NaCl on the accumulation of putrescine (one of the BAs often detected at high concentration in cheese) in experimental Cabrales-like cheeses containing Lactococcus lactis subsp. cremoris CECT 8666, a dairy strain that catabolises agmatine to putrescine via the agmatine deiminase (AGDI) pathway. The genes responsible for this pathway are grouped in the AGDI cluster. This comprises a regulatory gene (aguR) (transcribed independently), followed by the catabolic genes that together form an operon (aguBDAC). Reducing the NaCl concentration of the cheese led to increased putrescine accumulation. In contrast, increasing the NaCl concentration of both pH-uncontrolled and pH-controlled (pH 6) cultures of L. lactis subsp. cremoris CECT 8666 significantly inhibited its growth and the production of putrescine. Such production appeared to be inhibited via a reduction in the transcription of the aguBDAC operon; no effect on the transcription of aguR was recorded. The present results suggest that low-sodium cheeses are at risk of accumulating higher concentrations of putrescine.This work was funded by the Spanish Ministry of Economy and Competitiveness (AGL2013-45431-R) and by the GRUPIN14-137 project, which is co-financed by the Plan for Science, Technology and Innovation of the Principality of Asturias 2013–2017 and the European Regional Development Funds.Peer reviewe

Putrescine biosynthesis in Lactococcus lactis is transcriptionally activated at acidic pH and counteracts acidification of the cytosol

Lactococcus lactis subsp. cremoris CECT 8666 is a lactic acid bacterium that synthesizes the biogenic amine putrescine from agmatine via the agmatine deiminase (AGDI) pathway. The AGDI genes cluster includes aguR. This encodes a transmembrane protein that functions as a one-component signal transduction system, the job of which is to sense the agmatine concentration of the medium and accordingly regulate the transcription of the catabolic operon aguBDAC. The latter encodes the proteins necessary for agmatine uptake and its conversion into putrescine. This work reports the effect of extracellular pH on putrescine biosynthesis and on the genetic regulation of the AGDI pathway. Increased putrescine biosynthesis was detected at acidic pH (pH 5) compared to neutral pH. Acidic pH induced the transcription of the catabolic operon via the activation of the aguBDAC promoter P. However, the external pH had no significant effect on the activity of the aguR promoter P, or on the transcription of the aguR gene. The transcriptional activation of the AGDI pathway was also found to require a lower agmatine concentration at pH 5 than at neutral pH. Finally, the following of the AGDI pathway counteracted the acidification of the cytoplasm under acidic external conditions, suggesting it to provide protection against acid stress.This work was funded by the Spanish Ministry of Economy and Competitiveness (AGL2013-45431-R) and by the GRUPIN14-137 Project, which is co-financed by the Plan for Science, Technology and Innovation of the Principality of Asturias 2014–2017 and the European Regional Development Funds. The authors thank Adrian Burton for language and editing assistance.Peer Reviewe

Tyramine biosynthesis is transcriptionally induced at low pH and improves the fitness of Enterococcus faecalis in acidic environments

© 2014, Springer-Verlag Berlin Heidelberg. Enterococcus faecalis is a commensal bacterium of the human gut that requires the ability to pass through the stomach and therefore cope with low pH. E. faecalis has also been identified as one of the major tyramine producers in fermented food products, where they also encounter acidic environments. In the present work, we have constructed a non-tyramine-producing mutant to study the role of the tyramine biosynthetic pathway, which converts tyrosine to tyramine via amino acid decarboxylation. Wild-type strain showed higher survival in a system that mimics gastrointestinal stress, indicating that the tyramine biosynthetic pathway has a role in acid resistance. Transcriptional analyses of the E. faecalis V583 tyrosine decarboxylase cluster showed that an acidic pH, together with substrate availability, induces its expression and therefore the production of tyramine. The protective role of the tyramine pathway under acidic conditions appears to be exerted through the maintenance of the cytosolic pH. Tyramine production should be considered important in the adaptability of E. faecalis to acidic environments, such as fermented dairy foods, and to survive passage through the human gastrointestinal tract.This work was funded by the Ministry of Economy and Competitiveness, Spain (AGL2013-45431-R) and the Spanish National Research Council (CSIC201270E144). M.P. is beneficiary of an FPU fellowship from the Spanish Ministry of Education.Peer Reviewe

Is256 abolishes gelatinase activity and biofilm formation in a mutant of the nosocomial pathogen enterococcus faecalis v583

Enterococcus faecalis is one of the most controversial species of lactic acid bacteria. Some strains are used as probiotics, while others are associated with severe and life-threatening nosocomial infections. Their pathogenicity depends on the acqui-sition of multidrug resistance and virulence factors. Gelatinase, which is required in the first steps of biofilm formation, is an important virulence determinant involved in E. faecalis pathogenesis, including endocarditis and peritonitis. The gene that codes for gelatinase (gelE) is controlled by the Fsr quorum-sensing system, whose encoding genes (fsrA, fsrB, fsrC, and fsrD) are located immediately upstream of gelE. The integration of a DNA fragment into the fsr locus of a derived mutant of E. faecalis V583 suppressed the gelatinase activity and prevented biofilm formation. Sequence analysis indicated the presence of IS256 integrated into the fsrC gene at nucleotide position 321. Interestingly, IS256 is also associated with biofilm formation in Staphylococcus epidermidis and Staphylococcus aureus. This is the first description of an insertion sequence that prevents biofilm formation in E. faecalis. Copyright de NRC Research PressPeer Reviewe

Biofilm-forming capacity in biogenic amine-producing bacteria isolated from dairy products

Biofilms on the surface of food industry equipment are reservoirs of potentially food-contaminating bacteria-both spoilage and pathogenic. However, the capacity of biogenic amine (BA)-producers to form biofilms has remained largely unexamined. BAs are low molecular weight, biologically active compounds that in food can reach concentrations high enough to be a toxicological hazard. Fermented foods, especially some types of cheese, accumulate the highest BA concentrations of all. The present work examines the biofilm-forming capacity of 56 BA-producing strains belonging to three genera and 10 species (12 Enterococcus faecalis, 6 Enterococcus faecium, 6 Enterococcus durans, 1 Enterococcus hirae, 12 Lactococcus lactis, 7 Lactobacillus vaginalis, 2 Lactobacillus curvatus, 2 Lactobacillus brevis, 1 Lactobacillus reuteri, and 7 Lactobacillus parabuchneri), all isolated from dairy products. Strains of all the tested species - except for L. vaginalis-were able to produce biofilms on polystyrene and adhered to stainless steel. However, the biomass produced in biofilms was strain-dependent. These results suggest that biofilms may provide a route via which fermented foods can become contaminated by BA-producing microorganisms.This work was funded by the Spanish Ministry of the Economy and Competitiveness (AGL2013-45431-R) and the Plan for Science, Technology and Innovation 2013–2017 financed by the European Regional Development Fund and the Principality of Asturias (GRUPIN14-137). MD is a beneficiary of an FPI fellowship from the Spanish Ministry of the Economy and Competitiveness.Peer Reviewe

Genetic and functional analysis of biogenic amine production capacity among starter and non-starter lactic acid bacteria isolated from artisanal cheeses

© 2015, Springer-Verlag Berlin Heidelberg. This work reports the capacity of 137 strains of starter and non-starter LAB belonging to nine species of the genera Lactobacillus, Lactococcus, Streptococcus and Leuconostoc (all isolated from artisanal cheeses) to produce histamine, tyramine, putrescine and β-phenylethylamine, the biogenic amines (BA) most commonly found in dairy products. Production assays were performed in liquid media supplemented with the appropriate precursor amino acid; culture supernatants were then tested for BA by (U)HPLC. In addition, the presence of key genes involved in the biosynthetic pathways of the target BA, including the production of putrescine via the agmatine deiminase pathway, was assessed by PCR. Twenty strains were shown to have genes involved in the synthesis of BA; these belonged to the species Lactobacillus brevis (4), Lactobacillus curvatus (3), Lactococcus lactis (11) and Streptococcus thermophilus (2). With the exception of the two S. thermophilus strains, all those possessing genes involved in BA production synthesized the corresponding compound. Remarkably, all the putrescine-producing strains used the agmatine deiminase pathway. Four L. brevis and two L. curvatus strains were found able to produce both tyramine and putrescine. There is increasing interest in the use of autochthonous LAB strains in starter and adjunct cultures for producing dairy products with ‘particular geographic indication’ status. Such strains should not produce BA; the present results show that BA production capacity should be checked by (U)HPLC and PCR.This work was funded by the Ministry of Economy and Competitiveness, Spain (AGL2013-45431-R), the Fundación para el Fomento en Asturias de la Investigación Científica Aplicada y la Tecnología (FICYT), cofunded by FEDER (GRUPIN14-137) and the INIA (RM2011-00005-00-00).Peer Reviewe

Comparative analysis of the in vitro cytotoxicity of the dietary biogenic amines tyramine and histamine

Tyramine and histamine, the most toxic biogenic amines (BA), are often found in high concentrations in certain foods. Prompted by the limited knowledge of BA toxicity, and increasing awareness of the risks associated with high intakes of dietary BA, the in vitro cytotoxicity of tyramine and histamine was investigated. Tyramine and histamine were toxic for HT29 intestinal cell cultures at concentrations commonly found in BA-rich food, as determined by real-time cell analysis. Surprisingly, tyramine had a stronger and more rapid cytotoxic effect than histamine. Their mode of action was also different, while tyramine caused cell necrosis, histamine induced apoptosis. To avoid health risks, the BA content of foods should be reduced and legal limits established for tyramine.This work was funded by the Spanish Ministry of Economy and Competitiveness (AGL2013-45431-R) and by the GRUPIN14-137 project, which is co-financed by the Plan for Science, Technology and Innovation of the Principality of Asturias 2014-2017 and the European Regional Development Funds.Peer reviewe