Design and Expression of Specific Hybrid Lantibiotics Active Against Pathogenic Clostridium spp.

Clostridium difficile has been reported as the most common cause of nosocomial diarrhea (antibiotic-associated diarrhea), resulting in significant morbidity and mortality in hospitalized patients. The resistance of the clostridia' spores to antibiotics and their side effects on the gut microbiota are two factors related to the emergence of infection and its relapses. Lantibiotics provide an innovative alternative for cell growth inhibition due to their dual mechanism of action (membrane pore-forming and cell wall synthesis inhibition) and low resistance rate. Based on the fact that bacteriocins are usually active against bacteria closely related to the producer strains, a new dual approach combining genome mining and synthetic biology was performed, by designing new lantibiotics with high activity and specificity toward Clostridium. We first attempted the heterologous expression of putative lantibiotics identified following Clostridium genome mining. Subsequently, we designed new hybrid lantibiotics combining the start or end of the putative clostridial peptides and the start or end parts of nisin. The designed peptides were cloned and expressed using the nisin biosynthetic machinery in Lactococcus lactis. From the 20 initial peptides, only 1 fulfilled the requirements established in this work to be considered as a good candidate: high heterologous production level and high specificity/activity against clostridial species. The high specificity and activity observed for the peptide AMV10 makes it an interesting candidate as an alternative to traditional antibiotics in the treatment of C. difficile infections, avoiding side effects and protecting the normal gut microbiota

High-throughput screening for substrate specificity-adapted mutants of the nisin dehydratase NisB

Microbial lanthipeptides are formed by a two-step enzymatic introduction of (methyl)lanthionine rings. A dehydratase catalyzes the dehydration of serine and threonine residues, yielding dehydroalanine and dehydrobutyrine, respectively. Cyclase-catalyzed coupling of the formed dehydroresidues to cysteines forms (methyl)lanthionine rings in a peptide. Lanthipeptide biosynthetic systems allow discovery of target-specific, lanthionine-stabilized therapeutic peptides. However, the substrate specificity of existing modification enzymes impose limitations on installing lanthionines in non-natural substrates. The goal of the present study was to obtain a lanthipeptide dehydratase with the capacity to dehydrate substrates that are unsuitable for the nisin dehydratase NisB. We report high-throughput screening for tailored specificity of intracellular, genetically encoded NisB dehydratases. The principle is based on the screening of bacterially displayed lanthionine-constrained streptavidin ligands, which have a much higher affinity for streptavidin than linear ligands. The designed NisC-cyclizable high-affinity ligands can be formed via mutant NisB-catalyzed dehydration but less effectively via wild-type NisB activity. In Lactococcus lactis, a cell surface display precursor was designed comprising DSHPQFC. The Asp residue preceding the serine in this sequence disfavors its dehydration by wild-type NisB. The cell surface display vector was coexpressed with a mutant NisB library and NisTC. Subsequently, mutant NisB-containing bacteria that display cyclized strep ligands on the cell surface were selected via panning rounds with streptavidin-coupled magnetic beads. In this way, a NisB variant with a tailored capacity of dehydration was obtained, which was further evaluated with respect to its capacity to dehydrate nisin mutants. These results demonstrate a powerful method for selecting lanthipeptide modification enzymes with adapted substrate specificity

A new source of representative secondary PET nanoplastics. Obtention, characterization, and hazard evaluation

Acord transformatiu CRUE-CSICAltres ajuts: Juan de la Cierva IJC2020-2686IMicro and nanoplastics (MNPLs) are emergent environmental pollutants requiring urgent information on their potential risks to human health. One of the problems associated with the evaluation of their undesirable effects is the lack of representative samples, matching those resulting from the environmental degradation of plastic wastes. To such end, we propose an easy method to obtain polyethylene terephthalate nanoplastics from water plastic bottles (PET-NPLs) but, in principle, applicable to any other plastic goods sources. An extensive characterization indicates that the proposed process produces uniform samples of PET-NPLs of around 100 nm, as determined by using AF4 and multi-angle and dynamic light scattering methodologies. An important point to be highlighted is that to avoid the metal contamination resulting from methods using metal blades/burrs for milling, trituration, or sanding, we propose to use diamond burrs to produce metal-free samples. To visualize the toxicological profile of the produced PET-NPLs we have evaluated their ability to be internalized by cells, their cytotoxicity, their ability to induce oxidative stress, and induce DNA damage. In this preliminary approach, we have detected their cellular uptake, but without the induction of significant biological effects. Thus, no relevant increases in toxicity, reactive oxygen species (ROS) induction, or DNA damage -as detected with the comet assay- have been observed. The use of representative samples, as produced in this study, will generate relevant data in the discussion about the potential health risks associated with MNPLs exposures

Recomendaciones dirigidas a los familiares responsables del cuidado domiciliario de un paciente diagnosticado con Covid-19

The crisis that is causing the disease called COVID-19 is severely affecting the health system of many countries, which require alternative measures to care this large number of patients who are not able to be treated at medical centers. This is the reason why home care is considered an alternative, for which it must be very rigorous with the care of the sick person, since if it is not done in a proper way, the other members of the family home are under risk of contamination, which would increase the saturation of the health system. Indications such as the stay of the patient in one place, the use of protective barriers when entering this area, or the constant sanitization of hands, among other measures, can lead to the success of this type of actions, favoring the prompt recovery of the patient and the preservation of the health of the other members at home.La crisis que está ocasionando la enfermedad denominada COVID-19 está afectando severamente al sistema sanitario de diversos países, lo que obliga a tomar medidas alternativas para poder atender a una gran cantidad de pacientes que no pueden acceder a los centros de atención médica. Es por esta razón que se considera como una alternativa la atención domiciliaria para lo cual se debe ser muy riguroso en el cuidado de la persona afectada con COVID 19, ya que, si esto no se realiza de forma adecuada, los demás integrantes del hogar corren el riesgo de contaminarse, lo que incrementaría la saturación de los establecimientos de salud. Indicaciones como la permanencia del enfermo en un solo lugar, el uso de barreras de protección cuando se ingresa a esta área o la constante práctica de la higiene de manos, entre otras medidas, puede encaminar al éxito de este tipo de atención favoreciendo la pronta recuperación del paciente y la preservación de la salud de los demás habitantes del hogar.A atual crise que está causando a doença denominada COVID-19 está afetando gravemente o sistema de saúde de vários países, o que requer outras medidas a serem tomadas para poder atender este grande número de pacientes fora dos centros hospitalares. É por isso que a assistência domiciliar é considerada uma alternativa, para a qual deve ser muito rigorosa com o cuidado do doente, pois se não for realizada de forma adequada os demais membros do domicílio correm o risco de contaminar o que aumentaria a saturação das unidades de saúde. Indicações como a permanência do paciente em um único local, o uso de barreiras protetoras na entrada nesta área ou a prática constante de higienização das mãos, entre outras medidas, podem levar ao sucesso desse tipo de atendimento, favorecendo uma recuperação rápida do paciente e a preservação da saúde dos demais habitantes. &nbsp

Collaborative Database to Track Mass Mortality Events in the Mediterranean Sea

Anthropogenic climate change, and global warming in particular, has strong and increasing impacts on marine ecosystems (Poloczanska et al., 2013; Halpern et al., 2015; Smale et al., 2019). The Mediterranean Sea is considered a marine biodiversity hot-spot contributing to more than 7% of world's marine biodiversity including a high percentage of endemic species (Coll et al., 2010). The Mediterranean region is a climate change hotspot, where the respective impacts of warming are very pronounced and relatively well documented (Cramer et al., 2018). One of the major impacts of sea surface temperature rise in the marine coastal ecosystems is the occurrence of mass mortality events (MMEs). The first evidences of this phenomenon dated from the first half of'80 years affecting the Western Mediterranean and the Aegean Sea (Harmelin, 1984; Bavestrello and Boero, 1986; Gaino and Pronzato, 1989; Voultsiadou et al., 2011). The most impressive phenomenon happened in 1999 when an unprecedented large scale MME impacted populations of more than 30 species from different phyla along the French and Italian coasts (Cerrano et al., 2000; Perez et al., 2000). Following this event, several other large scale MMEs have been reported, along with numerous other minor ones, which are usually more restricted in geographic extend and/or number of affected species (Garrabou et al., 2009; Rivetti et al., 2014; Marbà et al., 2015; Rubio-Portillo et al., 2016, authors' personal observations). These events have generally been associated with strong and recurrent marine heat waves (Crisci et al., 2011; Kersting et al., 2013; Turicchia et al., 2018; Bensoussan et al., 2019) which are becoming more frequent globally (Smale et al., 2019). Both field observations and future projections using Regional Coupled Models (Adloff et al., 2015; Darmaraki et al., 2019) show the increase in Mediterranean sea surface temperature, with more frequent occurrence of extreme ocean warming events. As a result, new MMEs are expected during the coming years. To date, despite the efforts, neither updated nor comprehensive information can support scientific analysis of mortality events at a Mediterranean regional scale. Such information is vital to guide management and conservation strategies that can then inform adaptive management schemes that aim to face the impacts of climate change.MV-L was supported by a postdoctoral contract Juan de la Cierva-Incorporación (IJCI-2016-29329) of Ministerio de Ciencia, Innovación y Universidades. AI was supported by a Technical staff contract (PTA2015-10829-I) Ayudas Personal Técnico de Apoyo of Ministerio de Economía y Competitividad (2015). Interreg Med Programme (grant number Project MPA-Adapt 1MED15_3.2_M2_337) 85% cofunded by the European Regional Development Fund, the MIMOSA project funded by the Foundation Prince Albert II Monaco and the European Union's Horizon 2020 research and innovation programme under grant agreement no 689518 (MERCES). DG-G was supported by an FPU grant (FPU15/05457) from the Spanish Ministry of Education. J-BL was partially supported by the Strategic Funding UID/Multi/04423/2013 through national funds provided by FCT - Foundation for Science and Technology and European Regional Development Fund (ERDF), in the framework of the programme PT2020

Influence of Grasshopper Herbivory on Nitrogen Cycling in Northern Gulf of Mexico Black Needlerush Salt Marshes

Herbivory is a common process in salt marshes. However, the direct impact of marsh herbivory on nutrient cycling in this ecosystem is poorly understood. Using a 15N enrichment mesocosm study, we quantified nitrogen (N) cycling in sediment and plants of black needlerush (Juncus roemerianus) salt marshes, facilitated by litter decomposition and litter plus grasshopper feces decomposition. We found 15 times more 15N recovery in sediment with grasshopper herbivory compared to sediment with no grasshopper herbivory. In plants, even though we found three times and a half larger 15N recovery with grasshopper herbivory, we did not find significant differences. Thus, herbivory can enhance N cycling in black needlerush salt marshes sediments and elevate the role of these salt marshes as nutrient sinks.Fil: Montemayor Borsinger, Diana Ireri. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Marinas y Costeras. Universidad Nacional de Mar del Plata. Facultad de Ciencia Exactas y Naturales. Instituto de Investigaciones Marinas y Costeras; ArgentinaFil: Sparks, Eric L.. University of Mississippi; Estados Unidos. Alabama Sea Grant Consortium; Estados UnidosFil: Iribarne, Oscar Osvaldo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Marinas y Costeras. Universidad Nacional de Mar del Plata. Facultad de Ciencia Exactas y Naturales. Instituto de Investigaciones Marinas y Costeras; ArgentinaFil: Cebrian, Just. Dauphin Island Sea Lab; Estados Unidos. University of South Alabama; Estados Unido

Effects of nutrient enrichment and crab herbivory on a SW Atlantic salt marsh productivity

After intense debate it is now accepted that nutrients (a bottom-up process) and herbivores (a top-down process) are both important controls of plant productivity in many systems. Besides their direct effects, herbivores may also have profound positive or negative indirect effects that can be modulated by nutrients and time. The interactive relationships between time, nutrient availability and herbivore impacts (direct and indirect) on plant growth dynamics are an emerging research topic that merits further effort. Here we did several experiments in a SW Atlantic marsh to contribute towards that gap by focusing on the dominant plant, Spartina densiflora, and one of the dominant herbivores, the crab Neohelice (Chasmagnathus) granulata, in the marsh. Herbivory by the crab was highly seasonal, with most of the consumption occurring in fall. Even though crabs preferred nutrient enriched leaves, nitrogen content was not the driver of these seasonal variations. Crab herbivory had markedly indirect negative impacts on S. densiflora leaves, reducing their growth rates and increasing their senescence. These deleterious impacts may partially explain the seasonal decline in leaf growth and a net loss in leaf biomass observed in the fall. Fertilization did not seem to alter these processes. Adding nutrients increased leaf growth in the spring, where ambient herbivory was low, but it also increased herbivory in the fall, resulting in similar patterns as the ones observed under non-fertilized conditions. Herbivory by the crab also greatly affected the dynamics of S. densiflora stems. Increases in stem density in relation to initial conditions were larger in non-grazed than in grazed plots regardless of whether nutrients were added or not. Together, these results indicate that, in Southwestern marshes populated by S. densiflora and N. granulata, herbivory by the crab represents an important direct and indirect control of plant growth. Our results also emphasize the importance of considering impacts on growth rates and not only on biomass because not considering reduced growth after herbivory may lead to improper calculations of nutrient cycling or detritus production.Fil: Alberti, Juan. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Departamento de Biología. Laboratorio de Ecología; ArgentinaFil: Cebrian, Just. University of South Alabama; Estados Unidos. Dauphin Island Sea Lab; Estados UnidosFil: Mendez Casariego, Maria Agustina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Departamento de Biología. Laboratorio de Ecología; ArgentinaFil: Canepuccia, Alejandro Daniel. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Departamento de Biología. Laboratorio de Ecología; ArgentinaFil: Escapa, Carlos Mauricio. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Departamento de Biología. Laboratorio de Ecología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Iribarne, Oscar Osvaldo. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Departamento de Biología. Laboratorio de Ecología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentin

The blue carbon of southern southwest Atlantic salt marshes and their biotic and abiotic drivers

Abstract Coastal vegetated ecosystems are acknowledged for their capacity to sequester organic carbon (OC), known as blue C. Yet, blue C global accounting is incomplete, with major gaps in southern hemisphere data. It also shows a large variability suggesting that the interaction between environmental and biological drivers is important at the local scale. In southwest Atlantic salt marshes, to account for the space occupied by crab burrows, it is key to avoid overestimates. Here we found that southern southwest Atlantic salt marshes store on average 42.43 (SE = 27.56) Mg OC·ha−1 (40.74 (SE = 2.7) in belowground) and bury in average 47.62 g OC·m−2·yr−1 (ranging from 7.38 to 204.21). Accretion rates, granulometry, plant species and burrowing crabs were identified as the main factors in determining belowground OC stocks. These data lead to an updated global estimation for stocks in salt marshes of 185.89 Mg OC·ha−1 (n = 743; SE = 4.92) and a C burial rate of 199.61 g OC·m−2·yr−1 (n = 193; SE = 16.04), which are lower than previous estimates

Nutrient and herbivore alterations cause uncoupled changes in producer diversity, biomass and ecosystem function, but not in overall multifunctionality

Altered nutrient cycles and consumer populations are among the top anthropogenic influences on ecosystems. However, studies on the simultaneous impacts of human-driven environmental alterations on ecosystem functions, and the overall change in system multifunctionality are scarce. We used estuarine tidal flats to study the effects of changes in herbivore density and nutrient availability on benthic microalgae (diversity, abundance and biomass) and ecosystem functions (N2-fixation, denitrification, extracellular polymeric substances -EPS- as a proxy for sediment cohesiveness, sediment water content as a proxy of water retention capacity and sediment organic matter). We found consistent strong impacts of modified herbivory and weak effects of increased nutrient availability on the abundance, biomass and diversity of benthic microalgae. However, the effects on specific ecosystem functions were disparate. Some functions were independently affected by nutrient addition (N2-fixation), modified herbivory (sediment organic matter and water content), or their interaction (denitrification), while others were not affected (EPS). Overall system multifunction remained invariant despite changes in specific functions. This study reveals that anthropogenic pressures can induce decoupled effects between community structure and specific ecosystem functions. Our results highlight the need to address several ecosystem functions simultaneously for better ecosystem characterization and management.Fil: Alberti, Juan. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Marinas y Costeras. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Marinas y Costeras; ArgentinaFil: Cebrian, J.. Dauphin Island Sea Lab; Estados Unidos. University of South Alabama; Estados UnidosFil: Alvarez, F.. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Limnología "Dr. Raúl A. Ringuelet". Universidad Nacional de La Plata. Facultad de Ciencias Naturales y Museo. Instituto de Limnología; ArgentinaFil: Escapa, Mauricio. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Marinas y Costeras. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Marinas y Costeras; ArgentinaFil: Esquius, Karina Soledad. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Departamento de Biología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata; ArgentinaFil: Fanjul, Maria Eugenia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Marinas y Costeras. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Marinas y Costeras; ArgentinaFil: Sparks, E.L.. Mississippi State University Coastal Research and Extension; Estados Unidos. Mississippi-Alabama Sea Grant Consortium; Estados UnidosFil: Mortazavi, B.. Dauphin Island Sea Lab; Estados UnidosFil: Iribarne, Oscar Osvaldo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Marinas y Costeras. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Marinas y Costeras; Argentin