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Background & aims. Fragility fractures are those caused by low-energy trauma or falls from standing height. The mortality after the first year of suffering a hip fracture ranges from 15 to 30%. This problem has been addressed through the creation of interdisciplinary programs for fragility fracture early diagnosis and prevention. Describing the clinical characteristics of a cohort of patients diagnosed with fragility fractures in 10 fracture liaison services in Colombia. Methods. Led by the Asociaci n Colombiana de Osteoporosis y Metabolismo Mineral, this is a cross-sectional descriptive study, in patients with a diagnosis of fragility fracture in 10 fracture liaison services. Demographic and clinical variables were described for one year from the diagnosis of the fracture. Results. were analyzed 1699 records of patient fractures between 50 and 101 years of age, 1334 were women (76.5%), 581/1484 (39.1%) had previous fragility fractures, 570/1599 (35.7%) had a previous diagnosis of osteoporosis. Of these, 70/1051 (7%) received anti-osteoporosis medication, and 311/733 (42.4%) received it post-fracture. Of the total records with information, 65/707 (9.2%) died the following year of the fracture. Conclusions. Fracture prevention programs in institutions allow for de16 A. Medina et al. tecting patients with fragility fractures, clinical characteristics of these, in addition to strengthening the conditions of a centralized national registry, based on the parameters of the International Osteoporosis Foundation - Capture the Fracture program, through strategies with multidisciplinary personnel.

Summary Background Clostridioides difficile is transmitted through endospores. Most disinfection procedures for these structures deploy high concentrations of chlorine-derived compounds such as sodium hypochlorite (NaOCl) and sodium dichloroisocyanurate (NaDCC). However, these substances are linked to undesirable public health and environmental issues. Aim To compare the efficacy of NaCl-derived electrochemically activated solution (ECAS, 0.18% w/v NaOCl, pH=9.6–10.3), commercial bleach (5000 ppm, 2.83% w/v NaOCl, pH=5.6) and NaDCC (1000 ppm, pH=6.8) to inactivate C. difficile endospores on surfaces using a standard quantitative test (EPA MO-21-03). Findings Ten representative reference and field strains from multi-locus sequence typing Clades 1–5 were assayed (N=10). Irrespective of the phylogenetic background of the strains, ECAS showed comparable or better log reduction values [mean=3.22, 95% confidence interval (CI) 0.40–5.56] than bleach (mean=2.74, 95% CI 0.12–5.50) and NaDCC (mean=2.02, 95% CI 0.10–5.12). Cyclic voltammetry measurements revealed similar electrochemical behaviours and open-circuit potentials for ECAS and NaOCl. Congruently, similar morphologies for spores treated with these two compounds were observed by transmission electron microscopy. A factorial design demonstrated that exposure time, but not activation time, influenced the efficacy of ECAS. Conclusions ECAS and NaOC were found to have functional equivalence and may have a common mechanism of action.

Various antimicrobial agents are used in the poultry industry to treat microbial infections and prevent disease or as growth promoters. As a result, poultry litter (PL) can contain antibiotic residues (AR), antibiotic-resistant bacteria (ARB), and antibiotic resistance genes. Still, PL is used in many countries as a fertilizer and feed supplement for cattle. To evaluate whether usage of PL in agriculture leads to the accumulation of AR and ARB accumulate in the soil, we (i) measured the concentration of monensin, tylosin, ciprofloxacin, oxytetracycline, and chlortetracycline and the abundance of culturable monensin-, tylosin-, and ciprofloxacin-resistant bacteria in 15 commercial PL samples and (ii) exposed soil microcosms to two PL regimes and followed the persistence of PL-associated ARB for 128 days through cultivation on media containing antibiotics. The PL samples analyzed contained high concentrations of monensin (27–95 mg kg−1), tylosin (152–450 mg kg−1), ciprofloxacin (29–101 mg kg−1), and (oxy/chlor)tetracycline (13–87 mg kg−1). Congruently, they included large absolute and relative numbers of bacteria capable of growing on agar plates supplemented with 5 to 50 μg mL−1 monensin (medians, 107–109 CFU g−1, 0.6–45%) or 25 to 50 μg mL−1 tylosin (median, 108 CFU g−1, 14–26%). By contrast, the abundance of bacteria resistant to 25–250 μg mL−1 CP in the PL samples was much lower (median values ranging from 106 to less than 102 CFU g−1, relative abundances

Antimicrobial resistance (AMR) plays an important role in the pathogenesis and spread of Clostridioides difficile infection (CDI), the leading healthcare-related gastrointestinal infection in the world. An association between AMR and CDI outbreaks is well documented, however, data is limited to a few ‘epidemic’ strains in specific geographical regions. Here, through detailed analysis of 10 330 publicly-available C. difficile genomes from strains isolated worldwide (spanning 270 multilocus sequence types (STs) across all known evolutionary clades), this study provides the first species-wide snapshot of AMR genomic epidemiology in C. difficile . Of the 10 330 C . difficile genomes, 4532 (43.9 %) in 89 STs across clades 1–5 carried at least one genotypic AMR determinant, with 901 genomes (8.7 %) carrying AMR determinants for three or more antimicrobial classes (multidrug-resistant, MDR). No AMR genotype was identified in any strains belonging to the cryptic clades. C. difficile from Australia/New Zealand had the lowest AMR prevalence compared to strains from Asia, Europe and North America (PP C. difficile STs: ST1 (clade 2) with fluoroquinolone resistance (mainly T82I substitution in GyrA) (Ptet-family genes) (PB) resistance (mainly ermB) (PPtetM-positive transposon designated Tn6944 was identified, predominantly among clade 2 strains. This study provides a comprehensive review of AMR in the global C. difficile population which may aid in the early detection of drug-resistant C. difficile strains, and prevention of their dissemination worldwide.

Clostridioides difficile infection (CDI) remains an urgent global One Health threat. The genetic heterogeneity seen across C. difficile underscores its wide ecological versatility and has driven the significant changes in CDI epidemiology seen in the last 20 years. We analysed an international collection of over 12,000 C. difficile genomes spanning the eight currently defined phylogenetic clades. Through whole-genome average nucleotide identity, and pangenomic and Bayesian analyses, we identified major taxonomic incoherence with clear species boundaries for each of the recently described cryptic clades CI–III. The emergence of these three novel genomospecies predates clades C1–5 by millions of years, rewriting the global population structure of C. difficile specifically and taxonomy of the Peptostreptococcaceae in general. These genomospecies all show unique and highly divergent toxin gene architecture, advancing our understanding of the evolution of C. difficile and close relatives. Beyond the taxonomic ramifications, this work may impact the diagnosis of CDI.

Background When present in animal feedstuff, mycotoxins contaminants and antibiotic residues can have negative implications for animal production and Public Health, including the transmission of carcinogenic compounds and the selection of antibiotic resistant bacteria, respectively. So far there are no available methods in which both mycotoxins and antibiotic residues are analyzed using a parallel extraction approach. To address this issue, we developed a LC/MS methodology with high sensitivity (0.005 to 6.42 and 24.55 to 132.73 μg kg− 1 for mycotoxins and antimicrobials) and specificity (unique target ion mass/charge) that allows the detection of 26 mycotoxins and 23 antibiotic residues in animal feedstuff and validated it through the determination of these analytes in 294 animal feed and feed ingredient samples in the framework of a country-wide surveillance program. Two hundred and five of these samples were analyzed for mycotoxins and 89 for antibiotics. Findings Fumonisin was the most frequently toxin found, with FB1 and FB2 presenting prevalences of 50 and 52% and maximum concentrations of 14,927.61 and 8646.67 μg kg− 1, respectively. Other toxins, including diacetoxyscirpenol n = 4/101 (3.96%), fusarenon-X n = 2/101 (1.98%), citrinin n = 2 (1.98%), and patulin n = 1 (0.99%) were rarely found. Toxicologically relevant concentrations of toxin metabolites, such as HT-2 (6.38–485.49 μg kg− 1) and 3−/15-acetoxydeoxynivalenol (877.89–3236.56/5.44–1685.3 μg kg− 1), were also found. Few samples exceeded threshold mycotoxin concentrations defined in current EU guidelines. Dairy cattle and swine feeds included the higher number of samples exceeding guideline values (n = 6 and n = 5, respectively). From the total of samples analysed for antibiotics, 7.7% (n = 7/89) were classified as medicated for poultry and pigs. Unexpectedly, 57% of these medicated samples contained no detectable antibiotics (n = 4/7). The remaining 43% of the samples (n = 3/7) presented inconsistencies regarding the concentration of analytes declared on the labels or the antibiotics found. Likewise 74.6% (n = 50/67) of the non-medicated feed samples analyzed had antibiotic residues. Additionally, we analyzed commercial monensin standards for purity and evaluate batch-to-batch flushing feed industry practices. Conclusions Herein we report the results for a year-wide analysis for mycotoxins and antibiotics in feed samples. Mycotoxins, several metabolites, and the occurrence of these emerging contaminants were evaluated and antibiotic residues in non-medicated feed samples were found using a targeted MS-based LC approach. This validated multi-analyte method is expected to facilitate the monitoring and surveillance of contaminants, from natural and anthropogenic origin, in animal feed.

Resumen Objetivo Evaluar el nivel de implementacion de los protocolos asociados a la prevencion de la debilidad muscular adquirida en la unidad de cuidados intensivos (UCI), asi como la presencia del fisioterapeuta en distintas UCI de Espana. Metodo Estudio descriptivo, transversal realizado en 86 UCI de adultos de Espana entre los meses de marzo a junio de 2017. Se excluyeron UCI neuroquirurgicas y de grandes quemados. Se utilizo encuesta multirrespuesta que incluia preguntas sobre los protocolos de: control de glucemia, sedacion, valoracion del dolor, prevencion del delirium, manejo del delirium y movilizacion precoz. La encuesta fue rellenada a traves de un aplicativo protegido por usuario y contrasena. Analisis inferencial con t de Student o U de Mann-Whitney y de correlacion con Pearson o Rho de Spearman. Resultados El 89,5% de las UCI tenian protocolo de control de glucemia, con rango predominante de 110-140 mg/dl. El 74,4% evaluaban el nivel de sedacion, si bien solo tenian protocolos de sedacion el 36% de ellas. Con relacion a la valoracion del dolor se realizaba en el 73,7% de los pacientes comunicativos, mientras que en los no comunicativos solo era del 47,5%. Solo el 37,2% realizaban screening diario para detectar el delirium, y disponian de protocolos de prevencion del delirium el 31,4% de las UCI, del manejo del delirium el 26,7% y de movilizacion precoz el 14% de las UCI. En el 34,9% de los casos se solicita interconsulta al servicio de rehabilitacion. Conclusiones La implementacion de los diferentes protocolos asociados a la prevencion de la debilidad muscular adquirida ha sido elevada en relacion con los protocolos de control de glucemia, valoracion del nivel de sedacion y del dolor de pacientes comunicativos, mientras que baja en los de movilizacion precoz y screening y prevencion del delirio. Asimismo, es poco frecuente la presencia del fisioterapeuta en la UCI.

This study was funded by grants from the Ministerio de Economia, Industria y Competitividad y por el Fondo de Desarrollo Regional FEDER, Spain nº SAF2013-49019, SAF2017-85903-P, and from the Consejería de Innovación, Ciencia y Empresa, Junta de Andalucía (P07- CTS- 03135, P10- CTS- 5784, and CTS- 101), Spain. J.F. and L.M. have FPU fellowships from the Ministerio de Educación Cultura y Deporte, Spain. C.R.S. was a schorlarship holder from the Plan Propio de Investigación of the University of Granada., We wish to thank Michael O’Shea for proofreading the paper., Metabolic reprogramming, which is characteristic of cancer cells that rapidly adapt to the hypoxic microenvironment and is crucial for tumor growth and metastasis, is recognized as one of the major mechanisms underlying therapeutic resistance. Mitochondria, which are directly involved in metabolic reprogramming, are used to design novel mitochondria-targeted anticancer agents. Despite being targeted by melatonin, the functional role of mitochondria in melatonin's oncostatic activity remains unclear. In this study, we aim to investigate the role of melatonin in mitochondrial metabolism and its functional consequences in head and neck cancer. We analyzed the effects of melatonin on head and neck squamous cell carcinoma (HNSCC) cell lines (Cal-27 and SCC-9), which were treated with 100, 500, and 1500 mu M of melatonin for 1, 3, and 5 days, and found a connection between a change of metabolism following melatonin treatment and its effects on mitochondria. Our results demonstrate that melatonin induces a shift to an aerobic mitochondrial metabolism that is associated with changes in mitochondrial morphology, function, fusion, and fission in HNSCC. We found that melatonin increases oxidative phosphorylation (OXPHOS) and inhibits glycolysis in HNSCC, resulting in increased ROS production, apoptosis, and mitophagy, and decreased cell proliferation. Our findings highlight new molecular pathways involved in melatonin's oncostatic activity, suggesting that it could act as an adjuvant agent in a potential therapy for cancer patients. We also found that high doses of melatonin, such as those used in this study for its cytotoxic impact on HNSCC cells, might lead to additional effects through melatonin receptors., Ministerio de Economia, Industria y Competitividad y por el Fondo de Desarrollo Regional FEDER, Spain SAF2013-49019 SAF2017-85903-P, Junta de Andalucia P07-CTS-03135 P10-CTS-5784 CTS-101, Ministerio de Educacion Cultura y Deporte, Spain, Plan Propio de Investigacion of the University of Granada

Muscular aging is a complex process and underlying physiological mechanisms are not fully clear. In recent years, the participation of the NF-kB pathway and the NLRP3 inflammasome in the chronic inflammation process that accompanies the skeletal muscle’s aging has been confirmed. microRNAs (miRs) form part of a gene regulatory machinery, and they control numerous biological processes including inflammatory pathways. In this work, we studied the expression of four miRs, three of them are considered as inflammatory-related miRs (miR-21, miR-146a, and miR-223), and miR-483, which is related to the regulation of melatonin synthesis, among other targets. To investigate the changes of miRs expression in muscle along aging, the impact of inflammation, and the role of melatonin in aged skeletal muscle, we used the gastrocnemius muscle of wild type (WT) and NLRP3-knockout (NLRP3−) mice of 3, 12, and 24 months-old, with and without melatonin supplementation. The expression of miRs and pro-caspase-1, caspase-3, pro-IL-1β, bax, bcl-2, and p53, was investigated by qRT-PCR analysis. Histological examination of the gastrocnemius muscle was also done. The results showed that age increased the expression of miR-21 (p &lt, 0.01), miR-146a, and miR-223 (p &lt, 0.05, for both miRs) in WT mice, whereas the 24-months-old mutant mice revealed decline of miR-21 and miR-223 (p &lt, 0.05), compared to WT age. The lack of NLRP3 inflammasome also improved the skeletal muscle fibers arrangement and reduced the collagen deposits compared with WT muscle during aging. For the first time, we showed that melatonin significantly reduced the expression of miR-21, miR-146a, and miR-223 (p &lt, 0.05 for all ones, and p &lt, 0.01 for miR-21 at 24 months old) in aged WT mice, increased miR-223 in NLRP3− mice (p &lt, 0.05), and induced miR-483 expression in both mice strains, this increase being significant at 24 months of age.

Two Gram-positive, anaerobic bacteria, designated 27733 and 27737, were isolated from a soft tissue infection from a human patient. They were preliminarily identified asClostridium perfringensthrough a series of phenotypic tests, including Gram-staining, determination of lipase and hemolytic activities, MALDI-ToF profiling, and a commercial biochemical identification system. In line with these results, genomes obtained for both isolates were ~3.56 Mbp in size, showed a DNA G+C content of ~28.4%, and containedC. perfringensribosomal markers (i.e. 16S rRNA gene identity >99.0% toC. perfringensATCC13124T). A closer examination of these sequences; however, revealed low average Nucleotide Identity (~87%) and digital DNA-DNA Hybridization (~35%) values between isolates 27733/27737 andC. perfringensATCC13124T, as well as substantial differences in gene content to multipleC. perfringensstrains, indicating that they represent a novel species within the genusClostridium.Congruently, Bayesian dating analyses placed the divergence of this new species andC. perfringensfrom its common ancestor hundreds of thousands of years ago. Isolates 27733/27737 are not genomically identical (34-197 SNPs apart) and carry genes forC. perfringens-liketoxins (plc(alpha toxin),pfoA(perfringolysin O, theta-toxin),nagHIJKL(hyalorudinase, mu-toxin),nanHIJ(exo-alpha sialidase), andcloSI(alpha-clostripain). They do not have known antibiotic resistance genes but were catalogued as resistant to clindamycin through phenotypic tests. On the basis of the presented evidence, and due to its resemblance and potential confusion withC. perfringens,we herein propose the speciesC. perfringenosumsp. nov. and strain 27733 as its type strain.

Introduction. The coffee berry borer (Hypothenemus hampei Ferrari, CBB) is one of the most devastating pests on coffee plantations around the world. Although CBB is susceptible to the effect of δ–endotoxins of Bacillus thuringiensis subs. israelensis (Bti) at laboratory level, the efficacy of this control method is poor in the field, presumably due to the inactivation by digestive proteases different to those required for protoxin activation. Objective. To study whether the addition of a soybean flour extract enriched with protease inhibitors (PI), mixed with Bti crystals and spores (Bti-sc) in an artificial diet, could improve the toxicity of Bti against CBB. Materials and methods. This study was performed in San José, Costa Rica, between 2012 and 2013. A set of adult female CBB insects was exposed to a mixture containing different concentrations of a partially purified soybean meal extract with active PI and lyophilized Bti-sc, and were tested through a bioassay in artificial diet to estimate the sub-lethal concentration (LC50). The mortality results were validated by observing the dissected midgut, whose ultrastructure was analyzed by transmission electron microscopy. Results. The soybean extracts partially degraded the Bti-sc complex, it reduced its LC50 by almost four times (from 1.135 to 0.315 µg µl-1) and enhanced CBB mortality in a concentration-dependent manner. Histological analyses of the midgut confirmed this synergistic effect, since severe epithelial damage to the intestinal epithelium of CBB exposed to Bti-sc + PI was visualized compared to Bti-sc alone. Conclusions. The combination of a soybean extract enriched in PI and Bti-sc enhanced the mortality effect over CBB, which was confirmed by the midgut collapse. Soybean flour is a cost-effective supplement that could increase Bti effectiveness against CBB and delay the appearance of biological resistance. Resumen Introducción. La broca del café (Hypothenemus hampei, CBB) es una de las plagas más devastadoras en plantaciones de café alrededor del mundo. Aunque CBB es susceptible al efecto de las δ-endotoxinas del Bacillus thuringiensis subs. israelensis (Bti) a nivel de laboratorio, la eficacia de este método de control es deficiente en campo, posiblemente debido a la inactivación ocasionada por proteasas digestivas diferentes a las requeridas para la activación de las protoxinas. Objetivo. Determinar si la incorporación de un extracto de harina de soya con inhibidores de proteasas (PI) mezclado con cristales y esporas de Bti (Bti-sc) en una dieta artifical, podría mejorar la toxicidad de Bti contra CBB. Materiales y métodos. Este estudio se realizó en San José, Costa Rica entre 2012 y 2013. Se expuso un conjunto de insectos hembras adultas de CBB a una mezcla que incluyó diferentes concentraciones del extracto de soya parcialmente purificado con PI activo y un liofilizado de Bti-sc, y fueron evaluadas mediante un bioensayo en dieta artificial para estimar la concentración subletal (CL50). Los resultados de mortalidad se validaron mediante observación del intestino medio diseccionado, cuya ultraestructura se analizó mediante microscopía electrónica de transmisión. Resultados. El extracto de soya degradó parcialmente el complejo Bti-sc, redujo la CL50 en casi cuatro veces (de 1,135 a 0,315 µg µl-1) y potenció la mortalidad de CBB de manera concentración-dependiente. Los análisis histológicos del intestino medio confirmaron este efecto sinérgico, dado que se visualizaron daños severos en el epitelio intestinal de CBB expuestos a Bti-sc + PI comparado con Bti-sc solo. Conclusiones. La combinación del extracto de soya enriquecido con PI y Bti-sc potenció la letalidad sobre CBB, que se confirmó por el colapso intestinal. La harina de soya es un suplemento económico que podría aumentar la efectividad de Bti para controlar CBB y retrasar el desarrollo de resistencia biológica.

Here, we present the characterization of three iPSC lines derived from dermalfibroblasts of old healthy subjects. Fibroblasts were reprogrammed using Sendai viralvectors encoding OCT4, SOX2, KLF4 and c-MYC. The iPSCs expressed endogenous pluripotency markers, could generate the three germ layers (ectoderm, mesodermand endoderm), maintained a stable karyotype, and were free from Sendai vectors and reprogramming factors. These integration-free iPSCs can serve for establishingcontrol cell cultures in studies searching for phenotypes and mechanisms that could potentially be dysregulated in degenerative diseases., This work was funded by grants from the Spanish Ministerio de Economía y Competitividad (MINECO: SAF2013-47596-R and CIBERNED: CB06/05/0065) to C.V., from CIBERNED CB06/05/0055 to R.M, from CIBERNED CB06/05/0041 to J.K., and from Fundación Ramón Areces (CIVP18A3941) to C.V. and R.M. E.R.-T. was supported by a FPIFellowship from the MINECO.

Cancer is still a major public health burden because its incidence and mortality continue to increase worldwide. The limited treatment options for patients with advanced stages, the severe toxicity, the onset of multiple drug resistance and the high costs of current anticancer therapies favor poor prognosis and high mortality rates. Thus, the identification and development of preventive and cost effective therapeutic strategies to reverse cancer-associated morbidity and mortality are needed. Vegetable and fruit consumption is associated with decreased risk of cancer because of its chemopreventive and chemotherapeutic effects. The use of Punica granatum preparations has a long ethnomedical history and preclinical research has reported many pharmacological activities, including chemopreventive, chemosensitisation and chemotherapeutic effects. Many of these health beneficial effects are related to its complex chemical composition and synergistic interactions of its colonic microbial metabolites including ellagic acid, ellagitannins, punicic acid, flavonoids, anthocyanidins, anthocyanins, and estrogenic flavonols. This chapter summarizes the scientific evidence supporting anticancer effects of pomegranate constituents, focusing on its molecular targets and anticancer mechanisms of action, along with a critical evaluation of pomegranate polyphenols as future anticancer agents.

Hereditary angioedema as a result of deficiency of the C1 inhibitor (HAE-C1INH; MIM# 106100) is a rare autosomal disorder and affected individuals are generally heterozygous for dominant negative variants in theSERPING1gene. Homozygosity forSERPING1pathogenic variants was long considered to be embryonically lethal; however, five nonrelated families with a recessive HAE pattern have been described in the last decade. in this report, we functionally characterized two newly reported nonrelated, consanguineous families with a recessive presentation of HAE attributed toSERPING1variants in the reactive center loop (family D; S438F) and gate (family A; I379T) regions. S438F heterozygotes (family D) showed variable levels of intact 105-kDa and cleaved/inactive 96-kDa isoforms of C1INH, whereas their homozygous relative presented only the 96-kDa band. Functional studies showed that S438F reduced C1INH interaction with target proteases in heterozygous (C1s, 32-38% of controls and FXIIa, 28-35% of controls) and homozygous (C1s, 18-24% of controls and FXIIa, 4-8% of controls) carriers, which is consistent with the more severe presentation of HAE in the family and decreased C1q levels in homozygous patients. By contrast, plasma C1INH from I379T heterozygotes (family A) showed normal C1INH/C1s binding (84-94% of controls) and no significant reduction in C1INH/FXIIa complexes (50-70% of controls). However, the homozygote failed to inhibit both C1s (25-42% of controls) and FXIIa (14-18% of controls). This profile is concordant with the less severe presentation of HAE in the family and the conserved C4 and C1q levels in heterozygous and homozygous patients., [ER19P7AC7541/2019], We thank Dr Alper Ozdemir, Arda Kula, Suat Hopanci and Betul Hopanci for obtaining the patient's serum samples. Alberto Lopez Lera is supported by grant ER19P7AC7541/2019 from Centre for Biomedical Network Research or Rare Diseases (CIBERER).

Indexación: Scopus. Cfr is a radical S-adenosyl-L-methionine (SAM) enzyme that confers cross-resistance to antibiotics targeting the 23S rRNA through hypermethylation of nucleotide A2503. Three cfr-like genes implicated in antibiotic resistance have been described, two of which, cfr(B) and cfr(C), have been sporadically detected in Clostridium difficile. However, the methylase activity of Cfr(C) has not been confirmed. We found cfr(B), cfr(C), and a cfr-like gene that shows only 51 to 58% protein sequence identity to Cfr and Cfr-like enzymes in clinical C. difficile isolates recovered across nearly a decade in Mexico, Honduras, Costa Rica, and Chile. This new resistance gene was termed cfr(E). In agreement with the anticipated function of the cfrlike genes detected, all isolates exhibited high MIC values for several ribosome-targeting antibiotics. In addition, in vitro assays confirmed that Cfr(C) and Cfr(E) methylate Escherichia coli and, to a lesser extent, C. difficile 23S rRNA fragments at the expected positions. The analyzed isolates do not have mutations in 23S rRNA genes or genes encoding the ribosomal proteins L3 and L4 and lack poxtA, optrA, and pleuromutilin resistance genes. Moreover, these cfr-like genes were found in Tn6218-like transposons or integrative and conjugative elements (ICE) that could facilitate their transfer. These results indicate selection of potentially mobile cfr-like genes in C. difficile from Latin America and provide the first assessment of the methylation activity of Cfr(C) and Cfr(E), which belong to a cluster of Cfr-like proteins that does not include the functionally characterized enzymes Cfr, Cfr(B), and Cfr(D). https://journals.asm.org/doi/10.1128/AAC.01074-19

Clostridioides difficile is an emerging One Health pathogen and a common etiologic agent of diarrhea, both in healthcare settings and the community. This bacterial species is highly diverse, and its global population has been classified in eight clades by multilocus sequence typing (MLST). The C. difficile MLST Clade 2 includes the NAP1/RT027/ST01 strain, which is highly recognized due to its epidemicity and association with severe disease presentation and mortality. By contrast, the remaining 83 sequence types (STs) that compose this clade have received much less attention. In response to this shortcoming, we reviewed articles published in English between 1999 and 2020 and collected information for 27 Clade 2 STs, with an emphasis on STs 01, 67, 41 and 188/231/365. Our analysis provides evidence of large phenotypic differences that preclude support of the rather widespread notion that ST01 and Clade 2 strains are “hypervirulent”. Moreover, it revealed a profound lack of (meta)data for nearly 70% of the Clade 2 STs that have been identified in surveillance efforts. Targeted studies aiming to relate wet-lab and bioinformatics results to patient and clinical parameters should be performed to gain a more in-depth insight into the biology of this intriguing group of C. difficile isolates.

Antibiotics are widely used as growth promoters in animal husbandry; among them, the tetracyclines are a chemical group of relevance, due to their wide use in agriculture, surpassing in quantities applied almost every other antibiotic family. Seeing the considerable amounts of tetracyclines used worldwide, monitoring of these antibiotics is paramount. Advances must be made in the analysis of antibiotics to assess correct usage and dosage of tetracyclines in food and feedstuffs and possible residues in pertinent environmental samples. The tetracyclines are still considered a clinically relevant group of antibiotics, though dissemination of tolerance and resistance determinants have limited their use. This review focuses on four different aspects: (i) tetracyclines, usage, dosages, and regulatory issues that govern their food-related application, with particular attention to the prohibitions and restrictions that several countries have enforced in recent years by agencies from both the United States and the European Union, (ii) analytical methods for tetracyclines, determination, and residues thereof in feedstuffs and related matrices with an emphasis on the most relevant and novel techniques, including both screening and confirmatory methods, (iii) tetracycline resistance and tetracycline-resistant bacteria in feedstuff, and (iv) environmental and health risks accompanying the use of tetracyclines in animal nutrition. In the last two cases, we discuss the more relevant undesirable effects that tetracyclines exert over bacterial communities and nontarget species including unwanted effects in farmers. UCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias Agroalimentarias::Centro de Investigación en Nutrición Animal (CINA) UCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias de la Salud::Centro de Investigación en Enfermedades Tropicales (CIET)

Metabolic syndrome (MS) increases cardiovascular risk and is associated with cardiac dysfunction and arrhythmias, although the precise mechanisms are still under study. Chronic inflammation in MS has emerged as a possible cause of adverse cardiac events. Male Wistar rats fed with 30% sucrose in drinking water and standard chow for 25–27 weeks were compared to a control group. The MS group showed increased weight, visceral fat, blood pressure, and serum triglycerides. The most important increases in serum cytokines included IL-1β(7-fold), TNF-α(84%), IL-6 (41%), and leptin (2-fold), the latter also showing increased gene expression in heart tissue (35-fold). Heart function ex vivo in MS group showed a decreased mechanical performance response to isoproterenol challenge (ISO). Importantly, MS hearts under ISO showed nearly twofold the incidence of ventricular fibrillation. Healthy rat cardiomyocytes exposed to MS group serum displayed impaired contractile function and Ca2+handling during ISO treatment, showing slightly decreased cell shortening and Ca2+transient amplitude (23%), slower cytosolic calcium removal (17%), and more frequent spontaneous Ca2+release events (7.5-fold). As spontaneous Ca2+releases provide a substrate for ventricular arrhythmias, our study highlights the possible role of serum proinflammatory mediators in the development of arrhythmic events during MS.

When exposed to hostile environments such as radiation, physical injuries, chemicals, pollution, and microorganisms, the skin requires protective chemical molecules and pathways. Melatonin, a highly conserved ancient molecule, plays a crucial role in the maintenance of skin. As human skin has functional melatonin receptors and also acts as a complete system that is capable of producing and regulating melatonin synthesis, melatonin is a promising candidate for its maintenance and protection. Below, we review the studies of new metabolic pathways involved in the protective functions of melatonin in dermal cells. We also discuss the advantages of the topical use of melatonin for therapeutic purposes and skin protection. In our view, endogenous intracutaneous melatonin production, together with topically-applied exogenous melatonin and its metabolites, represent two of the most potent defense systems against external damage to the skin., Part of this review was co-funded by the Spanish Ministry of the Economy and Competitiveness, the FEDER Regional Development Fund (nos. SAF2013-49019 and SAF2017-85903), the Charles III Institute (no. CB/10/00238), and the Economy, Innovation, Science and Employment Council of the Junta de Andalucía (CTS-101). JF and LM are FPU (Professional University formation) Fellows of the Spanish Ministry of Education, Culture and Sport.

Background Intensive care unit–acquired muscle weakness (ICUAW) has an incidence of 40–46%. Early mobilisation is known to be a protective factor. Objective The aim of the study was to identify the incidence of ICUAW in Spain and to evaluate variables likely to contribute to the development of ICUAW. Methods A 4-month, prospective observational multicentre cohort study was conducted on patients receiving invasive mechanical ventilation for at least 48 h. Data were collected from ICU day 3 until ICU discharge. The primary outcome was presence of ICUAW (diagnosed using the Medical Research Council [MRC] scale). The secondary outcome was nurse–patient ratio, physiotherapist availability, analgesia, sedation and delirium management, glycaemic control, and daily level of mobility during the ICU stay as per the ICU Mobility Scale. A logistic regression model was constructed based exclusively on days 3–5 of the ICU stay. Results The data of 642 patients were analysed from 80 ICUs, accounting for 35% of all ICUs in Spain. The incidence of ICUAW was 58% (275 of 474 patients; 95% confidence interval [CI] [53–62]). The predictors for ICUAW were older age (odds ratio [OR] = 1.01; 95% CI [1.00–1.03]) and more days with renal replacement therapy (OR = 1.01; 95% CI [1.00–1.02]). The protective factors for ICUAW were male gender (OR = 0.58; 95% CI [0.38–0.89]), higher Barthel Index (showing prehospital functional independence) (OR = 0.97; 95% CI [0.95–0.99]), more days of being awake and cooperative (defined by a feasible MRC assessment) (OR = 0.98; 95% CI [0.97–0.99]), presence of delirium (OR = 0.98; 95% CI [0.97–0.99]), and more days with active mobilisation (ICU Mobility Scale ≥ 4) (OR = 0.98; 95% CI [0.97–0.99]). Conclusions The risk factors for ICUAW were functional dependence before admission, female gender, older age, and more days on renal replacement therapy. The protective factors for ICUAW were feasibility of MRC assessment, the presence of delirium, and being actively mobilised during the first 5 days in the ICU.

The L444P mutation in the GBA1 gene which encodes β-glucocerebrosidase-1, is a major risk factor for developing Parkinson's disease (PD) and dementia with Lewy bodies (DLB). We report the generation and characterization of an induced pluripotent stem cell (iPSC) line derived from a female PD patient carrying the L444P/wt mutation. The iPSC line presented a normal morphology, expressed endogenous pluripotency markers, could be differentiated into endodermal, mesodermal and ectodermal cells, was free from Sendai vectors and reprogramming factors, had a normal karyotype and maintained the original GBA1 genotype. Thus, this iPSC line can serve to establish cellular models of PD., We thank Drs. Marta Martínez and Miquel Vila for sharing with us PD5 fibroblasts. We also thank Mª José Román for technical assistance. This work was funded by grants from the Spanish Ministerio de Economía y Competitividad (MINECO: SAF2013-47596-R and CIBERNED CB06/05/0065) to C.V., from CIBERNED CB06/05/0055 to R.M, from CIBERNED CB06/05/0041 to J.K., and from Fundación Ramón Areces (CIVP18A3941) to C.V. and R.M. E.R.-T. was supported by a FPI Fellowship from the MINECO.

Head and neck cancer is the sixth leading cancer by incidence worldwide. Unfortunately, drug resistance and relapse are the principal limitations of clinical oncology for many patients, and the failure of conventional treatments is an extremely demoralizing experience. It is therefore crucial to find new therapeutic targets and drugs to enhance the cytotoxic effects of conventional treatments without potentiating or offsetting the adverse effects. Melatonin has oncostatic effects, although the mechanisms involved and doses required remain unclear. The purpose of this study is to determine the precise underlying mitochondrial mechanisms of melatonin, which increase the cytotoxicity of oncological treatments, and also to propose new melatonin treatments in order to alleviate and reverse radio- and chemoresistant processes. We analyzed the effects of melatonin on head and neck squamous cell carcinoma (HNSCC) cell lines (Cal-27 and SCC-9), which were treated with 0.1, 0.5, 1, and 1.5mM melatonin combined with 8 Gy irradiation or 10 μM cisplatin. Clonogenic and MTT assays, as well as autophagy and apoptosis, involving flow cytometry and western blot, were performed in order to determine the cytotoxic effects of the treatments. Mitochondrial function was evaluated by measuring mitochondrial respiration, mtDNA content (RT-PCR), and mitochondrial mass (NAO). ROS production, antioxidant enzyme activity, and GSH/GSSG levels were analyzed using a fluorometric method. We show that high concentrations of melatonin potentiate the cytotoxic effects of radiotherapy and CDDP in HNSCC, which are associated with increased mitochondrial function in these cells. In HNSCC, melatonin induces intracellular ROS, whose accumulation plays an upstream role in mitochondria-mediated apoptosis and autophagy. Our findings indicate that melatonin, at high concentrations, combined with cisplatin and radiotherapy to improve its effectiveness, is a potential adjuvant agent., This study was partially supported by grants from the Ministerio de Economía y Competitividad, Spain, and the FEDER Regional Development Fund (nos. SAF2013-49019 and SAF2017-85903), from the Instituto de Salud Carlos III (no. CB/10/00238), and from the Consejería de Economía, Innovación, Ciencia y Empleo, Junta de Andalucía (CTS-101).

Mutations in the GBA1 gene, which encodes the lysosomal enzyme Glucocerebrosidase1 are major risk factors for Parkinson's disease (PD) and dementia with Lewy bodies (DLB). We have generated induced pluripotent stem cells (iPSCs) from fibroblasts of four PD patients carrying the N370S/wt or the L444P/wt heterozygous mutations in GBA1. The iPSCs presented a normal morphology, expressed endogenous pluripotency markers and could be differentiated into endodermal, mesodermal and ectodermal cells. These iPSCs were free from Sendai viral vectors and reprogramming factors, had a normal karyotype and maintained the original GBA1 genotype., We thank Mª José Román for technical assistance. This work was funded by grants from the Spanish Ministerio de Economía y Competitividad (MINECO: SAF2013-47596-R and CIBERNED: CB06/ 05/0065) to C.V., from CIBERNED CB06/05/0055 to R.M, from CIBERNED CB06/05/0041 to J.K., and from Fundación Ramón Areces (CIVP18A3941) to C.V. and R.M. E.R.-T. was supported by a FPI Fellowship from the MINECO.

Music performance anxiety (MPA) is a phenomenon often encountered among professionals and students who make public appearances. This article presents the results of a study carried out on a sample of music students in superior music conservatories in Spain ( N = 434). Our goal was to analyze MPA on the basis of Barlow’s (2000) anxiety theory, supplementing it with further personality constructs such as dispositional optimism, general auto-efficacy, and sensitivity to reward and punishment. Our structural equation modeling (SEM) results reveal that several of those constructs exert their effect via the helplessness factor – the central construct in Barlow’s theory – and that they likewise exert a further series of direct effects on MPA. All in all, the variables taken into consideration account for 45.6% of variance in MPA in males and of 52.1% thereof in females. This study thus upholds Barlow’s theory of anxiety, while broadening it with further explanatory mechanisms.

Attention-deficit-hyperactivity disorder (ADHD) is a common neuro-behavioural disorder with onset in childhood. These children have impaired emotional self-control, self-regulation of drive and motivation. Numerous studies have reported cognitive disabilities in memory, executive functions, spatial abilities and language skills. The main objective of this work is to determine whether a socio-emotional intervention programme could improve executive functions in children with Attention Deficit Hyperactivity Disorder. The sample of this study consisted of 25 children (8 female and 17 male) aged between 8 and 12 years, diagnosed with ADHD and who were not taking any psychopharmacological treatment at the time of the study, and had not taken medication previously. Executive functioning was assessed through the Zoo Map test and Tower of Hanoi puzzle in pre-/post-test design. A socio-emotional intervention programme was implemented. The training consisted of 8 one-hour weekly sessions, on an individual basis. Results indicate that such a programme does lead to improved performance in the execution of tasks that evaluate executive functions. After the intervention, the children took less time to resolve the Zoo Map test. Results for the Hanoi Tower puzzle were also improved after intervention. The children needed a lower number of movements to complete the task.

Background: Hospital transmission of Clostridium difficile is fostered by contamination of surfaces and medical equipment with spores highly resistant to disinfectants and regular cleaning procedures. Despite the outbreaks and fatalities that C difficile causes, its epidemiology has not been studied in hospitals from middle- and low-income countries. To tackle this knowledge gap, the detection frequency of C difficile DNA on inert surfaces of a major Costa Rican hospital during and after an outbreak was compared. Methods: We used a presence-absence real-time polymerase chain reaction to detect a fragment of the tpi gene of C difficile on 21 surface samples collected during an outbreak and 54 surface samples taken 2 years later at the same hospital. Results: C difficile DNA was detected in 40% of the 75 environmental samples analyzed. Whereas 71% of the samples collected during the outbreak were positive, only 28% of the samples obtained 2 years after the outbreak gave the same result. This 2.5× ratio was maintained when the comparison was restricted to the wards that were sampled both during and after the outbreak (72% vs 35%, P = .016). Conclusions: Our results show that environmental surfaces in the hospital analyzed are continuously being contaminated with C difficile DNA and that their level of contamination is higher during an outbreak than after it UCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias de la Salud::Centro de Investigación en Enfermedades Tropicales (CIET)

In recent years, reports of NAP1/RT027/ST01 epidemic strains of Clostridium difficile producing outbreaks of healthcare-associated diarrhea have increased in America and Europe. We cultivated multidrug-resistant NAP1/RT027/ST01 strains from the FQR2 linage from TcdA/TcdB+ stool samples obtained from patients in two Honduran hospitals. The PFGE macrorestriction patterns of two of the isolates were new. These bacteria were toxigenic and induced with different magnitude classical cytopathic effects on HeLa cells. Besides their resistance to twelve antibiotics, including to clindamycin, fluoroquinolones, linezolid and tigecycline. In this regard, they show the gyrA mutation that typifies epidemic C. difficile genotypes and carry cfr-like genes in different molecular contexts, respectively. These results confirm the spread of multidrug-resistant NAP1/RT027/ST01 strains in Central America with potential idiosyncratic adaptations.

The population structure of Clostridium difficile currently comprises eight major genomic clades. For the highly divergent C-I clade, only two toxigenic strains have been reported, which lack the tcdA and tcdC genes and carry a complete locus for the binary toxin (CDT) next to an atypical TcdB monotoxin pathogenicity locus (PaLoc). As part of a routine surveillance of C. difficile in stool samples from diarrheic human patients, we discovered three isolates that consistently gave negative results in a PCR-based screening for tcdC. Through phenotypic assays, whole-genome sequencing, experiments in cell cultures, and infection biomodels we show that these three isolates (i) escape common laboratory diagnostic procedures, (ii) represent new ribotypes, PFGE-types, and sequence types within the Clade C-I, (iii) carry chromosomal or plasmidal TcdBs that induce classical or variant cytopathic effects (CPE), and (iv) cause different levels of cytotoxicity and hamster mortality rates. These results show that new strains of C. difficile can be detected by more refined techniques and raise questions on the origin, evolution, and distribution of the toxin loci of C. difficile and the mechanisms by which this emerging pathogen causes disease. Federal State of Lower Saxony/[(VWZN2889/3215/3266]/FSLS/Alemania German Center for Infection Research/[BMBF grant 8000-105-3]/DZIF/Alemania Universidad de Costa Rica/UCR/Costa Rica UCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias de la Salud::Centro de Investigación en Enfermedades Tropicales (CIET) UCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias de la Salud::Laboratorio de Ensayos Biológicos (LEBI)

Attention-deficit-hyperactivity disorder (ADHD) is a common neurobehavioral disorder with onset in childhood. Children with ADHD have been found to show impairment in emotional self-control, self-regulation and motivation. Numerous studies have reported cognitive disabilities in memory, executive functions, spatial abilities and language skills. The main objective of this research was to determine if the executive function intervention programme implemented could improve attention process and executive functions in children with Attention Deficit Hyperactivity Disorder. The sample consisted of 50 children (14 female and 36 male) aged between 8 and 12 years, diagnosed with ADHD and who were not under any psychopharmacological treatment at the time of the study, and had not taken medication to mitigate such disorder. Executive functioning was assessed through the Zoo Map Test and attention was evaluated by the “D2 Test of Attention” in a pretest-postest cuasi experimental design. The executive functioning intervention programme implemented was designed and tested for this research and consisted of 12 one-hour weekly sessions and the participants were trained individually. The data indicated that the intervention lead to improved performance in the execution of tasks that evaluate executive functions. After the intervention, the children took less time to resolve the Zoo Map Test, and the results in the “D2 Test of Attention” also improved after the training. Therefore, data showed that such executive functioning intervention programme could improve attention processes and executive functions in children with Attention Deficit Hyperactivity Disorder. Therefore, the programme used becomes effective intervention as well as the data found links the nature of the problem to a specific treatment management by objectives.

This study aimed to assess the molecular basis of the resistance to carbapenems in clinical isolates of Pseudomonas aeruginosa recovered from a tertiary-level health facility in San José, Costa Rica. A total of 198 non-duplicated isolates were evaluated for their susceptibility to β-lactams, aminoglycosides and fluoroquinolones. The production of metallo-β-lactamases (MBLs), the presence of MBL encoding genes (blaIMP, blaVIM and blaGIM-1) and the occurrence of these genes within class 1 integrons were investigated. In addition, an ERIC2 PCR fingerprinting method was used to elucidate the distribution of the detected MBL genes within the strain collection. Of the 198 isolates tested, 125 (63.1 %) were categorized as carbapenem-resistant. The majority (88.8 %) of the carbapemen-resistant isolates also showed resistance to ceftazidime, cefepime, aztreonam, ticarcillin/clavulanic acid, amikacin, gentamicin, tobramycin, ciprofloxacin and gatifloxacin. Among the carbapenem-resistant isolates, 102 (81.6 %) showed MBL activity. Strikingly, both blaIMP and blaVIM genes were simultaneously detected in most (94.1 %) of the 102 MBL producers. Five carbapenem-resistant MBL producers were positive only for blaIMP genes. Almost 70 % of the isolates examined harboured the intI1 gene, accompanied by the sul1 and qacEΔ1 genes in 136 (99 %) and 122 (89 %) isolates, respectively. The majority (94.4 %) of the carbapenem-resistant isolates carried the intI1 gene, in contrast to 26 % of the carbapenem-susceptible isolates. Ninety-three out of 96 (96.9 %) isolates carrying both blaIMP and blaVIM genes also harboured the intI1, sul1 and qacEΔ1 genes. Gene cassettes from carbapenem-susceptible and MBL-negative carbapenem-resistant isolates encoded aminoglycoside-resistance enzymes (aadA2, aadA4 and aadA6) as well as orfD and qacF genes. RAPD analysis distributed 126 of the isolates in 29 clusters. Eighty of the 90 blaIMP (+) blaVIM (+) isolates were sorted into 16 different clusters, suggesting that the blaIMP and blaVIM genes detected were located within a genetic element capable of lateral transfer. Carbapenem-resistant MBL-positive isolates were recovered from almost all hospital wards and were over-represented in samples obtained from the surgical emergency and intensive care therapy units. Remarkably, three carbapenem-resistant isolates, exhibiting MBL activity and carrying both blaIMP and blaVIM genes, were recovered from outpatients. Sequence analysis of both bla genes in various isolates revealed that they correspond to the alleles blaIMP-18 and blaVIM-2. To our knowledge, this is the first report of the combination of two metallo-β-lactamases encoded by the blaIMP-18 and blaVIM-2 genes in P. aeruginosa. Deutscher Akademischer Austauschdienst/[]/DAAD/Alemania UCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias de la Salud::Centro de Investigación en Enfermedades Tropicales (CIET)

Feed is a common vehicle for tetracycline administration in animal farming. However, veterinarians and farmers often overlook that common feed components, such as proteins and calcium, may interact with these drugs and thereby alter their properties. In this study, we aimed to quantify the effect of crude protein and calcium on the bioavailability of 14 first-, second-, and third-generation tetracyclines, tetracycline epimers that may arise during storage, and other tetracycline derivatives in 84 feedstuffs for shrimp, swine, tilapia, and poultry marketed in Costa Rica. To this end, crude protein was quantified with a Kjeldahl method, calcium was determined by flame atomic spectroscopy (FAAS), and tetracyclines were detected by high-performance liquid chromatography (HPLC) with fluorescence detection (FLD) and with two different whole-cell biosensors to discriminate matrix-bound, inactive tetracyclines from free, active fractions. Tilapia feed contained the highest amounts of HPLC-detectable tetracyclines (119–8365 mg kg−1). Poultry (78–438 mg kg−1) and swine feed (41–1076 mg kg−1) were characterized by intermediate concentrations, while shrimp feed showed the lowest amounts of these drugs (21–50 mg kg−1). Ten samples contained unauthorized antibiotics, including doxycycline, and another three had 4-epimers of tetracyclines, indicating degradation during prolonged storage. Biological detection of tetracyclines was inversely related to both feed crude protein (r2 = 0.58–0.75) and calcium content (r2 = 0.54–0.63). Moreover, only 12–55% of the concentrations determined by HPLC were appraised by the biosensors. Shrimp feed, followed by tilapia feed, showed the highest crude protein (14.5–52.2g 100 g−1) and calcium (0.89–5.90 mg kg−1) concentrations and, as predicted, the lowest bioavailable fractions of tetracyclines (12–40%). Finally, we provide equations that predict bioavailability as a function of crude protein, calcium, and tetracycline concentrations in feed. Our data reveals medication abnormalities in commercial feeds and provide novel biological information for several tetracyclines. UCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias de la Salud::Centro de Investigación en Enfermedades Tropicales (CIET) UCR::Vicerrectoría de Docencia::Salud::Facultad de Microbiología

41 p.-11 fig., C3 is the central component of the complement system. Upon activation, C3 sequentially generates various proteolytic fragments, C3a, C3b, iC3b, C3dg, each of them exposing novel surfaces, which are sites of interaction with other proteins. C3 and its fragments are therapeutic targets and markers of complement activation. We report the structural and functional characterization of four monoclonal antibodies (mAbs) generated by immunizing C3-deficient mice with a mixture of human C3b, iC3b and C3dg fragments, and discuss their potential applications. This collection includes three mAbs interacting with native C3 and inhibiting AP complement activation; two of them by blocking the cleavage of C3 by the AP C3-converase and one by impeding formation of the AP C3-convertase. The interaction sites of these mAbs in the target molecules were determined by resolving the structures of Fab fragments bound to C3b and/or iC3b using electron microscopy. A fourth mAb specifically recognizes the iC3b, C3dg, and C3d fragments. It binds to an evolutionary-conserved neoepitope generated after C3b cleavage by FI, detecting iC3b/C3dg deposition over opsonized surfaces by flow cytometry and immunohistochemistry in human and other species. Because well-characterized anti-complement mAbs are uncommon, the mAbs reported here may offer interesting therapeutic and diagnostic opportunities., Work in this report has been funded by the Spanish “Ministerio de Economía y Competitividad” (SAF2011-26583 and SAF2015-66287-R to SRdC and SAF2014- 52301-R to OL) and the Seventh Framework Programme European Union Project EURenOmics (305608) to SRdC. In addition, this work has been supported by a grant from the Autonomous Region of Madrid (S2010/BMD-2316) to SRdeC and OL.

The objective of this work was to evaluate the shear bond strength (SBS) and bond failure site of brackets bonded to teeth with fluorosis treated with and without microabrasion before placement of a self-etching primer (SEP) and a color-changing composite and to compare with a control group. This in vitro study included 120 premolars with and without dental fluorosis, divided into six groups: (1) healthy enamel etched for 15 sec and fifth generation primer, (2) healthy enamel, (3) mild fluorosis (MIF) enamel, (4) MIF enamel microabraded, (5) moderate fluorosis (MOF) enamel, and (6) MOF enamel microabraded. For groups 2–6, an SEP was used. All samples were evaluated with the SBS test and the modified adhesive remnant index (ARI). The highest SBS mean value was for healthy enamel etched for 15 sec and fifth generation primer and the lowest SBS mean value was for healthy enamel with SEP. Significant differences were found in the ARI between healthy and MOF groups. SBS of orthodontic brackets bonded on fluoro...

The familial form of Alzheimer's disease (FAD), which is caused by mutations in PRESENILIN 1 (PSEN1) and amyloid precursor protein (APP) genes, represents less than 5% of all AD cases and has an early-onset. We report the generation and characterization of an iPSC line derived from a FAD patient carrying the PSEN1-G206D mutation. The iPSC line maintained the original genotype, a normal karyotype, was free from Sendai viral vectors and reprogramming factors (OCT4, SOX2, KLF4 and c-MYC), presented a typical morphology, expressed endogenous pluripotency markers, and could be differentiated into ectodermal, mesodermal and endodermal cells, confirming its pluripotency., We thank Mª José Román for technical assistance. This work was funded by grants from the Spanish Ministerio de Economía y Competitividad (MINECO: SAF2016-80419-R and CIBERNED: CB06/05/0065, PI2013/01 and PI2015-2/02-4) to C.V.

Glioblastoma multiforme is the most frequent primary brain tumor, it has poor prognosis, and it remains refractory to current treatment. The success of temozolomide (TMZ) appears to be limited by the occurrence of chemoresistance. Recently, we report the use of pertussis toxin as adjuvant immunotherapy in a C6 glioma model; showing a decrease in tumoral size, it induced selective cell death in Treg cells, and it elicited less infiltration of tumoral macrophages. Here, we evaluated the cytotoxic effect of pertussis toxin in combination with TMZ for glioma treatment, both in vitro and in vivo RG2 glioma model.We determined cell viability, cell cycle, apoptosis, and autophagy on treated RG2 cells through flow cytometry, immunofluorescence, and Western blot assays. Twenty-eight rats were divided in four groups (n = 7) for each treatment. After intracranial implantation of RG2 cells, animals were treated with TMZ (10 mg/Kg/200 μl of apple juice), PTx (2 μg/200 μl of saline solution), and TMZ + PTx. Animals without treatment were considered as control.We found an induction of apoptosis in around 20 % of RG2 cells, in both single treatments and in their combination. Also, we determined the presence of autophagy vesicles, without any modifications in the cell cycle in the TMZ - PTx-treated groups. The survival analyses showed an increase due to individual treatments; while in the group treated with the combination TMZ - PTx, this effect was enhanced.We show that the concomitant use of pertussis toxin plus TMZ could represent an advantage to improve the glioma treatment.

The frequency of finding of clindamycin-resistant anaerobic bacteria in clinical samples has doubled from 2008 to 2010 in Costa Rica. To determine whether this increase is due to dissemination of erm genes aided by tetQ elements, we analyzed 100 isolates of Bacteroides or Parabacteroides from a regional hospital, a national hospital, and the community. Antimicrobial susceptibilities were recorded with a broth micro-dilution method and erm genes were detected by PCR and Southern blotting. In addition, plasmid isolation and mating experiments were performed to clarify the location and mobility of the detected erm genes. Resistance to clindamycin was by far more frequent in the regional hospital (72%) than in the national hospital (29%) and the community (26%). Resistance to tetracycline was even more common, with the community (85%) outweighing the hospitals (71–72%). While MIC of clindamycin were higher in the hospitals than in the community (P < 0.05), the opposite was seen for tetracycline (P < 0.0001). Of the sought-after genes, only ermG (n = 2), ermA (n = 1), and ermF (n = 1) were detected in the hospitals and ermF in the community (n = 2). In opposition to the low frequency of finding of erm genes, 71% of the isolates were positive for tetQ. None of the detected genes were encoded on plasmids. Only three isolates from the hospitals transferred their erm genes laterally. By contrast, 13 hospital isolates and two community isolates transferred tetQ. Despite the widespread finding of tetracycline-resistant tetQ-positive bacteria, mobile erm genes were rare in our bacterial collection. We conclude that the detected erm genes are likely not included in typical conjugative transposons of Bacteroides and Parabacteroides. Universidad de Costa Rica/[ED-2039]/UCR/Costa Rica Universidad de Costa Rica/[803-B1-142]/UCR/Costa Rica UCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias de la Salud::Centro de Investigación en Enfermedades Tropicales (CIET) UCR::Vicerrectoría de Docencia::Salud::Facultad de Microbiología

Aquaculture farmers commonly add tetracycline to fish feed or to their ponds to prevent or treat bacterial infections in their crops. To assess the short-term effect of tetracycline (TET) and of one of its reversible epimers, 4-epitetracycline (ETC), on the function and structure of a sediment microbial community from a tropical tilapia farm, we contrasted community-level physiological profiles (CLPP) and phospholipid fatty acid profiles (PLFA) obtained from microcosms exposed for 12 days to 5, 10, 50, or 75 mg kg(-1)of these antibiotics. Notwithstanding that the concentration of the antibiotics during the experiment decreased between 13-100% (TET) or 16-61% (ETC), both compounds provoked opposing metabolic responses that did not revert. TET displayed a tendency to inhibit respiration at concentrations50 mg kg(-1), whereas ETC showed the opposite effect. As revealed by the finding of the fatty acids 11:0 iso 3OH, 16:1w6c, and 18:1w6c, the sediment analyzed was predominantly colonized by Gram-negative bacteria. A marked decrease in fatty acid diversity accompanied the aforementioned metabolic responses, with TET concentrations50 mg kg(-1)leading to an enrichment of yeast and fungal biomarkers and both antibiotics at concentrations10 mg kg(-1)selecting for microorganisms with 11:0 iso 3OH. In agreement with CLPP data, differences between the PLFA profiles of control and treated microcosms were more pronounced for TET than for ETC. We conclude that high, yet field-relevant, concentrations of TET and ETC have the potential to modify the composition, and to a lesser extent, the functioning of a sediment microbial community. This study highlights the importance of considering antibiotic degradation products in ecotoxicological research.

Attention-deficit-hyperactivity disorder (ADHD) is a common neuro-behavioural disorder with onset in childhood. These children have impaired emotional self-control, self-regulation of drive and motivation. Numerous studies have reported cognitive disabilities in memory, executive functions, spatial abilities and language skills. The main objective of this work is to determine whether a socio-emotional intervention programme could improve executive functions in children with Attention Deficit Hyperactivity Disorder. The sample of this study consisted of 25 children (8 female and 17 male) aged between 8 and 12 years, diagnosed with ADHD and who were not taking any psychopharmacological treatment at the time of the study, and had not taken medication previously. Executive functioning was assessed through the Zoo Map test and Tower of Hanoi puzzle in pre-/post-test design. A socio-emotional intervention programme was implemented. The training consisted of 8 one-hour weekly sessions, on an individual basis. Results indicate that such a programme does lead to improved performance in the execution of tasks that evaluate executive functions. After the intervention, the children took less time to resolve the Zoo Map test. Results for the Hanoi Tower puzzle were also improved after intervention. The children needed a lower number of movements to complete the task.

The objective of this study was to determine the prevalence of hereditary ataxias in Cuba, with a special focus on the clinical and molecular features of SCA2. Clinical assessments were performed by neurological examinations and application of the SARA scale. Molecular analyses of genes SCA1–3, SCA6, SCA17 and DRPLA identified 753 patients with SCA and 7173 asymptomatic relatives, belonging to 200 unrelated families. 86.79% of all SCA patients were affected with SCA2. In the Holguin province, the average population prevalence of SCA2 is 40.18 × 105 inhabitants, with the remarkable figure of 141.66 × 105 in the Baguanos municipality. The high prevalence of the SCA2 mutation in Holguin reflects most likely a founder effect. The stabilization of the prevalence along time suggests the existence of premutated chromosomes with pure CAG, acting as reservoir for further expansions. CAG repeat length correlated inversely with age at onset, accounting for 80% of the variability. Genetic anticipation was observed in the 80% of transmissions. Repeat instability was greater in paternal transmissions whereas CAG expansions without anticipation was observed in 10.97% suggesting the effect of CAA interruptions in the CAG segment, which decrease the toxicity of the abnormal ataxin-2, and/or other protective factors.