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MONOGRAFÍA: ENSEÑANZA DE LA GRAMÁTICA EN ELE: RESULTADOS DE INVESTIGACIÓN Y NUEVAS PROPUESTAS El cometido del presente artículo es dar a conocer una propuesta didáctica que permita a los estudiantes de E/LE mejorar su competencia comunicativa a partir de la reflexión metalingüística sobre el uso de las construcciones interrogativas en español (en especial, de aquellas solo aparentemente interrogativas) y la comparación con el empleo que hacen de ellas en sus respectivas L1. Concretamente, nuestra propuesta constituye una secuencia didáctica para aprender gramática (Camps, 2003; Camps et al., 2005; Camps, 2010) que aspira a perfeccionar las habilidades expresivas y comprensivas de los estudiantes en diferentes niveles. Las secuencias didácticas constituyen unidades de programación que aportan coherencia a la enseñanza/aprendizaje de la lengua, ya que centran su objetivo en el desarrollo de actividades concebidas y secuenciadas con una finalidad comunicativa, en el trabajo de diferentes aspectos gramaticales. En definitiva, pretendemos ampliar la competencia lingüística del alumnado y afianzar su conocimiento sobre las construcciones interrogativas en español, especialmente en el registro coloquial. ABSTRACT The aim of this article is to present a didactic proposal that allows students of Spanish as foreign language to improve their communicative competence based on the metalinguistic reflection on the use of interrogative constructions in Spanish (especially those only apparently interrogative) and the comparison with the employment they make of them in their respective L1. Specifically, our proposal constitutes a didactic sequence for learning grammar (Camps, 2003; Camps and others, 2005; Camps, 2010) that aims to improve the expressive and comprehensive skills of students at different levels. The didactic sequences constitute programming units that contribute coherence to the teaching/learning of the language, since they focus on the development of activities conceived and sequenced for a communicative purpose, in the work of different grammatical aspects. In short, we intend to expand the linguistic competence of students and strengthen their knowledge about interrogative constructions in Spanish, especially in the colloquial register.

The capture of four specimens of Silvery John Dory (Zenopsis conchifer),a species recorded in the Mediterranean Sea for the first time in 2006, is reported from the Iberian coast (western Mediterranean). One of the specimens was caught near the Strait of Gibraltar and is probably a vagrant. Despite these catches, there is no evidence of a self–sustaining population, so this species should be considered as alien in the Mediterranean.

Background: Helicobacter pylori colonizes the human stomach and may affect the inflammatory response, hormone production related to energy regulation, and gastrointestinal microbiota composition. Previous studies have explored a potential association between H. pylori infection and pediatric obesity with varying results. Considering the immunomodulatory effects of early-life infection with H. pylori that can confer beneficial effects, we hypothesized that we would find an inverse relationship between H. pylori seropositivity and obesity among Danish children and adolescents., Methods: We assessed H. pylori seroprevalence in 713 subjects from an obesity clinic cohort and 990 subjects from a population-based cohort, aged 6 to 19 years, and examined its association with obesity and other cardiometabolic risk factors., Results: No association was found between H. pylori and body mass index standard deviation score (BMI SDS). H. pylori seropositivity was, however, significantly associated with higher fasting plasma glucose levels and the prevalence of hyperglycemia., Conclusion: While we did not find an association between H. pylori seropositivity and BMI SDS, we observed a significant association with higher fasting plasma glucose levels and increased prevalence of hyperglycemia, suggesting that H. pylori infection may contribute to impaired glucose regulation in Danish children and adolescents., (© 2024. The Author(s).)

Autophagy is an important metabolic pathway that can non-selectively recycle cellular material or lead to targeted degradation of protein aggregates or damaged organelles. Autophagosome formation starts with autophagy factors accumulating on lipid vesicles containing ATG9. These phagophores attach to donor membranes, expand via ATG2-mediated lipid transfer, capture cargo, and mature into autophagosomes, ultimately fusing with lysosomes for their degradation. Autophagy can be activated by nutrient stress, for example, by a reduction in the cellular levels of amino acids. In contrast, how autophagy is regulated by low cellular ATP levels via the AMP-activated protein kinase (AMPK), an important therapeutic target, is less clear. Using live-cell imaging and an automated image analysis pipeline, we systematically dissect how nutrient starvation regulates autophagosome biogenesis. We demonstrate that glucose starvation downregulates autophagosome maturation by AMPK-mediated inhibition of phagophore tethering to donor membrane. Our results clarify AMPKs regulatory role in autophagy and highlight its potential as a therapeutic target to reduce autophagy., (© 2024 Barnaba et al.)

Autophagy is a catabolic pathway required for the recycling of cytoplasmic materials. To define the mechanisms underlying autophagy it is critical to quantitatively characterize the dynamic behavior of autophagy factors in living cells. Using a panel of cell lines expressing HaloTagged autophagy factors from their endogenous loci, we analyzed the abundance, single-molecule dynamics, and autophagosome association kinetics of autophagy proteins involved in autophagosome biogenesis. We demonstrate that autophagosome formation is inefficient and ATG2-mediated tethering to donor membranes is a key commitment step in autophagosome formation. Furthermore, our observations support the model that phagophores are initiated by the accumulation of autophagy factors on mobile ATG9 vesicles, and that the ULK1 complex and PI3-kinase form a positive feedback loop required for autophagosome formation. Finally, we demonstrate that the duration of autophagosome biogenesis is ∼110 s. In total, our work provides quantitative insight into autophagosome biogenesis and establishes an experimental framework to analyze autophagy in human cells., (© 2023 Broadbent et al.)

Localization of a variety of RNAs to non-membrane-bound cellular compartments such as nucleoli and Cajal bodies is critical for their stability and function. The molecular mechanisms that underly the recruitment and exclusion of RNAs from these phase-separated organelles is incompletely understood. Telomerase is a ribonucleoprotein composed of the reverse transcriptase protein telomerase reverse transcriptase (TERT), the telomerase RNA (TR), and several auxiliary proteins, including TCAB1. Here we show that in the absence of TCAB1, a large fraction of TR is tightly bound to the nucleolus, while TERT is largely excluded from the nucleolus, reducing telomerase assembly. This suggests that nuclear compartmentalization by the non-membrane-bound nucleolus counteracts telomerase assembly, and TCAB1 is required to retain TR in the nucleoplasm. Our work provides insight into the mechanism and functional consequences of RNA recruitment to organelles formed by phase separation and demonstrates that TCAB1 plays an important role in telomerase assembly., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2023 The Author(s). Published by Elsevier Inc. All rights reserved.)

Under physiological conditions, phosphatidylserine (PS) predominantly localizes to the cytosolic leaflet of the plasma membrane of cells. During apoptosis, PS is exposed on the cell surface and serves as an "eat-me" signal for macrophages to prevent releasing self-immunogenic cellular components from dying cells which could potentially lead to autoimmunity. However, increasing evidence indicates that viable cells can also expose PS on their surface. Interestingly, tumor cell-derived extracellular vesicles (EVs) externalize PS. Recent studies have proposed PS-exposing EVs as a potential biomarker for the early detection of cancer and other diseases. However, there are confounding results regarding subtypes of PS-positive EVs, and knowledge of PS exposure on the EV surface requires further elucidation. In this study, we enriched small EVs (sEVs) and medium/large EVs (m/lEVs) from conditioned media of breast cancer cells (MDA-MB-231, MDA-MB-468) and non-cancerous cells (keratinocytes, fibroblasts). Since several PS-binding molecules are available to date, we compared recombinant proteins of annexin A5 and the carboxylated glutamic acid domain of Protein S (GlaS), also specific for PS, to detect PS-exposing EVs. Firstly, PS externalization in each EV fraction was analyzed using a bead-based EV assay, which combines EV capture using microbeads and analysis of PS-exposing EVs by flow cytometry. The bulk EV assay showed higher PS externalization in m/lEVs derived from MDA-MB-468 cells but not from MDA-MB-231 cells, while higher binding of GlaS was also observed in m/lEVs from fibroblasts. Second, using single EV flow cytometry, PS externalization was also analyzed on individual sEVs and m/lEVs. Significantly higher PS externalization was detected in m/lEVs (annexin A1 + ) derived from cancer cells compared to m/lEVs (annexin A1 + ) from non-cancerous cells. These results emphasize the significance of PS-exposing m/lEVs (annexin A1 + ) as an undervalued EV subtype for early cancer detection and provide a better understanding of PS externalization in disease-associated EV subtypes.

Current methods for characterizing the biodistribution of extracellular vesicles (EVs) are not sensitive enough to track EVs in vivo, despite significant advances over the past decade. Commonly used lipophilic fluorescent dyes are convenient, but lack specificity and yield inaccurate spatiotemporal images in the long-term tracking of EVs. In contrast, protein-based fluorescent or bioluminescent EV reporters have more accurately revealed their distribution in cells and mouse models. Here, we describe a red-shifted bioluminescence resonance energy transfer (BRET) EV reporter, PalmReNL, to analyze the trafficking of small EVs (<200 nm; sEVs) and medium/large EVs (>200 nm; m/lEVs) in mice. Its advantages are that (i) background signals in bioluminescence imaging (BLI) are negligible and (ii) the photons PalmReNL emits have spectral wavelengths longer than 600 nm and can more efficiently penetrate tissues than reporters emitting shorter wavelength light., (© 2023. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Cancer cells produce heterogeneous extracellular vesicles (EVs) as mediators of intercellular communication. This study focuses on a novel method to image EV subtypes and their biodistribution in vivo. A red-shifted bioluminescence resonance energy transfer (BRET) EV reporter is developed, called PalmReNL, which allows for highly sensitive EV tracking in vitro and in vivo. PalmReNL enables the authors to study the common surface molecules across EV subtypes that determine EV organotropism and their functional differences in cancer progression. Regardless of injection routes, whether retro-orbital or intraperitoneal, PalmReNL positive EVs, isolated from murine mammary carcinoma cells, localized to the lungs. The early appearance of metastatic foci in the lungs of mammary tumor-bearing mice following multiple intraperitoneal injections of the medium and large EV (m/lEV)-enriched fraction derived from mammary carcinoma cells is demonstrated. In addition, the results presented here show that tumor cell-derived m/lEVs act on distant tissues through upregulating LC3 expression within the lung., Competing Interests: The authors declare no conflict of interest., (© 2022 The Authors. Advanced Genetics published by Wiley Periodicals LLC.)

The recent pandemic caused by SARS-CoV-2 has now spread worldwide and caused more than 51,000 deaths, by April 2nd 2020. As predicted, there are several obstacles for medical and governmental authorities to efficiently manage this respiratory illness. In spite of appropriated supplies, most hospitals are suffering from a scarcity of free beds, protective masks, sanitizing liquids and even ECMO machines for patients with severe cases. Defeating this pandemic is impossible without united and coordinated international attempts shaped by all countries of the world. We believe that an international scaled-determination is required to diminish the complex impacts of pandemic. The most important priorities are supposed to be i) The development of potential vaccine candidates to provide protection and interrupt the transmission of SARS-CoV-2, ii) To ensure enough supplies for hospitals and their homogeneous distribution among the countries with the worst number of severe cases, iii) There is a need for more studies to identify potential treatments that are effective for the control of this viral infection and iv) It is imperative to provide easy access to diagnostic kits for all countries affected by this pandemic. In the light of these suggestions, it would be recommendable to at least temporarily abandon the political checkouts in both national and international levels; therefore, all partners will be potentially able to efficiently enforce their strategies for the elimination of this unique threat to the human populations., (© 2020 IMSS. Published by Elsevier Inc.)

Background: There is increasing evidence that the intestinal microbiota plays a crucial role in the maturation of the immune system and the prevention of diseases during childhood. Early-life short-course antibiotic use may affect the progression of subsequent disease conditions by changing both host microbiota and immunologic development. Epidemiologic studies provide evidence that early-life antibiotic exposures predispose to inflammatory bowel disease (IBD)., Methods: By using a murine model of dextran sodium sulfate (DSS)-induced colitis, we evaluated the effect on disease outcomes of early-life pulsed antibiotic treatment (PAT) using tylosin, a macrolide and amoxicillin, a beta-lactam. We evaluated microbiota effects at the 16S rRNA gene level, and intestinal T cells by flow cytometry. Antibiotic-perturbed or control microbiota were transferred to pups that then were challenged with DSS., Results: A single PAT course early-in-life exacerbated later DSS-induced colitis by both perturbing the microbial community and altering mucosal immune cell composition. By conventionalizing germ-free mice with either antibiotic-perturbed or control microbiota obtained 40 days after the challenge ended, we showed the transferrable and direct effect of the still-perturbed microbiota on colitis severity in the DSS model., Conclusions: The findings in this experimental model provide evidence that early-life microbiota perturbation may increase risk of colitis later in life.

Early-life antibiotic exposure may provoke long-lasting microbiota perturbation. Since a healthy gut microbiota confers resistance to enteric pathogens, we hypothesized that early-life antibiotic exposure would worsen the effects of a bacterial infection encountered as an adult. To test this hypothesis, C57BL/6 mice received a 5-day course of tylosin (macrolide), amoxicillin (β-lactam), or neither (control) early in life and were challenged with Citrobacter rodentium up to 80 days thereafter. The early-life antibiotic course led to persistent alterations in the intestinal microbiota and even with pathogen challenge 80 days later worsened the subsequent colitis. Compared to exposure to amoxicillin, exposure to tylosin led to greater disease severity and microbiota perturbation. Transferring the antibiotic-perturbed microbiota to germfree animals led to worsened colitis, indicating that the perturbed microbiota was sufficient for the increased disease susceptibility. These experiments highlight the long-term effects of early-life antibiotic exposure on susceptibility to acquired pathogens. IMPORTANCE The gastrointestinal microbiota protects hosts from enteric infections; while antibiotics, by altering the microbiota, may diminish this protection. We show that after early-life exposure to antibiotics host susceptibility to enhanced Citrobacter rodentium -induced colitis is persistent and that this enhanced disease susceptibility is transferable by the antibiotic-altered microbiota. These results strongly suggest that early-life antibiotics have long-term consequences on the gut microbiota and enteropathogen infection susceptibility., (Copyright © 2019 Roubaud-Baudron et al.)

An emerging approach for cancer treatment employs the use of extracellular vesicles, specifically exosomes and microvesicles, as delivery vehicles. We previously demonstrated that microvesicles can functionally deliver plasmid DNA to cells and showed that plasmid size and sequence, in part, determine the delivery efficiency. In this study, delivery vehicles comprised of microvesicles loaded with engineered minicircle (MC) DNA that encodes prodrug converting enzymes developed as a cancer therapy in mammary carcinoma models. We demonstrated that MCs can be loaded into shed microvesicles with greater efficiency than their parental plasmid counterparts and that microvesicle-mediated MC delivery led to significantly higher and more prolonged transgene expression in recipient cells than microvesicles loaded with the parental plasmid. Microvesicles loaded with MCs encoding a thymidine kinase (TK)/nitroreductase (NTR) fusion protein produced prolonged TK-NTR expression in mammary carcinoma cells. In vivo delivery of TK-NTR and administration of prodrugs led to the effective killing of both targeted cells and surrounding tumor cells via TK-NTR-mediated conversion of codelivered prodrugs into active cytotoxic agents. In vivo evaluation of the bystander effect in mouse models demonstrated that for effective therapy, at least 1% of tumor cells need to be delivered with TK-NTR-encoding MCs. These results suggest that MC delivery via microvesicles can mediate gene transfer to an extent that enables effective prodrug conversion and tumor cell death such that it comprises a promising approach to cancer therapy., (©2019 American Association for Cancer Research.)

Although the intestinal microbiome has been increasingly implicated in autoimmune diseases, much is unknown about its roles in Multiple Sclerosis (MS). Our aim was to compare the microbiome between treatment-naïve MS subjects early in their disease course and controls, and between Caucasian (CA), Hispanic (HA), and African American (AA) MS subjects. From fecal samples, we performed 16S rRNA V4 sequencing and analysis from 45 MS subjects (15 CA, 16 HA, 14 AA) and 44 matched healthy controls, and whole metagenomic shotgun sequencing from 24 MS subjects (all newly diagnosed, treatment-naïve, and steroid-free) and 24 controls. In all three ethnic groups, there was an increased relative abundance of the same single genus, Clostridium, compared to ethnicity-matched controls. Analysis of microbiota networks showed significant changes in the network characteristics between combined MS cohorts and controls, suggesting global differences not restricted to individual taxa. Metagenomic analysis revealed significant enrichment of individual species within Clostridia as well as particular functional pathways in the MS subjects. The increased relative abundance of Clostridia in all three early MS cohorts compared to controls provides candidate taxa for further study as biomarkers or as etiologic agents in MS.

Male infertility might be caused by genetic and/or environmental factors that impair spermatogenesis and epididymal sperm maturation. Here we report that heterozygous deletion of the nuclear receptor coactivator-5 (Ncoa5) gene resulted in decreased motility and progression of spermatozoa in the cauda epididymis, leading to infertility in male mice. Light microscopic and ultrastructural analysis revealed morphological defects in the spermatozoa collected from the cauda epididymis of Ncoa5 +/- mice. Immunohistochemistry showed that interleukin-6 (IL-6) expression in epithelial cells of Ncoa5 +/- epididymis was higher than wild type counterparts. Furthermore, heterozygous deletion of Il-6 gene in Ncoa5 +/- male mice partially improved spermatozoa motility and moderately rescued infertility phenotype. Our results uncover a previously unknown physiological role of NCOA5 in the regulation of epididymal sperm maturation and suggest that NCOA5 deficiency could cause male infertility through increased IL-6 expression in epididymis.

Chinese American immigrants are at increased risk for Helicobacter pylori infection and stomach cancer. Despite their increased risk, very few prevention strategies exist which target this vulnerable population. The purpose of this article is to present the stakeholder engaged development, review, assessment, refinement, and finalization of a H. pylori treatment adherence and stomach cancer prevention curriculum specifically designed to engage vulnerable, limited English proficient Chinese Americans in New York City.

During somatic cell nuclear transfer (SCNT), egg activation is required to initiate embryonic development. In zebrafish cloning, the reconstructed egg is activated by exposing it to hypotonic water. Egg activation using water-only is not capable of activating the same intracellular calcium release as fertilization which is required for proper embryonic development. Here we test whether the use of soluble sperm extract (SSE) can properly modulate the activation of reconstructed eggs during SCNT. We microinjected SSE from genomic-inactivated zebrafish sperm into unfertilized eggs and reconstructed eggs right after somatic cell nuclear transfer. We also evaluated the most effective approach for SSE microinjection. Microinjection of SSE (with 0.68 mg/ml of protein concentration) into non-activated eggs through the micropyle induced parthenogenetic development beyond the blastula stage, whereas all water-only activated eggs failed to enter the cleavage period. Microinjection of SSE at 1 mg/ml of protein concentration into non-activated reconstructed egg improved the developmental rate of cloned embryos in comparison to non-injected control clones. The cumulative survival time of cloned embryos injected with SSE was significantly longer than reconstructed eggs activated following sham injection (P<0.01). No significant difference was found among controls (P=0.32). SSE benefits both parthenogenesis and the survival cloned embryos which have never been reported in zebrafish. Further work is necessary to define the functional component(s) of SSE as well as the physiological pathway, to understand its principle of action and advance the utilization of SSE in cloning.

Helicobacter pylori is the most abundant bacterium in the gastric epithelium, and its presence has been associated with the risk of developing gastric cancer. As of 15 years ago, no other bacteria were associated with gastric epithelial colonization; but thanks to new methodologies, many other non- H. pylori bacteria have been identified. It is possible that non- H. pylori may have a significant role in the development of gastric cancer. Here, we discuss the specific role of H. pylori as a potential trigger for events that may be conducive to gastric cancer, and consider whether or not the rest of the gastric microbiota represent an additional risk in the development of this disease.

We dedicated this review to discuss Helicobacter pylori as one of the latest identified bacterial pathogens in humans and whether its role is mainly as a pathogen or a commensal. Diseases associated with this bacterium were highly prevalent during the 19th century and gradually have declined. Most diseases associated with H. pylori occurred in individuals older than 40 years of age. However, acquisition of H. pylori occurs mainly in young children inside the family setting. Prevalence and incidence of H. pylori has had a dramatic change in the last part of the 20th century and beginning of the 21th century. In developed countries there is a clear interruption of transmission and the lowest prevalence is observed in children younger than 10 years in these countries. A similar decline is observed but not at the same level in developing countries. Here we discuss the impact of the presence or absence of H. pylori in the health status of humans. We also discuss whether it is necessary or not to establish H. pylori eradication programs on light of the current decline in H. pylori prevalence.

Helicobacter pylori colonization is prevalent throughout the world, and is predominantly acquired during childhood. In developing countries, >70% of adult populations are colonized with H. pylori and >50% of children become colonized before the age of 10 years. However, the exact timing of acquisition is unknown. We assessed detection of H. pylori acquisition among a birth cohort of 105 children in Mirzapur, Bangladesh. Blood samples collected at time 0 (cord blood), and at 6, 12, 18, and 24 months of life were examined for the presence of IgG and IgA antibodies to whole cell H. pylori antigen and for IgG antibodies to the CagA antigen using specific ELISAs and immunoblotting. Breast milk samples were analyzed for H. pylori-specific IgA antibodies. Cord blood was used to establish maternal colonization status. H. pylori seroprevalence in the mothers was 92.8%. At the end of the two-year follow-up period, 50 (47.6%) of the 105 children were positive for H. pylori in more than one assay. Among the colonized children, CagA prevalence was 78.0%. A total of 58 children seroconverted: 50 children showed persistent colonization and 8 (7.6%) children showed transient seroconversion, but immunoblot analysis suggested that the transient seroconversion observed by ELISA may represent falsely positive results. Acquisition of H. pylori was not influenced by the mother H. pylori status in serum or breastmilk. In this population with high H. pylori prevalence, we confirmed that H. pylori in developing countries is detectable mainly after the first year of life.

Despite that the human gastrointestinal tract is the most populated ecological niche by bacteria in the human body, much is still unknown about its characteristics. This site is highly susceptible to the effects of many external factors that may affect in the quality and the quantity of the microbiome. Specific factors such as diet, personal hygiene, pharmacological drugs and the use of antibiotics can produce a significant impact on the gut microbiota. The effect of these factors is more relevant early in life, when the gut microbiota has not yet fully established. In this review, we discussed the effect of type and doses of the antibiotics on the gut microbiota and what the major consequences in the use and abuse of these antimicrobial agents., (Copyright © 2017 IMSS. Published by Elsevier Inc. All rights reserved.)

Broad-spectrum antibiotics are frequently prescribed to children. Early childhood represents a dynamic period for the intestinal microbial ecosystem, which is readily shaped by environmental cues; antibiotic-induced disruption of this sensitive community may have long-lasting host consequences. Here we demonstrate that a single pulsed macrolide antibiotic treatment (PAT) course early in life is sufficient to lead to durable alterations to the murine intestinal microbiota, ileal gene expression, specific intestinal T-cell populations, and secretory IgA expression. A PAT-perturbed microbial community is necessary for host effects and sufficient to transfer delayed secretory IgA expression. Additionally, early-life antibiotic exposure has lasting and transferable effects on microbial community network topology. Our results indicate that a single early-life macrolide course can alter the microbiota and modulate host immune phenotypes that persist long after exposure has ceased.High or multiple doses of macrolide antibiotics, when given early in life, can perturb the metabolic and immunological development of lab mice. Here, Ruiz et al. show that even a single macrolide course, given early in life, leads to long-lasting changes in the gut microbiota and immune system of mice.

To better characterize murine intestinal microbiota, a large number (187) of Gram-positive-staining, rod- and coccoid-shaped, and facultatively or strictly anaerobic bacteria were isolated from small and large intestinal contents from mice. Based on 16S rRNA gene sequencing, a total 115 isolates formed three phylogenetically distinct clusters located within the family Erysipelotrichaceae. Group 1, as represented by strain NYU-BL-A3T, was most closely related to Allobaculum stercoricanis, with 16S rRNA gene sequence similarity values of 87.7 %. A second group, represented by NYU-BL-A4T, was most closely related to Faecalibaculum rodentium, with 86.6 % 16S rRNA gene sequence similarity. A third group had a nearly identical 16S rRNA gene sequence (99.9 %) compared with the recently described Faecalibaculum rodentium, also recovered from a laboratory mouse; however, this strain had a few differences in biochemical characteristics, which are detailed in an emended description. The predominant (>10 %) cellular fatty acids of strain NYU-BL-A3T were C16 : 0 and C18 : 0, and those of strain NYU-BL-A4T were C10 : 0, C16 : 0, C18 : 0 and C18 : 1ω9c. The two groups could also be distinguished by multiple biochemical reactions, with the group represented by NYU-BL-A4T being considerably more active. Based on phylogenetic, biochemical and chemotaxonomic criteria, two novel genera are proposed, Ileibacterium valens gen. nov., sp. nov. with NYU-BL-A3T (=ATCC TSD-63T=DSM 103668T) as the type strain and Dubosiella newyorkensis gen. nov., sp. nov. with NYU-BL-A4T (=ATCC TSD-64T=DSM 103457T) as the type strain.

Background: Preeclampsia (PE), small for gestational age (SGA), and spontaneous preterm birth (PTB) each may be complications of impaired placental function in pregnancy. Although their exact pathogenesis is still unknown, certain infectious agents seem to play a role. Helicobacter pylori (H. pylori) colonization has been associated with increased risk for PE. Our aim was to assess the association between H. pylori colonization and PE, SGA, and PTB., Material and Methods: We measured IgG anti-H. pylori and CagA antibodies in serum of pregnant women (median 20.5 weeks, range 16.5-29.4) who participated in a population-based prospective cohort study. Delivery and medical records were assessed. Information on demographics, education, and maternal risk factors was collected by questionnaire. We used multivariate logistic regression analyses to assess associations between H. pylori colonization and PE, SGA, and PTB., Results: In total, 6348 pregnant women were assessed. H. pylori positivity was found in 2915 (46%) women, of whom 1023 (35%) also were CagA-positive. Pregnancy was complicated by PE, SGA, or PTB in 927 (15%) women. H. pylori colonization was associated with PE (aOR 1.51; 95%CI 1.03-2.25). Differentiation according to CagA status revealed the same risk. H. pylori was positively related with SGA, mainly explained by CagA-positive strains (aOR 1.34; 1.04-1.71). No association was observed between H. pylori and PTB., Conclusions: Our data suggest that H. pylori colonization may be a risk factor for PE and SGA. If these associations are confirmed by future studies and shown to be causal, H. pylori eradication may reduce related perinatal morbidity and mortality., (© 2016 John Wiley & Sons Ltd.)

Background: Helicobacter pylori colonizes the gastric mucosa of more than half of the world's population. There is increasing evidence H. pylori protects against the development of obesity and childhood asthma/allergies in which the development of these diseases coincide with transient dysbiosis. However, the mechanism underlying the association of H. pylori eradication with human metabolic and immunological disorders is not well-established. In this study, we aimed to investigate the local and systemic effects of H. pylori eradication through untargeted fecal lipidomics and plasma metabolomics approaches by liquid chromatography mass spectrometry (LC-MS). Results: Our study revealed that eradication of H. pylori eradication (i.e., loss of H. pylori and/or H. pylori eradication therapy) changed many global metabolite/lipid features, with the majority being down-regulated. Our findings primarily show that H. pylori eradication affects the host energy and lipid metabolism which may eventually lead to the development of metabolic disorders. Conclusion: These predictive metabolic signatures of metabolic and immunological disorders following H. pylori eradication can provide insights into dynamic local and systemic metabolism related to H. pylori eradication in modulating human health.

Helicobacter pylori remains one of the most common bacterial infections worldwide. Clarithromycin resistance is the most important cause of H. pylori eradication failures. Effective antibiotic therapies in H. pylori infection must be rapidly adapted to local resistance patterns. We investigated the prevalence of clarithromycin resistance due to mutations in positions 2142 and 2143 of 23SrRNA gene of H. pylori by fluorescence in situ hybridisation (FISH), and compared with culture and antimicrobial susceptibility testing in 234 adult patients with dyspepsia who were enrolled. Antrum and corpus biopsy specimens were obtained for rapid urease test, histopathology and culture. Epsilometer test was used to assess clarithromycin susceptibility. H. pylori presence and clarithromycin susceptibility were determined by FISH in paraffin-embedded biopsy specimens. We found that 164 (70.1%) patients were positive for H. pylori based on clinical criteria, 114 (69.5% CI 62.5-76.6%) were culture positive, and 137 (83.5% CI 77.8-89.2%) were FISH positive. Thus the sensitivity of FISH was significantly superior to that of culture. However specificity was not significantly different (91.4 versus 100.0%, respectively). The resistance rate to clarithromycin for both antrum and corpus was detected in H. pylori -positive patients; 20.2% by FISH and 28.0% by E-test.The concordance between E-test and FISH was only 89.5% due to the presence of point mutations different from A2143G, A2142G or A2142C. We conclude that FISH is significantly more sensitive than culture and the E-test for the detection of H. pylori and for rapid determinination of claritromycin susceptibility. The superior hybridisation efficiency of FISH is becoming an emerging molecular tool as a reliable, rapid and sensitive method for the detection and visualisation of H. pylori , especially when the management of H. pylori eradication therapy is necessary. This is particularly important for the treatment of patients with H. pylori eradication failure.

Background: More than 50 years after the discovery of antibiotics, bacterial infections have decreased substantially; however, antibiotics also may have negative effects such as increasing susceptibility to pathogens. An intact microbiome is an important line of defense against pathogens. We sought to determine the effect of orally administered antibiotics both on susceptibility to pathogens and on impact to the microbiome. We studied Campylobacter jejuni , one of the most common causes of human diarrhea, and Acinetobacter baumannii , which causes wound infections. We examined the effects of antibiotic treatment on the susceptibility of mice to those pathogens as well as their influence on the mouse gut microbiome., Results: In C57/BL6 mice models, we explored the effects of pathogen challenge, and antibiotic treatment on the intestinal microbiota. Mice were treated with either ciprofloxacin, penicillin, or water (control) for a 5-day period followed by a 5-day washout period prior to oral challenge with C. jejuni or A. baumannii to assess antibiotic effects on colonization susceptibility. Mice were successfully colonized with C. jejuni more than 118 days, but only transiently with A. baumannii . These challenges did not lead to any major effects on the composition of the gut microbiota. Although antibiotic pre-treatment did not modify pathogen colonization, it affected richness and community structure of the gut microbiome. However, the antibiotic dysbiosis was significantly reduced by pathogen challenge., Conclusions: We conclude that despite gut microbiota disturbance, susceptibility to gut colonization by these pathogens was unchanged. The major gut microbiome disturbance produced by antibiotic treatment may be reduced by colonization with specific microbial taxa.

Background and Aim: Helicobacter pylori can survive long incubation periods under anaerobic conditions, and should be possible to isolate under anaerobic conditions. Our aim was to isolate H. pylori in anaerobic conditions, from gastric biopsies of H. pylori infected patients., Methods: We enrolled 27 patients with bleeding (erosive) gastritis (mean age 36.3 years, 55.6% male) from Hanoi, Vietnam. H. pylori status was confirmed by qPCR., Results: H. pylori were recovered under anaerobic and micro-aerobic conditions from gastric biopsies in 16 patients. Anaerobic conditions yielded significantly higher H. pylori recovery rates than micro-aerobic conditions (81.3% vs. 31.3%, P= 0.01). H. pylori isolates were characterized by PCR for specific virulence markers and the genotypes were similar to those previously described in this region of the world., Conclusions: H. pylori can be isolated under anaerobic conditions. These findings may provide new insight into the physiology of this human pathogen and help to identify the route of H. pylori transmission., Competing Interests: All the authors declared that they do not have any conflict of interest., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Background: Highly virulent strains of the gastric pathogen Helicobacter pylori encode a type IV secretion system (T4SS) that delivers the effector protein CagA into gastric epithelial cells. Translocated CagA undergoes tyrosine phosphorylation by members of the oncogenic c-Src and c-Abl host kinases at EPIYA-sequence motifs A, B and D in East Asian-type strains. These phosphorylated EPIYA-motifs serve as recognition sites for various SH2-domains containing human proteins, mediating interactions of CagA with host signaling factors to manipulate signal transduction pathways. Recognition of phospho-CagA is mainly based on the use of commercial pan-phosphotyrosine antibodies that were originally designed to detect phosphotyrosines in mammalian proteins. Specific anti-phospho-EPIYA antibodies for each of the three sites in CagA are not forthcoming., Results: This study was designed to systematically analyze the detection preferences of each phosphorylated East Asian CagA EPIYA-motif by pan-phosphotyrosine antibodies and to determine a minimal recognition sequence. We synthesized phospho- and non-phosphopeptides derived from each predominant EPIYA-site, and determined the recognition patterns by seven different pan-phosphotyrosine antibodies using Western blotting, and also investigated representative East Asian H. pylori isolates during infection. The results indicate that a total of only 9-11 amino acids containing the phosphorylated East Asian EPIYA-types are required and sufficient to detect the phosphopeptides with high specificity. However, the sequence recognition by the different antibodies was found to bear high variability. From the seven antibodies used, only four recognized all three phosphorylated EPIYA-motifs A, B and D similarly well. Two of the phosphotyrosine antibodies preferentially bound primarily to the phosphorylated motif A and D, while the seventh antibody failed to react with any of the phosphorylated EPIYA-motifs. Control experiments confirmed that none of the antibodies reacted with non-phospho-CagA peptides and in accordance were able to recognize phosphotyrosine proteins in human cells., Conclusions: The results of this study disclose the various binding preferences of commercial anti-phosphotyrosine antibodies for phospho-EPIYA-motifs, and are valuable in the application for further characterization of CagA phosphorylation events during infection with H. pylori and risk prediction for gastric disease development.

The epidemiology of Helicobacter pylori (H. pylori) infection is related to human poverty with marked differences between developing and developed countries. Socioeconomic factors and living standards are the main determinants of the age-dependent acquisition rate of H. pylori, and consequently its prevalence. The aim of this study was to assess the risk and sero-prevalence of H. pylori colonization among Orang Asli in Peninsula Malaysia. This cross-sectional study was conducted on Orang Asli subjects in seven isolated settlements spanning across all three major tribes (Negrito, Proto Malay and Senoi) in Malaysia. Socio-demographic characteristics of the subjects were obtained through interview. Subjects were tested for H. pylori colonization based on CagA and whole cell (WC) antigen serological assays. A total of 275 subjects participated in this study. Among these subjects, 115 (44.7%) were H. pylori sero-positive with highest sero-prevalence among Negrito (65.7%). Among subjects who were H. pylori sero-positive, CagA sero positivity was also significantly higher among Negrito. The highest proportion of respondents reported to be H. pylori sero-positive was from age group 30 years old and below (57.9%), males (56.2%), Negrito (48.6%) and live in bamboo house (92.3%). The highest proportion of respondents reported to be CagA sero-positive was from age group 30 years old and below (41.4%), males (35.6%) and Negrito (48.6%). The results of this study demonstrate that H. pylori colonization can be related to age, gender, tribes and house materials and CagA sero-positive stain closely associated with age, gender and tribes.

Early childhood is a critical stage for the foundation and development of both the microbiome and host. Early-life antibiotic exposures, cesarean section, and formula feeding could disrupt microbiome establishment and adversely affect health later in life. We profiled microbial development during the first 2 years of life in a cohort of 43 U.S. infants and identified multiple disturbances associated with antibiotic exposures, cesarean section, and formula feeding. These exposures contributed to altered establishment of maternal bacteria, delayed microbiome development, and altered α-diversity. These findings illustrate the complexity of early-life microbiome development and its sensitivity to perturbation., (Copyright © 2016, American Association for the Advancement of Science.)

Unlabelled: We studied skin microbiota present in three skin sites (forearm, axilla, scalp) in men from six ethnic groups living in New York City., Methods: Samples were obtained at baseline and after four days following use of neutral soap and stopping regular hygiene products, including shampoos and deodorants. DNA was extracted using the MoBio Power Lyzer kit and 16S rRNA gene sequences determined on the IIlumina MiSeq platform, using QIIME for analysis., Results: Our analysis confirmed skin swabbing as a useful method for sampling different areas of the skin because DNA concentrations and number of sequences obtained across subject libraries were similar. We confirmed that skin location was the main factor determining the composition of bacterial communities. Alpha diversity, expressed as number of species observed, was greater in arm than on scalp or axilla in all studied groups. We observed an unexpected increase in α-diversity on arm, with similar tendency on scalp, in the South Asian group after subjects stopped using their regular shampoos and deodorants. Significant differences at phylum and genus levels were observed between subjects of the different ethnic origins at all skin sites., Conclusions: We conclude that ethnicity and particular soap and shampoo practices are secondary factors compared to the ecological zone of the human body in determining cutaneous microbiota composition.