Determinación de marcas epigenéticas en genesimplicados en la respuesta temprana a Botrytis cinerea de Arabidopsis thaliana y Solanum lycopersicum

Existen en estudios recientes realizados en plantas, evidencias que vinculan la presencia de modificaciones epigenéticas con su respuesta a estreses bióticos. Sin embargo, dentro de estos estreses no se encuentra a penas información sobre patógenos necrótrofos. En esta tesis se exploran las posibles alteraciones en la estructura cromatinica que plantas de las especies Arabidopsis thaliana y Solanum lycopersicum (tomate) podrían experimentar en varios de sus genes expresados en la respuesta a la infección por Botrytis cinerea, un hongo necrótrofo. Se plantea para ellos la puesta a punto de un protocolo de extracción de cromatina y de inmunoprecipitación de la misma (ChIP) y, se relatan las dificultades derivadas de un patosistema de estas características, en las que el patógeno genera tejido necrótico a medida que avanza. Una vez obtenidos los protocolos funcionales, se analizan diversas modificaciones epigenéticas; concretamente de la hisona 3, entre ellas: algunas que se han relacionado con activación transcripcional como H3K4me3, H3K9ac y otras relacionadas con represión como H3K27me3; por otreo lado se estudia la presencia de la RNAPII en algunas condiciones. Los resultados muestran, en plantas de A. thaliana infectada a dos tiempos, un enriquecimiento de estas marcas activadoras en la zona del cuerpo y el promotor del gen AtPR1 (marcador de activación de una de las principales rutas defensivas), así como empobrecimiento de la represora. Resultados similares se observan con otros genes inducidos como AtCYP71A13 o AtELI3, implicados en la síntesis de la fitoalexina camalexina y del polímero lignina respectivamente. Por otro lado, resultados diferentes se observan en genes con demostrada expresión reprimida durante la infección, como son AtEXL7 o AtBGLU23. Tras resolver ciertos problemas, se consigue también analizar algunas de estas modificaciones (las activadoras) en plantas de S. lycopersicum y, adicionalmente, se examina la presencia de la RNAPII. Se examinan también genes inducidos por la infección de este hongo, entre los que cabe destacar los involucrados en la síntesis de las oxilipinas (con importantes implicaciones defensivas), SlDES, SlLOXD y SlDOX1. Similarmente a lo visto en A. thaliana se ve un incremento en las marcas activadoras en distintas zonas de estos genes y, además, se observa como la presencia de la RNAPII a menudo acompaña a estas marcas. Estos resultados pueden ser utilizados para la determinación de otras modificaciones epigenéticas y otros genes, además del estudio de la posible herencia transgeneracional de dichas modificaciones. Por otro lado, muchos de los genes examinados en ambas especies son biomarcadores potenciales de infección temprana en el patosistema S. lycopesicum-B. cinerea, lo que podría contribuir a su pronta detección en este cultivo y otros similares.There are recent studies carried out on plants, tha show evidences that link the presence of epigenetic modifications with their response to biotic stresses. However, within these stresses there is hardly any information about necrotrophic pathogens. This thesis explores the possible alterations in the chromatin structure that plants of the species Arabidopsis thaliana and Solanum lycopersicum (tomato) could experience in several of their genes expressed in response to the infection by Botrytis cinerea, a necrotrophic fungus. The development of a chromatin extraction protocol and its immunoprecipitation (ChIP) is proposed in order to achieve this goal, and the difficulties derived from a pathosystem of these characteristics are reported, in which the pathogen generates necrotic tissue as it advances . Once the functional protocols have been obtained, various epigenetic modifications are analyzed; specifically of hisona 3, among them: some that have been related to transcriptional activation such as H3K4me3, H3K9ac and others related to repression such as H3K27me3; On the other hand, the presence of RNAPII is studied in some conditions. The results show, in plants of A. thaliana whose samples have been collected in different stages of the infection, an enrichment of these activating marks in the area of ​​the body and the promoter of the AtPR1 gene (marker of activation of one of the main defensive pathways), as well as impoverishment of the repressor. Similar results are observed with other genes induced as AtCYP71A13 or AtELI3, involved in the synthesis of the phytoalexin camalexin and the lignin polymer respectively. On the other hand, different results are observed in genes with demonstrated repressed expression during infection, such as AtEXL7 or AtBGLU23. After solving certain problems, it is also possible to analyze some of these modifications (activators) in plants of S. lycopersicum and, additionally, the presence of RNAPII is examined. Genes induced by the infection of this fungus are also examined, among which are those involved in the synthesis of oxilipins (molecules with important defensive implications), SlDES, SlLOXD and SlDOX1. Similar to that seen in A. thaliana, there is an increase in the activating marks in different areas of these genes and, in addition, it is observed how the presence of the RNAPII often accompanies these marks. These results can be used for the determination of other epigenetic modifications and other genes, besides the study of the possible transgenerational inheritance of said modifications. On the other hand, many of the genes examined in both species are potential biomarkers of early infection in the S. lycopesicum-B patosystem. cinerea, which could contribute to its early detection in this crop and other similar ones

Absence of Cu–Zn superoxide dismutase BCSOD1 reduces Botrytiscinerea virulence in Arabidopsis and tomato plants, revealinginterplay among reactive oxygen species, callose and signallingpathways

Plants activate responses against pathogens, including the oxida-tive burst. Necrotrophic pathogens can produce reactive oxygenspecies (ROS) that benefit the colonization process. Previously, wehave demonstrated that tomato plants challenged with Botrytiscinerea accumulate ROS and callose, together with the inductionof genes involved in defence, signalling and oxidative metabolism.Here, we studied the infection phenotype of the Dbcs od1 strain inboth tomato and Arabidopsis plants. This mutant lacks bcsod1,which encodes Cu–Zn superoxide dismutase (SOD). This enzymecatalyses the conversion of superoxide ion (O–2) into hydrogen per-oxide (H2O2). ROS play a protective role and act as signals inplants. Dbcsod1 displayed reduced virulence compared with wild-type B05.10 in both species. Plants infected with Dbcsod1 accu-mulated less H2O2and more O–2than those infected with B05.10,which is associated with an increase in the defensive polymer cal-lose. This supports a major role of fungal SOD in H2O2productionduring the plant–pathogen interaction. The early induction of thecallose synthase gene PMR4 suggested that changes in ROSaltered plant defensive responses at the transcriptional level. Themetabolites and genes involved in signalling and in response tooxidative stress were differentially expressed on Dbcsod1 infec-tion, supporting the notion that plants perceive changes in ROSbalance and activate defence responses. A higher O2–/H2O2ratioseems to be beneficial for plant protection against this necrotroph.Our results highlight the relevance of callose and the oxylipin 12-oxo-phytodienoic acid (OPDA) in the response to changes in theoxidative environment, and clarify the mechanisms that underliethe responses to Botrytis in Arabidopsis and tomato plants.This work was supported by grants from the Spanish Ministry of Scienceand Innovation (AGL2010-22300-C03-01-02 and AGL2013-49023-C03-01-02), co-funded by the European Regional Development Funds (ERDF)and by Generalitat Valenciana Grupos de Excelencia PROMETEO/2012/066. J.L.-C. and O.C.-S. were recipients of research contracts from thePROMETEO/2012/066 grant. E.F.-C. was the recipient of research con-tracts from grant AGL2010-22300-C03-01. J.L.-C. is the recipient of aresearch contract from grant AGL2013-49023-C03-01

Plan estratégico de recursos humanos que contribuya a mejorar el clima organizacional de los empleados de la empresa. Ciudad de San Miguel, año 2019: caso de estudio: Hotel Trópico INN

RESUMEN: El estudio del clima organizacional realizado en El Hotel Trópico Inn, hace propio el objetivo de analizar la situación real y consciente, sobre las actividades cotidianas desarrolladas al interior y que desmejoran cualquier esfuerzo realizado por sus empleados ameritó que se desarrollara un estudio dedicado y minucioso para contrarrestar los efectos tanto internos como externos. El ambiente donde toda persona se desarrolla es de vital importancia, ya que este tiene un efecto directo en los comportamientos, decisiones, motivaciones, etc. Dentro de las empresas o instituciones ya sean públicas o privadas, este ambiente es llamado Clima Organizacional. El clima organizacional es influido por varios aspectos y algunos de ellos son las relaciones interpersonales, comunicación, liderazgo, estrés y motivación: El Clima Organizacional es uno de los aspectos más importantes que la mayoría de las organizaciones ignora, El Hotel Trópico Inn no es la excepción; es aquí donde surge la necesidad de realizar el estudio de un: “Plan estratégico de recursos humanos que contribuya a mejorar el clima organizacional de los empleados de la empresa Hotel Trópico Inn de la ciudad de san miguel” Se realizó un diagnóstico de análisis interno y externo para determinar los factores que afectan el clima organizacional de la empresa. Los aspectos que contiene la guía de clima organizacional son; técnicas y métodos de motivación con el que se busca incentivar al personal, otro aspecto es el estrés en el cual se ayuda al empleado sobre el Página | 154 manejo del mismo, el liderazgo, el cual ayuda en la toma de decisiones y actitudes de los colaboradores. ABSTRACT: The study of the organizational climate carried out at El Hotel Trópico Inn, makes its own the objective of analyzing the real and conscious situation, about the daily activities carried out inside and that deteriorate any effort made by its employees, merited the development of a dedicated study and thorough to counteract both internal and external effects. The environment where everyone develops is of vital importance, since it has a direct effect on behaviors, decisions, motivations, etc. Within companies or institutions, whether public or private, this environment is called Organizational Climate. The organizational climate is influenced by several aspects and some of them are interpersonal relationships, communication, leadership, stress and motivation: The Organizational Climate is one of the most important aspects that most organizations ignore, The Hotel Trópico Inn is not the exception; It is here where the need arises to carry out the study of a: "Strategic human resources plan that contributes to improving the organizational climate of the employees of the Hotel Trópico Inn company in the city of San Miguel". An internal analysis diagnosis and external to determine the factors that affect the organizational climate of the company. The aspects contained in the organizational climate guide are; Motivation techniques and methods with which it is sought to incentivize the staff, another aspect is stress in which the employee is helped on the management of the same, the leadership, which helps in the decision making and attitudes of the collaborators

The Histone Marks Signature in Exonic and Intronic Regions Is Relevant in Early Response of Tomato Genes to Botrytis cinerea and in miRNA Regulation

Research into the relationship between epigenetic regulation and resistance to biotic stresses provides alternatives for plant protection and crop improvement. To unravel the mechanisms underlying tomato responses to Botrytis cinerea, we performed a chromatin immunoprecipitation (ChIP) analysis showing the increase in H3K9ac mark along the early induced genes SlyDES, SlyDOX1, and SlyLoxD encoding oxylipin-pathway enzymes, and SlyWRKY75 coding for a transcriptional regulator of hormonal signaling. This histone mark showed a more distinct distribution than the previously studied H3K4me3. The RNAPol-ChIP analysis reflected the actual gene transcription associated with increased histone modifications. A different pattern of marks in the oxylipin-related genes against P. syringae supported a pathogen-specific profile, while no significant differences occurred in SlyWRKY75. The epigenetic regulation of SlyWRKY75 by the intron-binding miR1127-3p was supported by the presence of SlyWRKY75 pre-mRNA in control plants. Interestingly, mRNA was found to be accumulated in response to B. cinerea and P. syringae, while reduction in miRNA only occurred against B. cinerea. The intronic region presented a similar pattern of marks than the rest of the gene in both pathosystems, except for H3K4me3 in the miRNA binding site upon B. cinerea. We located the gene encoding Sly-miR1127-3p, which presented reduced H3K4me3 on its promoter against B. cinerea

Identification of stress associated microRNAs in Solanum lycopersicum by high-throughput Sequencing

Tomato (Solanum lycopersicum) is one of the most important crops around the world and also a model plant to study response to stress. High-throughput sequencing was used to analyse the microRNA (miRNA) profile of tomato plants undergoing five biotic and abiotic stress conditions (drought, heat, P. syringae infection, B. cinerea infection, and herbivore insect attack with Leptinotarsa decemlineata larvae) and one chemical treatment with a plant defence inducer, hexanoic acid. We identified 104 conserved miRNAs belonging to 37 families and we predicted 61 novel tomato miRNAs. Among those 165 miRNAs, 41 were stress-responsive. Reverse transcription quantitative PCR (RT-qPCR) was used to validate high-throughput expression analysis data, confirming the expression profiles of 10 out of 11 randomly selected miRNAs. Most of the differentially expressed miRNAs were stress-specific, except for sly-miR167c-3p upregulated in B. cinerea and P. syringae infection, sly-newmiR26-3p upregulated in drought and Hx treatment samples, and sly-newmiR33-3p, sly-newmiR6-3p and sly-newmiR8-3p differentially expressed both in biotic and abiotic stresses. From mature miRNAs sequences of the 41 stress-responsive miRNAs 279 targets were predicted. An inverse correlation between the expression profiles of 4 selected miRNAs (sly-miR171a, sly-miR172c, sly-newmiR22-3p and sly-miR167c-3p) and their target genes (Kinesin, PPR, GRAS40, ABC transporter, GDP and RLP1) was confirmed by RT-qPCR. Altogether, our analysis of miRNAs in different biotic and abiotic stress conditions highlight the interest to understand the functional role of miRNAs in tomato stress response as well as their putative targets which could help to elucidate plants molecular and physiological adaptation to stress

Different HCV Exposure Drives Specific miRNA Profile in PBMCs of HIV Patients

Micro RNAs (miRNAs) are essential players in HIV and HCV infections, as both viruses modulate cellular miRNAs and interact with the miRNA-mediated host response. We aim to analyze the miRNA profile of HIV patients with different exposure to HCV to explore specific signatures in the miRNA profile of PBMCs for each type of infection. We massively sequenced small RNAs of PBMCs from 117 HIV+ infected patients: 45 HIV+ patients chronically infected with HCV (HIV/HCV+), 36 HIV+ that spontaneously clarified HCV after acute infection (HIV/HCV-) and 36 HIV+ patients without previous HCV infection (HIV). Thirty-two healthy patients were used as healthy controls (HC). Differential expression analysis showed significantly differentially expressed (SDE) miRNAs in HIV/HCV+ (n = 153), HIV/HCV- (n = 169) and HIV (n = 153) patients. We found putative dysregulated pathways, such as infectious-related and PI3K signaling pathways, common in all contrasts. Specifically, putatively targeted genes involved in antifolate resistance (HIV/HV+), cancer-related pathways (HIV/HCV-) and HIF-signaling (HIV) were identified, among others. Our findings revealed that HCV strongly influences the expression profile of PBMCs from HIV patients through the disruption of its miRNome. Thus, different HCV exposure can be identified by specific miRNA signatures in PBMCs.This work has been supported by grants from Institute of Health Carlos III, [PI15CIII/00031 and PI18CIII/00020/ to AFR and VB] and the Foundation Universidad Alfonso X el Sabio-Santander [grant number 1.010.932 to AFR] and the Spanish AIDS Research Network (RD16CIII/0002/0002), and Centro de Investigación Biomédica en Red (CIBER) en Enfermedades Infecciosas (CB21/13/00044). AFR is supported by the Miguel Servet programme from Fondo de Investigación Sanitaria (ISCIII) [CP14/CIII/00010 and CPII20CIII/0001].info:eu-repo/semantics/publishedVersio

Tratamientos psicológicos empíricamente apoyados para adultos: Una revisión selectiva

Antecedentes: los tratamientos psicológicos han mostrado su eficacia, efectividad y eficiencia para el abordaje de los trastornos mentales; no obstante, considerando el conocimiento científico generado en los últimos años, no se dispone de trabajos de actualización en español sobre cuáles son los tratamientos psicológicos con respaldo empírico. El objetivo fue realizar una revisión selectiva de los principales tratamientos psicológicos empíricamente apoyados para el abordaje de trastornos mentales en personas adultas. Método: se recogen niveles de evidencia y grados de recomendación en función de los criterios propuestos por el Sistema Nacional de Salud de España (en las Guías de Práctica Clínica) para diferentes trastornos psicológicos. Resultados: los resultados sugieren que los tratamientos psicológicos disponen de apoyo empírico para el abordaje de un amplio elenco de trastornos psicológicos. El grado de apoyo empírico oscila de bajo a alto en función del trastorno psicológico analizado. La revisión sugiere que ciertos campos de intervención necesitan una mayor investigación. Conclusiones: a partir de esta revisión selectiva, los profesionales de la psicología podrán disponer de información rigurosa y actualizada que les permita tomar decisiones informadas a la hora de implementar aquellos procedimientos psicoterapéuticos empíricamente fundamentados en función de las características de las personas que demandan ayuda. Background: Psychological treatments have shown their efficacy, effectiveness, and efficiency in dealing with mental disorders. However, considering the scientific knowledge generated in recent years, in the Spanish context, there are no updating studies about empirically supported psychological treatments. The main goal was to carry out a selective review of the main empirically supported psychological treatments for mental disorders in adults. Method: Levels of evidence and degrees of recommendation were collected based on the criteria proposed by the Spanish National Health System (Clinical Practice Guidelines) for different psychological disorders. Results: The results indicate that psychological treatments have empirical support for the approach to a wide range of psychological disorders. These levels of empirical evidence gathered range from low to high depending on the psychological disorder analysed. The review indicates the existence of certain fields of intervention that need further investigation. Conclusions: Based on this selective review, psychology professionals will be able to have rigorous, up-to-date information that allows them to make informed decisions when implementing empirically based psychotherapeutic procedures based on the characteristics of the people who require help

Role of age and comorbidities in mortality of patients with infective endocarditis

[Purpose]: The aim of this study was to analyse the characteristics of patients with IE in three groups of age and to assess the ability of age and the Charlson Comorbidity Index (CCI) to predict mortality. [Methods]: Prospective cohort study of all patients with IE included in the GAMES Spanish database between 2008 and 2015.Patients were stratified into three age groups:<65 years,65 to 80 years,and ≥ 80 years.The area under the receiver-operating characteristic (AUROC) curve was calculated to quantify the diagnostic accuracy of the CCI to predict mortality risk. [Results]: A total of 3120 patients with IE (1327 < 65 years;1291 65-80 years;502 ≥ 80 years) were enrolled.Fever and heart failure were the most common presentations of IE, with no differences among age groups.Patients ≥80 years who underwent surgery were significantly lower compared with other age groups (14.3%,65 years; 20.5%,65-79 years; 31.3%,≥80 years). In-hospital mortality was lower in the <65-year group (20.3%,<65 years;30.1%,65-79 years;34.7%,≥80 years;p < 0.001) as well as 1-year mortality (3.2%, <65 years; 5.5%, 65-80 years;7.6%,≥80 years; p = 0.003).Independent predictors of mortality were age ≥ 80 years (hazard ratio [HR]:2.78;95% confidence interval [CI]:2.32–3.34), CCI ≥ 3 (HR:1.62; 95% CI:1.39–1.88),and non-performed surgery (HR:1.64;95% CI:11.16–1.58).When the three age groups were compared,the AUROC curve for CCI was significantly larger for patients aged <65 years(p < 0.001) for both in-hospital and 1-year mortality. [Conclusion]: There were no differences in the clinical presentation of IE between the groups. Age ≥ 80 years, high comorbidity (measured by CCI),and non-performance of surgery were independent predictors of mortality in patients with IE.CCI could help to identify those patients with IE and surgical indication who present a lower risk of in-hospital and 1-year mortality after surgery, especially in the <65-year group

Absence of endo-1,4-β-glucanase KOR1 alters the Jasmonate-dependent defence response to Pseudomonas syringae in Arabidopsis

During plant–pathogen interactions, the plant cell wall forms part of active defence against invaders. In recent years, cell wall-editing enzymes, associated with growth and development, have been related to plant susceptibility or resistance. Our previous work identified a role for several tomato and Arabidopsis endo-1,4-β-glucanases (EGs) in plant–pathogen interactions. Here we studied the response of the Arabidopsis thaliana T-DNA insertion mutant lacking EG Korrigan1 (KOR1) infected with Pseudomonas syringae. KOR1 is predicted to be an EG which is thought to participate in cellulose biosynthesis. We found that kor1-1 plants were more susceptible to P. syringae, and displayed severe disease symptoms and enhanced bacterial growth if compared to Wassilewskija (Ws) wild-type plants. Hormonal and gene expression analyses revealed that the jasmonic acid (JA) pathway was activated more in kor1-1 plants with an increase in the JA-biosynthesis gene LOX3 and a greater accumulation of JA. Upon infection the accumulation of JA and JA-isoleucine (JA-Ile) was higher than in wild-type plants and increased the induction of LOX3 and the JA-responsive PDF1.2 gene. In addition, the increase of salicylic acid (SA) in healthy and infected kor1-1 may reflect the complex interaction between JA and SA, which results in the more susceptible phenotype displayed by the infected mutant plants. Callose deposition was enhanced in infected kor1-1 and an increase in pathogen-induced hydrogen peroxide took place. The susceptible phenotype displayed by KOR1-deficient plants was coronatine-independent. No significant changes were detected in the hormonal profile of the kor1-1 plants infected by coronatine-deficient P. syringae cmaA, which supports that absence of EG KOR1 alters per se the plant response to infection. We previously reported increased resistance of kor1-1 to B. cinerea, hence, the lack of this EG alters cell wall properties and plant responses in such a way that benefits P. syringae colonisation but restricts B. cinerea invasion

Epigenetic regulation of the expression of WRKY75 transcription factor in response to biotic and abiotic stresses in Solanaceae plants

Key message SlyWRKY75: gene expression was induced in response to biotic stresses, especially in Botrytis cinerea-infected tomato plants, in which Sly-miR1127-3p is a putative SlyWRKY75 regulator and epigenetic marks were detected. Abstract WRKY75 transcription factor involved in Pi homeostasis was recently found also induced in defense against necrotrophic pathogens. In this study, we analyzed by RT-qPCR the expression of SlyWRKY75 gene in tomato plants in response to abiotic stresses (drought or heat) and biotic stresses (Colorado potato beetle larvae infestation, Pseudomonas syringae or Botrytis cinerea infection) being only differentially expressed following biotic stresses, especially upon B. cinerea infection (55-fold induction). JA and JA-Ile levels were significantly increased in tomato plants under biotic stresses compared with control plants, indicating that SlyWRKY75 might be a transcriptional regulator of the JA pathway. The contribution of miRNAs and epigenetic molecular mechanisms to the regulation of this gene in B. cinerea-infected tomato plants was explored. We identified a putative Sly-miR1127-3p miRNA predicted to bind the intronic region of the SlyWRKY75 genomic sequence. Sly-miR1127-3p miRNA was repressed in infected plants (0.4-fold) supporting that it might act as an epigenetic regulation factor of SlyWRKY75 gene expression rather than via the post-transcriptional mechanisms of canonical miRNAs. It has been proposed that certain miRNAs can mediate DNA methylation in the plant nucleus broadening miRNA functions with transcriptional gene silencing by targeting intron-containing pre-mRNAs. Histone modifications analysis by chromatin immunoprecipitation (ChIP) demonstrated the presence of the activator histone modification H3K4me3 on SlyWRKY75 transcription start site and gene body. The induction of this gene in response to B. cinerea correlates with the presence of an activator mark. Thus, miRNAs and chromatin modifications might cooperate as epigenetic factors to modulate SlyWRKY75 gene expression