Caracterización "in vitro" e "in vivo" de los vectores atenuados de poxvirus MVA y NYVAC como candidatos vacunales frente al VIH-SIDA

Tesis Doctoral inédita leída en la Universidad Autónoma de Madrid, Facultad de Ciencias, Departamento de Biología Molecular. Fecha de lectura: 23-11-200

Correlación directa entre volumen de infarto cerebral isquémico medido en resonancia magnética y niveles séricos de neurofilamento

Antecedentes La concentración de neurofilamento (NfL) en sangre ha sido propuesta como un biomarcador de daño neuroaxonal. Distintas condiciones neurológicas pueden elevar sus niveles, pero su importancia en el infarto cerebral isquémico aún permanece indefinida. Con el objetivo de describir su comportamiento en esta patología, en este estudio se realizó una correlación entre los niveles de NfL y el volumen de infarto cerebral. Métodos Estudio retrospectivo. Se utilizaron muestras de sangre almacenadas e información de los pacientes de una cohorte longitudinal prospectiva. Se incluyeron un total de 59 pacientes con diagnóstico de infarto cerebral isquémico. Los niveles de NfL se analizaron en sangre con el kit LS-F6701 mediante técnica ELISA. El volumen de infarto cerebral fue medido en resonancia magnética utilizando el software 3D-Slicer. Los niveles de NfL fueron correlacionados con (I) volumen de infarto cerebral, (II) severidad del infarto cerebral calculado con la escala National Institute of Health Stroke Scale (NIHSS), (III) pronóstico funcional a los tres meses medido con la escala de Rankin modificada. El análisis de las variables se realizó utilizando el coeficiente de correlación de Spearman. Resultados Los niveles de NfL en sangre se correlacionaron de manera positiva con el volumen de infarto cerebral (r. Spearman 0.25, p=0.05) y con la escala de severidad clínica NIHSS (r. Spearman 0.27, p=0.039). El pronóstico funcional a los tres meses después del infarto cerebral no fue asociado con los niveles de NfL. Conclusión La presencia y la cantidad de daño neuroaxonal estimado por los niveles en sangre de NfL correlacionaron con el volumen de infarto cerebral y la severidad clínica. No encontramos asociación entre NfL y el pronóstico a los tres meses

Diversity in viral anti-PKR mechanisms: A remarkable case of evolutionary convergence

Most viruses express during infection products that prevent or neutralize the effect of the host dsRNA activated protein kinase (PKR). Translation of Sindbis virus (SINV) mRNA escapes to PKR activation and eIF2 phosphorylation in infected cells by a mechanism that requires a stem loop structure in viral 26S mRNA termed DLP to initiate translation in the absence of functional eIF2. Unlike the rest of viruses tested, we found that Alphavirus infection allowed a strong PKR activation and eIF2α phosphorylation in vitro and in infected animals so that the presence of DLP structure in mRNA was critical for translation and replication of SINV. Interestingly, infection of MEFs with some viruses that express PKR inhibitors prevented eIF2α phosphorylation after superinfection with SINV, suggesting that viral anti-PKR mechanisms could be exchangeable. Thus, translation of SINV mutant lacking the DLP structure (ΔDLP) in 26S mRNA was partially rescued in cells expressing vaccinia virus (VV) E3 protein, a known inhibitor of PKR. This case of heterotypic complementation among evolutionary distant viruses confirmed experimentally a remarkable case of convergent evolution in viral anti-PKR mechanisms. Our data reinforce the critical role of PKR in regulating virus-host interaction and reveal the versatility of viruses to find different solutions to solve the same conflict.This work was supported in part from the VIRUS-HOST interaction programme (Comunidad de Madrid) and by grants from the Ministerio de Ciencia e Innovación (SAF2006-09810; SAF2008-02036) and the Fundación Mutua Madrileña (FMM 2008). The institutional support from Fundación Ramón Areces is also acknowledged. E.D was a recipient of VIRUS-HOST programme postdoctoral contract. R.T. was a recipient of the SAF2006-09810 contract and I.V. was a researcher of Ramón y Cajal Programme.Peer Reviewe

Suppression of NYVAC infection in hela cells requires RNase L but is independent of protein kinase R activity

Protein kinase R (PKR) and RNase L are host cell components that function to contain viral spread after infections. In this study, we analyzed the role of both proteins in the abortive infection of human HeLa cells with the poxvirus strain NYVAC, for which an inhibition of viral A27L and B5R gene expression is described. Specifically, the translation of these viral genes is independent of PKR activation, but their expression is dependent on the RNase L activityThis work was supported by grants from the Spanish Ministry of Health (FIS2011-00127) and Bayer Group Grants4Grants (2013-08-0982 to S.G., SAF2008-02036 to M.E., and AGL2010-15495 to D.R.

Molecular mechanisms of resveratrol as chemo and radiosensitizer in cancer

One of the primary diseases that cause death worldwide is cancer. Cancer cells can be intrinsically resistant or acquire resistance to therapies and drugs used for cancer treatment through multiple mechanisms of action that favor cell survival and proliferation, becoming one of the leading causes of treatment failure against cancer. A promising strategy to overcome chemoresistance and radioresistance is the co-administration of anticancer agents and natural compounds with anticancer properties, such as the polyphenolic compound resveratrol (RSV). RSV has been reported to be able to sensitize cancer cells to chemotherapeutic agents and radiotherapy, promoting cancer cell death. This review describes the reported molecular mechanisms by which RSV sensitizes tumor cells to radiotherapy and chemotherapy treatment

LED-enhanced dietary and organoleptic qualities in postharvest tomato fruit

Tomato fruit possess high lycopene concentrations, which increases after reaching the breaker stage of ripeness. Light emitting diode (LED) systems have emerged as a clean and efficient artificial lighting technique for use in horticulture. The objective of this research was to examine the effect of LEDs on postharvest and lycopene quality parameters. The effects on quality parameters were significantly different depending on the tomato fruit cultivar. The commercial and organoleptic quality parameters most affected were firmness and titratable acidity. The lycopene concentration in all tomato cultivars increased between 30% and 60% when they were exposed to LED light compared with dark-exposed fruit. One hour of LED light exposure per day during the postharvest phase of fruit increased commercial and organoleptic parameters, and increased the lycopene concentration. Fruit exposed to LED light with a high red:far red (R:FR) light ratio had increased firmness compared with those exposed to other types of white LED light. The highest R:FR ratio used in this study resulted in fruit with higher titratable acidity than those exposed to lower R:FR ratios. Overall, the results of this study demonstrated that the postharvest exposure of tomato fruit to LED light with a high R:FR ratio induces lycopene synthesis, with lycopene concentrations being 41% higher in comparison with exposure to darkness and 24% higher in comparison with exposure to other LED lighting conditions

Control of Macrophomina pseudophaseolina Crous, Sarr & Ndiaye with Trichoderma spp. and botanical and chemical pesticides

Chili is the most important traditionally consumed vegetable in Mexico; however, production is reduced by soil fungi. The objective of the research was to evaluate in in vitro, greenhouse and field conditions, the effectiveness of biological, botanical and chemical pesticides for the control of Macrophomina pseudophaseolina. In vitro, greenhouse and field experiments were carried out to evaluate the control effect of different pesticides for the control of M. pseudophaseolina. It was determined that all the evaluated strains of Trichoderma spp. They had a fungistatic effect against M. pseudophaseolina, and T. reesei showed the greatest antagonism and antibiosis against M. pseudophaseolina. High, medium and low doses of NeemAcar® and high and medium doses of Regalia® Maxx inhibited 100% the growth of M. pseudophaseolina mycelium. In the greenhouse, the lowest percentage of severity was obtained in the treatment with Regalia® Maxx + T. reesei. In the field, the lowest severity was determined with the application of NeemAcar® CE + Headline®. The preventive or curative application of biological, botanical and chemical pesticides did not influence the severity of the disease.Objective: The objective of the research was to evaluate in in vitro, greenhouse and field conditions, the effectiveness of biological, botanical and chemical pesticides for the control of Macrophomina pseudophaseolina. Design/methodology/approach: In vitro, greenhouse and field experiments were carried out to evaluate the control effect of different pesticides for the control of M. pseudophaseolina. Results: It was determined that all the evaluated strains of Trichoderma spp. they had a fungistatic effect against M. pseudophaseolina, and T. reesei showed the greatest antagonism and antibiosis against M. pseudophaseolina. High, medium and low doses of NeemAcar® and high and medium doses of Regalia® Maxx inhibited 100% the growth of M. pseudophaseolina mycelium. In the greenhouse, the lowest percentage of severity was obtained in the treatment with Regalia® Maxx + T. reesei. In the field, the lowest severity was determined with the application of NeemAcar® CE + Headline®. Limitations on study/implications: Our results are essential for the management of this disease by producers. Findings/conclusions: The implementation of the use of Trichoderma spp., botanical pesticides and chemical insecticides is recommended for the control of M. pseudophaseolina,

Remote Substituent Effects on the Stereoselectivity and Organocatalytic Activity of Densely Substituted Unnatural Proline Esters in Aldol Reactions

The organocatalytic activities of highly substituted proline esters obtained through asymmetric [3+2] cycloadditions of azomethine ylides derived from glycine iminoesters have been analyzed by 19F NMR and through kinetic isotope effects. Kinetic rate constants have been determined for unnatural proline esters incorporating different substituents. It has been found that exo-L and endo-L unnatural proline methyl esters yield opposite enantiomers in aldol reactions between cyclic ketones and aromatic aldehydes. The combined results reported in this study show subtle and remote effects that determine the organocatalytic behavior of these synthetic but readily available amino acid derivatives. These data can be used as design criteria for the development of new pyrrolidine-based organocatalysts.Financial support was provided by the Spanish Ministerio de Economía y Competitividad (MINECO) and the Fondo Europeo de Desarrollo Regional (FEDER) (projects CTQ2010-16959/BQU, CTQ2012-35535, CTQ2013-40855-R, CTQ2007-62771/BQU, CTQ2010-20387, CTQ2010-17436, and Consolider-Ingenio CSD2007-00006), the University of the Basque Country (UPV/EHU, UFI11/22 QOSYC), the Basque Government (GV/EJ, grant IT-324-07), the Generalitat Valenciana-FEDER (PROMETEO/2009/039), the Gobierno de Aragón-FSE (research group E40), and the University of Alicante. M. d. G. R. thanks the Donostia International Physics Center (DIPC) for a postdoctoral contract. M. S. and L. C. gratefully thank MINECO for a contract funding their PhD projects

The impact of extensive grazing in the behavior of soluble sugars in Prosopis laevigata (Humb. & Bonpl. ex Willd.) M.C.Johnst. trees

Objective: To evaluate the impact of extensive grazing in the accumulation of soluble sugars in Prosopis laevigata trees, whose leaves and fruits are directly consumed by cattle. Design/Methodology/Approach: The ejido Emiliano Zapata in Durango was the study area. Stem and root samples were collected from a stand of extensive grazing and a stand without cattle. The sampling was carried out in three growth stages: March (flowering), June (fruition), and October (leaf fall). The samples were frozen in liquid nitrogen and were lyophilized. Afterwards, they were ground and 10 mg of dry matter were weighted in microtubes. The total soluble sugars (TSS) concentration was determined following the Van Handel methodology, using a spectrophotometer at 625 nm. The statistical analysis was carried out using an ANOVA and the Tukey’s test. Results: In March, the grazing area had lower TSS concentrations during regrowth than the area without grazing, both at root and stem levels. Study Limitations/Implications: The intensity of grazing and the pasture rotation should be regulated to favor carbohydrate accumulation in trees, which is required for the formation of the meristematic tissues. Finding/Conclusions: Extensive grazing has an impact on the synthesis and accumulation of TSS in mesquite trees. Therefore, the consumption of branches, leaves, and fruits decreases TSS concentrations in the stem and the root

Disease surveillance during the reintroduction of the Iberian Lynx (Lynx pardinus) in southwestern Spain

This article belongs to the Section Wildlife.The restoration of Iberian lynx (Lynx pardinus) populations in Extremadura (Southwestern Spain) have been carried out since 2014. One of the measures to ensure the success of this program is to examine the effects that diseases may have on reintroduction. Since diseases may be greatly located at certain sites because of the specific ecological requirements of the pathogens and/or vectors, reintroduced individuals may present a risk of infection once released. To determine which pathogens the reintroduced individuals may encounter, we performed a molecular and sero-epidemiological survey in reintroduced and wild-born lynxes. From 2015 to 2019, 69 Iberian lynxes (40 reintroduced and 29 wild-born) were sampled and screened against 10 viral, bacterial and piroplasmid agents. In parallel, 195 sympatric carnivores from the families Canidae, Felidae, Viverridae, Herpestidae and Mustelidae were tested against current or past infections to six common canine/feline viruses. In the Iberian lynx, low contact rates of active infection were obtained for the feline leukemia provirus (FeLV: 1.5%; 1/67), feline parvovirus (FPV: 1.5%; 1/67) and Cytauxzoon sp. (6.7%; 1/15). We also confirmed the emergence of Aujeszky’s disease (suid herpesvirus-1) in this population (SuHV-1: 11.8%; 2/17). Evidence of previous exposure was detected for canine distemper virus (CDV: 5.8%; 3/52), feline coronavirus (1.9%; 1/52), FPV (7.7%; 1/13) and feline calicivirus (FCV: 5.3%; 1/19). From 25 recovered lynx carcasses, we could confirm infectious etiology involvement in the death of four individuals (SuHV-1 in two individuals, coinfection of Cytauxzoon spp. and Aeromonas veronii in one lynx and a Streptococcus canis myositis in another lynx). We confirmed the circulation of CDV, FPV, FeLV, FCV and the feline immunodeficiency virus within the sympatric carnivore community. Due to the low contact rate of infectious agents in such a small, endangered population, we recommend continuing a disease surveillance program to determine the prognostic factors of survival, understand the role that disease may play during the reintroduction and anticipate disease outbreaks that may pose a risk for the entire reintroduced population.[Simple Summary]: The restoration of Iberian lynx (Lynx pardinus) populations in Extremadura (Southwestern Spain) have been carried out since 2014. To evaluate the effect that infectious diseases may have on their reintroduction, we performed a molecular and sero-epidemiological survey in reintroduced and wild-born lynxes and sympatric carnivores. From 2015 to 2019, 69 Iberian lynxes were screened against 10 viral, bacterial and piroplasmid agents. In parallel, 195 sympatric carnivores were tested against current or past infections to six common canine/feline viruses. In the Iberian lynx, low contact rates of active infection were obtained for feline leukemia provirus (FeLV: 1.5%; 1/67), feline parvovirus (FPV: 1.5%; 1/67) and Cytauxzoon sp. (6.7%; 1/15). We confirmed the emergence of Aujeszky’s disease (suid herpesvirus-1) in this population (SuHV-1: 11.8%; 2/17). We detected the circulation of FeLV, parvovirus, canine distemper virus (CDV), feline calicivirus (FCV) and feline immunodeficiency virus within the sympatric carnivore community and FCV, FPV, CDV and feline coronavirus in lynxes. Due to the low contact rate of infectious agents in such a small, endangered population, we recommend continuing a disease surveillance program to determine the prognostic factors of survival, understand the role that disease may play during the reintroduction and anticipate disease outbreaks that may pose a risk for the entire reintroduced population.This research was funded by the European Union through its LIFE project Life+IBERLINCE (LIFE+10NAT/ES/570) “Recuperación de la distribución histórica del lince ibérico (Lynx pardinus) en España y Portugal”.Peer reviewe