The Role of Extracellular Vesicles in Modulating the Host Immune Response during Parasitic Infections.

Parasites are the cause of major diseases affecting billions of people. As the inflictions caused by these parasites affect mainly developing countries, they are considered as neglected diseases. These parasitic infections are often chronic and lead to significant immunomodulation of the host immune response by the parasite, which could benefit both the parasite and the host and are the result of millions of years of co-evolution. The description of parasite extracellular vesicles (EVs) in protozoa and helminths suggests that they may play an important role in host-parasite communication. In this review, recent studies on parasitic (protozoa and helminths) EVs are presented and their potential use as novel therapeutical approaches is discussed.

Surface analysis of Dicrocoelium dendriticum. The molecular characterization of exosomes reveals the presence of miRNAs.

With the aim of characterizing the molecules involved in the interaction of Dicrocoelium dendriticum adults and the host, we have performed proteomic analyses of the external surface of the parasite using the currently available datasets including the transcriptome of the related species Echinostoma caproni. We have identified 182 parasite proteins on the outermost surface of D. dendriticum. The presence of exosome-like vesicles in the ESP of D. dendriticum and their components has also been characterized. Using proteomic approaches, we have characterized 84 proteins in these vesicles. Interestingly, we have detected miRNA in D. dendriticum exosomes, thus representing the first report of miRNA in helminth exosomes. BIOLOGICAL SIGNIFICANCE: In order to identify potential targets for intervention against parasitic helminths, we have analyzed the surface of the parasitic helminth Dicrocoelium dendriticum. Along with the proteomic analyses of the outermost layer of the parasite, our work describes the molecular characterization of the exosomes of D. dendriticum. Our proteomic data confirm the improvement of protein identification from "non-model organisms" like helminths, when using different search engines against a combination of available databases. In addition, this work represents the first report of miRNAs in parasitic helminth exosomes. These vesicles can pack specific proteins and RNAs providing stability and resistance to RNAse digestion in body fluids, and provide a way to regulate host-parasite interplay. The present data should provide a solid foundation for the development of novel methods to control this non-model organism and related parasites. This article is part of a Special Issue entitled: Proteomics of non-model organisms.

Cuantificación relativa de genes relacionados con la respuesta inmune antiviral en una población venezolana de Aedes (stegomyia) aegypti / Relative quantification of Antiviral immune response related genes in an Aedes (stemgoyia) aegypti Venezuelan population

El virus DENV-3 es el responsable del segundo mayor porcentaje de afecciones hemorrágicas severas causadas por este virus, solo superado por el virus DENV-2. La virulencia de este serotipo, causante de brotes epidémicos a gran escala en países como Venezuela donde circula activamente durante todo el año, es la razón principal del desarrollo investigativo en el campo de las interacciones virus-vector en la búsqueda de un control epidémico efectivo. En el presente trabajo, se evaluó la expresión de dos genes, RNAi (dcr-2 y ago-2), que codifican para proteínas de respuesta antiviral en mosquitos con la finalidad de estudiar el cambio en la expresión de los mismos en el vector una vez infectado con el DENV-3. Para ello, se infectó artificialmente una población de Aedes (stegomyia) aegypti de Trujillo, elaborándose dos grupos experimentales (15dpi y 20dpi) y un control; posteriormente se aisló, cuantificó y se realizó transcripción reversa al RNA total aislado de los grupos. La infección en los grupos experimentales se evidenció por la detección de bandas de productos de PCR de 511pb para DENV y 290pb para DENV-3 mediante electroforesis en gel de agarosa al 2%, y se realizó la cuantificación relativa de la expresión genética por PCR en tiempo real. Como resultado, la expresión de los genes no presentó cambios con respecto a los mosquitos no infectados (grupo control) (p<0.05). Estos resultados indican que el virus DENV-3 pudiera estar mostrando mecanismos de evasión sobre las vías de RNAi dentro del cuerpo del vector, y esto puede estar relacionado al hecho de que la replicación de los miembros del genero Flavivirus, entre ellos el DENV, se lleva a cabo en el sistema de membranas y vesículas, lo que les permite evadir el encuentro en el citoplasma con las proteínas asociadas al complejo de RNAi.

DENV-3 is the responsible for the second major percentage of hemorrhagic severe affections caused by this virus, only overcome by DENV-2. The virulence of this serotype is the cause of broad scale fever outbreaks in different countries, especially in Venezuela where it circulates actively throughout the year. This is the main reason of investigative developments in the virus-vector interactions field to find an effective epidemic control. In this study, we evaluated the expression of dcr-2 and ago-2, two RNAi antiviral pathway genes in mosquitoes to study the fold change in the expression of these genes in the mosquito vector Aedes (stegomyia) aegypti infected with DENV-3. We artificially infected a population of mosquitoes from Trujillo state (Venezuela) and prepared three (3) experimental groups (15dpi, 20dpi and a control group). Later, we isolated, quantified and applied a reverse transcription to the total RNA obtained of mosquitoes and the infection in the experimental groups was detected performing a 2% agarose gel electrophoresis of the PCR products of the total RNA (511bp for DENV and 290bp for DENV3). In addition, the infection in the experimental groups was confirmed by the detection of PCR products. The fold change in dcr-2 and ago-2 expression genes was quantify by Real Time PCR. As results, the fold change expression in the ago-2 and dcr-2 were equal to the non-infected mosquitoes “control group” (p<0.05). These results indicate that DENV-3 could have evasion mechanisms of RNAi pathway inside vector’s body, and this can be related to the fact that dengue virus replication is accomplished in the vesicle system and membrane system as well (endoplasmic reticulum and golgi complex) avoiding cytoplasmic encounter with the RNAi pathway proteins.