Role of the complement system in antibody-dependent enhancement of flavivirus infections

Flavivirus infections have increased dramatically in the last decades in tropical and subtropical regions of the world. Antibody-dependent enhancement of dengue virus infections has been one of the main hypotheses to explain severity of disease and one of the major challenges to safe and effective vaccine development. In the presence of cross-reactive sub-neutralizing concentrations of anti-dengue antibodies, immune complexes can amplify viral infection in mononuclear phagocytic cells, triggering a cytokine cascade and activating the complement system that leads to severe disease. The complement system comprises a family of plasma and cellular surface proteins that recognize pathogen associated molecular patterns, modified ligands and immune complexes, interacting in a regulated manner and forming an enzymatic cascade. Pathogenic as well as protective effects of complement have been reported in flavivirus infections. This review provides updated knowledge on complement activation during flavivirus infection, including antiviral effects of complement and its regulation, as well as mechanisms of complement evasion and dysregulation of complement activity during viral infection leading to pathogenesis. Particularly, insights into classical pathway activation and its protective role on antibody-dependent enhancement of flavivirus infections are highlighted.Fil: Byrne, Alana Brooke. Gobierno de la Ciudad de Buenos Aires. Hospital General de Niños "Ricardo Gutiérrez"; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Talarico, Laura Beatriz. Gobierno de la Ciudad de Buenos Aires. Hospital General de Niños "Ricardo Gutiérrez"; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentin

Age-associated differences in clinical manifestations and laboratory parameters during a dengue virus type 4 outbreak in Argentina

Infection by any of the four dengue virus (DENV) serotypes produces a wide spectrum of clinical illness in humans. Differences in clinical manifestation and severity have been associated with secondary heterologous infection, patient age, and virus serotype. In this context, this retrospective study sought to analyze the presentation of dengue in patients during the 2014 DENV-4 outbreak affecting the City of Orán, Salta Province, Argentina. Demographic data, clinical manifestations, and laboratory abnormalities of laboratory-confirmed dengue patients were compared between age groups and between patients with and without warning signs. Of 301 patients with laboratory-confirmed dengue, 37.9% presented dengue with warning signs. Although nearly half of all patients had secondary DENV infections, no severe dengue cases, or deaths were reported. Furthermore, no association was found between incidence of warning signs and pre-existing immunity to DENV. Pediatric patients were least likely to present warning signs and showed significantly decreased risk of fever, retro-orbital pain, arthalgia, diarrhea and thrombocytopenia, and higher risk of rash compared to older patients. Female patients of all ages were also at higher risk of developing several symptoms. The characterization of DENV-4 infection in humans, a DENV serotype recently reported in Argentina, revealed differences in clinical manifestations, laboratory parameters and the presence/absence of warning signs based on age group. Further investigation of these age-related differences should contribute to better assessment of dengue disease in at risk populations.Fil: Byrne, Alana Brooke. Fundación para la Investigación en Infectología Infantil; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Gutierrez, Francisco Guillermo. Hospital San Vicente de Paul; Argentina. Universidad Nacional de Tucumán. Facultad de Medicina; ArgentinaFil: Bruno, Agostina Alejandra. Hospital San Vicente de Paul; ArgentinaFil: Cordoba, María Teresa. Hospital San Vicente de Paul; ArgentinaFil: Bono, María M.. Hospital San Vicente de Paul; ArgentinaFil: Polack, Fernando Pedro. Fundación para la Investigación en Infectología Infantil; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Talarico, Laura Beatriz. Fundación para la Investigación en Infectología Infantil; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Quipildor, Marcelo Omar. Hospital San Vicente de Paul; Argentin

A murine model of dengue virus infection in suckling C57BL/6 and BALB/c mice

Dengue is a significant public health concern across tropical and subtropical regions worldwide, principally causing disease in children. Very young children are at increased risk of severe manifestations of dengue infection. The mechanism of dengue disease in this population is not fully understood. In this study, we present a murine model of dengue virus primary infection in suckling C57BL/6 and BALB/c mice in order to investigate disease pathogenesis. Three‐day‐old C57BL/6 mice intraperitoneally infected with DENV‐2 NGC were more susceptible to infection than BALB/c mice, showing increased liver enzymes, extended viremia, dissemination to organs and histological alterations in liver and small intestine. Furthermore, the immune response in DENV‐infected C57BL/6 mice exhibited a marked Th1 bias compared to BALB/c mice. These findings highlight the possibility of establishing an immunocompetent mouse model of DENV‐2 infection in suckling mice that reproduces certain signs of disease observed in humans and that could be used to further study age‐related mechanisms of dengue pathogenesis.Fil: Byrne, Alana Brooke. Fundación para la Investigación en Infectología Infantil; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: García, Ayelén G.. Fundación para la Investigación en Infectología Infantil; ArgentinaFil: Brahamian, Jorge Martin. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales; Argentina. Fundación para la Investigación en Infectología Infantil; ArgentinaFil: Mauri, Aldana. Fundación para la Investigación en Infectología Infantil; ArgentinaFil: Ferretti, Adrián. Fundación para la Investigación en Infectología Infantil; ArgentinaFil: Polack, Fernando Pedro. Fundación para la Investigación en Infectología Infantil; ArgentinaFil: Talarico, Laura Beatriz. Fundación para la Investigación en Infectología Infantil; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentin

Interference in dengue virus adsorption and uncoating by carrageenans

This study demonstrated that the λ- and ι-carrageenans, sulfated polysaccharides containing linear chains of galactopyranosyl residues, are potent inhibitors of dengue virus type 2 (DENV-2) and 3 (DENV-3) multiplication in Vero and HepG2 cells, with values of effective concentration 50% from 0.14 to 4.1 μg/ml. This activity was assayed by plaque reduction, virus yield inhibition and antigen expression tests, and was independent of the input multiplicity of infection in the range 0.001-1. The inhibitory action of the λ-carrageenan, an heparan sulfate (HS)-imitative compound, was exerted by a dual interference with virus adsorption and internalization of nucleocapsid into the cytoplasm. Although virus particles may enter the cell when compound was added after DENV-2 adsorption, as shown by intracellular uptake of radiolabeled DENV-2 particles and quantitative RT-PCR, infectious center and virion uncoating assays have shown that carrageenan-treated virions cannot be released from the endosomes. Viral protein synthesis, the first step of macromolecular synthesis after DENV entry to the host cell, was not affected by the carrageenan. Furthermore, no inhibition of virus multiplication was detected when the entry process was bypassed through DENV-2 RNA transfection into the cell. The dual sites of action of an HS-like molecule suggest that, at least in monkey kidney and human hepatic cells, the HS residues in the cell membrane appear to act as mediators for DENV-2 entry, an interesting alternative target for flavivirus therapy.Fil: Talarico, Laura Beatriz. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Química Biológica. Laboratorio de Virología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Damonte, Elsa Beatriz. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Química Biológica. Laboratorio de Virología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentin

Cell entry of dengue virus

Dengue virus is an expanding public health problem in tropical and subtropical regions of the world, mainly owing to failure in the maintenance of control programs for the mosquito vector Aedes aegypti and increasing and unplanned urbanization. It has been estimated that over 50 million dengue virus infections of varying severity occur globally each year. making this virus the most significant mosquito-borne human pathogen. However, there is no specific antiviral therapy or vaccine for treatment or prevention. This review focuses on recent data describing the putative molecules and mechanisms involved in the complex process of dengue virus binding and entry into mosquito and mammalian cells in primary infections. Furthermore, the perspectives of these early events in the virus life cycle as a target for antidengue therapeutic strategies are also considered.Fil: Acosta, Eliana Gisela. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Química Biológica. Laboratorio de Virología; ArgentinaFil: Talarico, Laura Beatriz. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Química Biológica. Laboratorio de Virología; ArgentinaFil: Damonte, Elsa Beatriz. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Química Biológica. Laboratorio de Virología; Argentin

Murine models of dengue virus infection for novel drug discovery

Introduction: Dengue virus (DENV) is the causative agent of the most prevalent human disease transmitted by mosquitoes in tropical and subtropical regions worldwide. At present, no antiviral drug is available and the difficulties to develop highly protective vaccines against the four DENV serotypes maintain the requirement of effective options for dengue chemotherapy. Areas covered: The availability of animal models that reproduce human disease is a very valuable tool for the preclinical evaluation of potential antivirals. Here, the main murine models of dengue infection are described, including immunocompetent wild-type mice, immunocompromised mice deficient in diverse components of the interferon (IFN) pathway and humanized mice. The main findings in antiviral testing of DENV inhibitory compounds in murine models are also presented. Expert opinion: At present, there is no murine model that fully recapitulates human disease. However, immunocompromised mice deficient in IFN-α/β and -γ receptors, with their limitations, have shown to be the most suitable system for antiviral preclinical testing. In fact, the AG129 mouse model allowed the identification of celgosivir, an inhibitor of cellular glucosidases, as a promising option for DENV therapy. However, clinical trials still were not successful, emphasizing the difficulties in the transition from preclinical testing to human treatment.Fil: Byrne, Alana Brooke. Gobierno de la Ciudad Autónoma de Buenos Aires. Hospital General de Niños "Ricardo Gutiérrez". Departamento de Medicina; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Garcia, Cybele. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales; ArgentinaFil: Damonte, Elsa Beatriz. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales; ArgentinaFil: Talarico, Laura Beatriz. Gobierno de la Ciudad Autónoma de Buenos Aires. Hospital General de Niños "Ricardo Gutiérrez". Departamento de Medicina; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentin

Changes in antiviral susceptibility to entry inhibitors and endocytic uptake of dengue-2 virus serially passaged in Vero and C6/36 cells

The aim of the present study was to analyze the influence of virus origin, mammalian or mosquito cell-derived, on antiviral susceptibility of DENV-2 to entry inhibitors and the association of this effect with any alteration in the mode of entry into the cell. To this end, ten serial passages of DENV-2 were performed in mosquito C6/36 cells or monkey Vero cells and the antiviral susceptibility of each virus passage to sulfated polysaccharides (SPs), like heparin and carrageenans, was evaluated by a virus plaque reduction assay. After serial passaging in Vero cells, DENV-2 became increasingly resistant to SP inhibition whereas the antiviral susceptibility was not altered in virus propagated in C6/36 cells. The change in antiviral susceptibility was associated to a differential mode of entry into the host cell. The route of endocytic entry for productive Vero cell infection was altered from a non-classical clathrin independent pathway for C6/36-grown virus to a clathrin-mediated endocytosis when the virus was serially propagated in Vero cells. Our results show the impact of the cellular system used for successive propagation of DENV on the initial interaction between the host cell and the virion in the next round of infection and the relevant consequences it might have during the in vitro evaluation of entry inhibitors.Fil: Acosta, Eliana Gisela. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Química Biológica. Laboratorio de Virología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Piccini, Luana Érica. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Química Biológica. Laboratorio de Virología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales; ArgentinaFil: Talarico, Laura Beatriz. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Química Biológica. Laboratorio de Virología; ArgentinaFil: Castilla, Viviana. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Química Biológica. Laboratorio de Virología; ArgentinaFil: Damonte, Elsa Beatriz. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales; Argentina. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Química Biológica. Laboratorio de Virología; Argentin

An algal-derived DL-galactan hybrid is an efficient preventing agent for in vitro dengue virus infection

The DL-galactan hybrid C2S-3, isolated from the red seaweed Cryptonemia crenulata (Halymeniaceae, Halymeniales), is a potent and selective inhibitor of the multiplication of diverse strains of DENV-2 in Vero cells with higher effectiveness than the reference polysaccharide heparin. The presence of the compound either only at virus adsorption or at virus internalization exerted a significant and dose-dependent inhibition in DENV-2 plaque number. The compound failed to inactivate DENV-2 directly by incubation of virus before cell infection as well as to induce a refractory state by cell pretreatment. Thus, the inhibitory effect was exclusively exerted through a blockade in virus multiplication during the infectious process. When the entry of DENV-2 particles into the cell is bypassed, as occurs in virus RNA transfection, the polysaccharide C2S-3 failed to block the completion of the multiplication cycle. Furthermore, the antiviral properties of C2S-3 are not correlated with anticoagulant activity.Fil: Talarico, Laura Beatriz. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales; ArgentinaFil: Duarte, María E. R.. Universidade Federal do Paraná; BrasilFil: Zibetti, Rosiane G. M.. Universidade Federal do Paraná; Brasil. Instituto de Ensino Superior Pequeno Príncipe; BrasilFil: Noseda, Miguel D.. Universidade Federal do Paraná; BrasilFil: Damonte, Elsa Beatriz. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales; Argentin

Antiviral activity of an N-allyl acridone against dengue virus

Dengue virus (DENV), a member of the family Flaviviridae, is at present the most widespread causative agent of a human viral disease transmitted by mosquitoes. Despite the increasing incidence of this pathogen, there are no antiviral drugs or vaccines currently available for treatment or prevention. In a previous screening assay, we identified a group of N-allyl acridones as effective virus inhibitors. Here, the antiviral activity and mode of action targeted to viral RNA replication of one of the most active DENV-2 inhibitors was further characterized.Results: The compound 10-allyl-7-chloro-9(10H)-acridone, designated 3b, was active to inhibit the in vitro infection of Vero cells with the four DENV serotypes, with effective concentration 50% (EC50) values in the range 12.5-27.1 μM, as determined by virus yield inhibition assays. The compound was also effective in human HeLa cells. No cytotoxicity was detected at 3b concentrations up to 1000 μM. Mechanistic studies demonstrated that virus entry into the host cell was not affected, whereas viral RNA synthesis was strongly inhibited, as quantified by real time RT-PCR. The addition of exogenous guanosine together with 3b rescued only partially the infectivity of DENV-2.Conclusions: The acridone derivative 3b selectively inhibits the infection of Vero cells with the four DENV serotypes without a direct interaction with the host cell or the virion but interfering specifically with the intracellular virus multiplication. The mode of antiviral action for this acridone apparently involves the cellular enzyme inosine-monophospahe dehydrogenase together with another still unidentified target related to DENV RNA synthesis.Fil: Mazzucco, María Belén. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Química Biológica; ArgentinaFil: Talarico, Laura Beatriz. Fundacion Para la Invest.en Infectologia Infantil; Argentina. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Química Biológica. Laboratorio de Virología; ArgentinaFil: Vatansever, Sezen. Koc University; TurquíaFil: Carro, Ana Clara. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Química Biológica. Laboratorio de Virología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales; ArgentinaFil: Fascio, Mirta Liliana. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales; ArgentinaFil: D'Accorso, Norma Beatriz. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Química Orgánica; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Centro de Investigaciones en Hidratos de Carbono; ArgentinaFil: Garcia, Cybele. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Química Biológica. Laboratorio de Virología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales; ArgentinaFil: Damonte, Elsa Beatriz. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Química Biológica. Laboratorio de Virología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Ciudad Universitaria. Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales; Argentin

Chemical structure and antiviral activity of carrageenans from Meristiella gelidium against herpes simplex and dengue virus

Chemical and spectroscopic methods showed that the major KCl-precipitated galactans from Meristiella gelidium (Solieriaceae) are iota/kappa/nu-hybrid carrageenans with the former one in higher proportion. These carrageenans showed, by HPSEC-MALLS analysis, unimodal symmetrical peaks with MW of 425.6–956.7 kDa. The effectiveness of the crude extracts from M. gelidium against HSV-2 was higher than the corresponding extract from G. griffithsiae, previously determined. However, when considering the homogeneous carrageenans, the fractions obtained from both seaweeds showed the same level of activity. The extracts and carrageenan derived from M. gelidium were more effective inhibitors of DENV-2 if compared with G. griffithsiae samples and reference polysaccharides. The most active fraction obtained from M. gelidium showed a selectivity index against HSV-2 of 25,000, a value high enough to consider this carrageenan as a promising agent to be evaluated for the treatment of genital HSV-2 infections.Fil: De S.F. Tischer, Paula Cristina. Universidade Federal do Paraná; BrasilFil: Talarico, Laura Beatriz. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Química Biológica. Laboratorio de Virología; ArgentinaFil: Noseda, Miguel D.. Universidade Federal do Paraná; BrasilFil: Pita B. Guimarães, Silvia Maria. Instituto de Botânica; BrasilFil: Damonte, Elsa Beatriz. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales. Departamento de Química Biológica. Laboratorio de Virología; ArgentinaFil: Duarte, Maria Eugênia R.. Universidade Federal do Paraná; Brasi