Clinical aspects and detection of Zika virus RNA in several tissues of experimentally infected BALB/cAn mice / Aspectos clínicos e detecção de RNA do vírus Zika em diferentes tecidos de camundongos BALB/cAn infectados experimentalmente

Our group infected BALB/cAn mice with Zika virus to evaluate clinical signs and viral load in several tissues at three different kinetic points. We inoculated fifteen mice with a 100µl of a viral solution to collect nine different tissues, from each animal, for RNA extraction and quantification. Infections caused no lethality. Some of them, however, showed great agitation, hair bristling, and itchy skin. Viral RNA was detected in one sample of heart, eight of the spleen, and two of skeletal muscle. Seven positive detections were from the third day after infection. Only spleen yielded positive results at a later time.

Morphological Aspects and Viremia Analysis of BALB/c Murine Model Experimentally Infected with Dengue Virus Serotype 4

Ever since its brief introduction in the Brazilian territory in 1981, dengue virus serotype 4 (DENV-4) remained absent from the national epidemiological scenario for almost 25 years. The emergence of DENV-4 in 2010 resulted in epidemics in most Brazilian states. DENV-4, however, remains one of the least studied among the four DENV serotypes. Despite being known as a mild serotype, DENV-4 is associated with severe cases and deaths and deserves to be investigated; however, the lack of suitable experimental animal models is a limiting factor for pathogenesis studies. Here, we aimed to investigate the susceptibility and potential tropism of DENV-4 for liver, lung and heart of an immunocompetent mice model, and to evaluate and investigate the resulting morphological and ultrastructural alterations upon viral infection. BALB/c mice were inoculated intravenously with non-neuroadapted doses of DENV-4 isolated from a human case. The histopathological analysis of liver revealed typical alterations of DENV, such as microsteatosis, edema and vascular congestion, while in lung, widespread areas of hemorrhage and interstitial pneumonia were observed. While milder alterations were present in heart, characterized by limited hemorrhage and discrete presence of inflammatory infiltrate, the disorganization of the structure of the intercalated disc is of particular interest. DENV-4 RNA was detected in liver, lung, heart and serum of BALB/c mice through qRT-PCR, while the NS3 viral protein was observed in all of the aforementioned organs through immunohistochemistry. These findings indicate the susceptibility of the model to the serotype and further reinforce the usefulness of BALB/c mice in studying the many alterations caused by DENV

Estudo do tropismo do vírus dengue tipo 4 em modelo murino BALB/c: infecção experimental, análises morfológicas e de viremia

Made available in DSpace on 2018-07-10T18:52:12Z (GMT). No. of bitstreams: 2 license.txt: 1748 bytes, checksum: 8a4605be74aa9ea9d79846c1fba20a33 (MD5) arthur_rasinhas_ioc_mest_2017.pdf: 8690113 bytes, checksum: b72e84d9b2ba4ee76c6cc2af74d12d40 (MD5) Previous issue date: 2017Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Rio de Janeiro, RJ, Brasil.Desde sua introdução no território brasileiro, no ano de 1981, o DENV-4 permaneceu ausente do cenário epidemiológico nacional por quase 25 anos, até sua reintrodução em 2010. Esse evento marca a cocirculação dos 4 sorotipos conhecidos no Brasil e embora os mecanismos por trás da patogênese do vírus ainda não sejam completamente compreendidos, a interação entre anticorpos oriundos de infecções heterotípicas é um fator essencial para a evolução de quadros brandos de DEN em quadros hemorrágicos e de choque. A dificuldade no estudo da interação do vírus com o hospedeiro se deve principalmente à ausência de um modelo animal experimental que reproduza adequadamente a infecção por DENV como observada em casos humanos Modelos atuais fazem uso de animais imunodeficientes, vias de inoculação invasivas e inóculos virais neuroadaptados, condições que não apenas dificultam a compreensão da patogênese como também impossibilitam a utilização de tais modelos para testes de candidatos a vacinas e fármacos. O presente estudo tem como objetivo analisar o potencial tropismo do DENV-4 por fígado, pulmão e coração de camundongos da linhagem BALB/c e avaliar as alterações morfológicas e ultraestruturais geradas pelo vírus nestes órgãos. Para tal, camundongos da linhagem imunocompetente BALB/c foram inoculados por via intravenosa com doses não neuroadaptadas de DENV-4 isolado de caso humano. Alterações observadas nos tecidos estudados apresentaram perfil semelhante ao caracterizado em casos humanos de DEN, e a detecção do vírus nos tecidos revelou a susceptibilidade do modelo ao sorotipo.Ever since its introduction in the Brazilian territory, in 1981, DENV-4 has remained absent from the national epidemiological scene for almost 25 years, until its reintroduction in 2010. This event marks the cocirculation of the 4 known serotypes in Brazil, and although the mechanisms behind the virus\2019 pathogenesis are not yet fully understood, the interaction between antibodies originated from heterotypical infections is an essential factor for the evolution of mild DEN cases into hemorrhagic and shock cases. The difficulty in studying the interaction between virus and host is mainly due to the absence of an experimental animal model that adequately replicates the DENV infection as observed in humans Current models utilize immunodeficient animals, invasive inoculation routes, and neuroadapted viral inocula, conditions that not only hamper the comprehension of the pathogenesis but also prevent the use of such models for testing vaccine and drug candidates. The present study aims to analyze the potential tropism of DENV-4 for hepatic, pulmonary and cardiac tissue and evaluate the morphological and ultrastructural alterations caused by the virus in said tissues. To achieve this goal, immunocompetent mice of the BALB/c line were inoculated via intravenous route with non neuroadapted doses of DENV-4 isolated from human case. Alterations observed in the analyzed tissue presented similar profile to that shown in human cases of DEN, and the detection of the virus within the tissue show the susceptibility of the model to the serotype

First detection of dengue virus in the saliva of immunocompetent murine model

<div><p>The lack of an experimental animal model for the study of dengue pathogenesis is a limiting factor for the development of vaccines and drugs. In previous studies, our group demonstrated the susceptibility of BALB/c mice to infection by dengue virus (DENV) 1 and 2, and the virus was successfully isolated in several organs. In this study, BALB/c mice were experimentally infected intravenously with DENV-4, and samples of their saliva were collected. Viral RNA extracted from the saliva samples was subjected to qRT-PCR, with a detection limit of 0.002 PFU/mL. The presence of DENV-4 viral RNA was detected in the saliva of two mice, presenting viral titers of 109 RNA/mL. The detection of DENV RNA via saliva sampling is not a common practice in dengue diagnosis, due to the lower detection rates in human patients. However, the results observed in this study seem to indicate that, as in humans, detection rates of DENV RNA in mouse saliva are also low, correlating the infection in both cases. This study reports the first DENV detection in the saliva of BALB/c immunocompetent mice experimentally infected with non-neuroadapted DENV-4.</p></div

Immunocompetent Mice Infected by Two Lineages of Dengue Virus Type 2: Observations on the Pathology of the Lung, Heart and Skeletal Muscle

Dengue virus (DENV) infection by one of the four serotypes (DENV-1 to 4) may result in a wide spectrum of clinical manifestations, with unpredictable evolution and organ involvement. Due to its association with severe epidemics and clinical manifestations, DENV-2 has been substantially investigated. In fact, the first emergence of a new lineage of the DENV-2 Asian/American genotype in Brazil (Lineage II) in 2008 was associated with severe cases and increased mortality related to organ involvement. A major challenge for dengue pathogenesis studies has been a suitable animal model, but the use of immune-competent mice, although sometimes controversial, has proven to be useful, as histological observations in infected animals reveal tissue alterations consistent to those observed in dengue human cases. Here, we aimed to investigate the outcomes caused by two distinct lineages of the DENV-2 Asian/American genotype in the lung, heart and skeletal muscle tissues of infected BALB/c mice. Tissues were submitted to histopathology, immunohistochemistry, histomorphometry and transmission electron microscopy (TEM) analysis. The viral genome was detected in heart and skeletal muscle samples. The viral antigen was detected in cardiomyocytes and endothelial cells of heart tissue. Heart and lung tissue samples presented morphological alterations comparable to those seen in dengue human cases. Creatine kinase serum levels were higher in mice infected with both lineages of DENV-2. Additionally, statistically significant differences, concerning alveolar septa thickening and heart weight, were observed between BALB/c mice infected with both DENV-2 lineages, which was demonstrated to be an appropriate experimental model for dengue pathogenesis studies on lung, heart and skeletal muscle tissues