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.

Retention of a recombinant GFP protein expressed by the yellow fever 17D virus in the E/NS1 intergenic region in the endoplasmic reticulum

The flaviviral envelope proteins, E protein and precursor membrane protein, are mainly associated with the endoplasmic reticulum (ER) through two transmembrane (TM) domains that are exposed to the luminal face of this compartment. Their retention is associated with the viral assembly process. ER-retrieval motifs were mapped at the carboxy terminus of these envelope proteins. A recombinant yellow fever (YF) 17D virus expressing the reporter green fluorescent protein (GFP) with the stem-anchor (SA) region of E protein fused to its carboxy terminus was subjected to distinct genetic mutations in the SA sequence to investigate their effect on ER retention. Initially, we introduced progressive deletions of the stem elements (H1, CS and H2). In a second set of mutants, the effect of a length increase for the first TM anchor region was evaluated either by replacing it with the longer TM of human LAMP-1 or by the insertion of the VALLLVA sequence into its carboxy terminus. We did not detect any effect on the GFP localisation in the cell, which remained associated with the ER. Further studies should be undertaken to elucidate the causes of the ER retention of recombinant proteins expressed at the intergenic E/NS1 region of the YF 17D virus polyprotein

Crime (In)Segurança e Mediatização

info:eu-repo/semantics/publishedVersio

Structural investigation of C6/36 and Vero cell cultures infected with a Brazilian Zika virus - Fig 6

<p><b>C6/36 cells infected with ZIKV analyzed by transmission electron microscopy (TEM) at different time points post-infection (A-B: 48 hr p.i., C-D: 72 hr p.i.).</b> Several large viroplasm-like perinuclear compartments (V) (A-B) and ZIKV particles (*) measuring approximately 40–50 nm in diameter in the endoplasmic reticulum cisternae (RER) (A, B) and in lysosomes (L) (C-D) were observed. Nucleocapsids were observed inside the rER (D). Thickening of the nuclear membrane and rough endoplasmic reticulum cisternae (rER) (black head arrow) (C), numerous lysosomes (L) (C, D) and vesicular compartments associated with rER (arrow) (C) measuring approximately 100 nm in diameter were observed. Nucleus (N).</p

Placental histopatology and clinical presentation of severe congenital Zika Syndrome in a human immunodeficiency virus-exposed uninfected infant

Submitted by Sandra Infurna ([email protected]) on 2018-01-16T13:40:19Z No. of bitstreams: 1 flavia_santos_etal_IOC_2017.pdf: 3809350 bytes, checksum: 4d9ef1ac3e72042bfdc5b1a8428c23d5 (MD5)Approved for entry into archive by Sandra Infurna ([email protected]) on 2018-01-16T13:59:35Z (GMT) No. of bitstreams: 1 flavia_santos_etal_IOC_2017.pdf: 3809350 bytes, checksum: 4d9ef1ac3e72042bfdc5b1a8428c23d5 (MD5)Made available in DSpace on 2018-01-16T13:59:35Z (GMT). No. of bitstreams: 1 flavia_santos_etal_IOC_2017.pdf: 3809350 bytes, checksum: 4d9ef1ac3e72042bfdc5b1a8428c23d5 (MD5) Previous issue date: 2017Universidade do Estado do Rio de Janeiro. Laboratório de Ultraestrutura e Biologia Tecidual. Rio de Janeiro, RJ, Brasil.Faculdade de Medicina de Campos. Campos dos Goytacazes, RJ, Brasil / Universidade Estadual do Norte Fluminense. Laboratório de Biotecnologia. Campos dos Goytacazes, RJ, Brasil.Faculdade de Medicina de Campos. Campos dos Goytacazes, RJ, Brasil.Faculdade de Medicina de Campos. Campos dos Goytacazes, RJ, Brasil / Universidade Estadual do Norte Fluminense. Laboratório de Biotecnologia. Campos dos Goytacazes, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Imunologia Viral. Rio de Janeiro, RJ. Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Imunologia Viral. Rio de Janeiro, RJ. Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório de Imunologia Viral. Rio de Janeiro, RJ. Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório Interdisciplinar de Pesquisas Médicas. Rio de Janeiro, RJ. Brasil.Fundação Oswaldo Cruz. Biomanguinhos. Laboratório de Tecnologia Virológica. Rio de Janeiro, RJ. Brasil.Universidade Federal do Estado do Rio de Janeiro. Anatomia Patológica. Rio de Janeiro, RJ, Brasil.Universidade do Estado do Rio de Janeiro. Laboratório de Ultraestrutura e Biologia Tecidual. Rio de Janeiro, RJ, Brasil.Universidade Estadual do Norte Fluminense. Laboratório de Biotecnologia. Campos dos Goytacazes, RJ, Brasil.Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Laboratório Interdisciplinar de Pesquisas Médicas. Rio de Janeiro, RJ. Brasil.In the large Zika virus (ZIKV) epidemic that occurred in Brazil in 2015, the intrauterine fetal exposure to ZIKV was associated with a significant risk of developing microcephaly and neurological disorders in the infected infants. ZIKV-associated disease has since been reported in 24 countries in the Americas. At present, definitive evidence is lacking regarding the intrauterine co-exposure to ZIKV and other viral infections and whether the coinfection impacts the risk of acquiring either infection or disease severity. Here, we provide evidence of intrauterine exposure to both ZIKV and human immunodeficiency virus (HIV) infections, causing congenital Zika syndrome in an HIV-exposed uninfected infant. Clinical, imaging and laboratory examinations of the pregnant woman and the newborn were performed. Histopathology, ZIKV/HIV-specific immunoassays, and ultrastructural evaluation of the placenta were performed. The Zika-asymptomatic, HIV-positive pregnant woman underwent ultrasounds revealing fetal cerebral ventriculomegaly, microcephaly, and brain atrophy. Her baby girl was born small for gestational age and with the neurological sequelae of congenital Zika syndrome. The evaluation of the abnormally large term placenta revealed severe damage to the maternal decidua and chorionic villi, cells positive for ZIKV-specific antigens but not for HIV antigens, and intracellular membranous clusters of virus-like particles approximately 25 nm in diameter. The rapid progression and severity of the congenital Zika syndrome may be related to the uncontrolled HIV disease in the mother. The poor inflammatory response observed in the placenta may have reduced the inherent risk of mother-to-child transmission of HIV

Detection of ZIKV RNA and viral load quantification from C6/36 and Vero cell supernatants at 24, 48 and 72 hr p.i. by quantitative real-time RT-PCR.

<p>Detection of ZIKV RNA and viral load quantification from C6/36 and Vero cell supernatants at 24, 48 and 72 hr p.i. by quantitative real-time RT-PCR.</p

Uninfected C6/36 cells (negative controls) at different time points in culture.

<p><b>No cellular ultrastructural alterations were observed.</b> (A–B) 24 hr of culture; (C) 48 hr of culture; (D) 72 hr of culture. Rough endoplasmic reticulum cisternae (rER), nucleus (N).</p

Immunolocalization of ZIKV E protein (4G2, green) [arrow] inside cytoplasm of C6/36 cells infected with ZIKV at different time points post infection.

<p>(A) Uninfected cells; (B) 24 hr p.i.;(C/D) 48 hr p. i.; (E) 72 hr p.i. The nuclei (N) were counterstained with DAPI (blue). (F) The graph represents the mean ± standard deviation of the infection percentage (%). The highest number of infected cells was observed at 72 hr p.i.</p

Immunolocalization of ZIKV E protein (4G2, green) [arrow] in the cytoplasm of Vero cells infected with ZIKV at different time points post-infection.

<p>(A) Uninfected cells; (B) 24 hr p.i; (C) 48 hr p.i.; (D) 72 hr p.i. The nuclei were counterstained with DAPI (blue). (E) The graph represents the mean ± standard deviation of the infection percentage (%). A higher number of infected cells was observed at 24 hr p.i.</p