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ACD can configure probes for the various manual and automated assays for INS for RNAscope Assay, or for Basescope Assay compatible for your species of interest.

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Zika virus infects human testicular tissue and germ cells

J Clin Invest.

2018 Jul 31

Matusali G, Houzet L, Satie AP, Mahé D, Aubry F, Couderc T, Frouard J, Bourgeau S, Bensalah K, Lavoué S, Joguet G, Bujan L, Cabié A, Avelar GF, Lecuit M, Le Tortorec A, Dejucq-Rainsford N.
PMID: 30063220 | DOI: 10.1172/JCI121735

Zika virus (ZIKV) is a teratogenic mosquito-borne flavivirus which can be sexually transmitted from man to woman. High viral loads and prolonged viral shedding in semen suggest that ZIKV replicates within the human male genital tract, but its target organs are unknown. Using ex vivo infection of organotypic cultures, we demonstrated here that ZIKV replicates in human testicular tissue and infects a broad range of cell types, including germ cells, which we also identified as infected in the semen from ZIKV-infected donors. ZIKV had no major deleterious effect on the morphology and hormonal production of the human testis explants. Infection induced a broad antiviral response but no interferon up-regulation and minimal pro-inflammatory response in testis explants, with no cytopathic effect. Finally, we studied ZIKV infection in mouse testis, and compared it to human infection. This study provides key insights into how ZIKV may persist in semen and alter semen parameters, as well as a valuable tool for testing antiviral agents.

Forebrain neural precursor cells are differentially vulnerable to Zika virus infection

eNeuro

2021 Jul 16

Shelton, SM;Soucy, AR;Kurzion, R;Zeldich, E;Connor, JH;Haydar, TF;
PMID: 34272257 | DOI: 10.1523/ENEURO.0108-21.2021

Prenatal exposure to Zika virus (ZIKV) can result in microencephaly and congenital Zika syndrome, though some brain cells and structures are spared by the virus for unknown reasons. Here, a novel murine model of fetal ZIKV infection incorporating intraventricular infection and cell type specific in utero electroporation was used to identify the time course of ZIKV infection and to determine the identity of cells that are initially infected or spared during neocortical neurogenesis. In vivo time course studies revealed the presence of ZIKV in apical radial glial cells (aRGCs) at early time points following virus exposure, while basal intermediate progenitor cells (bIPCs) became maximally (ZIKV+) after 3 days of virus exposure. ZIKV-infected fetal brains exhibited microencephaly as early as one day following infection, regardless of developmental age. This change in brain size was caused in part by apoptosis and reduced proliferation that persisted until birth. While 60% of aRGC basal fibers were perturbed during infection, 40% retained normal morphology, indicating that aRGCs are not uniformly vulnerable to ZIKV infection. To investigate this heterogeneous vulnerability, we performed genetic fate mapping using cell type-specific probes derived from a mouse E15.5 neocortical wall single cell RNA-Seq dataset. The results indicate that one class of aRGCs preferentially express the putative ZIKV entry receptor AXL, and that these cells are more vulnerable to ZIKV infection than other aRGC subtypes with low AXL expression. Together, these data uncover crucial temporal and cellular details of ZIKV fetal brain infection for prevention strategies and for management of congenital Zika syndrome.Significance StatementThe transcriptional signatures of neural precursor cells were utilized for the first time to test Zika virus susceptibility in a direct fetal brain infection model. This novel methodology allowed for elucidation of time point specific differences in neural precursor cell susceptibility that have been debated in the field. Additionally, elucidation of cell morphological features using in utero electroporation revealed substantial but incomplete interruption of basal fibers, a finding that implies interference with neuronal migration. The model presented here, allows for assessment of pre-natal development after exposure to a variety of viruses. The improved specificity of apical radial glial cell labeling afforded by the cell-specific labeling tools uncover functional differences between apical radial glial cell types that will have important implications for children exposed to ZIKV as well as for understanding corticogenesis.
Timing of Gestational Exposure to Zika Virus is Associated with Postnatal Growth Restriction in a Murine Model.

Am J Obstet Gynecol.

2018 Jun 11

Valentine GC, Seferovic MD, Fowler SW, Major AM, Gorchakov R, Berry R, Swennes AG, Murray KO, Suter MA, Aagaard KM.
PMID: 29902449 | DOI: 10.1016/j.ajog.2018.06.005

Abstract

BACKGROUND:

Vertical transmission of Zika virus (ZIKV) leads to infection of neuroprogenitor cells and destruction of brain parenchyma. Recent evidence suggests that the timing of infection as well as host factors may affect vertical transmission. As a result, congenital ZIKV infection may only become clinically apparent in the postnatal period.

OBJECTIVES:

We sought to develop an outbred mouse model of ZIKV vertical transmission to determine if the timing of gestational ZIKV exposure yields phenotypic differences at birth and through adolescence. We hypothesized that later gestational inoculations would only become apparent in adolescence.

METHODS:

To better recapitulate human exposures, timed pregnant Swiss-Webster dams (n=15) were subcutaneously inoculated with 1x104PFU of first passage contemporary ZIKV HN16 strain or a mock injection on embryonic day 4, 8, or 12 with bioactive anti-interferon alpha receptor antibody administered in days preceding and proceeding inoculation. The antibody was given to prevent the robust type I interferon signaling cascade that make mice inherently resistant to ZIKV infection. At birth and adolescence (6 weeks of age) offspring were assessed for growth, brain weight and biparietal head diameters (BPD), and ZIKV viral levels by RT-PCR or in situ hybridization.

RESULTS:

Pups of ZIKV-infected dams infected at e4 and e8 but not e12 were growth restricted (p<0.003). Brain weights were significantly smaller at birth (p=0.01) for e8 ZIKV-exposed offspring. At 6 weeks of age, biparietal diameters (BPD) were smaller for all ZIKV exposed males and females (p<0.05), with e8 exposed males smallest by BPD and growth restriction measurements (weight >2 SD, p=0.0007). All pups and adolescent mice were assessed for ZIKV infection by RT-PCR. Analysis of all underweight pups reveled one to be positive for neuronal ZIKV infection by in situ hybridization, while a second moribund animal was diffusely positive at 8 days of age by ZIKV infectivity throughout the brain, kidneys and intestine.

CONCLUSION:

These findings demonstrate that postnatal effects of infection occurring at single time points continue to be detrimental to offspring in the postnatal period in a subset of littermates and subject to a window of gestational susceptibility coinciding with placentation. This model recapitulates frequently encountered clinical scenarios in non-endemic regions, including the majority of the U.S., where travel related exposure occurs in short and well-defined windows of gestation. Our low rate of infection and relatively rare evidence of congenital Zika syndrome parallels human population-based data.

Experimental Zika Virus Infection in the Pregnant Common Marmoset Induces Spontaneous Fetal Loss and Neurodevelopmental Abnormalities.

Sci Rep.

2018 May 01

Seferovic M, Martín CS, Tardif SD, Rutherford J, Castro ECC, Li T, Hodara VL, Parodi LM, Giavedoni L, Layne-Colon D, Tamhankar M, Yagi S, Martyn C, Reyes K, Suter MA, Aagaard KM, Chiu CY, Patterson JL.
PMID: 29717225 | DOI: 10.1038/s41598-018-25205-1

During its most recent outbreak across the Americas, Zika virus (ZIKV) was surprisingly shown to cause fetal loss and congenital malformations in acutely and chronically infected pregnant women. However, understanding the underlying pathogenesis of ZIKV congenital disease has been hampered by a lack of relevant in vivo experimental models. Here we present a candidate New World monkey model of ZIKV infection in pregnant marmosets that faithfully recapitulates human disease. ZIKV inoculation at the human-equivalent of early gestation caused an asymptomatic seroconversion, induction of type I/II interferon-associated genes and proinflammatory cytokines, and persistent viremia and viruria. Spontaneous pregnancy loss was observed 16-18 days post-infection, with extensive active placental viral replication and fetal neurocellular disorganization similar to that seen in humans. These findings underscore the key role of the placenta as a conduit for fetal infection, and demonstrate the utility of marmosets as a highly relevant model for studying congenital ZIKV disease and pregnancy loss.

Zika virus has oncolytic activity against glioblastoma stem cells

Journal of Experimental Medicine

2017 Sep 05

Zhu Z, Gorman MJ, McKenzie LD, Chai JN, Hubert CG, Prager BC, Fernandez E, Richner JM, Zhang R, Shan C, Wang X, Shi PY, Diamond MS, Rich JN, Chheda MG.
PMID: 28874392 | DOI: 10.1084/jem.20171093

Glioblastoma is a highly lethal brain cancer that frequently recurs in proximity to the original resection cavity. We explored the use of oncolytic virus therapy against glioblastoma with Zika virus (ZIKV), a flavivirus that induces cell death and differentiation of neural precursor cells in the developing fetus. ZIKV preferentially infected and killed glioblastoma stem cells (GSCs) relative to differentiated tumor progeny or normal neuronal cells. The effects against GSCs were not a general property of neurotropic flaviviruses, as West Nile virus indiscriminately killed both tumor and normal neural cells. ZIKV potently depleted patient-derived GSCs grown in culture and in organoids. Moreover, mice with glioblastoma survived substantially longer and at greater rates when the tumor was inoculated with a mouse-adapted strain of ZIKV. Our results suggest that ZIKV is an oncolytic virus that can preferentially target GSCs; thus, genetically modified strains that further optimize safety could have therapeutic efficacy for adult glioblastoma patients.

Lineage-dependent differences in the disease progression of Zika virus infection in type-I interferon receptor knockout (A129) mice

PLoS Negl Trop Dis.

2017 Jul 03

Dowall SD, Graham VA, Rayner E, Hunter L, Atkinson B, Pearson G, Dennis M, Hewson R.
PMID: 28672028 | DOI: 10.1371/journal.pntd.0005704

Zika virus (ZIKV) falls into two lineages: African (ZIKVAF) and Asian (ZIKVAS). These lineages have not been tested comprehensively in parallel for disease progression using an animal model system. Here, using the established type-I interferon receptor knockout (A129) mouse model, it is first demonstrated that ZIKVAF causes lethal infection, with different kinetics of disease manifestations according to the challenge dose. Animals challenged with a low dose of 10 plaque-forming units (pfu) developed more neurological symptoms than those challenged with 5-log higher doses. By contrast, animals challenged with ZIKVAS displayed no clinical signs or mortality, even at doses of 106 pfu. However, viral RNA was detected in the tissues of animals infected with ZIKV strains from both lineages and similar histological changes were observed. The present study highlights strain specific virulence differences between the African and Asian lineages in a ZIKV mouse model.

Neutralizing human antibodies prevent Zika virus replication and fetal disease in mice.

Nature.

2016 Nov 07

Sapparapu G, Fernandez E, Kose N, Cao B, Fox JM, Bombardi RG, Zhao H, Nelson CA, Bryan AL, Barnes T, Davidson E, Mysorekar IU, Fremont DH, Doranz BJ, Diamond MS, Crowe JE.
PMID: 27819683 | DOI: 10.1038/nature20564

Zika virus (ZIKV) is an emerging mosquito-transmitted flavivirus that can cause severe disease, including congenital birth defects during pregnancy1. To develop candidate therapeutic agents against ZIKV, we isolated a panel of human monoclonal antibodies (mAbs) from subjects with prior ZIKV infection. A subset of mAbs recognized diverse epitopes on the envelope (E) protein and exhibited potently neutralizing activity. One of the most inhibitory mAbs, ZIKV-117, broadly neutralized infection of ZIKV strains corresponding to African, Asian, and American lineages. Epitope mapping studies revealed that ZIKV-117 recognized a unique quaternary epitope on the E protein dimer-dimer interface. We evaluated the therapeutic efficacy of ZIKV-117 in pregnant and non-pregnant mice. mAb treatment markedly reduced tissue pathology, placental and fetal infection, and mortality in mice. Thus, neutralizing human mAbs can protect against maternal-fetal transmission, infection and disease, and reveal important determinants for structure-based rational vaccine design efforts.

Neuropathogenesis of Zika Virus in a Highly Susceptible Immunocompetent Mouse Model after Antibody Blockade of Type I Interferon

PLoS Negl Trop Dis.

2017 Jan 09

Smith DR, Hollidge B, Daye S, Zeng X, Blancett C, Kuszpit K, Bocan T, Koehler JW, Coyne S, Minogue T, Kenny T, Chi X, Yim S, Miller L, Schmaljohn C, Bavari S, Golden JW.
PMID: 28068342 | DOI: 10.1371/journal.pntd.0005296

Animal models are needed to better understand the pathogenic mechanisms of Zika virus (ZIKV) and to evaluate candidate medical countermeasures. Adult mice infected with ZIKV develop a transient viremia, but do not demonstrate signs of morbidity or mortality. Mice deficient in type I or a combination of type I and type II interferon (IFN) responses are highly susceptible to ZIKV infection; however, the absence of a competent immune system limits their usefulness for studying medical countermeasures. Here we employ a murine model for ZIKV using wild-type C57BL/6 mice treated with an antibody to disrupt type I IFN signaling to study ZIKV pathogenesis. We observed 40% mortality in antibody treated mice exposed to ZIKV subcutaneously whereas mice exposed by intraperitoneal inoculation were highlysusceptible incurring 100% mortality. Mice infected by both exposure routes experienced weight loss, high viremia, and severe neuropathologic changes. The most significant histopathological findings occurred in the central nervous system where lesions represent an acute to subacute encephalitis/encephalomyelitis that is characterized by neuronal death, astrogliosis, microgliosis, scattered necrotic cellular debris, and inflammatory cell infiltrates. This model of ZIKV pathogenesis will be valuable for evaluating medical countermeasures and the pathogenic mechanisms of ZIKV because it allows immune responses to be elicited in immunologically competent mice with IFN I blockade only induced at the time of infection.

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Description
sense
Example: Hs-LAG3-sense
Standard probes for RNA detection are in antisense. Sense probe is reverse complent to the corresponding antisense probe.
Intron#
Example: Mm-Htt-intron2
Probe targets the indicated intron in the target gene, commonly used for pre-mRNA detection
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Example: Hs-CD3-pool (Hs-CD3D, Hs-CD3E, Hs-CD3G)
A mixture of multiple probe sets targeting multiple genes or transcripts
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Example: Hs-PDGFB-No-XMm
Does not cross detect with the species (Sp)
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Example: Rn-Pde9a-XMm
designed to cross detect with the species (Sp)
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Example: Mm-Islr-O1
Alternative design targeting different regions of the same transcript or isoforms
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Example: Hs-SLC31A-CDS
Probe targets the protein-coding sequence only
EnEmProbe targets exons n and m
En-EmProbe targets region from exon n to exon m
Retired Nomenclature
tvn
Example: Hs-LEPR-tv1
Designed to target transcript variant n
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Example: Hs-ACVRL1-ORF
Probe targets open reading frame
UTR
Example: Hs-HTT-UTR-C3
Probe targets the untranslated region (non-protein-coding region) only
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Example: Hs-GNRHR-5UTR
Probe targets the 5' untranslated region only
3UTR
Example: Rn-Npy1r-3UTR
Probe targets the 3' untranslated region only
Pan
Example: Pool
A mixture of multiple probe sets targeting multiple genes or transcripts

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