An epidemic Zika virus isolate suppresses antiviral immunity by disrupting antigen presentation pathways

Zika virus (ZIKV) has emerged as an important global health threat, with the recently acquired capacity to cause severe neurological symptoms and to persist within host tissues. We previously demonstrated that an early Asian lineage ZIKV isolate induces a highly activated CD8 T cell response specific for an immunodominant epitope in the ZIKV envelope protein in wild-type mice. Here we show that a contemporary ZIKV isolate from the Brazilian outbreak severely limits CD8 T cell immunity in mice and blocks generation of the immunodominant CD8 T cell response. This is associated with a more sustained infection that is cleared between 7- and 14-days post-infection. Mechanistically, we demonstrate that infection with the Brazilian ZIKV isolate reduces the cross-presentation capacity of dendritic cells and fails to fully activate the immunoproteasome. Thus, our study provides an isolate-specific mechanism of host immune evasion by one Brazilian ZIKV isolate, which differs from the early Asian lineage isolate and provides potential insight into viral persistence associated with recent ZIKV outbreaks

Cyclophilin D Regulates Antiviral CD8+ T Cell Survival in a Cell-Extrinsic Manner

CD8+ T cell–mediated immunity is critical for host defense against viruses and requires mitochondria-mediated type I IFN (IFN-I) signaling for optimal protection. Cyclophilin D (CypD) is a mitochondrial matrix protein that modulates the mitochondrial permeability transition pore, but its role in IFN-I signaling and CD8+ T cell responses to viral infection has not been previously explored. In this study, we demonstrate that CypD plays a critical extrinsic role in the survival of Ag-specific CD8+ T cell following acute viral infection with lymphocytic choriomeningitis virus in mice. CypD deficiency resulted in reduced IFN-I and increased CD8+ T cell death, resulting in a reduced antiviral CD8+ T cell response. In addition, CypD deficiency was associated with an increase in pathogen burden at an early time-point following infection. Furthermore, our data demonstrate that transfer of wild-type macrophages (expressing CypD) to CypD-deficient mice can partially restore CD8+ T cell responses. These results establish that CypD plays an extrinsic role in regulating optimal effector CD8+ T cell responses to viral infection. Furthermore, this suggests that, under certain circumstances, inhibition of CypD function may have a detrimental impact on the host’s ability to respond to viral infection

Protective to a T: The Role of T Cells during Zika Virus Infection

CD4 and CD8 T cells are an important part of the host’s capacity to defend itself against viral infections. During flavivirus infections, T cells have been implicated in both protective and pathogenic responses. Given the recent emergence of Zika virus (ZIKV) as a prominent global health threat, the question remains as to how T cells contribute to anti-ZIKV immunity. Furthermore, high homology between ZIKV and other, co-circulating flaviviruses opens the possibility of positive or negative effects of cross-reactivity due to pre-existing immunity. In this review, we will discuss the CD4 and CD8 T cell responses to ZIKV, and the lessons we have learned from both mouse and human infections. In addition, we will consider the possibility of whether T cells, in the context of flavivirus-naïve and flavivirus-immune subjects, play a role in promoting ZIKV pathogenesis during infection

Zika Virus Pathogenesis: From Early Case Reports to Epidemics

For the first 60 years following its isolation, Zika virus (ZIKV) remained a relatively poorly described member of the Flaviviridae family. However, since 2007, it has caused a series of increasingly severe outbreaks and is now associated with neurological symptoms such as Guillain-Barré syndrome and congenital Zika syndrome (CZS). A number of reports have improved our understanding of rare complications that may be associated with ZIKV infection in adults, the areas of the body to which it spreads, and viral persistence in various tissues. Likewise, studies on the effect of ZIKV infection during pregnancy have identified risk factors for CZS and the impact this syndrome has on early childhood. Understanding these outcomes and the factors that drive ZIKV pathogenesis are key to developing vaccination and therapeutic approaches to avoid these severe and potentially debilitating symptoms

Analysis of the T Cell Response to Zika Virus and Identification of a Novel CD8⁺ T Cell Epitope in Immunocompetent Mice

<div><p>Zika virus (ZIKV) is an emerging arbovirus of the <i>Flaviviridae</i> family. Although ZIKV infection is typically mild and self-limiting in healthy adults, infection has been associated with neurological symptoms such as Guillain-Barré syndrome, and a causal link has been established between fetal microcephaly and ZIKV infection during pregnancy. These risks, and the magnitude of the ongoing ZIKV pandemic, have created an urgent need for the development of animal models to study the immune response to ZIKV infection. Previous animal models have primarily focused on pathogenesis in immunocompromised mice. In this study, we provide a model of ZIKV infection in wild-type immunocompetent C57BL/6 mice, and have provided an analysis of the immune response to infection. We evaluated the activation of several innate immune cell types, and studied the kinetics, phenotype, and functionality of T cell responses to ZIKV infection. Our results demonstrate that ZIKV infection is mild in wild-type immunocompetent C57BL/6 mice, resulting in minimal morbidity. Our data establish that at the peak of the adaptive response, antigen-experienced CD4⁺ T cells polarize to a Th1 phenotype, and antigen-experienced CD8⁺ T cells exhibit an activated effector phenotype, producing both effector cytokines and cytolytic molecules. Furthermore, we have identified a novel ZIKV CD8⁺ T cell epitope in the envelope protein that is recognized by the majority of responding cells. Our model provides an important reference point that will help dissect the impact of polymorphisms in the circulating ZIKV strains on the immune response and ZIKV pathogenesis. In addition, the identification of a ZIKV epitope will allow for the design of tetramers to study epitope-specific T cell responses, and will have important implications for the design and development of ZIKV vaccine strategies.</p></div

ZIKV infection kinetics.

<p><b>(A)</b> Viral RNA was quantified in the spleens of C57BL/6 mice infected i.v. with 10⁶ PFU of ZIKV at 6, 12, 24, 48 and 72 h post-infection using qRT-PCR analysis. <b>(B)</b> Viral RNA was quantified in the spleens of C57BL/6 mice infected i.v. with 10⁶ PFU of ZIKV or an equivalent dose of UV-inactivated virus at 12 h post-infection using qRT-PCR analysis. Data are presented as plaque forming unit (PFU) equivalents per gram of tissue after normalization to a standard curve (<a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1006184#ppat.1006184.s002" target="_blank">S2 Fig</a>). The dotted line indicates limit of detection based on the average number of PFU equivalents per gram of tissue from the spleens of mock-infected mice at 48 h post-infection <b>(A)</b> or 12 h post-infection <b>(B)</b>. Error bars represent mean ± SEM. Data are pooled from two independent experiments, n = 3 mice per group per experiment.</p

ZIKV infection induces a prototypical effector CD8⁺ T cell response in immunocompetent mice.

<p>Representative plots of IFN-γ <b>(A)</b>, TNF-α <b>(B)</b>, and IL-2 <b>(C)</b> production from splenic CD8α^loCD11a^hi CD8⁺ T cells from ZIKV-infected mice 7 dpi. Total splenocytes were incubated in media alone in the presence of Brefeldin A (top) or stimulated with PMA and ionomycin in the presence of Brefeldin A for 3 h at 37°C (bottom). Percentage <b>(D)</b>, number <b>(E)</b>, and gMFI <b>(F)</b> of cytokine-producing CD8α^loCD11a^hi CD8⁺ T cells from the spleens of ZIKV-infected mice 7 dpi. <b>(G)</b> Representative histograms of T-bet expression in splenic CD8α^loCD11a^hi CD8⁺ T cells from mock- and ZIKV-infected mice 7 dpi. Shaded histograms represent isotype control. Percentage <b>(H)</b> and gMFI <b>(I)</b> of T-bet expression in splenic CD8α^loCD11a^hi CD8⁺ T cells from mock- and ZIKV-infected mice 7 dpi. Error bars represent mean ± SEM. Data are pooled from two independent experiments, n = 3–5 mice per group per experiment. Data in <b>(H</b> and <b>I)</b> were analyzed with a two-tailed, unpaired Student’s t test. ****p<0.0001.</p

ZIKV induces innate immune cell activation.

<p>Representative histograms and geometric mean fluorescence intensity (gMFI) of CD40 <b>(A</b>-<b>B)</b>, CD80 <b>(C</b>-<b>D)</b>, and CD86 <b>(E-F)</b> on splenic dendritic cells (DCs) at 2 dpi with 10⁶ PFU of ZIKV i.v. (open histograms with solid line), an equivalent dose of UV-inactivated ZIKV (open histograms with dashed line) or mock infection (shaded histograms). Histograms represent DCs gated as CD3^- CD19^- NK1.1^- MHC-II⁺ CD11c⁺ cells. Representative histogram <b>(G)</b>, percentage <b>(H)</b>, and number <b>(I)</b> of CD69⁺ NK1.1⁺CD3^- natural killer cells from the spleens of ZIKV- (open histogram with solid line), UV-inactivated ZIKV- (open histogram with dashed line) and mock-infected (shaded histogram) mice 2 dpi. Number on histogram indicates percentage of CD69⁺ cells from ZIKV-infected mouse. <b>(J)</b> Mice were infected with 10⁶ PFU of ZIKV or an equivalent dose of UV-inactivated virus, spleens were harvested 12 h post-infection and total RNA was extracted. IFN-α and IFN-β mRNA expression was assessed by qRT-PCR, normalized to TATA-binding protein mRNA expression, and expressed as fold change over mRNA expression in mock-infected mice. Error bars represent mean ± SEM. Data in (<b>A</b>-<b>I</b>) are pooled from two independent experiments, n = 3 mice per group per experiment. Data in (<b>J</b>) are pooled from three independent experiments, n = 3 mice per group per experiment. Data in <b>(B</b>, <b>D</b>, <b>F</b>, <b>H</b> and <b>I)</b> were analyzed by one-way ANOVA with Tukey’s post-test of multiple comparisons. ****p<0.0001.</p

ZIKV infection induces a Th1 CD4⁺ T cell response.

<p>Representative plots of IFN-γ <b>(A)</b>, TNF-α <b>(B)</b>, and IL-2 <b>(C)</b> production from splenic CD11a⁺CD49d⁺ CD4⁺ T cells from ZIKV-infected mice 7 dpi. Total splenocytes were incubated in media alone in the presence of Brefeldin A (top) or stimulated with PMA and ionomycin in the presence of Brefeldin A for 3 h at 37°C (bottom). Percentage <b>(D)</b>, number <b>(E)</b>, and gMFI <b>(F)</b> of cytokine-producing CD11a⁺CD49d⁺ CD4⁺ T cells from the spleens of ZIKV-infected mice 7 dpi. <b>(G)</b> Representative histograms of T-bet expression in splenic CD11a⁺CD49d⁺ CD4⁺ T cells from mock- and ZIKV-infected mice 7 dpi. Shaded histograms represent isotype control. Percentage <b>(H)</b> and gMFI <b>(I)</b> of T-bet expression in splenic CD11a⁺CD49d⁺ CD4⁺ T cells from mock- and ZIKV-infected mice 7 dpi. Error bars represent mean ± SEM. Data are pooled from two independent experiments, n = 3–5 mice per group per experiment. Data in <b>(H</b> and <b>I)</b> were analyzed with a two-tailed, unpaired Student’s t test. ****p<0.0001.</p