Acute Zika Virus Infection Research Articles

Comprehensive analysis of early T cell responses to acute Zika Virus infection during the first epidemic in Bahia, Brazil

Background In most cases, Zika virus (ZIKV) causes a self-limited acute illness in adults, characterized by mild clinical symptoms that resolve within a few days. Immune responses, both innate and adaptive, play a central role in controlling and eliminating virus-infected cells during the early stages of infection. Aim To test the hypothesis that circulating T cells exhibit phenotypic and functional activation characteristics during the viremic phase of ZIKV infection. Methods A comprehensive analysis using mass cytometry was performed on peripheral blood mononuclear cells obtained from patients with acute ZIKV infection (as confirmed by RT-PCR) and compared with that from healthy donors (HD). The frequency of IFN-γ-producing T cells in response to peptide pools covering immunogenic regions of structural and nonstructural ZIKV proteins was quantified using an ELISpot assay. Results Circulating CD4+ and CD8+ T lymphocytes from ZIKV-infected patients expressed higher levels of IFN-γ and pSTAT-5, as well as cell surface markers associated with proliferation (Ki-67), activation ((HLA-DR, CD38) or exhaustion (PD1 and CTLA-4), compared to those from HD. Activation of CD4+ and CD8+ memory T cell subsets, including Transitional Memory T Cells (TTM), Effector Memory T cells (TEM), and Effector Memory T cells Re-expressing CD45RA (TEMRA), was prominent among CD4+ T cell subset of ZIKV-infected patients and was associated with increased levels of IFN-γ, pSTAT-5, Ki-67, CTLA-4, and PD1, as compared to HD. Additionally, approximately 30% of ZIKV-infected patients exhibited a T cell response primarily directed against the ZIKV NS5 protein. Conclusion Circulating T lymphocytes spontaneously produce IFN-γ and express elevated levels of pSTAT-5 during the early phase of ZIKV infection whereas recognition of ZIKV antigen results in the generation of virus-specific IFN-γ-producing T cells.

Prior dengue virus serotype 3 infection modulates subsequent plasmablast responses to Zika virus infection in rhesus macaques.

The Zika virus epidemic of 2015-2016 in the Americas revealed that this mosquito-transmitted virus could be congenitally transmitted during pregnancy and cause birth defects in newborns. Currently, there are no interventions to mitigate this disease and Zika virus is likely to re-emerge. Understanding how protective antibody responses are generated against Zika virus can help in the development of a safe and effective vaccine. One main challenge is that Zika virus co-circulates with related viruses like dengue, such that prior exposure to one can generate cross-reactive antibodies against the other which may enhance infection and disease from the second virus. In this study, we sought to understand how prior dengue virus infection impacts subsequent immunity to Zika virus by single-cell sequencing of antibody producing cells in a second Zika virus infection. Identifying specific qualities of Zika virus immunity that are modulated by prior dengue virus immunity will enable optimal immunization strategies.

Development of a quantitative NS1 antigen enzyme-linked immunosorbent assay (ELISA) for Zika virus detection using a novel virus-specific mAb

Viruses from the Flaviviridae family, such as Dengue virus (DENV), Yellow fever virus (YFV), and Zika virus (ZIKV) are notorious global public health problems. ZIKV emergence in Polynesia and the Americas from 2013 to 2016 raised concerns as new distinguishing features set it apart from previous outbreaks, including its association with neurological complications and heightened disease severity. Virus detection is impaired as cross-reactivity to other closely related orthoflaviviruses is common among commercially available diagnostic kits. While non-structural protein 1 (NS1) has been used as an early marker of DENV and West Nile virus (WNV) infection, little is known about NS1 expression during ZIKV infection. In the present work, we developed a NS1 capture ELISA using a novel ZIKV-specific monoclonal antibody to study NS1 expression dynamics in vitro in mosquito and human cell lines. While detectable in culture supernatants, higher concentrations of NS1 were predominantly cell-associated. To our knowledge, this is the first report of NS1 detection in human cells despite viral clearance over time. Tests with human samples need to be conducted to validate the applicability of NS1 detection for diagnosis, but overall, the tools developed in this work are promising for specific detection of acute ZIKV infection.

Does the Presence or a High Titer of Yellow Fever Virus Antibodies Interfere with Pregnancy Outcomes in Women with Zika Virus Infection?

Zika virus (ZIKV) and yellow fever virus (YFV) originated in Africa and expanded to the Americas, where both are co-circulated. It is hypothesized that in areas of high circulation and vaccination coverage against YFV, children of pregnant women have a lower risk of microcephaly. We evaluated the presence and titers of antibodies and outcomes in women who had ZIKV infection during pregnancy. Pregnancy outcomes were classified as severe, moderate, and without any important outcome. An outcome was defined as severe if miscarriage, stillbirth, or microcephaly occurred, and moderate if low birth weight and/or preterm delivery occurred. If none of these events were identified, the pregnancy was defined as having no adverse effects. A sample of 172 pregnant women with an acute ZIKV infection confirmed during pregnancy were collected throughout 2016. About 89% (150 of 169) of them presented immunity against YFV, including 100% (09 of 09) of those who had severe outcomes, 84% (16 of 19) of those who had moderate outcomes, and 89% (125 of 141) of those who had non-outcomes. There was no difference between groups regarding the presence of anti-YFV antibodies (p = 0.65) and YFV titers (p = 0.6). We were unable to demonstrate a protective association between the presence or titers of YFV antibodies and protection against serious adverse outcomes from exposure to ZIKV in utero.

Cryptic Zika virus infections unmasked from suspected malaria cases in Northeastern Nigeria

Introduction Although environmental and human behavioral factors in countries with Zika virus (ZIKV) outbreaks are also common in Nigeria, such an outbreak has not yet been reported probably due to misdiagnosis. The atypical symptoms of malaria and ZIKV infections at the initial phase could leverage their misdiagnosis. This study randomly recruited 496 malaria-suspected patients who visited selected health institutions in Adamawa, Bauchi, and Borno states for malaria tests. These patients’ sera were analyzed for ZIKV antibodies using ELISA and plaque reduction neutralization tests (PRNT) at 90% endpoint. About 13.8% of Zika virus-neutralizing antibodies (nAb) did not cross-react with dengue, yellow fever, and West Nile viruses suggesting possible monotypic infections. However, 86% of the sera with ZIKV nAb also neutralized other related viruses at varied degrees: dengue viruses (60.7%), West Nile viruses (23.2%), yellow fever virus (7.1%) and 39.3% were co-infections with chikungunya viruses. Notably, the cross-reactions could also reflect co-infections as these viruses are also endemic in the country. The serum dilution that neutralized 90–100% ZIKV infectivity ranged from 1:8 to 1:128. Also, our findings suggest distinct protection against the ZIKV between different collection sites studied. As indicated by nAb, acute ZIKV infection was detected in 1.7% of IgM-positive patients while past infections occurred in 8.5% of IgM-negatives in the three states. In Borno State, 9.4% of IgG neutralized ZIKV denoting past infections while 13.5% were non-neutralizing IgM and IgG indicating other related virus infections. The age, gender, and occupation of the patients and ZIKV nAb were not significantly different. ZIKV nAb from samples collected within 1–7 days after the onset of symptoms was not significantly different from those of 7–10 days. A wider interval with the same techniques in this study may probably give better diagnostic outcomes. ZIKV nAb was significantly distinct among recipients and non-recipients of antibiotic/antimalaria treatments before seeking malaria tests. The inhibiting effect of these drugs on ZIKV infection progression may probably contribute to the absence of neurological disorders associated with the virus despite being endemic in the environment for several decades. Also, protection against ZIKV as marked by the nAb was different among the vaccinated and unvaccinated YF vaccine recipients. Thus, the YF vaccine may be a good alternative to the Zika vaccine in resource-constrained countries. Conclusion The cryptic ZIKV infections underscore the need for differential diagnosis of malaria-suspected febrile patients for arboviruses, especially the Zika virus. The absence of systemic surveillance for the virus is worrisome because of its association with neurological disorders in newborns. Co-infections with other arboviruses may impact adversely on the management of these diseases individually.

Neonatal Zika virus infection causes transient perineuronal net degradation.

Perineuronal nets (PNNs) form a specialized extracellular matrix that predominantly surrounds parvalbumin (PV)-expressing GABAergic inhibitory interneurons and help regulate neuronal activity. Their formation early in the postnatal period is regulated by neuronal signaling and glial activation raising concerns that part of the long-term effects ascribed to perinatal viral infections could be mediated by altered PNN formation. Previously, we developed a model of neonatal Zika virus (ZIKV) infection where mice have lifelong neurological sequelae that includes motor disfunction and reduced anxiety coupled with a persistent low-grade expression in proinflammatory markers despite resolving the acute infection. Here, we demonstrate that ZIKV infection to P1 neonatal mice results in a reduction of PNN formation during the acute disease with significant reduction in Wisteria floribunda agglutinin (WFA) staining at the peak of infection [15 days post infection (dpi)] that persisted after the symptoms resolved (30 dpi). At 60 dpi, when there is residual inflammation in the CNS, the number of WFA+ cells and the level of WFA staining as well as levels of aggrecan and brevican in the brains of convalescent mice were not different from those in uninfected controls, however, there was increased frequency of PNNs with an immature phenotype. Over time the impact of the perinatal infection became less evident and there were no clear differences in PNN morphology between the groups at 1 year post infection. Of note, the reduction in PNNs during acute ZIKV infection was not associated with decreased mRNA levels of aggrecan or brevican, but increased levels of degraded aggrecan and brevican indicating increased PNN degradation. These changes were associated with increased expression of matrix metalloproteinase 12 (MMP12) and MMP19, but not MMP9, a disintegrin and metalloproteinase with thrombospondin motifs 4 (ADAMTS4) or ADAMTS5. Together our findings indicate that infection at the time of PNN development interferes with PNN formation, but the nets can reform once the infection and inflammation subside.

Analyses of Early ZIKV Genomes Are Consistent with Viral Spread from Northeast Brazil to the Americas.

The Americas, particularly Brazil, were greatly impacted by the widespread Zika virus (ZIKV) outbreak in 2015 and 2016. Efforts were made to implement genomic surveillance of ZIKV as part of the public health responses. The accuracy of spatiotemporal reconstructions of the epidemic spread relies on the unbiased sampling of the transmission process. In the early stages of the outbreak, we recruited patients exhibiting clinical symptoms of arbovirus-like infection from Salvador and Campo Formoso, Bahia, in Northeast Brazil. Between May 2015 and June 2016, we identified 21 cases of acute ZIKV infection and subsequently recovered 14 near full-length sequences using the amplicon tiling multiplex approach with nanopore sequencing. We performed a time-calibrated discrete phylogeographic analysis to trace the spread and migration history of the ZIKV. Our phylogenetic analysis supports a consistent relationship between ZIKV migration from Northeast to Southeast Brazil and its subsequent dissemination beyond Brazil. Additionally, our analysis provides insights into the migration of ZIKV from Brazil to Haiti and the role Brazil played in the spread of ZIKV to other countries, such as Singapore, the USA, and the Dominican Republic. The data generated by this study enhances our understanding of ZIKV dynamics and supports the existing knowledge, which can aid in future surveillance efforts against the virus.

Transcriptional and post-transcriptional mechanisms that regulate the genetic program in Zika virus-infected macrophages.

Besides our understanding of the effects of ZIKA virus (ZIKV) infection on neural progenitors' cells the pathogenesis of this RNA virus also involves antigen-presenting cells, including macrophages. However, the molecular mechanisms that control gene activation and repression associated with the macrophage response to acute ZIKV infection are not fully understood. We approached the issue by RNA-seq and miRNA-seq datasets to understand the genetic program of ZIKV-infected macrophages. Results indicate that macrophage activates a regulatory program, involving 1067 differentially expressed genes. These genetic programs induced an inflammatory response mediated by chemokines as well as an interferon-independent anti-viral response, presumptively activated by IL-27. Additionally, the pathogenetic process involves changes in other signaling pathways such as cellular stress, cell signaling, metabolism, and cell differentiation. Furthermore, transcriptional control analysis revealed regulatory functions of key transcription factors principally, NFκB and STAT1, as well as HIF1A, ETV7, and PRMD1 that are associated with metabolic reprogramming during viral infection. We also noted six long-noncoding RNAs (lncRNAs) that may act in the regulation of gene expression, including MROCKI and ZC2HC1A-2, that are involved in the inflammatory response and expression of the cytokines, respectively. On the other hand, post-transcriptional control by miRNAs, including miR-155-5p and miR-146a-5p, are associated with modulation of genes related to inflammatory and antiviral responses. Relevant to the post-transcriptional control, our data unveiled the role of RNA binding proteins that have diverse functions such as ribonucleases (PNPT1, ZC3H12A, and ZC3HAV1), splicing factors (SSB, RBM11, and RAVER2), and RNA modifiers (PARP10 and PARP14). Overall, the results establish an unbiased approach to discerning the wiring of a regulatory mechanism controlling the genetic program in ZIKV-infected macrophages.

Pathogenesis and Manifestations of Zika Virus-Associated Ocular Diseases.

Zika virus (ZIKV) is mosquito-borne flavivirus that caused a significant public health concern in French Polynesia and South America. The two major complications that gained the most media attention during the ZIKV outbreak were Guillain–Barré syndrome (GBS) and microcephaly in newborn infants. The two modes of ZIKV transmission are the vector-borne and non-vector borne modes of transmission. Aedes aegypti and Aedes albopictus are the most important vectors of ZIKV. ZIKV binds to surface receptors on permissive cells that support infection and replication, such as neural progenitor cells, dendritic cells, dermal fibroblasts, retinal pigment epithelial cells, endothelial cells, macrophages, epidermal keratinocytes, and trophoblasts to cause infection. The innate immune response to ZIKV infection is mediated by interferons and natural killer cells, whereas the adaptive immune response is mediated by CD8+T cells, Th1 cells, and neutralizing antibodies. The non-structural proteins of ZIKV, such as non-structural protein 5, are involved in the evasion of the host’s immune defense mechanisms. Ocular manifestations of ZIKV arise from the virus’ ability to cross both the blood–brain barrier and blood-retinal barrier, as well as the blood-aqueous barrier. Most notably, this results in the development of GBS, a rare neurological complication in acute ZIKV infection. This can yield ocular symptoms and signs. Additionally, infants to whom ZIKV is transmitted congenitally develop congenital Zika syndrome (CZS). The ocular manifestations are widely variable, and include nonpurulent conjunctivitis, anterior uveitis, keratitis, trabeculitis, congenital glaucoma, microphthalmia, hypoplastic optic disc, and optic nerve pallor. There are currently no FDA approved therapeutic agents for treating ZIKV infections and, as such, a meticulous ocular examination is an important aspect of the diagnosis. This review utilized several published articles regarding the ocular findings of ZIKV, antiviral immune responses to ZIKV infection, and the pathogenesis of ocular manifestations in individuals with ZIKV infection. This review summarizes the current knowledge on the viral immunology of ZIKV, interactions between ZIKV and the host’s immune defense mechanism, pathological mechanisms, as well as anterior and posterior segment findings associated with ZIKV infection.

Myopericarditis associated with acute Zika virus infection: a case report

BackgroundZika virus infection is commonly described as a mild and self-limiting illness. However, cardiac complications were associated with acute Zika virus infection.Case presentationA 46-year-old woman without previous comorbidities with a 1-day history of symptoms tested positive for ZIKV by real time reverse transcriptase polymerase chain reaction (rRT-PCR). She was admitted two days after with clinical worsening, cardiac enzymes elevated, and cardiac imaging findings, and the diagnosis of myopericarditis was made. The patient was treated and presented significant clinical improvement after one year.ConclusionsCardiac complication following ZIKV infection appears to be infrequent. Here, we report a rare case of viral myopericarditis caused by ZIKV infection.

Increased Serum Levels of Growth-Differentiation Factor 3 (GDF3) and Inflammasome-Related Markers in Pregnant Women during Acute Zika Virus Infection.

The systemic inflammatory response elicited by acute Zika virus (ZIKV) infection during pregnancy plays a key role in the clinical outcomes in mothers and congenitally infected offspring. The present study aimed to evaluate the serum levels of GDF-3 and inflammasome-related markers in pregnant women during acute ZIKV infection. Serum samples from pregnant (n = 18) and non-pregnant (n = 22) women with acute ZIKV infection were assessed for NLRP3, IL-1β, IL-18, and GDF3 markers through an enzyme-linked immunosorbent assay. ZIKV-negative pregnant (n = 18) and non-pregnant women (n = 15) were used as control groups. All serum markers were highly elevated in the ZIKV-infected groups in comparison with control groups (p < 0.0001). Among the ZIKV-infected groups, the serum markers were significantly augmented in the pregnant women in comparison with non-pregnant women (NLRP3 p < 0.001; IL-1β, IL-18, and GDF3 p < 0.0001). The IL-18 marker was found at significantly higher levels (p < 0.05) in the third trimester of pregnancy. Bivariate and multivariate analyses showed a strong positive correlation between GDF3 and NLRP3 markers among ZIKV-infected pregnant women (r = 0.91, p < 0.0001). The findings indicated that acute ZIKV infection during pregnancy induces the overexpression of GDF-3 and inflammasome-related markers, which may contribute to congenital disorders and harmful pregnancy outcomes.

A cytotoxic-skewed immune set point predicts low neutralizing antibody levels after Zika virus infection.

SUMMARYAlthough generating high neutralizing antibody levels is a key component of protective immunity after acute viral infection or vaccination, little is known about why some individuals generate high versus low neutralizing antibody titers. Here, we leverage the high-dimensional single-cell profiling capacity of mass cytometry to characterize the longitudinal cellular immune response to Zika virus (ZIKV) infection in viremic blood donors in Puerto Rico. During acute ZIKV infection, we identify widely coordinated responses across innate and adaptive immune cell lineages. High frequencies of multiple activated cell types during acute infection are associated with high titers of ZIKV neutralizing antibodies 6 months post-infection, while stable immune features suggesting a cytotoxic-skewed immune set point are associated with low titers. Our study offers insight into the coordination of immune responses and identifies candidate cellular biomarkers that may offer predictive value in vaccine efficacy trials aimed at inducing high levels of antiviral neutralizing antibodies.

Molecular Approaches for the Validation of the Baboon as a Nonhuman Primate Model for the Study of Zika Virus Infection.

Nonhuman primates (NHP) are particularly important for modeling infections with viruses that do not naturally replicate in rodent cells. Zika virus (ZIKV) has been responsible for sporadic epidemics, but in 2015 a disseminated outbreak of ZIKV resulted in the World Health Organization declaring it a global health emergency. Since the advent of this last epidemic, several NHP species, including the baboon, have been utilized for modeling and understanding the complications of ZIKV infection in humans; several health issues related to the outcome of infection have not been resolved yet and require further investigation. This study was designed to validate, in baboons, the molecular signatures that have previously been identified in ZIKV-infected humans and macaque models. We performed a comprehensive molecular analysis of baboons during acute ZIKV infection, including flow cytometry, cytokine, immunological, and transcriptomic analyses. We show here that, similar to most human cases, ZIKV infection of male baboons tends to be subclinical, but is associated with a rapid and transient antiviral interferon-based response signature that induces a detectable humoral and cell-mediated immune response. This immunity against the virus protects animals from challenge with a divergent ZIKV strain, as evidenced by undetectable viremia but clear anamnestic responses. These results provide additional support for the use of baboons as an alternative animal model to macaques and validate omic techniques that could help identify the molecular basis of complications associated with ZIKV infections in humans.

Involvement of Th1Th17 Cell Subpopulations in the Immune Responses of Mothers Who Gave Birth to Children with Congenital Zika Syndrome (CZS).

High levels of T helper 17 cell (Th17)-related cytokines have been shown in acute Zika virus (ZIKV) infection. We hypothesized that the high levels of Th17-related cytokines, associated with a regulatory environment during pregnancy, create a favorable milieu for the differentiation of CD4+Th17 cells. We present data from a cross-sectional study on mothers who confirmed ZIKV infection by qRT-PCR and their children. We also recruited non-pregnant women infected with ZIKV in the same period. ZIKV infection occurred between 2015 and 2017. We collected samples for this study between 2018 and 2019, years after the initial infection. We highlight that, after in vitro stimulation with ZIKV CD4 megapool (ZIKV MP), we found a lower frequency of IL-17-producing CD4+ T cells (Th17), especially in the mothers, confirmed by the decrease in IL-17 production in the supernatant. However, a higher frequency of CD4+ IL-17+ IFN-γ+ T cells (Th1Th17) responding to the ZIKV MP was observed in the cells of the mothers and children but not in those of the non-pregnant women. Our data indicate that the priming of CD4 T cells of the Th1Th17 phenotype occurred preferentially in the mothers who gave birth to children with CZS and in the children.

Zika virus infection and microcephaly: spatial analysis and socio-environmental determinants in a region of high Aedes aegypti infestation in the Central-West Region of Brazil

BackgroundMore than 5 years after the Zika virus (ZIKV) epidemic, Zika infection remains a major concern in regions with high Aedes infestation. The objectives of this study were (i) to identify clusters of ZIKV infection and microcephaly, and/or central nervous system (CNS) alterations associated with congenital infection during the epidemic peak in 2016 and subsequently, in 2017 and 2018; (ii) to measure the non-spatial correlation between ZIKV infection and microcephaly and/or CNS alterations associated with congenital infection; and (iii) to analyse the sociodemographic/economic, health, and environmental determinants associated with the incidence of ZIKV in a region of high infestation by Aedes aegypti in the Central-West Region of Brazil.MethodsThis ecological study analysed 246 municipalities in the state of Goiás (6.9 million inhabitants). The data were obtained from the Information System for Notifiable Diseases (ZIKV cases) and the Public Health Event Registry (microcephaly and/or CNS alterations associated with congenital infection). Incidence rates and prevalence of ZIKA infection were smoothed by an empirical Bayesian estimator (LEbayes), producing the local empirical Bayesian rate (LEBR). In the spatial analysis, ZIKV infection and microcephaly cases were georeferenced by the municipality of residence for 2016 and grouped for 2017 and 2018. Global Moran's I and the Hot Spot Analysis tool (Getis-Ord Gi* statistics) were used to analyse the spatial autocorrelation and clusters of ZIKV infection and microcephaly, respectively. A generalised linear model from the Poisson family was used to assess the association between ecological determinants and the smoothing incidence rate of ZIKV infection.ResultsA total of 9892 cases of acute ZIKV infection and 121 cases of microcephaly were confirmed. The mean LEBR of the ZIKV infection in the 246 municipalities was 22.3 cases/100,000 inhabitants in 2016, and 10.3 cases/100,000 inhabitants in 2017 and 2018. The LEBR of the prevalence rate of microcephaly and/or CNS alterations associated with congenital infection was 7 cases/10,000 live births in 2016 and 2 cases/10,000 live births during 2017–2018. Hotspots of ZIKV infection and microcephaly cases were identified in the capital and neighbouring municipalities in 2016, with new clusters in the following years. In a multiple regression Poisson analysis, ZIKV infection was associated with higher population density, the incidence of dengue, Aedes larvae infestation index, and average rainfall. The important determinant of ZIKV infection incidence reduction was the increase in households attended by endemic disease control agents.ConclusionsOur analyses were able to capture, in a more granular way, aspects that make it possible to inform public managers of the sentinel areas identified in the post-epidemic hotspots.

Clinical and Epidemiological Features of Acute Zika Virus Infections in León, Nicaragua

The American Zika virus (ZIKV) epidemic has highlighted the need to gain a better understanding of this emerging virus. The goal of this study was to describe the clinical symptoms, laboratory findings, and risk factors for symptomatic ZIKV infection in an area with ongoing transmission of other arboviral infections. We recruited patients at least 2 years of age seeking care at public health centers in León, Nicaragua, between January 2016 and August 2017, for fever, maculopapular rash, and/or nonsuppurative conjunctivitis with a duration of less than 1 week. A laboratory diagnosis of ZIKV was established using a combination of molecular and serological tests. Clinical and laboratory findings and potential risk factors were compared between participants with and without acute ZIKV infection. Fifty-eight (26%) of the 225 participants included in the analysis were found to have acute ZIKV infection. Pregnancy and reports of previous arboviral infection were associated with a higher risk of ZIKV infection. Rash, conjunctivitis, sore throat, and lower absolute neutrophil counts were associated with acute ZIKV infection. The clinical characteristics and risk factors identified were consistent with those identified by previous studies; however, we found sore throat to be a feature of ZIKV infection. We also found that neutrophil counts were lower in ZIKV-infected subjects. These clinical symptoms and laboratory data may help clinicians suspect ZIKV infection during future outbreaks.

A Systematic Evaluation of IgM and IgG Antibody Assay Accuracy in Diagnosing Acute Zika Virus Infection in Brazil: Lessons Relevant to Emerging Infections.

Accurate diagnostics underpin effective public health responses to emerging viruses. For viruses, such as Zika virus (ZIKV), where the viremia clears quickly, antibody-based (IgM or IgG) diagnostics are recommended for patients who present 7 days after symptom onset. However, cross-reactive antibody responses can complicate test interpretation among populations where closely related viruses circulate. We examined the accuracy (proportion of samples correctly categorized as Zika positive or negative) for antibody-based diagnostics among Brazilian residents (Rio de Janeiro) during the ZIKV outbreak. Four ZIKV enzyme-linked immunosorbent assays (ELISAs; IgM and IgG Euroimmun, IgM Novagnost, and CDC MAC), two dengue ELISAs (IgM and IgG Panbio), and the ZIKV plaque reduction neutralization test (PRNT) were evaluated. Positive samples were ZIKV PCR confirmed clinical cases collected in 2015-2016 (n = 169); negative samples (n = 236) were collected before ZIKV was present in Brazil (≤2013). Among serum samples collected ≥7 days from symptom onset, PRNT exhibited the highest accuracy (93.7%), followed by the Euroimmun IgG ELISA (77.9%). All IgM assays exhibited lower accuracy (<75%). IgG was detected more consistently than IgM among ZIKV cases using Euroimmun ELISAs (68% versus 22%). Anti-dengue virus IgM ELISA was positive in 41.1% of confirmed ZIKV samples tested. The Euroimmun IgG assay, although misdiagnosing 22% of samples, provided the most accurate ELISA. Anti-ZIKV IgG was detected more reliably than IgM among ZIKV patients, suggesting a secondary antibody response to assay antigens following ZIKV infection. Antibody ELISAs need careful evaluation in their target population to optimize use and minimize misdiagnosis, prior to widespread deployment, particularly where related viruses cocirculate.

Phenotypical characterization of regulatory T cells in acute Zika infection

Zika virus (ZIKV), alongside Dengue virus (DENV), Chikungunya virus (CHIKV), and Yellow Fever Virus (YFV) are prevalent arboviruses in the Americas. Each of these infections is associated with the development of associated disease immunopathology. Immunopathological processes are an outcome of counter-balancing impacts between effector and regulatory immune mechanisms. In this context, regulatory T cells (Tregs) are key in modulating the immune response and, therefore, in tissue damage control. However, to date, Treg phenotypes and mechanisms during acute infection of the ZIKV in humans have not been fully investigated. The main aim of this work was to characterize Tregs and their immunological profile related to cytokine production and molecules that are capable of controlling the exacerbated inflammatory profile in acute Zika infected patients. Using whole blood analyses of infected patients, an ex vivo phenotypical characterization of Tregs, circulating during acute Zika virus infection, was conducted by flow cytometry. We found that though there are no differences in absolute Treg frequency between infected and healthy control groups. However, pro-inflammatory cytokine up-regulation such as IFN-γ and LAP was observed in the acute disease. Furthermore, acute ZIKV patients expressed increased levels of CD39/CD73, perforin/granzyme B, PD-1, and CTLA-4, all markers involved in mechanisms used by Tregs to attempt to control strong inflammatory responses. Thus, the data indicates a potential contribution of Tregs during the inflammatory ZIKV infection response.

Pregnancy Alters Innate and Adaptive Immune Responses to Zika Virus Infection in the Reproductive Tract.

Recent outbreaks of Zika virus (ZIKV) have been associated with birth defects, including microcephaly and neurologic impairment. However, the mechanisms that confer potential susceptibility to ZIKV during pregnancy remain unclear. We hypothesized that poor outcomes from ZIKV infection during pregnancy are due in part to pregnancy-induced alteration of innate immune cell frequencies and cytokine expression. To examine the impact of pregnancy on innate immune responses, we inoculated immunocompetent pregnant and nonpregnant female C57BL/6 mice with 5 × 105 focus-forming units of ZIKV intravaginally. Innate immune cell frequencies and cytokine expression were measured by flow cytometry at day 3 postinfection. Compared with nonpregnant mice, pregnant mice exhibited higher frequencies of uterine macrophages (CD68+) and CD11c+ CD103+ and CD11c+ CD11b+ dendritic cells. Additionally, ZIKV-infected pregnant mice had lower frequencies of CD45+ IL-12+ and CD11b+ IL-12+ cells in the uterus and spleen. Next, we measured the frequencies of Ag-experienced CD4 (CD4+ CD11a+ CD49d+) and CD8 (CD8lo CD11ahi) T cells at day 10 postinfection to determine the impact of pregnancy-associated changes in innate cellular IL-12 responses on the adaptive immune response. We found that pregnant mice had lower frequencies of uterine Ag-experienced CD4 T cells and ZIKV-infected pregnant mice had lower frequencies of uterine Ag-experienced CD8 T cells compared with ZIKV-infected nonpregnant mice. These data show that pregnancy results in altered innate and adaptive immune responses to ZIKV infection in the reproductive tract of mice and that pregnancy-associated immune modulation may play an important role in the severity of acute ZIKV infection.

Association of Anterior Uveitis With Acute Zika Virus Infection in Adults

Acute anterior uveitis has been reported to be associated with Zika virus (ZIKV) infection in case reports and retrospective studies based on systemic clinical signs of ZIKV and positive serologic tests. To describe the ophthalmic findings associated with systemic ZIKV infection during the 2016 ZIKV outbreak in Guadeloupe in the French West Indies. This cohort study assessed adult patients with red eye and virologic or serologic confirmation of acute ZIKV infection who were admitted to the Pointe-à-Pitre University Hospital from January 1, 2016, to January 1, 2017, for a systematic ophthalmologic examination. All patients with anterior uveitis also underwent a complete uveitis workup to rule out other causes of uveitis. Patients were excluded from the study if they had active uveitis and a history of uveitis or positive serology for other infectious diseases including dengue fever virus and chikungunya virus. Data were analyzed from January 1, 2016, to January 1, 2017. Patients with confirmed ZIKV infection underwent full ophthalmic examination at study inception and 3, 6, and 12 months. Description of the presenting ocular characteristics, management, and outcome. Of 62 total patients with a red eye, 32 (51.6%) had no uveitis (mean [SD] age, 27.8 [3.7] years; 26 women [81.3%]) and 30 (48.4%) had anterior uveitis (mean [SD] age, 27.5 [3.8] years; 26 women [86.7%]), which was bilateral in all cases. Acute anterior uveitis was nongranulomatous and without synechiae, exhibiting mild anterior chamber reaction with small keratic precipitates. Ocular hypertension occurred in 25 patients (83.3%) with uveitis at presentation and persisted in 15 patients (50%) at 1 year despite antiglaucoma therapy and control of uveitis. Uveitis responded to topical steroids in all cases. Intraocular pressure was within normal limits for all patients without uveitis throughout the follow-up period. Anterior uveitis was present in almost half of patients with a red eye and acute systemic ZIKV infection; the uveitis was usually bilateral and often associated with increased intraocular pressure. These results suggest that ophthalmic evaluation should be performed in patients with red eye and acute ZIKV infection to potentially detect and manage hypertensive anterior uveitis.