Dengue virus surveillance in Nepal yields the first on-site whole genome sequences of isolates from the 2022 outbreak.

BACKGROUND: The 4 serotypes of dengue virus (DENV1-4) can each cause potentially deadly dengue disease, and are spreading globally from tropical and subtropical areas to more temperate ones. Nepal provides a microcosm of this global phenomenon, having met each of these grim benchmarks. To better understand DENV transmission dynamics and spread into new areas, we chose to study dengue in Nepal and, in so doing, to build the onsite infrastructure needed to manage future, larger studies. METHODS AND RESULTS: During the 2022 dengue season, we enrolled 384 patients presenting at a hospital in Kathmandu with dengue-like symptoms; 79% of the study participants had active or recent DENV infection (NS1 antigen and IgM). To identify circulating serotypes, we screened serum from 50 of the NS1+ participants by RT-PCR and identified DENV1, 2, and 3 - with DENV1 and 3 codominant. We also performed whole-genome sequencing of DENV, for the first time in Nepal, using our new on-site capacity. Sequencing analysis demonstrated the DENV1 and 3 genomes clustered with sequences reported from India in 2019, and the DENV2 genome clustered with a sequence reported from China in 2018. CONCLUSION: These findings highlight DENV's geographic expansion from neighboring countries, identify China and India as the likely origin of the 2022 DENV cases in Nepal, and demonstrate the feasibility of building onsite capacity for more rapid genomic surveillance of circulating DENV. These ongoing efforts promise to protect populations in Nepal and beyond by informing the development and deployment of DENV drugs and vaccines in real time.

CD4+ T cells promote humoral immunity and viral control during Zika virus infection.

Several Zika virus (ZIKV) vaccines designed to elicit protective antibody (Ab) responses are currently under rapid development, but the underlying mechanisms that control the magnitude and quality of the Ab response remain unclear. Here, we investigated the CD4+ T cell response to primary intravenous and intravaginal infection with ZIKV. Using the LysMCre+Ifnar1fl/fl (myeloid type I IFN receptor-deficient) C57BL/6 mouse models, we identified six I-Ab-restricted ZIKV epitopes that stimulated CD4+ T cells with a predominantly cytotoxic Th1 phenotype in mice primed with ZIKV. Intravenous and intravaginal infection with ZIKV effectively induced follicular helper and regulatory CD4+ T cells. Treatment of mice with a CD4+ T cell-depleting Ab reduced the plasma cell, germinal center B cell, and IgG responses to ZIKV without affecting the CD8+ T cell response. CD4+ T cells were required to protect mice from a lethal dose of ZIKV after infection intravaginally, but not intravenously. However, adoptive transfer and peptide immunization experiments showed a role for memory CD4+ T cells in ZIKV clearance in mice challenged intravenously. These results demonstrate that CD4+ T cells are required mainly for the generation of a ZIKV-specific humoral response but not for an efficient CD8+ T cell response. Thus, CD4+ T cells could be important mediators of protection against ZIKV, depending on the infection or vaccination context.

Correction: CD4+ T cells promote humoral immunity and viral control during Zika virus infection.

[This corrects the article DOI: 10.1371/journal.ppat.1007474.].

Neuropathologic, phenotypic and functional analyses of Mucosal Associated Invariant T cells in Multiple Sclerosis.

BACKGROUND: The involvement of Mucosal Associated Invariant T (MAIT) cells, which are anti-microbial semi-invariant T cells, remains elusive in Multiple Sclerosis (MS). OBJECTIVE: Deciphering the potential involvement of MAIT cells in the MS inflammatory process. METHODS: By flow cytometry, blood MAIT cells from similar cohorts of MS patients and healthy volunteers (HV) were compared for frequency, phenotype, activation potential after in vitro TCR engagement by bacterial ligands and transmigration abilities through an in vitro model of blood-brain barrier. MS CNS samples were also studied by immunofluorescent staining and quantitative PCR. RESULTS AND CONCLUSION: Blood MAIT cells from relapsing-remitting MS patients and HV presented similar frequency, ex vivo effector phenotype and activation abilities. MAIT cells represented 0.5% of the total infiltrating T cells on 39 MS CNS lesions. This is low as compared to blood frequency (p<0.001), but consistent with their low transmigration rate. Finally, transcriptional over-expression of MR1 - which presents cognate antigens to MAIT cells - and of the activating cytokines IL-18 and IL-23 was evidenced in MS lesions, suggesting that the CNS microenvironment is suited to activate the few infiltrating MAIT cells. Taken together, these data place MAIT cells from MS patients as minor components of the inflammatory pathological process.

Unaltered regulatory B-cell frequency and function in patients with multiple sclerosis.

Multiple sclerosis (MS) is a chronic disease of the central nervous system (CNS) typically characterized by the recruitment of T cells into the CNS. However, certain subsets of B cells have been shown to negatively regulate autoimmune diseases and some data support a prominent role for B cells in MS physiopathology. For B cells in MS patients we analyzed subset frequency, cytokine secretion ability and suppressive properties. No differences in the frequencies of the B-cell subsets or in their ability to secrete cytokines were observed between MS and healthy volunteers (HV). Prestimulated B cells from MS patients also inhibited CD4(+)CD25(-) T cell proliferation with a similar efficiency as B cells from HV. Altogether, our data show that, in our MS patient cohort, regulatory B cells have conserved frequency and function.

Human immunodeficiency virus-1 genome from patient with fever, Nepal.

A patient with fever presented to the referral infectious disease hospital in Kathmandu, Nepal. Metagenomic sequencing of the patient's serum recovered a near-complete genome of human immunodeficiency virus-1 (HIV-1), distinct from previous HIV-1 genomes from Nepal in GenBank. It shared 92.48% nucleotide identity with an HIV-1 subtype C isolate from India.

Gemykibivirus detection in acute encephalitis patients from Nepal.

Acute encephalitis syndrome (AES) causes significant morbidity and mortality worldwide. In Nepal, Japanese encephalitis virus (JEV) accounts for ~5-20% of AES cases, but ~75% of AES cases are of unknown etiology. We identified a gemykibivirus in CSF collected in 2020 from an 8-year-old male patient with AES using metagenomic next-generation sequencing. Gemykibiviruses are single stranded, circular DNA viruses in the family Genomoviridae. The complete genome of 2,211 nucleotides was sequenced, which shared 98.69% nucleotide identity to its closest relative, Human associated gemykibivirus 2 isolate SAfia-449D. Two real-time PCR assays were designed, and screening of 337 cerebrospinal fluid (CSF) and 164 serum samples from AES patients in Nepal collected in 2020 and 2022 yielded 11 CSF and 1 serum sample that were positive in both PCR assays. Complete genomes of seven of the positives were sequenced. These results identify a potential candidate etiologic agent of encephalitis in Nepal. IMPORTANCE: Viral encephalitis is a devastating disease, but unfortunately, worldwide, the causative virus in many cases is unknown. Therefore, it is important to identify viruses that could be responsible for cases of human encephalitis. Here, using metagenomic sequencing of CSF, we identified a gemykibivirus in a male child from Nepal with acute encephalitis syndrome (AES). We subsequently detected gemykibivirus DNA in CSF or serum of 12 more encephalitis patients by real-time PCR. The virus genomes we identified are highly similar to gemykibiviruses previously detected in CSF of three encephalitis patients from Sri Lanka. These results raise the possibility that gemykibivirus could be an underrecognized human pathogen.

Human coronavirus OC43-elicited CD4+ T cells protect against SARS-CoV-2 in HLA transgenic mice.

SARS-CoV-2-reactive T cells are detected in some healthy unexposed individuals. Human studies indicate these T cells could be elicited by the common cold coronavirus OC43. To directly test this assumption and define the role of OC43-elicited T cells that are cross-reactive with SARS-CoV-2, we develop a model of sequential infections with OC43 followed by SARS-CoV-2 in HLA-B*0702 and HLA-DRB1*0101 Ifnar1-/- transgenic mice. We find that OC43 infection can elicit polyfunctional CD8+ and CD4+ effector T cells that cross-react with SARS-CoV-2 peptides. Furthermore, pre-exposure to OC43 reduces subsequent SARS-CoV-2 infection and disease in the lung for a short-term in HLA-DRB1*0101 Ifnar1-/- transgenic mice, and a longer-term in HLA-B*0702 Ifnar1-/- transgenic mice. Depletion of CD4+ T cells in HLA-DRB1*0101 Ifnar1-/- transgenic mice with prior OC43 exposure results in increased viral burden in the lung but no change in virus-induced lung damage following infection with SARS-CoV-2 (versus CD4+ T cell-sufficient mice), demonstrating that the OC43-elicited SARS-CoV-2 cross-reactive T cell-mediated cross-protection against SARS-CoV-2 is partially dependent on CD4+ T cells. These findings contribute to our understanding of the origin of pre-existing SARS-CoV-2-reactive T cells and their effects on SARS-CoV-2 clinical outcomes, and also carry implications for development of broadly protective betacoronavirus vaccines.

Influence of Th1 versus Th2 immune bias on viral, pathological, and immunological dynamics in SARS-CoV-2 variant-infected human ACE2 knock-in mice.

BACKGROUND: Mouse models that recapitulate key features of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection are important tools for understanding complex interactions between host genetics, immune responses, and SARS-CoV-2 pathogenesis. Little is known about how predominantly cellular (Th1 type) versus humoral (Th2 type) immune responses influence SARS-CoV-2 dynamics, including infectivity and disease course. METHODS: We generated knock-in (KI) mice expressing human ACE2 (hACE2) and/or human TMPRSS2 (hTMPRSS2) on Th1-biased (C57BL/6; B6) and Th2-biased (BALB/c) genetic backgrounds. Mice were infected intranasally with SARS-CoV-2 Delta (B.1.617.2) or Omicron BA.1 (B.1.1.529) variants, followed by assessment of disease course, respiratory tract infection, lung histopathology, and humoral and cellular immune responses. FINDINGS: In both B6 and BALB/c mice, hACE2 expression was required for infection of the lungs with Delta, but not Omicron BA.1. Disease severity was greater in Omicron BA.1-infected hTMPRSS2-KI and double-KI BALB/c mice compared with B6 mice, and in Delta-infected double-KI B6 and BALB/c mice compared with hACE2-KI mice. hACE2-KI B6 mice developed more severe lung pathology and more robust SARS-CoV-2-specific splenic CD8 T cell responses compared with hACE2-KI BALB/c mice. There were no notable differences between the two genetic backgrounds in plasma cell, germinal center B cell, or antibody responses to SARS-CoV-2. INTERPRETATION: SARS-CoV-2 Delta and Omicron BA.1 infection, disease course, and CD8 T cell response are influenced by the host genetic background. These humanized mice hold promise as important tools for investigating the mechanisms underlying the heterogeneity of SARS-CoV-2-induced pathogenesis and immune response. FUNDING: This work was funded by NIH U19 AI142790-02S1, the GHR Foundation, the Arvin Gottleib Foundation, and the Overton family (to SS and EOS); Prebys Foundation (to SS); NIH R44 AI157900 (to KJ); and by an American Association of Immunologists Career Reentry Fellowship (FASB).

Frequency of circulating autoreactive T cells committed to myelin determinants in relapsing-remitting multiple sclerosis patients.

Multiple sclerosis (MS) is considered as an autoimmune disease in which T cell reactivity to self-antigens expressed in the brain, particularly myelin antigens, plays a pivotal role. Various myelin-derived peptides, including peptides of myelin basic protein (MBP), proteolipid protein (PLP) and myelin oligodendrocyte glycoprotein (MOG) have been studied as putative target in MS. However, CD4(+) and CD8(+) T cells recognizing autoantigens from brain have been detected in the blood of MS patients as well as the blood of normal individuals. Here we review and discuss studies focused on the assessment of the frequency of autoreactive T cells responding to a given antigen using different assays including LDA, IFNγ-ELISPOT and TRAP (T cell Recognition of Antigen Presenting Cells by Protein transfer) in MS.

Acute-phase Serum Cytokine Levels and Correlation with Clinical Outcomes in Children and Adults with Primary and Secondary Dengue Virus Infection in Myanmar between 2017 and 2019.

The dengue virus (DENV) has been endemic in Myanmar since 1970, causing outbreaks every 2-3 years. DENV infection symptoms range from mild fever to lethal hemorrhage. Clinical biomarkers must be identified to facilitate patient risk stratification in the early stages of infection. We analyzed 45 cytokines and other factors in serum samples from the acute phase of DENV infection (within 3-5 days of symptom onset) from 167 patients in Yangon, Myanmar, between 2017 and 2019. All of the patients tested positive for serum DENV nonstructural protein 1 antigen (NS1 Ag); 78.4% and 62.9% were positive for immunoglobulin M (IgM) and G (IgG), respectively; and 18.0%, 19.8%, and 11.9% tested positive for serotypes 1, 3, and 4, respectively. Although the DENV-4 viral load was significantly higher than those of DENV-1 or DENV-3, disease severity was not associated with viral load or serotype. Significant correlations were identified between disease severity and CCL5, SCF, PDGF-BB, IL-10, and TNF-α levels; between NS1 Ag and SCF, CCL5, IFN-α, IL-1α, and IL-22 levels; between thrombocytopenia and IL-2, TNF-α, VEGF-D, and IL-6 levels; and between primary or secondary infection and IL-2, IL-6, IL-31, IL-12p70, and MIP-1ß levels. These circulating factors may represent leading signatures in acute DENV infections, reflecting the clinical outcomes in the dengue endemic region, Myanmar.

T cell recognition of self-antigen presenting cells by protein transfer assay reveals a high frequency of anti-myelin T cells in multiple sclerosis.

Although peripheral blood myelin-autoreactive T cells are thought to play a key role in multiple sclerosis, they are generally considered to have qualitative differences rather than quantitative ones when compared to those found in healthy individuals. Here, we revisited the assessment of myelin-autoreactive T cells in a new approach based on their combined ability to acquire membrane proteins from autologous antigen presenting cells, and to respond to whole myelin extract as the stimulating autoantigen. Using this approach, the myelin-autoreactive T cell frequency in patients with multiple sclerosis was found to be unexpectedly high (n = 22, subtracted values median 2.08%, range 0-6%; background median 1%, range 0-4%) and to exceed that of age/gender-matched healthy individuals significantly (n = 18, subtracted values median 0.1%, range 0-5.3%, P < 0.0001; background median 1.45%, range 0.1-4%). Higher anti-myelin autoreactivity was stable in patients with multiple sclerosis after several months. These data correlated with whole myelin-induced gamma interferon-enzyme-linked immunosorbent spot assay performed under the same conditions, although the values obtained with enzyme-linked immunosorbent spot assay under all conditions were 58 times lower than with this new method. The myelin-autoreactive T cells were memory T cells expressing CD40L with a CD62(low) phenotype, suggesting their ability for homing to tissues. Collectively, these new data show a higher frequency of autoreactive T cells during multiple sclerosis than in age/gender-matched healthy individuals, and support an autoimmune aetiology in multiple sclerosis.

Repeated exposure to dengue virus elicits robust cross neutralizing antibodies against Zika virus in residents of Northeastern Thailand.

Zika virus (ZIKV) and dengue virus (DENV) are antigenically related mosquito-borne flaviviruses. ZIKV is becoming increasingly prevalent in DENV-endemic regions, raising the possibility that pre-existing immunity to one virus could modulate the response to a heterologous virus, although whether this would be beneficial or detrimental is unclear. Here, we analyzed sera from residents of a DENV-endemic region of Thailand to determine the prevalence of DENV-elicited antibodies capable of cross-neutralizing ZIKV. Sixty-one participants who were asymptomatic and unselected for viral serostatus were enrolled. Among them, 52 and 51 were seropositive for IgG antibody against DENV or ZIKV E proteins (ELISA assay), respectively. Notably, 44.23% (23/52) of DENV seropositive participants had serological evidence of multiple exposures to DENV, and these subjects had strikingly higher titers and broader reactivities of neutralizing antibodies (NAbs) against ZIKV and DENV heterotypes compared with participants with serological evidence of a single DENV infection (25/52, 48.1%). In total, 17 of the 61 participants (27.9%) had NAbs against ZIKV and all four DENV serotypes, and an additional 9 (14.8%) had NAbs against ZIKV and DENV1, 2, and 3. NAbs against DENV2 were the most prevalent (44/61, 72.1%) followed by DENV3 (38/61, 62.3%) and DENV1 (36/61, 59.0%). Of note, anti-ZIKV NAbs were more prevalent than anti-DENV4 NAbs (27/61, 44.3% and 21/61, 34.4%, respectively). Primary ZIKV infection was detected in two participants, confirming that ZIKV co-circulates in this region. Thus, residents of DENV-endemic regions with repeated exposure to DENV have higher titers of NAbs against ZIKV than individuals with only a single DENV exposure.

Whole Genome Sequencing of Dengue Virus Serotype 2 from Two Clinical Isolates and Serological Profile of Dengue in the 2015-2016 Nepal Outbreak.

Dengue virus (DENV) is the cause of one of the most prevalent neglected tropical diseases, and up to half of the world's population is at risk for infection. Recent results from clinical trials have shown that DENV vaccination can induce the development of severe dengue disease and/or prolong hospitalization after natural infection in certain naive populations. Thus, it is crucial that vaccine development takes into account the history of DENV exposure in the targeted population. In Nepal, DENV infection was first documented in 2004, and despite the increasing prevalence of DENV infection, the population remains relatively naive. However, it is not known which of the four DENV serotypes circulate in Nepal or whether there is evidence of repeated exposure to DENV in the Nepali population. To address this, we studied 112 patients who presented with symptomology suspicious for DENV infection at clinics throughout Nepal during late 2015 and early 2016. Of the 112 patients examined, 39 showed serological and/or genetic evidence of primary or secondary DENV infection: 30 were positive for DENV exposure by IgM/IgG ELISA, two by real-time reverse-transcription PCR (RT-PCR), and seven by both methods. Dengue virus 1-3, but not DENV4, serotypes were detected by RT-PCR. Whole genome sequencing of two DENV2 strains isolated from patients with primary and secondary infections suggests that DENV was introduced into Nepal through India, with which it shares a porous border. Further study is needed to better define the DENV epidemic in Nepal, a country with limited scientific resources and infrastructure.

CD8+ T cells mediate protection against Zika virus induced by an NS3-based vaccine.

Zika virus (ZIKV) is associated with congenital malformations in infants born to infected mothers, and with Guillain-Barré syndrome in infected adults. Development of ZIKV vaccines has focused predominantly on the induction of neutralizing antibodies, although a suboptimal antibody response may theoretically enhance disease severity through antibody-dependent enhancement (ADE). Here, we report induction of a protective anti-ZIKV CD8+ T cell response in the HLA-B*0702 Ifnar1-/- transgenic mice using an alphavirus-based replicon RNA vaccine expressing ZIKV nonstructural protein NS3, a potent T cell antigen. The NS3 vaccine did not induce a neutralizing antibody response but elicited polyfunctional CD8+ T cells that were necessary and sufficient for preventing death in lethally infected adult mice and fetal growth restriction in infected pregnant mice. These data identify CD8+ T cells as the major mediators of ZIKV NS3 vaccine-induced protection and suggest a new strategy to develop safe and effective anti-flavivirus vaccines.

Cross-Reactive T Cell Immunity to Dengue and Zika Viruses: New Insights Into Vaccine Development.

Dengue virus (DENV) is a member of the Flavivirus family that includes Zika virus (ZIKV), West Nile virus, Japanese encephalitis virus, and yellow fever virus. As the most prevalent of the flaviviruses, DENV is responsible for tens of millions of infections each year. The clinical manifestations of infection with one of the four DENV serotypes (DENV1-4) range from no symptoms to hemorrhagic fever and shock ("severe dengue"), which is fatal in ~25,000 patients annually. Many factors contribute to the development of severe dengue, including the DENV serotype and host expression of certain HLA alleles; however, it now seems clear that pre-existing immunity to DENV-and possibly other flaviviruses-is a major precipitating factor. While primary infection with one DENV serotype elicits strong cellular and humoral immune responses that likely confer long-lived protection against the same serotype, subsequent infection with a different serotype carries an increased risk of developing severe dengue. Thus, primary DENV infection elicits cross-reactive immunity that may be protective or pathogenic, depending on the context of the subsequent infection. Many flaviviruses share high sequence homology, raising the possibility that cross-reactive immunity to one virus may contribute to protection against or pathogenesis of a second virus in a similar manner. In addition, several flaviviruses are now endemic in overlapping geographic regions, underscoring the need to gain more knowledge about the mechanisms underlying cross-reactive immunity to different DENV serotypes and flaviviruses. Here, we review our current understanding of T cell immunity to DENV, focusing on cross-reactivity with other serotypes and flaviviruses such as ZIKV, and the role of DENV-elicited CD4+ and CD8+ T cells in protection. Recent work in this area supports a beneficial role for cross-reactive T cells and provides new insights into the design of safe and efficient flavivirus/pan-flavivirus vaccines.

Human Polyclonal Antibodies Prevent Lethal Zika Virus Infection in Mice.

Zika virus (ZIKV) is an emerging mosquito-borne flavivirus that represents a major threat to global health. ZIKV infections in adults are generally asymptomatic or present with mild symptoms. However, recent outbreaks of ZIKV have revealed that it can cause Congenital Zika Syndrome in neonates and Guillain-Barré syndrome in adults. Currently, no ZIKV-specific vaccines or antiviral treatments are available. In this study, we tested the efficacy of convalescent plasma IgG hyperimmune product (ZIKV-IG) isolated from individuals with high neutralizing anti-ZIKV titers as a therapeutic candidate against ZIKV infection using a model of ZIKV infection in Ifnar1-/- mice. ZIKV-IG successfully protected mice from lethal ZIKV challenge. In particular, ZIKV-IG treatment at 24 hours after lethal ZIKV infection improved survival by reducing weight loss and tissue viral burden and improving clinical score. Additionally, ZIKV-IG eliminated ZIKV-induced tissue damage and inflammation in the brain and liver. These results indicate that ZIKV-IG is efficacious against ZIKV, suggesting this human polyclonal antibody is a viable candidate for further development as a treatment against human ZIKV infection.

Cross-reactive Dengue virus-specific CD8+ T cells protect against Zika virus during pregnancy.

As Zika virus (ZIKV) emerges into Dengue virus (DENV)-endemic areas, cases of ZIKV infection in DENV-immune pregnant women may rise. Here we show that prior DENV immunity affects maternal and fetal ZIKV infection in pregnancy using sequential DENV and ZIKV infection models. Fetuses in ZIKV-infected DENV-immune dams were normal sized, whereas fetal demise occurred in non-immune dams. Moreover, reduced ZIKV RNA is present in the placenta and fetuses of ZIKV-infected DENV-immune dams. DENV cross-reactive CD8+ T cells expand in the maternal spleen and decidua of ZIKV-infected dams, their depletion increases ZIKV infection in the placenta and fetus, and results in fetal demise. The inducement of cross-reactive CD8+ T cells via peptide immunization or adoptive transfer results in decreased ZIKV infection in the placenta. Prior DENV immunity can protect against ZIKV infection during pregnancy in mice, and CD8+ T cells are sufficient for this cross-protection. This has implications for understanding the natural history of ZIKV in DENV-endemic areas and the development of optimal ZIKV vaccines.

Mapping and Role of the CD8+ T Cell Response During Primary Zika Virus Infection in Mice.

CD8+ T cells may play a dual role in protection against and pathogenesis of flaviviruses, including Zika virus (ZIKV). We evaluated the CD8+ T cell response in ZIKV-infected LysMCre+IFNARfl/fl C57BL/6 (H-2b) mice lacking the type I interferon receptor in a subset of myeloid cells. In total, 26 and 15 CD8+ T cell-reactive peptides for ZIKV African (MR766) and Asian (FSS13025) lineage strains, respectively, were identified and validated. CD8+ T cells from infected mice were polyfunctional and mediated cytotoxicity. Adoptive transfer of ZIKV-immune CD8+ T cells reduced viral burdens, whereas their depletion led to higher tissue burdens, and CD8-/- mice displayed higher mortality with ZIKV infection. Collectively, these results demonstrate that CD8+ T cells protect against ZIKV infection. Further, this study provides a T cell competent mouse model for investigating ZIKV-specific T cell responses.

Dengue virus-reactive CD8+ T cells mediate cross-protection against subsequent Zika virus challenge.

Zika virus (ZIKV) and dengue virus (DENV) are antigenically related flaviviruses that share cross-reactivity in antibody and T cell responses, and co-circulate in increasing numbers of countries. Whether pre-existing DENV immunity can cross-protect or enhance ZIKV infection during sequential infection of the same host is unknown. Here, we show that DENV-immune Ifnar1 -/- or wild-type C57BL/6 mice infected with ZIKV have cross-reactive immunity to subsequent ZIKV infection and pathogenesis. Adoptive transfer and cell depletion studies demonstrate that DENV-immune CD8+ T cells predominantly mediate cross-protective responses to ZIKV. In contrast, passive transfer studies suggest that DENV-immune serum does not protect against ZIKV infection. Thus, CD8+ T cell immunity generated during primary DENV infection can confer protection against secondary ZIKV infection in mice. Further optimization of current DENV vaccines for T cell responses might confer cross-protection and prevent antibody-mediated enhancement of ZIKV infection.