Publications by authors named "Sean A Diehl"

45 Publications

UV decontamination of personal protective equipment with idle laboratory biosafety cabinets during the COVID-19 pandemic.

PLoS One 2021 26;16(7):e0241734. Epub 2021 Jul 26.

Cleveland Clinic Lerner Research Institute and Case Western Reserve University School of Medicine, Cleveland, OH, United States of America.

Personal protective equipment (PPE) is crucially important to the safety of both patients and medical personnel, particularly in the event of an infectious pandemic. As the incidence of Coronavirus Disease 2019 (COVID-19) increases exponentially in the United States and many parts of the world, healthcare provider demand for these necessities is currently outpacing supply. In the midst of the current pandemic, there has been a concerted effort to identify viable ways to conserve PPE, including decontamination after use. In this study, we outline a procedure by which PPE may be decontaminated using ultraviolet (UV) radiation in biosafety cabinets (BSCs), a common element of many academic, public health, and hospital laboratories. According to the literature, effective decontamination of N95 respirator masks or surgical masks requires UV-C doses of greater than 1 Jcm-2, which was achieved after 4.3 hours per side when placing the N95 at the bottom of the BSCs tested in this study. We then demonstrated complete inactivation of the human coronavirus NL63 on N95 mask material after 15 minutes of UV-C exposure at 61 cm (232 μWcm-2). Our results provide support to healthcare organizations looking for methods to extend their reserves of PPE.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0241734PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8312969PMC
August 2021

Immunotranscriptomic profiling the acute and clearance phases of a human challenge dengue virus serotype 2 infection model.

Nat Commun 2021 05 24;12(1):3054. Epub 2021 May 24.

Department of Microbiology and Molecular Genetics, Larner College of Medicine, University of Vermont, Burlington, VT, USA.

About 20-25% of dengue virus (DENV) infections become symptomatic ranging from self-limiting fever to shock. Immune gene expression changes during progression to severe dengue have been documented in hospitalized patients; however, baseline or kinetic information is difficult to standardize in natural infection. Here we profile the host immunotranscriptome response in humans before, during, and after infection with a partially attenuated rDEN2Δ30 challenge virus (ClinicalTrials.gov NCT02021968). Inflammatory genes including type I interferon and viral restriction pathways are induced during DENV2 viremia and return to baseline after viral clearance, while others including myeloid, migratory, humoral, and growth factor immune regulation factors pathways are found at non-baseline levels post-viremia. Furthermore, pre-infection baseline gene expression is useful to predict rDEN2Δ30-induced immune responses and the development of rash. Our results suggest a distinct immunological profile for mild rDEN2Δ30 infection and offer new potential biomarkers for characterizing primary DENV infection.
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http://dx.doi.org/10.1038/s41467-021-22930-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8144425PMC
May 2021

A tetravalent live attenuated dengue virus vaccine stimulates balanced immunity to multiple serotypes in humans.

Nat Commun 2021 02 17;12(1):1102. Epub 2021 Feb 17.

Department of Microbiology and Immunology, University of North Carolina School of Medicine, Chapel Hill, NC, USA.

The four-dengue virus (DENV) serotypes infect several hundred million people annually. For the greatest safety and efficacy, tetravalent DENV vaccines are designed to stimulate balanced protective immunity to all four serotypes. However, this has been difficult to achieve. Clinical trials with a leading vaccine demonstrated that unbalanced replication and immunodominance of one vaccine component over others can lead to low efficacy and vaccine enhanced severe disease. The Laboratory of Infectious Diseases at the National Institutes of Health has developed a live attenuated tetravalent DENV vaccine (TV003), which is currently being tested in phase 3 clinical trials. Here we report, our study to determine if TV003 stimulate balanced and serotype-specific (TS) neutralizing antibody (nAb) responses to each serotype. Serum samples from twenty-one dengue-naive individuals participated under study protocol CIR287 (ClinicalTrials.gov NCT02021968) are analyzed 6 months after vaccination. Most subjects (76%) develop TS nAbs to 3 or 4 DENV serotypes, indicating immunity is induced by each vaccine component. Vaccine-induced TS nAbs map to epitopes known to be targets of nAbs in people infected with wild type DENVs. Following challenge with a partially attenuated strain of DENV2, all 21 subjects are protected from the efficacy endpoints. However, some vaccinated individuals develop post challenge nAb boost, while others mount post-challenge antibody responses that are consistent with sterilizing immunity. TV003 vaccine induced DENV2 TS nAbs are associated with sterilizing immunity. Our results indicate that nAbs to TS epitopes on each serotype may be a better correlate than total levels of nAbs currently used for guiding DENV vaccine development.
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http://dx.doi.org/10.1038/s41467-021-21384-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7889627PMC
February 2021

A Novel Antigenic Site Spanning Domains I and III of the Zika Virus Envelope Glycoprotein Is the Target of Strongly Neutralizing Human Monoclonal Antibodies.

J Virol 2021 04 12;95(9). Epub 2021 Apr 12.

Department of Medicine, Division of Infectious Disease, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, USA

Zika virus (ZIKV), a mosquito-transmitted flavivirus, caused a large epidemic in Latin America between 2015 and 2017. Effective ZIKV vaccines and treatments are urgently needed to prevent future epidemics and severe disease sequelae. People infected with ZIKV develop strongly neutralizing antibodies linked to viral clearance and durable protective immunity. To understand the mechanisms of protective immunity and to support the development of ZIKV vaccines, we characterize here a strongly neutralizing antibody, B11F, isolated from a patient who recovered from ZIKV. Our results indicate that B11F targets a complex epitope on the virus that spans domains I and III of the envelope glycoprotein. While previous studies point to quaternary epitopes centered on domain II of the ZIKV E glycoprotein as targets of strongly neutralizing and protective human antibodies, we uncover a new site spanning domains I and III as a target of strongly neutralizing human antibodies. People infected with Zika virus develop durable neutralizing antibodies that prevent repeat infections. In the current study, we characterize a ZIKV-neutralizing human monoclonal antibody isolated from a patient after recovery. Our studies establish a novel site on the viral envelope that is targeted by human neutralizing antibodies. Our results are relevant to understanding how antibodies block infection and to guiding the design and evaluation of candidate vaccines.
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http://dx.doi.org/10.1128/JVI.02423-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8104094PMC
April 2021

Stimulation of B Cell Immunity in Flavivirus-Naive Individuals by the Tetravalent Live Attenuated Dengue Vaccine TV003.

Cell Rep Med 2020 Dec 22;1(9):100155. Epub 2020 Dec 22.

Department of Microbiology and Molecular Genetics, Vaccine Testing Center, Larner College of Medicine, University of Vermont, Burlington, VT 05405, USA.

The tetravalent live attenuated dengue vaccine candidate TV003 induces neutralizing antibodies against all four dengue virus serotypes (DENV1-DENV4) and protects against experimental challenge with DENV2 in humans. Here, we track vaccine viremia and B and T cell responses to this vaccination/challenge model to understand how vaccine viremia links adaptive immunity and development of protective antibody responses. TV003 viremia triggers an acute plasmablast response that, in combination with DENV-specific CD4 T cells, correlates with serum neutralizing antibodies. TV003 vaccinees develop DENV2-reactive memory B cells, including serotype-specific and multivalent specificities in line with the composition of serum antibodies. There is no post-challenge plasmablast response in vaccinees, although stronger and earlier post-TV003 plasmablast responses associate with sterile humoral protection from DENV2 challenge. TV003 vaccine triggers plasmablasts and memory B cells, which, with support from CD4 T cells, functionally link early vaccine viremia and the serum antibody responses.
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http://dx.doi.org/10.1016/j.xcrm.2020.100155DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7762770PMC
December 2020

Jobs, Housing, and Mask Wearing: Cross-Sectional Study of Risk Factors for COVID-19.

JMIR Public Health Surveill 2021 01 11;7(1):e24320. Epub 2021 Jan 11.

Department of Medicine, Larner College of Medicine, University of Vermont, Burlington, VT, United States.

Background: Many studies have focused on the characteristics of symptomatic patients with COVID-19 and clinical risk factors. This study reports the prevalence of COVID-19 in an asymptomatic population of a hospital service area (HSA) and identifies factors that affect exposure to the virus.

Objective: The aim of this study is to measure the prevalence of COVID-19 in an HSA, identify factors that may increase or decrease the risk of infection, and analyze factors that increase the number of daily contacts.

Methods: This study surveyed 1694 patients between April 30 and May 13, 2020, about their work and living situations, income, behavior, sociodemographic characteristics, and prepandemic health characteristics. This data was linked to testing data for 454 of these patients, including polymerase chain reaction test results and two different serologic assays. Positivity rate was used to calculate approximate prevalence, hospitalization rate, and infection fatality rate (IFR). Survey data was used to analyze risk factors, including the number of contacts reported by study participants. The data was also used to identify factors increasing the number of daily contacts, such as mask wearing and living environment.

Results: We found a positivity rate of 2.2%, a hospitalization rate of 1.2%, and an adjusted IFR of 0.55%. A higher number of daily contacts with adults and older adults increases the probability of becoming infected. Occupation, living in an apartment versus a house, and wearing a face mask outside work increased the number of daily contacts.

Conclusions: Studying prevalence in an asymptomatic population revealed estimates of unreported COVID-19 cases. Occupational, living situation, and behavioral data about COVID-19-protective behaviors such as wearing a mask may aid in the identification of nonclinical factors affecting the number of daily contacts, which may increase SARS-CoV-2 exposure.
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http://dx.doi.org/10.2196/24320DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7800904PMC
January 2021

Kinetics and isotype assessment of antibodies targeting the spike protein receptor-binding domain of severe acute respiratory syndrome-coronavirus-2 in COVID-19 patients as a function of age, biological sex and disease severity.

Clin Transl Immunology 2020 7;9(10):e1189. Epub 2020 Oct 7.

Department of Microbiology and Molecular Genetics Larner College of Medicine, University of Vermont Burlington VT USA.

Objectives: There is an incomplete understanding of the host humoral immune response to severe acute respiratory syndrome (SARS)-coronavirus (CoV)-2, which underlies COVID-19, during acute infection. Host factors such as age and sex as well as the kinetics and functionality of antibody responses are important factors to consider as vaccine development proceeds. The receptor-binding domain of the CoV spike (RBD-S) protein mediates host cell binding and infection and is a major target for vaccine design to elicit neutralising antibodies.

Methods: We assessed serum anti-SARS-CoV-2 RBD-S IgG, IgM and IgA antibodies by a two-step ELISA and neutralising antibodies in a cross-sectional study of hospitalised COVID-19 patients of varying disease severities. Anti-RBD-S IgG levels were also determined in asymptomatic seropositives.

Results: We found equivalent levels of anti-RBD-S antibodies in male and female patients and no age-related deficiencies even out to 93 years of age. The anti-RBD-S response was evident as little as 6 days after onset of symptoms and for at least 5 weeks after symptom onset. Anti-RBD-S IgG, IgM and IgA responses were simultaneously induced within 10 days after onset, with anti-RBD-S IgG sustained over a 5-week period. Anti-RBD-S antibodies strongly correlated with neutralising activity. Lastly, anti-RBD-S IgG responses were higher in symptomatic COVID-19 patients during acute infection compared with asymptomatic seropositive donors.

Conclusion: Our results suggest that anti-RBD-S IgG reflect functional immune responses to SARS-CoV-2, but do not completely explain age- and sex-related disparities in COVID-19 fatalities.
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http://dx.doi.org/10.1002/cti2.1189DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7541824PMC
October 2020

Direct RT-qPCR detection of SARS-CoV-2 RNA from patient nasopharyngeal swabs without an RNA extraction step.

PLoS Biol 2020 10 2;18(10):e3000896. Epub 2020 Oct 2.

Division of Immunobiology, Department of Medicine, Robert Larner, M.D. College of Medicine, University of Vermont, Burlington, Vermont, United States of America.

The ongoing COVID-19 pandemic has created an unprecedented need for rapid diagnostic testing. The World Health Organization (WHO) recommends a standard assay that includes an RNA extraction step from a nasopharyngeal (NP) swab followed by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) to detect the purified SARS-CoV-2 RNA. The current global shortage of RNA extraction kits has caused a severe bottleneck to COVID-19 testing. The goal of this study was to determine whether SARS-CoV-2 RNA could be detected from NP samples via a direct RT-qPCR assay that omits the RNA extraction step altogether. The direct RT-qPCR approach correctly identified 92% of a reference set of blinded NP samples (n = 155) demonstrated to be positive for SARS-CoV-2 RNA by traditional clinical diagnostic RT-qPCR that included an RNA extraction. Importantly, the direct method had sufficient sensitivity to reliably detect those patients with viral loads that correlate with the presence of infectious virus. Thus, this strategy has the potential to ease supply choke points to substantially expand COVID-19 testing and screening capacity and should be applicable throughout the world.
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http://dx.doi.org/10.1371/journal.pbio.3000896DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7556528PMC
October 2020

Kinetics and Isotype Assessment of Antibodies Targeting the Spike Protein Receptor Binding Domain of SARS-CoV-2 In COVID-19 Patients as a function of Age and Biological Sex.

medRxiv 2020 Jul 16. Epub 2020 Jul 16.

SARS-CoV-2 is the newly emerged virus responsible for the global COVID-19 pandemic. There is an incomplete understanding of the host humoral immune response to SARS-CoV-2 during acute infection. Host factors such as age and sex as well the kinetics and functionality of antibody responses are important factors to consider as vaccine development proceeds. The receptor-binding domain of the CoV spike (RBD-S) protein is important in host cell recognition and infection and antibodies targeting this domain are often neutralizing. In a cross-sectional study of anti-RBD-S antibodies in COVID-19 patients we found equivalent levels in male and female patients and no age-related deficiencies even out to 93 years of age. The anti-RBD-S response was evident as little as 6 days after onset of symptoms and for at least 5 weeks after symptom onset. Anti-RBD-S IgG, IgM, and IgA responses were simultaneously induced within 10 days after onset, but isotype-specific kinetics differed such that anti-RBD-S IgG was most sustained over a 5-week period. The kinetics and magnitude of neutralizing antibody formation strongly correlated with that seen for anti-RBD-S antibodies. Our results suggest age- and sex- related disparities in COVID-19 fatalities are not explained by anti-RBD-S responses. The multi-isotype anti-RBD-S response induced by live virus infection could serve as a potential marker by which to monitor vaccine-induced responses.
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http://dx.doi.org/10.1101/2020.07.15.20154443DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7386516PMC
July 2020

DIRECT RT-qPCR DETECTION OF SARS-CoV-2 RNA FROM PATIENT NASOPHARYNGEAL SWABS WITHOUT AN RNA EXTRACTION STEP.

bioRxiv 2020 Apr 6. Epub 2020 Apr 6.

Department of Medicine, Division of Immunobiology, Robert Larner, M.D. College of Medicine, University of Vermont, Burlington VT, 05405, USA.

The ongoing COVID-19 pandemic has caused an unprecedented need for rapid diagnostic testing. The Centers for Disease Control and Prevention (CDC) and the World Health Organization (WHO) recommend a standard assay that includes an RNA extraction step from a nasopharyngeal (NP) swab followed by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) to detect the purified SARS-CoV-2 RNA. The current global shortage of RNA extraction kits has caused a severe bottleneck to COVID-19 testing. We hypothesized that SARS-CoV-2 RNA could be detected from NP samples via a direct RT-qPCR assay that omits the RNA extraction step altogether, and tested this hypothesis on a series of blinded clinical samples. The direct RT-qPCR approach correctly identified 92% of NP samples (n = 155) demonstrated to be positive for SARS-CoV-2 RNA by traditional clinical diagnostic RT-qPCR that included an RNA extraction. Thus, direct RT-qPCR could be a front-line approach to identify the substantial majority of COVID-19 patients, reserving a repeat test with RNA extraction for those individuals with high suspicion of infection but an initial negative result. This strategy would drastically ease supply chokepoints of COVID-19 testing and should be applicable throughout the world.
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http://dx.doi.org/10.1101/2020.03.20.001008DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7239058PMC
April 2020

Rapid Induction and Maintenance of Virus-Specific CD8 T and CD4 T Cells Following Protective Vaccination Against Dengue Virus Challenge in Humans.

Front Immunol 2020 24;11:479. Epub 2020 Mar 24.

Department of Microbiology and Molecular Genetics, Larner College of Medicine, University of Vermont, Burlington, VT, United States.

Dengue virus (DENV) is a mosquito-borne flavivirus that causes serious human disease. The current lack of an effective vaccine to simultaneously protect against the four serotypes of DENV in seronegative individuals is a major unmet medical need. Further, the immunological basis for protective immunity in the setting of DENV infection or vaccination is not fully understood. Our team has developed a live attenuated tetravalent dengue virus vaccine that provides complete protection in a human model of dengue virus challenge. The goal of this study was to define, in the context of protective human vaccination, the quality of vaccine-induced DENV-specific CD8 and CD4 T cells and the temporal dynamics associated with their formation and maintenance. Multifunctional, DENV-specific CD8 and CD4 T cells developed 8-14 days after vaccination and were maintained for at least 6 months. Virus-specific CD8 T cells were a mixture of effector memory T cells (T) and effector memory T cells re-expressing CD45RA (T), with T cells predominating until day 21 post-vaccination and T cells thereafter. The majority of virus-specific CD4 T cells were T with a small fraction being T. The frequency of virus-specific CD8 and CD4 T cells were further skewed to the T phenotype following either a second dose of the tetravalent vaccine or challenge with a single serotype of DENV. Collectively, our study has defined the phenotypic profile of antiviral CD8 and CD4 T cells associated with protective immunity to DENV infection and the kinetics of their formation and maintenance.
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http://dx.doi.org/10.3389/fimmu.2020.00479DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7105617PMC
March 2021

T Cell Responses Induced by Attenuated Flavivirus Vaccination Are Specific and Show Limited Cross-Reactivity with Other Flavivirus Species.

J Virol 2020 05 4;94(10). Epub 2020 May 4.

Division of Vaccine Discovery, La Jolla Institute for Immunology, La Jolla, California, USA

Members of the flavivirus genus share a high level of sequence similarity and often circulate in the same geographical regions. However, whether T cells induced by one viral species cross-react with other related flaviviruses has not been globally addressed. In this study, we tested pools of epitopes derived from dengue (DENV), Zika (ZIKV), Japanese encephalitis (JEV), West Nile (WNV), and yellow fever (YFV) viruses by intracellular cytokine staining (ICS) using peripheral blood mononuclear cells (PBMCs) of individuals naturally exposed to DENV or immunized with DENV (TV005) or YF17D vaccine. CD8 T cell responses recognized epitopes from multiple flaviviruses; however, the magnitude of cross-reactive responses was consistently severalfold lower than those to the autologous epitope pools and was associated with lower expression of activation markers such as CD40L, CD69, and CD137. Next, we characterized the antigen sensitivity of short-term T cell lines (TCL) representing 29 different individual epitope/donor combinations. TCL derived from DENV monovalent vaccinees induced CD8 and CD4 T cells that cross-reacted within the DENV serocomplex but were consistently associated with >100-fold-lower antigen sensitivity for most other flaviviruses, with no cross-recognition of YFV-derived peptides. CD8 and CD4 TCL from YF17D vaccinees were associated with very limited cross-reactivity with any other flaviviruses and in five out of eight cases >1,000-fold-lower antigen sensitivity. Overall, our data suggest limited cross-reactivity for both CD4 and CD8 T cell responses between flaviviruses and have implications for understanding immunity elicited by natural infection and strategies to develop live attenuated vaccines against flaviviral species. The envelope (E) protein is the dominant target of neutralizing antibodies for dengue virus (DENV) and yellow fever virus (YFV). Accordingly, several DENV vaccine constructs use the E protein in a live attenuated vaccine format, utilizing a backbone derived from a heterologous flavivirus (such as YF) as a delivery vector. This backbone comprises the nonstructural (NS) and capsid (C) antigens, which are dominant targets of T cell responses. Here, we demonstrate that cross-reactivity at the level of T cell responses among different flaviviruses is very limited, despite high levels of sequence homology. Thus, the use of heterologous flavivirus species as a live attenuated vaccine vector is not likely to generate optimal T cell responses and might thus impair vaccine performance.
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http://dx.doi.org/10.1128/JVI.00089-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7199411PMC
May 2020

Human antibody response to Zika targets type-specific quaternary structure epitopes.

JCI Insight 2019 04 18;4(8). Epub 2019 Apr 18.

Department of Microbiology and Immunology, University of North Carolina School of Medicine, Chapel Hill, North Carolina, USA.

The recent Zika virus (ZIKV) epidemic in the Americas has revealed rare but serious manifestations of infection. ZIKV has emerged in regions endemic for dengue virus (DENV), a closely related mosquito-borne flavivirus. Cross-reactive antibodies confound studies of ZIKV epidemiology and pathogenesis. The immune responses to ZIKV may be different in people, depending on their DENV immune status. Here, we focus on the human B cell and antibody response to ZIKV as a primary flavivirus infection to define the properties of neutralizing and protective antibodies generated in the absence of preexisting immunity to DENV. The plasma antibody and memory B cell response is highly ZIKV type-specific, and ZIKV-neutralizing antibodies mainly target quaternary structure epitopes on the viral envelope. To map viral epitopes targeted by protective antibodies, we isolated 2 type-specific monoclonal antibodies (mAbs) from a ZIKV case. Both mAbs were strongly neutralizing in vitro and protective in vivo. The mAbs recognize distinct epitopes centered on domains I and II of the envelope protein. We also demonstrate that the epitopes of these mAbs define antigenic regions commonly targeted by plasma antibodies in individuals from endemic and nonendemic regions who have recovered from ZIKV infections.
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http://dx.doi.org/10.1172/jci.insight.124588DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6538335PMC
April 2019

Longitudinal analysis of acute and convalescent B cell responses in a human primary dengue serotype 2 infection model.

EBioMedicine 2019 Mar 8;41:465-478. Epub 2019 Mar 8.

Department of Microbiology and Molecular Genetics, Vaccine Testing Center, Larner College of Medicine, University of Vermont, Burlington, VT 05405, USA. Electronic address:

Background: Acute viral infections induce a rapid and transient increase in antibody-secreting plasmablasts. At convalescence, memory B cells (MBC) and long-lived plasma cells (LLPC) are responsible for long-term humoral immunity. Following an acute viral infection, the specific properties and relationships between antibodies produced by these B cell compartments are poorly understood.

Methods: We utilized a controlled human challenge model of primary dengue virus serotype 2 (DENV2) infection to study acute and convalescent B-cell responses.

Findings: The level of DENV2 replication was correlated with the magnitude of the plasmablast response. Functional analysis of plasmablast-derived monoclonal antibodies showed that the DENV2-specific response was dominated by cells producing DENV2 serotype-specific antibodies. DENV2-neutralizing antibodies targeted quaternary structure epitopes centered on domain III of the viral envelope protein (EDIII). Functional analysis of MBC and serum antibodies from the same subjects six months post-challenge revealed maintenance of the serotype-specific response in both compartments. The serum response mainly targeted DENV2 serotype-specific epitopes on EDIII.

Interpretation: Our data suggest overall functional alignment of DENV2-specific responses from the plasmablast, through the MBC and LLPC compartments following primary DENV2 inflection. These results provide enhanced resolution of the temporal and specificity of the B cell compartment in viral infection and serve as framework for evaluation of B cell responses in challenge models.

Funding: This study was supported by the Bill and Melinda Gates Foundation and the National Institutes of Health.
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http://dx.doi.org/10.1016/j.ebiom.2019.02.060DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6444124PMC
March 2019

Human megakaryocytes possess intrinsic antiviral immunity through regulated induction of IFITM3.

Blood 2019 05 5;133(19):2013-2026. Epub 2019 Feb 5.

University of Utah Molecular Medicine Program, Salt Lake City, UT.

Evolving evidence indicates that platelets and megakaryocytes (MKs) have unexpected activities in inflammation and infection; whether viral infections upregulate biologically active, antiviral immune genes in platelets and MKs is unknown, however. We examined antiviral immune genes in these cells in dengue and influenza infections, viruses that are global public health threats. Using complementary biochemical, pharmacological, and genetic approaches, we examined the regulation and function of interferon-induced transmembrane protein 3 (IFITM3), an antiviral immune effector gene not previously studied in human platelets and MKs. IFITM3 was markedly upregulated in platelets isolated from patients during clinical influenza and dengue virus (DENV) infections. Lower IFITM3 expression in platelets correlated with increased illness severity and mortality in patients. Administering a live, attenuated DENV vaccine to healthy subjects significantly increased platelet IFITM3 expression. Infecting human MKs with DENV selectively increased type I interferons and IFITM3. Overexpression of IFITM3 in MKs was sufficient to prevent DENV infection. In naturally occurring, genetic loss-of-function studies, MKs from healthy subjects harboring a homozygous mutation in IFITM3 (rs12252-C, a common single-nucleotide polymorphism in areas of the world where DENV is endemic) were significantly more susceptible to DENV infection. DENV-induced MK secretion of interferons prevented infection of bystander MKs and hematopoietic stem cells. Thus, viral infections upregulate IFITM3 in human platelets and MKs, and IFITM3 expression is associated with adverse clinical outcomes. These observations establish, for the first time, that human MKs possess antiviral functions, preventing DENV infection of MKs and hematopoietic stem cells after local immune signaling.
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http://dx.doi.org/10.1182/blood-2018-09-873984DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6509546PMC
May 2019

Neonatal vitamin A supplementation and immune responses to oral polio vaccine in Zimbabwean infants.

Trans R Soc Trop Med Hyg 2019 03;113(3):110-115

Zvitambo Institute for Maternal and Child Health Research, 16 Lauchlan Avenue, Harare, Zimbabwe.

Background: Micronutrient deficiencies may contribute to reduced oral vaccine immunogenicity in developing countries. We hypothesised that neonatal vitamin A supplementation (NVAS) would improve oral vaccine responses.

Methods: We performed a cross-sectional study of infants recruited at birth to the Zimbabwe Vitamin A for Mothers and Babies (ZVITAMBO) trial, a randomised controlled trial of single, high-dose NVAS vs placebo conducted in Zimbabwe between 1997-2001. We measured poliovirus-specific IgA to type 1-3 polio strains by semiquantitative capture ELISA in cryopreserved plasma samples collected at 6 months of age.

Results: A total of 181 infants fulfilled inclusion criteria, of whom 80 were randomised to NVAS and 101 to placebo. There were no significant differences in baseline characteristics between groups. At 6 months of age, median (IQR) vaccine titres for infants randomised to NVAS vs placebo were 932 (421-3001) vs 1774 (711-5431) for Sabin-1 (p=0.04); 1361 (705-3402) vs 2309 (1081-4283) for Sabin-2 (p=0.15); and 1584 (796-4216) vs 2260 (996-5723) for Sabin-3 (p=0.14), respectively. After adjusting for breast feeding status, birth weight, season and infant sex in a linear regression model, there was only weak evidence of difference in log mean titres between vitamin A and placebo groups for Sabin-1 (p=0.08) and no evidence of difference in log mean titres for Sabin-2 and Sabin-3.

Conclusions: NVAS did not augment oral polio vaccine responses in Zimbabwean infants. Further research is required to understand the impact of NVAS on responses to other oral vaccines.The trial is registered with clinicaltrials.gov identifier: NCT00198718.
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http://dx.doi.org/10.1093/trstmh/try126DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6391935PMC
March 2019

Viridot: An automated virus plaque (immunofocus) counter for the measurement of serological neutralizing responses with application to dengue virus.

PLoS Negl Trop Dis 2018 10 24;12(10):e0006862. Epub 2018 Oct 24.

Department of Biology, University of Florida, Gainesville, FL, United States.

The gold-standard method for quantifying neutralizing antibody responses to many viruses, including dengue virus (DENV), is the plaque reduction neutralization test (PRNT, also called the immunofocus reduction neutralization test). The PRNT conducted on 96-well plates is high-throughput and requires a smaller volume of antiserum than on 6- or 24-well plates, but manual plaque counting is challenging and existing automated plaque counters are expensive or difficult to optimize. We have developed Viridot (Viridot package), a program for R with a user interface in shiny, that counts viral plaques of a variety of phenotypes, estimates neutralizing antibody titers, and performs other calculations of use to virologists. The Viridot plaque counter includes an automatic parameter identification mode (misses <10 plaques/well for 87% of diverse DENV strains [n = 1521]) and a mode that allows the user to fine-tune the parameters used for counting plaques. We compared standardized manual and Viridot plaque counting methods applied to the same wells by two analyses and found that Viridot plaque counts were as similar to the same analyst's manual count (Lin's concordance correlation coefficient, ρc = 0.99 [95% confidence interval: 0.99-1.00]) as manual counts between analysts (ρc = 0.99 [95% CI: 0.98-0.99]). The average ratio of neutralizing antibody titers based on manual counted plaques to Viridot counted plaques was 1.05 (95% CI: 0.98-1.14), similar to the average ratio of antibody titers based on manual plaque counts by the two analysts (1.06 [95% CI: 0.84-1.34]). Across diverse DENV and ZIKV strains (n = 14), manual and Viridot plaque counts were mostly consistent (range of ρc = 0.74 to 1.00) and the average ratio of antibody titers based on manual and Viridot counted plaques was close to 1 (0.94 [0.86-1.02]). Thus, Viridot can be used for plaque counting and neutralizing antibody titer estimation of diverse DENV strains and potentially other viruses on 96-well plates as well as for formalization of plaque-counting rules for standardization across experiments and analysts.
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http://dx.doi.org/10.1371/journal.pntd.0006862DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6226209PMC
October 2018

Rotavirus-Specific Immunoglobulin A Responses Are Impaired and Serve as a Suboptimal Correlate of Protection Among Infants in Bangladesh.

Clin Infect Dis 2018 07;67(2):186-192

Department of Medicine, Vaccine Testing Center, University of Vermont Larner College of Medicine, Burlington.

Background: Rotavirus (RV)-specific immunoglobulin A (IgA) responses following oral RV vaccination are impaired in low-income countries, where the utility of RV-IgA as a correlate of protection (CoP) remains unclear. In a monovalent oral RV vaccine (Rotarix) efficacy trial among infants in Dhaka, Bangladesh, we identified factors associated with poor RV-IgA responses and explored the utility of RV-IgA as a CoP.

Methods: Infants were randomized to receive Rotarix or no Rotarix at 10 and 17 weeks of life and followed with active diarrheal surveillance. RV-IgA concentration, seroconversion, and seropositivity were determined at 18 weeks of life and analyzed for correlation(s) with rotavirus diarrhea (RVD) and for contribution to Rotarix vaccine effect.

Results: Among vaccinated infants, overall RV-IgA geometric mean concentration was 21 U/mL; only 27% seroconverted and 32% were seropositive after vaccination. Increased RV-specific maternal antibodies significantly impaired immunogenicity. Seroconversion was associated with reduced risk of RVD through 1 year of life, but RV-IgA seropositivity only explained 7.8% of the vaccine effect demonstrated by the clinical endpoint (RVD).

Conclusions: RV-IgA responses were low among infants in Bangladesh and were significantly impaired by maternal antibodies. RV-IgA is a suboptimal CoP in this setting; an improved CoP for RV in low-income countries is needed.

Clinical Trials Registration: NCT01375647.
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http://dx.doi.org/10.1093/cid/ciy076DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6030840PMC
July 2018

Histo-Blood Group Antigen Phenotype Determines Susceptibility to Genotype-Specific Rotavirus Infections and Impacts Measures of Rotavirus Vaccine Efficacy.

J Infect Dis 2018 04;217(9):1399-1407

Department of Medicine, University of Vermont Larner College of Medicine, Burlington.

Background: Lewis and secretor histo-blood group antigens (HBGAs) have been associated with decreased susceptibility to P[8] genotype rotavirus (RV) infections. Efficacy of vaccines containing attenuated P[8] strains is decreased in low-income countries. Host phenotype might impact vaccine efficacy (VE) by altering susceptibility to vaccination or RV diarrhea (RVD). We performed a substudy in a monovalent RV vaccine (RV1) efficacy trial in Bangladesh to determine the impact of Lewis and secretor status on risk of RVD and VE.

Methods: In infants randomized to receive RV1 or no RV1 at 10 and 17 weeks with 1 year of complete active diarrheal surveillance, we performed Lewis and secretor phenotyping and genotyped the infecting strain of each episode of RVD.

Results: A vaccine containing P[8] RV protected secretors and nonsecretors similarly. However, unvaccinated nonsecretors had a reduced risk of RVD (relative risk, 0.53 [95% confidence interval, .36-.79]) mediated by complete protection from P[4] but not P[8] RVs. This effect reduced VE in nonsecretors to 31.7%, compared to 56.2% among secretors, and decreased VE for the overall cohort.

Conclusions: Host HBGA status may impact VE estimates by altering susceptibility to RV in unvaccinated children; future trials should therefore account for HBGA status.

Clinical Trials Registration: NCT01375647.
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http://dx.doi.org/10.1093/infdis/jiy054DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5894073PMC
April 2018

Global Assessment of Dengue Virus-Specific CD4 T Cell Responses in Dengue-Endemic Areas.

Front Immunol 2017 13;8:1309. Epub 2017 Oct 13.

Division of Clinical Immunology and Allergy, School of Medicine, University of São Paulo, São Paulo, Brazil.

Background: Dengue is a major public health problem worldwide. Assessment of adaptive immunity is important to understanding immunopathology and to define correlates of protection against dengue virus (DENV). To enable global assessment of CD4 T cell responses, we mapped HLA-DRB1-restricted DENV-specific CD4 T cell epitopes in individuals previously exposed to DENV in the general population of the dengue-endemic region of Managua, Nicaragua.

Methods: HLA class II epitopes in the population of Managua were identified by an IFNγ ELISPOT assay. CD4 T cells purified by magnetic bead negative selection were stimulated with HLA-matched epitope pools in the presence of autologous antigen-presenting cells, followed by pool deconvolution to identify specific epitopes. The epitopes identified in this study were combined with those previously identified in the DENV endemic region of Sri Lanka, to generate a "megapool" (MP) consisting of 180 peptides specifically designed to achieve balanced HLA and DENV serotype coverage. The DENV CD4MP was validated by intracellular cytokine staining assays.

Results: We detected responses directed against a total of 431 epitopes, representing all 4 DENV serotypes, restricted by 15 different HLA-DRB1 alleles. The responses were associated with a similar pattern of protein immunodominance, overall higher magnitude of responses, as compared to what was observed previously in the Sri Lanka region. Based on these epitope mapping studies, we designed a DENV CD4 MP with higher and more consistent coverage, which allowed the detection of CD4 T cell DENV responses in various cohorts of DENV exposed donors worldwide, including donors from Nicaragua, Brazil, Singapore, Sri Lanka, and U.S. domestic flavivirus-naïve subjects immunized with Tetravalent Dengue Live-Attenuated Vaccine (TV005). This broad reactivity reflects that the 21 HLA-DRB1 alleles analyzed in this and previous studies account for more than 80% of alleles present with a phenotypic frequency ≥5% worldwide, corresponding to 92% phenotypic coverage of the general population (i.e., 92% of individuals express at least one of these alleles).

Conclusion: The DENV CD4 MP can be utilized to measure responses to DENV irrespective of geographical location.
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http://dx.doi.org/10.3389/fimmu.2017.01309DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5646259PMC
October 2017

Prior Dengue Virus Exposure Shapes T Cell Immunity to Zika Virus in Humans.

J Virol 2017 12 30;91(24). Epub 2017 Nov 30.

Blood Systems Research Institute, San Francisco, California, USA.

While progress has been made in characterizing humoral immunity to Zika virus (ZIKV) in humans, little is known regarding the corresponding T cell responses to ZIKV. Here, we investigate the kinetics and viral epitopes targeted by T cells responding to ZIKV and address the critical question of whether preexisting dengue virus (DENV) T cell immunity modulates these responses. We find that memory T cell responses elicited by prior infection with DENV or vaccination with tetravalent dengue attenuated vaccines (TDLAV) recognize ZIKV-derived peptides. This cross-reactivity is explained by the sequence similarity of the two viruses, as the ZIKV peptides recognized by DENV-elicited memory T cells are identical or highly conserved in DENV and ZIKV. DENV exposure prior to ZIKV infection also influences the timing and magnitude of the T cell response. ZIKV-reactive T cells in the acute phase of infection are detected earlier and in greater magnitude in DENV-immune patients. Conversely, the frequency of ZIKV-reactive T cells continues to rise in the convalescent phase in DENV-naive donors but declines in DENV-preexposed donors, compatible with more efficient control of ZIKV replication and/or clearance of ZIKV antigen. The quality of responses is also influenced by previous DENV exposure, and ZIKV-specific CD8 T cells from DENV-preexposed donors selectively upregulated granzyme B and PD1, unlike DENV-naive donors. Finally, we discovered that ZIKV structural proteins (E, prM, and C) are major targets of both the CD4 and CD8 T cell responses, whereas DENV T cell epitopes are found primarily in nonstructural proteins. The issue of potential ZIKV and DENV cross-reactivity and how preexisting DENV T cell immunity modulates Zika T cell responses is of great relevance, as the two viruses often cocirculate and Zika virus has been spreading in geographical regions where DENV is endemic or hyperendemic. Our data show that memory T cell responses elicited by prior infection with DENV recognize ZIKV-derived peptides and that DENV exposure prior to ZIKV infection influences the timing, magnitude, and quality of the T cell response. Additionally, we show that ZIKV-specific responses target different proteins than DENV-specific responses, pointing toward important implications for vaccine design against this global threat.
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http://dx.doi.org/10.1128/JVI.01469-17DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5709580PMC
December 2017

Immune responses to oral poliovirus vaccine in HIV-exposed uninfected Zimbabwean infants.

Hum Vaccin Immunother 2017 11 31;13(11):2543-2547. Epub 2017 Aug 31.

a Zvitambo Institute for Maternal and Child Health Research , Harare , Zimbabwe.

It remains uncertain whether HIV-exposed uninfected (HEU) infants have impaired responses to oral vaccines. We performed a cross-sectional study of 6-month-old infants recruited at birth to the ZVITAMBO trial in Zimbabwe between 1997-2001, before introduction of prevention of mother-to-child transmission interventions. We measured poliovirus-specific IgA to type 1-3 polio strains by semi-quantitative capture ELISA in cryopreserved serum samples collected from 85 HEU and 101 HIV-unexposed infants at 6 months of age, one month after their last immunisation with trivalent OPV. Almost all infants were breastfed, with the majority in both groups mixed breastfed (70.6% HEU versus 71.3% HIV-unexposed). Median (IQR) vaccine titers for HEU and HIV-unexposed infants were 1592 (618-4896) vs. 1774 (711-5431) for Sabin 1 (P = 0.46); 1895 (810-4398) vs. 2308 (1081-4283) for Sabin 2 (P = 0.52); and 1798 (774-4192) vs. 2260 (996-5723) for Sabin 3 (P = 0.18). There were no significant differences in vaccine titers between HEU and HIV-unexposed infants, suggesting that vertical HIV exposure does not impact oral poliovirus vaccine immunogenicity.
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http://dx.doi.org/10.1080/21645515.2017.1359454DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5703368PMC
November 2017

In a randomized trial, the live attenuated tetravalent dengue vaccine TV003 is well-tolerated and highly immunogenic in subjects with flavivirus exposure prior to vaccination.

PLoS Negl Trop Dis 2017 05 8;11(5):e0005584. Epub 2017 May 8.

Vaccine Testing Center, Department of Medicine, University of Vermont Larner College of Medicine, Burlington, Vermont, United States of America.

Infection caused by the four serotypes of dengue virus (DENV-1-4) is a leading cause of mosquito-borne disease. Clinically-severe dengue disease is more common when secondary dengue infection occurs following prior infection with a heterologous dengue serotype. Other flaviviruses such as yellow fever virus, Japanese encephalitis virus, and Zika virus, can also elicit antibodies which are cross-reactive to DENV. As candidate dengue vaccines become available in endemic settings and for individuals who have received other flavivirus vaccines, it is important to examine vaccine safety and immunogenicity in these flavivirus-experienced populations. We performed a randomized, controlled trial of the National Institutes of Health live attenuated tetravalent dengue vaccine candidate (TV003) in fifty-eight individuals with prior exposure to flavivirus infection or vaccine. As in prior studies of this vaccine in flavivirus-naive volunteers, flavivirus-experienced subjects received two doses of vaccine six months apart and were followed closely for clinical events, laboratory changes, viremia, and neutralizing antibody titers. TV003 was well tolerated with few adverse events other than rash, which was predominately mild. Following one dose, 87% of vaccinees had an antibody response to all four serotypes (tetravalent response), suggesting a robust immune response. In addition, 76% of vaccinees were viremic; mean peak titers ranged from 0.68–1.1 log10 PFU/mL and did not differ by serotype. The second dose of TV003 was not associated with viremia, rash, or a sustained boost in antibody titers indicating that a single dose of the vaccine is likely sufficient to prevent viral replication and thus protect against disease. In comparison to the viremia and neutralizing antibody response elicited by TV003 in flavivirus-naïve subjects from prior studies, we found that subjects who were flavivirus-exposed prior to vaccination exhibited slightly higher DENV-3 viremia, higher neutralizing antibody titers to DENV-2, -3, and -4, and a higher tetravalent response frequency after TV003 administration. In summary, we demonstrate that the NIH tetravalent dengue vaccine TV003 is well-tolerated in flavivirus-experienced individuals and elicits robust post-vaccination neutralizing antibody titers.

Trial Registration: ClinicalTrials.gov NCT01506570.
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http://dx.doi.org/10.1371/journal.pntd.0005584DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5436874PMC
May 2017

Patterns of Cellular Immunity Associated with Experimental Infection with rDEN2Δ30 (Tonga/74) Support Its Suitability as a Human Dengue Virus Challenge Strain.

J Virol 2017 Apr 29;91(8). Epub 2017 Mar 29.

Division of Vaccine Discovery, La Jolla Institute for Allergy and Immunology, La Jolla, California, USA

A deletion variant of the dengue virus (DENV) serotype 2 (DENV2) Tonga/74 strain lacking 30 nucleotides from its 3' untranslated region (rDEN2Δ30) has previously been established for use in a controlled human DENV challenge model. To evaluate if this model is appropriate for the derivation of correlates of protection for DENV vaccines on the basis of cellular immunity, we wanted to compare the cellular immune response to this challenge strain to the response induced by natural infection. To achieve this, we predicted HLA class I- and class II-restricted peptides from rDEN2Δ30 and used them in a gamma interferon enzyme-linked immunosorbent spot assay to interrogate CD8 and CD4 T cell responses in healthy volunteers infected with rDEN2Δ30. At the level of CD8 responses, vigorous responses were detected in approximately 80% of donors. These responses were similar in terms of the magnitude and the numbers of epitopes recognized to the responses previously observed in peripheral blood mononuclear cells from donors from regions where DENV is hyperendemic. The similarity extended to the immunodominance hierarchy of the DENV nonstructural proteins, with NS3, NS5, and NS1 being dominant in both donor cohorts. At the CD4 level, the responses to rDEN2Δ30 vaccination were less vigorous than those to natural DENV infection and were more focused on nonstructural proteins. The epitopes recognized following rDEN2Δ30 infection and natural infection were largely overlapping for both the CD8 (100%) and CD4 (85%) responses. Finally, rDEN2Δ30 induced stronger CD8 responses than other, more attenuated DENV isolates. The lack of a known correlate of protection and the failure of a neutralizing antibody to correlate with protection against dengue virus have highlighted the need for a human DENV challenge model to better evaluate the candidate live attenuated dengue vaccines. In this study, we sought to characterize the immune profiles of rDEN2Δ30-infected subjects and to compare the profiles with those for subjects from areas where DENV is hyperendemic. Our data demonstrate that T cell responses to rDENV2Δ30 are largely similar to those to natural infection in terms of specificity, highlighting that the response to this virus in humans is appropriate as a model for the T cell response to primary DENV2 infection.
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http://dx.doi.org/10.1128/JVI.02133-16DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5375662PMC
April 2017

Human CD4 T Cell Responses to an Attenuated Tetravalent Dengue Vaccine Parallel Those Induced by Natural Infection in Magnitude, HLA Restriction, and Antigen Specificity.

J Virol 2017 03 14;91(5). Epub 2017 Feb 14.

Division of Vaccine Discovery, La Jolla Institute for Allergy and Immunology, La Jolla, California, USA

Dengue virus (DENV) is responsible for growing numbers of infections worldwide and has proven to be a significant challenge for vaccine development. We previously demonstrated that CD8 T cell responses elicited by a dengue live attenuated virus (DLAV) vaccine resemble those observed after natural infection. In this study, we screened peripheral blood mononuclear cells (PBMCs) from donors vaccinated with a tetravalent DLAV vaccine (TV005) with pools of dengue virus-derived predicted major histocompatibility complex (MHC) class II binding peptides. The definition of CD4 T cell responses after live vaccination is important because CD4 T cells are known contributors to host immunity, including cytokine production, help for CD8 T and B cells, and direct cytotoxicity against infected cells. While responses to all antigens were observed, DENV-specific CD4 T cells were focused predominantly on the capsid and nonstructural NS3 and NS5 antigens. Importantly, CD4 T cell responses in vaccinees were similar in magnitude and breadth to those after natural infection, recognized the same antigen hierarchy, and had similar profiles of HLA restriction. We conclude that TV005 vaccination has the capacity to elicit CD4 cell responses closely mirroring those observed in a population associated with natural immunity. The development of effective vaccination strategies against dengue virus infection is of high global public health interest. Here we study the CD4 T cell responses elicited by a tetravalent live attenuated dengue vaccine and show that they resemble responses seen in humans naturally exposed to dengue virus. This is an important issue, since it is likely that optimal immunity induced by a vaccine requires induction of CD4 responses against the same antigens as those recognized as dominant in natural infection. Detailed knowledge of the T cell response may further contribute to the identification of robust correlates of protection against dengue virus.
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http://dx.doi.org/10.1128/JVI.02147-16DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5309943PMC
March 2017

Delayed Dosing of Oral Rotavirus Vaccine Demonstrates Decreased Risk of Rotavirus Gastroenteritis Associated With Serum Zinc: A Randomized Controlled Trial.

Clin Infect Dis 2016 Sep 23;63(5):634-41. Epub 2016 May 23.

Department of Medicine, Vaccine Testing Center and Unit of Infectious Diseases, University of Vermont College of Medicine, Burlington.

Background: Rotavirus is the world's leading cause of childhood diarrheal death. Despite successes, oral rotavirus vaccines are less effective in developing countries. In an urban slum of Dhaka, we performed active diarrhea surveillance to evaluate monovalent G1P[8] rotavirus vaccine (RV1) efficacy and understand variables contributing to risk of rotavirus diarrhea (RVD).

Methods: We performed a randomized controlled trial of monovalent oral rotavirus vaccine (RV1). Seven hundred healthy infants received RV1 or no RV1 (1:1) using delayed dosing (10 and 17 weeks) and were followed for 1 year. Intensive diarrhea surveillance was performed. The primary outcome was ≥1 episode of RVD. Nutritional, socioeconomic, and immunologic factors were assessed by logistic regression best-subsets analysis for association with risk of RVD and interactions with vaccine arm.

Results: Incidence of all RVD was 38.3 cases per 100 person-years. Per-protocol RV1 efficacy was 73.5% (95% confidence interval [CI], 45.8%-87.0%) against severe RVD and 51% (95% CI, 33.8%-63.7%) against all RVD. Serum zinc level (odds ratio [OR], 0.77; P = .002) and lack of rotavirus immunoglobulin A (IgA) seroconversion (OR, 1.95; P = .018) were associated with risk of RVD, independent of vaccination status. Water treatment and exclusive breastfeeding were of borderline significance. Factors not associated with RVD included height for age at 10 weeks, vitamin D, retinol binding protein, maternal education, household income, and sex.

Conclusions: In an urban slum with high incidence of RVD, the efficacy of RV1 against severe RVD was higher than anticipated in the setting of delayed dosing. Lower serum zinc level and lack of IgA seroconversion were associated with increased risk of RVD independent of vaccination.

Clinical Trials Registration: NCT01375647.
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http://dx.doi.org/10.1093/cid/ciw346DOI Listing
September 2016

The live attenuated dengue vaccine TV003 elicits complete protection against dengue in a human challenge model.

Sci Transl Med 2016 Mar 16;8(330):330ra36. Epub 2016 Mar 16.

Center for Immunization Research, Johns Hopkins Bloomberg School of Public Health, Baltimore, MD 21205, USA.

A dengue human challenge model can be an important tool to identify candidate dengue vaccines that should be further evaluated in large efficacy trials in endemic areas. Dengue is responsible for about 390 million infections annually. Protective efficacy results for the most advanced dengue vaccine candidate (CYD) were disappointing despite its ability to induce neutralizing antibodies against all four dengue virus (DENV) serotypes. TV003 is a live attenuated tetravalent DENV vaccine currently in phase 2 evaluation. To better assess the protective efficacy of TV003, a randomized double-blind, placebo-controlled trial in which recipients of TV003 or placebo were challenged 6 months later with a DENV-2 strain, rDEN2Δ30, was conducted. The primary endpoint of the trial was protection against dengue infection, defined as rDEN2Δ30 viremia. Secondary endpoints were protection against rash and neutropenia. All 21 recipients of TV003 who were challenged with rDEN2Δ30 were protected from infection with rDEN2Δ30. None developed viremia, rash, or neutropenia after challenge. In contrast, 100% of the 20 placebo recipients who were challenged with rDEN2Δ30 developed viremia, 80% developed rash, and 20% developed neutropenia. TV003 induced complete protection against challenge with rDEN2Δ30 administered 6 months after vaccination. TV003 will be further evaluated in dengue-endemic areas. The controlled dengue human challenge model can accelerate vaccine development by evaluating the protection afforded by the vaccine, thereby eliminating poor candidates from further consideration before the initiation of large efficacy trials.
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http://dx.doi.org/10.1126/scitranslmed.aaf1517DOI Listing
March 2016

Mitochondrial Ca²⁺ and membrane potential, an alternative pathway for Interleukin 6 to regulate CD4 cell effector function.

Elife 2015 May 14;4. Epub 2015 May 14.

Department of Medicine, Immunobiology Program, University of Vermont, Burlington, United States.

IL-6 plays an important role in determining the fate of effector CD4 cells and the cytokines that these cells produce. Here we identify a novel molecular mechanism by which IL-6 regulates CD4 cell effector function. We show that IL-6-dependent signal facilitates the formation of mitochondrial respiratory chain supercomplexes to sustain high mitochondrial membrane potential late during activation of CD4 cells. Mitochondrial hyperpolarization caused by IL-6 is uncoupled from the production of ATP by oxidative phosphorylation. However, it is a mechanism to raise the levels of mitochondrial Ca(2+) late during activation of CD4 cells. Increased levels of mitochondrial Ca(2+) in the presence of IL-6 are used to prolong Il4 and Il21 expression in effector CD4 cells. Thus, the effect of IL-6 on mitochondrial membrane potential and mitochondrial Ca(2+) is an alternative pathway by which IL-6 regulates effector function of CD4 cells and it could contribute to the pathogenesis of inflammatory diseases.
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http://dx.doi.org/10.7554/eLife.06376DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4447996PMC
May 2015
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