Immunogenicity and protection of a variant nanoparticle vaccine that confers broad neutralization against SARS-CoV-2 variants.

SARS-CoV-2 variants have emerged with elevated transmission and a higher risk of infection for vaccinated individuals. We demonstrate that a recombinant prefusion-stabilized spike (rS) protein vaccine based on Beta/B.1.351 (rS-Beta) produces a robust anamnestic response in baboons against SARS-CoV-2 variants when given as a booster one year after immunization with NVX-CoV2373. Additionally, rS-Beta is highly immunogenic in mice and produces neutralizing antibodies against WA1/2020, Beta/B.1.351, and Omicron/BA.1. Mice vaccinated with two doses of Novavax prototype NVX-CoV2373 (rS-WU1) or rS-Beta alone, in combination, or heterologous prime-boost, are protected from challenge. Virus titer is undetectable in lungs in all vaccinated mice, and Th1-skewed cellular responses are observed. We tested sera from a panel of variant spike protein vaccines and find broad neutralization and inhibition of spike:ACE2 binding from the rS-Beta and rS-Delta vaccines against a variety of variants including Omicron. This study demonstrates that rS-Beta vaccine alone or in combination with rS-WU1 induces antibody-and cell-mediated responses that are protective against challenge with SARS-CoV-2 variants and offers broader neutralizing capacity than a rS-WU1 prime/boost regimen alone. Together, these nonhuman primate and murine data suggest a Beta variant booster dose could elicit a broad immune response to fight new and future SARS-CoV-2 variants.

Zika virus spreads through infection of lymph node-resident macrophages.

To disseminate through the body, Zika virus (ZIKV) is thought to exploit the mobility of myeloid cells, in particular monocytes and dendritic cells. However, the timing and mechanisms underlying shuttling of the virus by immune cells remains unclear. To understand the early steps in ZIKV transit from the skin, at different time points, we spatially mapped ZIKV infection in lymph nodes (LNs), an intermediary site en route to the blood. Contrary to prevailing hypotheses, migratory immune cells are not required for the virus to reach the LNs or blood. Instead, ZIKV rapidly infects a subset of sessile CD169+ macrophages in the LNs, which release the virus to infect downstream LNs. Infection of CD169+ macrophages alone is sufficient to initiate viremia. Overall, our experiments indicate that macrophages that reside in the LNs contribute to initial ZIKV spread. These studies enhance our understanding of ZIKV dissemination and identify another anatomical site for potential antiviral intervention.

Safety and immunogenicity following a homologous booster dose of a SARS-CoV-2 recombinant spike protein vaccine (NVX-CoV2373): a secondary analysis of a randomised, placebo-controlled, phase 2 trial.

BACKGROUND: Emerging SARS-CoV-2 variants and evidence of waning vaccine efficacy present substantial obstacles towards controlling the COVID-19 pandemic. Booster doses of SARS-CoV-2 vaccines might address these concerns by amplifying and broadening the immune responses seen with initial vaccination regimens. We aimed to assess the immunogenicity and safety of a homologous booster dose of a SARS-CoV-2 recombinant spike protein vaccine (NVX-CoV2373). METHODS: This secondary analysis of a phase 2, randomised study assessed a single booster dose of a SARS-CoV-2 recombinant spike protein vaccine with Matrix-M adjuvant (NVX-CoV2373) in healthy adults aged 18-84 years, recruited from 17 clinical centres in the USA and Australia. Eligible participants had a BMI of 17-35 kg/m2 and, for women, were heterosexually inactive or using contraception. Participants who had a history of SARS-CoV or SARS-CoV-2, confirmed diagnosis of COVID-19, serious chronic medical conditions, or were pregnant or breastfeeding were excluded. Approximately 6 months following their primary two-dose vaccination series (administered day 0 and day 21), participants who received placebo for their primary vaccination series received a placebo booster (group A) and participants who received NVX-CoV2373 for their primary vaccination series (group B) were randomly assigned (1:1) again, via centralised interactive response technology system, to receive either placebo (group B1) or a single booster dose of NVX-CoV2373 (5 µg SARS-CoV-2 rS with 50 µg Matrix-M adjuvant; group B2) via intramuscular injection; randomisation was stratified by age and study site. Vaccinations were administered by designated site personnel who were masked to treatment assignment, and participants and other site staff were also masked. Administration personnel also assessed the outcome. The primary endpoints are safety (unsolicited adverse events) and reactogenicity (solicited local and systemic) events and immunogenicity (serum IgG antibody concentrations for the SARS-CoV-2 rS protein antigen) assessed 14 days after the primary vaccination series (day 35) and 28 days following booster (day 217). Safety was analysed in all participants in groups A, B1, and B2, according to the treatment received; immunogenicity was analysed in the per-protocol population (ie, participants in groups A, B1, and B2) who received all assigned doses and who did not test SARS-CoV-2-positive or received an authorised vaccine, analysed according to treatment assignment). This trial is registered with ClinicalTrials.gov, NCT04368988. FINDINGS: 1610 participants were screened from Aug 24, 2020, to Sept 25, 2020. 1282 participants were enrolled, of whom 173 were assigned again to placebo (group A), 106 were re-randomised to NVX-CoV2373-placebo (group B1), and 104 were re-randomised to NVX-CoV2373-NVX-CoV2373 (group B2); after accounting for exclusions and incorrect administration, 172 participants in group A, 102 in group B1, and 105 in group B2 were analysed for safety. Following the active booster, the proportion of participants with available data reporting local (80 [82%] of 97 participants had any adverse event; 13 [13%] had a grade ≥3 event) and systemic (75 [77%] of 98 participants had any adverse event; 15 [15%] had a grade ≥3 event) reactions was higher than after primary vaccination (175 [70%] of 250 participants had any local adverse event, 13 [5%] had a grade ≥3 event; 132 [53%] of 250 had any systemic adverse event, 14 [6%] had a grade ≥3 event). Local and systemic events were transient in nature (median duration 1·0-2·5 days). In the per-protocol immunogenicity population at day 217 (167 participants in group A, 101 participants in group B1, 101 participants in group B2), IgG geometric mean titres (GMT) had increased by 4·7-fold and MN50 GMT by 4·1-fold for the ancestral SARS-CoV-2 strain compared with the day 35 titres. INTERPRETATION: Administration of a booster dose of NVX-CoV2373 resulted in an incremental increase in reactogenicity. For both the prototype strain and all variants evaluated, immune responses following the booster were similar to or higher than those associated with high levels of efficacy in phase 3 studies of the vaccine. These data support the use of NVX-CoV2373 in booster programmes. FUNDING: Novavax and the Coalition for Epidemic Preparedness Innovations.

Fab and Fc contribute to maximal protection against SARS-CoV-2 following NVX-CoV2373 subunit vaccine with Matrix-M vaccination.

Recently approved vaccines have shown remarkable efficacy in limiting SARS-CoV-2-associated disease. However, with the variety of vaccines, immunization strategies, and waning antibody titers, defining the correlates of immunity across a spectrum of antibody titers is urgently required. Thus, we profiled the humoral immune response in a cohort of non-human primates immunized with a recombinant SARS-CoV-2 spike glycoprotein (NVX-CoV2373) at two doses, administered as a single- or two-dose regimen. Both antigen dose and boosting significantly altered neutralization titers and Fc-effector profiles, driving unique vaccine-induced antibody fingerprints. Combined differences in antibody effector functions and neutralization were associated with distinct levels of protection in the upper and lower respiratory tract. Moreover, NVX-CoV2373 elicited antibodies that functionally targeted emerging SARS-CoV-2 variants. Collectively, the data presented here suggest that a single dose may prevent disease via combined Fc/Fab functions but that two doses may be essential to block further transmission of SARS-CoV-2 and emerging variants.

Limited Flavivirus Cross-Reactive Antibody Responses Elicited by a Zika Virus Deoxyribonucleic Acid Vaccine Candidate in Humans.

Zika virus (ZIKV) deoxyribonucleic acid vaccine VRC5283 encoding viral structural genes has been shown to be immunogenic in humans. Recognizing that antigenically related flaviviruses cocirculate in regions with ZIKV activity, we explored the degree of antibody cross-reactivity elicited by this vaccine candidate using genetically diverse flaviviruses. The antibody response of vaccinated individuals with no evidence of prior flavivirus infection or vaccine experience had a limited capacity to bind heterologous viruses. In contrast, vaccine-elicited antibodies from individuals with prior flavivirus experience had a greater capacity to bind, but not neutralize, distantly related flaviviruses. These findings suggest that prior flavivirus exposure shapes the humoral immune response to vaccination.

Collaboration between the Fab and Fc contribute to maximal protection against SARS-CoV-2 following NVX-CoV2373 subunit vaccine with Matrix-M™ vaccination.

Recently approved vaccines have already shown remarkable protection in limiting SARS-CoV-2 associated disease. However, immunologic mechanism(s) of protection, as well as how boosting alters immunity to wildtype and newly emerging strains, remain incompletely understood. Here we deeply profiled the humoral immune response in a cohort of non-human primates immunized with a stable recombinant full-length SARS-CoV-2 spike (S) glycoprotein (NVX-CoV2373) at two dose levels, administered as a single or two-dose regimen with a saponin-based adjuvant Matrix-M™. While antigen dose had some effect on Fc-effector profiles, both antigen dose and boosting significantly altered overall titers, neutralization and Fc-effector profiles, driving unique vaccine-induced antibody fingerprints. Combined differences in antibody effector functions and neutralization were strongly associated with distinct levels of protection in the upper and lower respiratory tract, pointing to the presence of combined, but distinct, compartment-specific neutralization and Fc-mechanisms as key determinants of protective immunity against infection. Moreover, NVX-CoV2373 elicited antibodies functionally target emerging SARS-CoV-2 variants, collectively pointing to the critical collaborative role for Fab and Fc in driving maximal protection against SARS-CoV-2. Collectively, the data presented here suggest that a single dose may prevent disease, but that two doses may be essential to block further transmission of SARS-CoV-2 and emerging variants.

Collaboration between the Fab and Fc contribute to maximal protection against SARS-CoV-2 in nonhuman primates following NVX-CoV2373 subunit vaccine with Matrix-M™ vaccination.

Recently approved vaccines have already shown remarkable protection in limiting SARS-CoV-2 associated disease. However, immunologic mechanism(s) of protection, as well as how boosting alters immunity to wildtype and newly emerging strains, remain incompletely understood. Here we deeply profiled the humoral immune response in a cohort of non-human primates immunized with a stable recombinant full-length SARS-CoV-2 spike (S) glycoprotein (NVX-CoV2373) at two dose levels, administered as a single or two-dose regimen with a saponin-based adjuvant Matrix-M™. While antigen dose had some effect on Fc-effector profiles, both antigen dose and boosting significantly altered overall titers, neutralization and Fc-effector profiles, driving unique vaccine-induced antibody fingerprints. Combined differences in antibody effector functions and neutralization were strongly associated with distinct levels of protection in the upper and lower respiratory tract, pointing to the presence of combined, but distinct, compartment-specific neutralization and Fc-mechanisms as key determinants of protective immunity against infection. Moreover, NVX-CoV2373 elicited antibodies functionally target emerging SARS-CoV-2 variants, collectively pointing to the critical collaborative role for Fab and Fc in driving maximal protection against SARS-CoV-2. Collectively, the data presented here suggest that a single dose may prevent disease, but that two doses may be essential to block further transmission of SARS-CoV-2 and emerging variants. HIGHLIGHTS: NVX-CoV2373 subunit vaccine elicits receptor blocking, virus neutralizing antibodies, and Fc-effector functional antibodies.The vaccine protects against respiratory tract infection and virus shedding in non-human primates (NHPs).Both neutralizing and Fc-effector functions contribute to protection, potentially through different mechanisms in the upper and lower respiratory tract.Both macaque and human vaccine-induced antibodies exhibit altered Fc-receptor binding to emerging mutants.

SARS-CoV-2 spike glycoprotein vaccine candidate NVX-CoV2373 immunogenicity in baboons and protection in mice.

The COVID-19 pandemic continues to spread throughout the world with an urgent need for a safe and protective vaccine to effectuate herd protection and control the spread of SARS-CoV-2. Here, we report the development of a SARS-CoV-2 subunit vaccine (NVX-CoV2373) from the full-length spike (S) protein that is stable in the prefusion conformation. NVX-CoV2373 S form 27.2-nm nanoparticles that are thermostable and bind with high affinity to the human angiotensin-converting enzyme 2 (hACE2) receptor. In mice, low-dose NVX-CoV2373 with saponin-based Matrix-M adjuvant elicit high titer anti-S IgG that blocks hACE2 receptor binding, neutralize virus, and protects against SARS-CoV-2 challenge with no evidence of vaccine-associated enhanced respiratory disease. NVX-CoV2373 also elicits multifunctional CD4+ and CD8+ T cells, CD4+ follicular helper T cells (Tfh), and antigen-specific germinal center (GC) B cells in the spleen. In baboons, low-dose levels of NVX-CoV2373 with Matrix-M was also highly immunogenic and elicited high titer anti-S antibodies and functional antibodies that block S-protein binding to hACE2 and neutralize virus infection and antigen-specific T cells. These results support the ongoing phase 1/2 clinical evaluation of the safety and immunogenicity of NVX-CoV2373 with Matrix-M (NCT04368988).

NVX-CoV2373 vaccine protects cynomolgus macaque upper and lower airways against SARS-CoV-2 challenge.

There is an urgent need for a safe and protective vaccine to control the global spread of SARS-CoV-2 and prevent COVID-19. Here, we report the immunogenicity and protective efficacy of a SARS-CoV-2 subunit vaccine (NVX-CoV2373) produced from the full-length SARS-CoV-2 spike (S) glycoprotein stabilized in the prefusion conformation. Cynomolgus macaques (Macaca fascicularis) immunized with NVX-CoV2373 and the saponin-based Matrix-M™ adjuvant induced anti-S antibody that was neutralizing and blocked binding to the human angiotensin-converting enzyme 2 (hACE2) receptor. Following intranasal and intratracheal challenge with SARS-CoV-2, immunized macaques were protected against upper and lower infection and pulmonary disease. These results support ongoing phase 1/2 clinical studies of the safety and immunogenicity of NVX-CoV2327 vaccine (NCT04368988).

Distinct neutralizing antibody correlates of protection among related Zika virus vaccines identify a role for antibody quality.

The emergence of Zika virus (ZIKV) in the Americas stimulated the development of multiple ZIKV vaccine candidates. We previously developed two related DNA vaccine candidates encoding ZIKV structural proteins that were immunogenic in animal models and humans. We sought to identify neutralizing antibody (NAb) properties induced by each vaccine that correlated with protection in nonhuman primates (NHPs). Despite eliciting equivalent NAb titers in NHPs, these vaccines were not equally protective. The transfer of equivalent titers of vaccine-elicited NAb into AG129 mice also revealed nonequivalent protection, indicating qualitative differences among antibodies (Abs) elicited by these vaccines. Both vaccines elicited Abs with similar binding titers against envelope protein monomers and those incorporated into virus-like particles, as well as a comparable capacity to orchestrate phagocytosis. Functional analysis of vaccine-elicited NAbs from NHPs and humans revealed a capacity to neutralize the structurally mature form of the ZIKV virion that varied in magnitude among vaccine candidates. Conversely, sensitivity to the virion maturation state was not a characteristic of NAbs induced by natural or experimental infection. Passive transfer experiments in mice revealed that neutralization of mature ZIKV virions more accurately predicts protection from ZIKV infection. These findings demonstrate that NAb correlates of protection may differ among vaccine antigens when assayed using standard neutralization platforms and suggest that measurements of Ab quality, including the capacity to neutralize mature virions, will be critical for defining correlates of ZIKV vaccine-induced immunity.

Cross-Reactive Flavivirus Antibody: Friend and Foe?

The introduction of Zika virus (ZIKV) into the Americas sparked interest in the influence of heterologous antibodies on sequential ZIKV and dengue virus (DENV) infections. In this issue of Cell Host & Microbe, Fowler et al. (2018) demonstrate in mice that maternally acquired anti-ZIKV antibodies exacerbate disease following DENV challenge.

VPg unlinkase/TDP2 in cardiovirus infected cells: Re-localization and proteolytic cleavage.

Cardioviruses cause diseases in many animals including, in rare cases, humans. Although they share common features with all picornaviruses, cardioviruses have unique properties that distinguish them from other family members, including enteroviruses. One feature shared by all picornaviruses is the covalent attachment of VPg to the 5' end of genomic RNA via a phosphotyrosyl linkage. For enteroviruses, this linkage is cleaved by a host cell protein, TDP2. Since TDP2 is divergently required during enterovirus infections, we determined if TDP2 is necessary during infection by the prototype cardiovirus, EMCV. We found that EMCV yields are reduced in the absence of TDP2. We observed a decrease in viral protein accumulation and viral RNA replication in the absence of TDP2. In contrast to enterovirus infections, we found that TDP2 is modified at peak times of EMCV infection. This finding suggests a unique mechanism for cardioviruses to regulate TDP2 activity during infection.

Divergent Requirement for a DNA Repair Enzyme during Enterovirus Infections.

UNLABELLED: Viruses of the Enterovirus genus of picornaviruses, including poliovirus, coxsackievirus B3 (CVB3), and human rhinovirus, commandeer the functions of host cell proteins to aid in the replication of their small viral genomic RNAs during infection. One of these host proteins is a cellular DNA repair enzyme known as 5' tyrosyl-DNA phosphodiesterase 2 (TDP2). TDP2 was previously demonstrated to mediate the cleavage of a unique covalent linkage between a viral protein (VPg) and the 5' end of picornavirus RNAs. Although VPg is absent from actively translating poliovirus mRNAs, the removal of VPg is not required for the in vitro translation and replication of the RNA. However, TDP2 appears to be excluded from replication and encapsidation sites during peak times of poliovirus infection of HeLa cells, suggesting a role for TDP2 during the viral replication cycle. Using a mouse embryonic fibroblast cell line lacking TDP2, we found that TDP2 is differentially required among enteroviruses. Our single-cycle viral growth analysis shows that CVB3 replication has a greater dependency on TDP2 than does poliovirus or human rhinovirus replication. During infection, CVB3 protein accumulation is undetectable (by Western blot analysis) in the absence of TDP2, whereas poliovirus protein accumulation is reduced but still detectable. Using an infectious CVB3 RNA with a reporter, CVB3 RNA could still be replicated in the absence of TDP2 following transfection, albeit at reduced levels. Overall, these results indicate that TDP2 potentiates viral replication during enterovirus infections of cultured cells, making TDP2 a potential target for antiviral development for picornavirus infections. IMPORTANCE: Picornaviruses are one of the most prevalent groups of viruses that infect humans and livestock worldwide. These viruses include the human pathogens belonging to the Enterovirus genus, such as poliovirus, coxsackievirus B3 (CVB3), and human rhinovirus. Diseases caused by enteroviruses pose a major problem for public health and have significant economic impact. Poliovirus can cause paralytic poliomyelitis. CVB3 can cause hand, foot, and mouth disease and myocarditis. Human rhinovirus is the causative agent of the common cold, which has a severe economic impact due to lost productivity and severe health consequences in individuals with respiratory dysfunction, such as asthma. By gaining a better understanding of the enterovirus replication cycle, antiviral drugs against enteroviruses may be developed. Here, we report that the absence of the cellular enzyme TDP2 can significantly decrease viral yields of poliovirus, CVB3, and human rhinovirus, making TDP2 a potential target for an antiviral against enterovirus infections.

Activating receptor NKG2D targets RAE-1-expressing allogeneic neural precursor cells in a viral model of multiple sclerosis.

Transplantation of major histocompatibility complex-mismatched mouse neural precursor cells (NPCs) into mice persistently infected with the neurotropic JHM strain of mouse hepatitis virus (JHMV) results in rapid rejection that is mediated, in part, by T cells. However, the contribution of the innate immune response to allograft rejection in a model of viral-induced neurological disease has not been well defined. Herein, we demonstrate that the natural killer (NK) cell-expressing-activating receptor NKG2D participates in transplanted allogeneic NPC rejection in mice persistently infected with JHMV. Cultured NPCs derived from C57BL/6 (H-2(b) ) mice express the NKG2D ligand retinoic acid early precursor transcript (RAE)-1 but expression was dramatically reduced upon differentiation into either glia or neurons. RAE-1(+) NPCs were susceptible to NK cell-mediated killing whereas RAE-1(-) cells were resistant to lysis. Transplantation of C57BL/6-derived NPCs into JHMV-infected BALB/c (H-2(d) ) mice resulted in infiltration of NKG2D(+) CD49b(+) NK cells and treatment with blocking antibody specific for NKG2D increased survival of allogeneic NPCs. Furthermore, transplantation of differentiated RAE-1(-) allogeneic NPCs into JHMV-infected BALB/c mice resulted in enhanced survival, highlighting a role for the NKG2D/RAE-1 signaling axis in allograft rejection. We also demonstrate that transplantation of allogeneic NPCs into JHMV-infected mice resulted in infection of the transplanted cells suggesting that these cells may be targets for infection. Viral infection of cultured cells increased RAE-1 expression, resulting in enhanced NK cell-mediated killing through NKG2D recognition. Collectively, these results show that in a viral-induced demyelination model, NK cells contribute to rejection of allogeneic NPCs through an NKG2D signaling pathway.

Modification of picornavirus genomic RNA using 'click' chemistry shows that unlinking of the VPg peptide is dispensable for translation and replication of the incoming viral RNA.

Picornaviruses constitute a large group of viruses comprising medically and economically important pathogens such as poliovirus, coxsackievirus, rhinovirus, enterovirus 71 and foot-and-mouth disease virus. A unique characteristic of these viruses is the use of a viral peptide (VPg) as primer for viral RNA synthesis. As a consequence, all newly formed viral RNA molecules possess a covalently linked VPg peptide. It is known that VPg is enzymatically released from the incoming viral RNA by a host protein, called TDP2, but it is still unclear whether the release of VPg is necessary to initiate RNA translation. To study the possible requirement of VPg release for RNA translation, we developed a novel method to modify the genomic viral RNA with VPg linked via a 'non-cleavable' bond. We coupled an azide-modified VPg peptide to an RNA primer harboring a cyclooctyne [bicyclo[6.1.0]nonyne (BCN)] by a copper-free 'click' reaction, leading to a VPg-triazole-RNA construct that was 'non-cleavable' by TDP2. We successfully ligated the VPg-RNA complex to the viral genomic RNA, directed by base pairing. We show that the lack of VPg unlinkase does not influence RNA translation or replication. Thus, the release of the VPg from the incoming viral RNA is not a prerequisite for RNA translation or replication.

Neural stem cell depletion and CNS developmental defects after enteroviral infection.

Coxsackieviruses are significant human pathogens causing myocarditis, meningitis, and encephalitis. We previously demonstrated the ability of coxsackievirus B3 (CVB3) to persist within the neonatal central nervous system (CNS) and to target neural stem cells. Given that CVB3 is a cytolytic virus and may therefore damage target cells, we characterized the potential reduction in neurogenesis within the developing brain and the subsequent developmental defects that occurred after the loss of these essential neural stem cells. Neonatal mice were inoculated with a recombinant CVB3 expressing eGFP (eGFP-CVB3), and alterations in neurogenesis and brain development were evaluated over time. We observed a reduction in proliferating cells in CNS neurogenic regions simultaneously with the presence of nestin(+) cells undergoing apoptosis. The size of the brain appeared smaller by histology, and a permanent decrease in brain wet weight was observed after eGFP-CVB3 infection. We also observed an inverse relationship between the amount of virus material and brain wet weight up to day 30 postinfection. In addition, signs of astrogliosis and a compaction of the cortical layers were observed at 90 days postinfection. Intriguingly, partial brain wet weight recovery was observed in mice treated with the antiviral drug ribavirin during the persistent stage of infection. Hence, long-term neurological sequelae might be expected after neonatal enteroviral infections, yet antiviral treatment initiated long after the end of acute infection might limit virus-mediated neuropathology.

The Effect of H-bonding and Proton Transfer on the Voltammetry of 2,3,5,6-Tetramethyl-p-phenylenediamine in Acetonitrile. An Unexpectedly Complex Mechanism for a Simple Redox Couple.

The voltammetry of 2,3,5,6-tetramethyl-p-phenylenediamine, H2PD, has been studied and compared to that of its isomer N,N,N'N'-tetramethyl-p-phenylenediamine, Me2PD. Both undergo two reversible electron transfer processes in acetonitrile that nominally correspond to 1e- oxidation to the radical cations, Me2PD+ and H2PD+, and a second 1e- oxidation at more positive potentials to the quinonediimine dications, Me2PD2+ and H2PD2+. While the voltammetry of Me2PD agrees with this simple mechanism, that of H2PD does not. The second voltammetric wave is too small. UV/Vis spectroelectrochemical experiments indicate that the second wave does correspond to oxidation of H2PD+ to H2PD2+ in solution. The fact that the second wave is not present at all at the lowest concentrations (5 µM), and that it increases at longer times and higher concentrations, indicates that H2PD+ is not the initial solution product of the first oxidation. A number of lines of evidence suggest instead that the initial product is a mixed valent, H-bonded dimer between one H2PD in the the full reduced, fully protonated state, H4PD2+, and another in the fully oxidized, fully deprotonated state, PD. A mechanism is proposed in which this dimer is formed on the electrode surface through proton transfer and H-bonding. Once desorbed into solution, it breaks apart via reaction with other H2PD's, to give 2 H2PD+, which is the thermodynamically favored species in solution.

Viral persistence and chronic immunopathology in the adult central nervous system following Coxsackievirus infection during the neonatal period.

Coxsackieviruses are significant human pathogens, and the neonatal central nervous system (CNS) is a major target for infection. Despite the extreme susceptibility of newborn infants to coxsackievirus infection and viral tropism for the CNS, few studies have been aimed at determining the long-term consequences of infection on the developing CNS. We previously described a neonatal mouse model of coxsackievirus B3 (CVB3) infection and determined that proliferating stem cells in the CNS were preferentially targeted. Here, we describe later stages of infection, the ensuing inflammatory response, and subsequent lesions which remain in the adult CNS of surviving animals. High levels of type I interferons and chemokines (in particular MCP-5, IP10, and RANTES) were upregulated following infection and remained at high levels up to day 10 postinfection (p.i). Chronic inflammation and lesions were observed in the hippocampus and cortex of surviving mice for up to 9 months p.i. CVB3 RNA was detected in the CNS up to 3 months p.i at high abundance ( approximately 10(6) genomes/mouse brain), and viral genomic material remained detectable in culture after two rounds of in vitro passage. These data suggest that CVB3 may persist in the CNS as a low-level, noncytolytic infection, causing ongoing inflammatory lesions. Thus, the effects of a relatively common infection during the neonatal period may be long lasting, and the prognosis for newborn infants recovering from acute infection should be reexplored.