Primary infection with varicella zoster virus (VZV) results in varicella (more commonly known as chickenpox) after which VZV establishes latency in sensory ganglia. VZV can reactivate to cause herpes zoster (shingles), a debilitating disease that affects one million individuals in the US alone annually. Current vaccines against varicella (Varivax) and herpes zoster (Zostavax) are not 100% efficacious. Specifically, studies have shown that 1 dose of varivax can lead to breakthrough varicella, albeit rarely, in children and a 2-dose regimen is now recommended. Similarly, although Zostavax results in a 50% reduction in HZ cases, a significant number of recipients remain at risk. To design more efficacious vaccines, we need a better understanding of the immune response to VZV. Clinical observations suggest that T cell immunity plays a more critical role in the protection against VZV primary infection and reactivation. However, no studies to date have directly tested this hypothesis due to the scarcity of animal models that recapitulate the immune response to VZV. We have recently shown that SVV infection of rhesus macaques models the hallmarks of primary VZV infection in children. In this study, we used this model to experimentally determine the role of CD4, CD8 and B cell responses in the resolution of primary SVV infection in unvaccinated animals. Data presented in this manuscript show that while CD20 depletion leads to a significant delay and decrease in the antibody response to SVV, loss of B cells does not alter the severity of varicella or the kinetics/magnitude of the T cell response. Loss of CD8 T cells resulted in slightly higher viral loads and prolonged viremia. In contrast, CD4 depletion led to higher viral loads, prolonged viremia and disseminated varicella. CD4 depleted animals also had delayed and reduced antibody and CD8 T cell responses. These results are similar to clinical observations that children with agammaglobulinemia have uncomplicated varicella whereas children with T cell deficiencies are at increased risk of progressive varicella with significant complications. Moreover, our studies indicate that CD4 T cell responses to SVV play a more critical role than antibody or CD8 T cell responses in the control of primary SVV infection and suggest that one potential mechanism for enhancing the efficacy of VZV vaccines is by eliciting robust CD4 T cell responses.

Varicella zoster virus (VZV) is a neurotropic α-herpesvirus that causes chickenpox during primary infection and establishes latency in sensory ganglia. Reactivation of VZV results in herpes zoster and other neurological complications. Our understanding of the VZV transcriptome during acute and latent infection in immune competent individuals remains incomplete. Infection of rhesus macaques with the homologous simian varicella virus (SVV) recapitulates the hallmarks of VZV infection. We therefore characterized the SVV transcriptome by quantitative real-time reverse transcriptase PCR (RT-qPCR) during acute infection in bronchial alveolar lavage (BAL) cells and peripheral blood mononuclear cells (PBMC), and during latency in sensory ganglia obtained from the same rhesus macaques. During acute infection, all known SVV open reading frames (ORFs) were detected and the most abundantly expressed ORFs are involved in virus replication and assembly such as the transcriptional activator ORF 63 and the structural proteins ORF 41 and ORF 49. In contrast, latent SVV gene expression is highly restricted. ORF 61, a viral transactivator and latency-associated transcript, is the most prevalent transcript detected in sensory ganglia. We also detected ORFs A, B, 4, 10, 63, 64, 65, 66 and 68 though significantly less frequently than ORF 61. This comprehensive analysis has revealed genes that potentially play a role in the establishment and/or maintenance of SVV latency.

dVaricella zoster virus (VZV) is a neurotropic alphaherpesvirus that causes chickenpox during primary infection and establishes latency in sensory ganglia. Infection of rhesus macaques (RM) with the homologous simian varicella virus (SVV) recapitulates hallmarks of VZV infection. We have shown that an antisense transcript of SVV open reading frame 61 (ORF61), a viral transactivator, was detected most frequently in latently infected RM sensory ganglia. In this study, we compared disease progression, viral replication, immune response, and the establishment of latency following intrabronchial infection with a recombinant SVV lacking ORF61 (SVV⌬ORF61) to those following infection with wild-type (WT) SVV. Varicella severity and viral latency within sensory ganglia were comparable in RMs infected with SVV⌬ORF61 and WT SVV. In contrast, viral loads, B and T cell responses, and plasma inflammatory cytokine levels were decreased in RMs infected with SVV⌬ORF61. To investigate the mechanisms underlying the reduced adaptive immune response, we compared acute SVV gene expression, frequency and proliferation of dendritic cell (DC) subsets, and the expression of innate antiviral genes in bronchoalveolar lavage (BAL) samples. The abundance of SVV transcripts in all kinetic classes was significantly decreased in RMs infected with SVV⌬ORF61. In addition, we detected a higher frequency and proliferation of plasmacytoid dendritic cells in BAL fluid at 3 days postinfection in RMs infected with SVV⌬ORF61, which was accompanied by a slight increase in type I interferon gene expression. Taken together, our data suggest that ORF61 plays an important role in orchestrating viral gene expression in vivo and interferes with the host antiviral interferon response. V aricella zoster virus (VZV) is a neurotropic human alphaherpesvirus that causes chickenpox (varicella) during primary infection. VZV establishes a life-long latent infection in sensory ganglia, including the trigeminal and dorsal root ganglia. Reactivation of VZV leads to herpes zoster (HZ; shingles), which is estimated to affect 1 million individuals each year in the United States and can result in significant morbidity and occasionally mortality in aged and immunocompromised individuals (1-4). Reactivation of VZV is generally believed to be due to a decline in T cell immunity (5-10); however, the viral genes that control the switch between latent and lytic replication remain unknown. This is due in part to the fact that VZV is strictly a human pathogen, and animal models of VZV infection recapitulate only certain aspects of pathogenesis. We have previously shown that rhesus macaques (RMs) infected with simian varicella virus (SVV) display the hallmarks of VZV infection in humans, including generalized varicella, development of cellular and humoral immunity, and establishment of latency (11).Utilizing this model, we previously found that SVV open reading frame 61 (ORF61) was the most prevalent transcript detected during latency in sensory ganglia (12). Interestingly, SVV ORF61 ...

S imian varicella virus (SVV) and varicella-zoster virus (VZV)are neurotropic alphaherpesviruses that establish latency within the sensory ganglia during primary infection. Viral latency is generally characterized by a quiescent state with limited gene expression; for example, during herpes simplex virus (HSV) latency, only the latency-associated transcripts (LATs) are abundantly expressed (1). However, VZV latency can be associated with the transcription of multiple genes, including open reading frames (ORFs) 4, 11, 18, 21, 29, 40, 41, 43, 57, 62, 63, 66, and 68 (2-9). We recently examined the SVV latent transcriptome from juvenile rhesus macaques (RMs) and found that SVV latency is characterized by a restricted gene expression profile (10). SVV ORF 61 was the most prevalent transcript detected during latent infection, present in at least one sensory ganglion in all RMs tested and in 10 of 16 sensory ganglia total. We also detected SVV ORFs A, B, 4, 10, 55, 63, 64, 65, 66, 67, and 68, although less frequently, in 1 to 4 of 16 latently infected sensory ganglia from juvenile RMs.Several clinical observations highlight the importance of cellmediated immune responses in controlling VZV infection and reactivation. Specifically, a lack of immunoglobulin production due to agammaglobulinemia does not complicate the outcome of varicella in children (11,12). In contrast, T cell deficiencies, including congenital deficiencies or those induced by HIV infection or immune suppression, lead to severe and disseminated varicella (13-17). Similarly, a higher incidence of herpes zoster (HZ) in aged patients is associated with diminished T cell proliferation to VZV antigens in vitro, while antibody titers remain stable (18). Moreover, HIV patients are more susceptible to HZ when their absolute numbers of CD4 T cells decline to less than 500 cells per microliter (19)(20)(21). Finally, the frequency of VZV reactivation is related to the intensity of immune suppression, with higher incidence in patients receiving combined therapy than in patients receiving chemotherapy or radiotherapy alone (22).We have recently shown that the resolution of acute SVV infection is also dependent on cellular immunity. Specifically, loss of CD4 T cells during acute SVV infection in juvenile RMs resulted in higher peak viral loads, prolonged viremia, and disseminated varicella compared to these parameters in controls (23). CD8-depleted RMs had slightly higher viral loads and more prolonged varicella rash than controls. Lastly, CD20 depletion did not alter the severity of varicella or the kinetics and magnitude of the T cell response. Our data indicate that CD4 T cell immunity is critical in controlling acute SVV infection in RMs and are in agreement with clinical observations during acute VZV infection.To improve HZ-associated morbidity, it is critical to understand the virological and immunological parameters that control latency and reactivation. In the present study, we extend our previous reports (10, 23) and investigate the impact of T cell and B c...