Analysis of the specificity and kinetics of neutralizing antibodies (nAbs) elicited by SARS-CoV-2 infection is crucial for understanding immune protection and identifying targets for vaccine design. In a cohort of 647 SARS-CoV-2-infected subjects we found that both the magnitude of Ab responses to SARS-CoV-2 spike (S) and nucleoprotein and nAb titers correlate with clinical scores. The receptor-binding domain (RBD) is immunodominant and the target of 90% of the neutralizing activity present in SARS-CoV-2 immune sera. Whereas overall RBD-specific serum IgG titers waned with a half-life of 49 days, nAb titers and avidity increased over time for some individuals, consistent with affinity maturation. We structurally defined an RBD antigenic map and serologically quantified serum Abs specific for distinct RBD epitopes leading to the identification of two major receptor-binding motif antigenic sites. Our results explain the immunodominance of the receptor-binding motif and will guide the design of COVID-19 vaccines and therapeutics.

The spillovers of β-coronaviruses in humans and the emergence of SARS-CoV-2 variants highlight the need for broad coronavirus countermeasures. We describe five monoclonal antibodies (mAbs) cross-reacting with the stem helix of multiple β-coronavirus spike glycoproteins isolated from COVID-19 convalescent individuals. Using structural and functional studies we show that the mAb with the greatest breadth (S2P6) neutralizes pseudotyped viruses from three different subgenera through inhibition of membrane fusion and delineate the molecular basis for its cross-reactivity. S2P6 reduces viral burden in hamsters challenged with SARS-CoV-2 through viral neutralization and Fc-mediated effector functions. Stem helix antibodies are rare, oftentimes of narrow specificity and can acquire neutralization breadth through somatic mutations. These data provide a framework for structure-guided design of pan-β-coronavirus vaccines eliciting broad protection.

The transcriptional program associated with herpesvirus latency and the viral genes regulating entry into and exit from latency are poorly understood and controversial. Here, we developed and validated a targeted enrichment platform and conducted large-scale transcriptome analyses of human cytomegalovirus (HCMV) infection. We used both an experimental hematopoietic cell model of latency and cells from naturally infected, healthy human subjects (clinical) to define the breadth of viral genes expressed. The viral transcriptome derived from experimental infection was highly correlated with that from clinical infection, validating our experimental latency model. These transcriptomes revealed a broader profile of gene expression during infection in hematopoietic cells than previously appreciated. Further, using recombinant viruses that establish a nonreactivating, latent-like or a replicative infection in CD34 hematopoietic progenitor cells, we defined classes of low to moderately expressed genes that are differentially regulated in latent vs. replicative states of infection. Most of these genes have yet to be studied in depth. By contrast, genes that were highly expressed, were expressed similarly in both latent and replicative infection. From these findings, a model emerges whereby low or moderately expressed genes may have the greatest impact on regulating the switch between viral latency and replication. The core set of viral genes expressed in natural infection and differentially regulated depending on the pattern of infection provides insight into the HCMV transcriptome associated with latency in the host and a resource for investigating virus-host interactions underlying persistence.

Naïve T cell responses are eroded with aging. We and others have recently shown that unimmunized old mice lose ≥70% of Ag-specific CD8 T cell precursors and that many of the remaining precursors acquire a “virtual (central) memory” (VM; CD44hiCD62Lhi) phenotype. Here, we demonstrate that unimmunized T cell receptor (TCR) transgenic (Tg) mice also undergo massive VM conversion with age, exhibiting rapid effector function upon both TCR and cytokine triggering. Age-related VM conversion in TCRTg mice directly depended upon replacement of the original TCRTg specificity by endogenous TCRα rearrangements, indicating that TCR signals must be critical in VM conversion. Importantly, we found that VM conversion had adverse functional effects in both old wild type and old TCRTg mice - old VM, but not old true naïve, T cells exhibited blunted TCR-mediated, but not IL-15-mediated, proliferation. This selective proliferative senescence correlated with increased apoptosis in old VM cells in response to peptide, but decreased apoptosis in response to homeostatic cytokines IL-7 & IL-15. Our results identify TCR as the key factor in differential maintenance and function of Ag-specific precursors in unimmunized mice with aging, and demonstrate that two separate age-related defects – drastic reduction in true naïve T cell precursors and impaired proliferative capacity of their VM cousins –combine to reduce naïve T cell responses with aging.

Immunosenescence, defined as the age-associated dysregulation and dysfunction of the immune system, is characterized by impaired protective immunity and decreased efficacy of vaccines. An increasing number of immunological, clinical and epidemiological studies suggest that persistent Cytomegalovirus (CMV) infection is associated with accelerated aging of the immune system and with several age-related diseases. However, current evidence on whether and how human CMV (HCMV) infection is implicated in immunosenescence and in age-related diseases remains incomplete and many aspects of CMV involvement in immune aging remain controversial. The attendees of the 4th International Workshop on "CMV & Immunosenescence", held in Parma, Italy, 25-27th March, 2013, presented and discussed data related to these open questions, which are reported in this commentary.

Persistent CMV infection has been associated with immune senescence. To address the causal impact of life-long persistent viral infection on immune homeostasis and defense, we infected young mice systemically with HSV-1, MCMV or both viruses, and studied their T-cell homeostasis and function. Herpesvirus+ mice exhibited increased all-cause mortality compared to controls. Upon Listeria-OVA infection, 23 month-old animals that had experienced life-long herpesvirus infections showed impaired bacterial control and CD8 T-cell function, along with distinct alterations in the T-cell repertoire both before and after Listeria challenge, compared to age-matched, herpesvirus-free controls. Herpesvirus infection was associated with reduced naïve CD8 T-cell precursors, above the loss attributable to aging. Moreover, the OVA-specific CD8 T-cell repertoire recruited after Listeria challenge was entirely non-overlapping between control and herpesvirus+ mice. Our study for the first time causally links life-long herpesvirus infection to all-cause mortality in mice and to disturbances in the T-cell repertoire, which themselves correspond to impaired immunity to a new infection in aging.

Studies of CD8 T cell responses to vaccination or infection with various pathogens in both animal models and human subjects have revealed a markedly consistent array of age-related defects. In general, recent work shows that aged CD8 T cell responses are decreased in magnitude, and show poor differentiation into effector cells, with a reduced arsenal of effector functions. Here we review potential mechanisms underlying these defects. We specifically address phenotypic and numeric changes to the naïve CD8 T cell precursor pool, the impact of persistent viral infection(s) and inflammation, and contributions of the aging environment in which these cells are activated.

Summary Aging is accompanied by altered immunity, resulting in a variable state of poorly understood immunodeficiency. While both the numbers and the functionality of naïve T-cells are decreased by aging, the impact of these changes upon immune defense against bacterial pathogens in vivo remains understudied. Using a model of Listeria monocytogenes (Lm), where the primary CD8 T-cell response is critically important for immune defense, we show that C57BL/6 (B6) mice exhibit an age-dependent reduction in survival, with delayed bacterial clearance in old animals. Kinetic analysis of antigen-specific CD8 T-cell expansion showed that CD8 effectors begin dividing at the same time in old and adult mice, but that the proliferative burst remained incomplete during discrete windows of time and was coupled with increased effector apoptosis in old mice. Further, antilisterial CD8 T-cells in old mice showed altered expression of key phenotypic and effector molecules and diminished polyfunctionality, measured by the ability to simultaneously produce multiple effector molecules. These results suggest that defects in functional maturation of CD8 cells in aged mice, compounded by (or perhaps coupled to) their reduced expansion in response to infection, yield effector CD8 T-cell populations insufficient in size and capability to effectively clear newly encountered intracellular pathogens.