High seroprevalence of human herpesviruses in HIV-infected individuals attending primary healthcare facilities in rural South Africa

Seroprevalence data of human herpesviruses (HHVs) are limited for sub-Saharan Africa. These are important to provide an indication of potential burden of HHV-related disease, in particular in human immunodeficiency virus (HIV)-infected individuals who are known to be at increased risk of these conditions in the Western world. In this cross-sectional study among 405 HIV-infected and antiretroviral therapy naïve individuals in rural South Africa the seroprevalence of HHVs was: herpes simplex virus type 1 (HSV-1) (98%), herpes simplex virus type 2 (HSV-2) (87%), varicella zoster virus (VZV) (89%), and 100% for both Epstein-Barr virus (EBV) and cytomegalovirus (CMV). Independent factors associated with VZV seropositivity were low educational status and having children. Lack of in-house access to drinking water was independently associated with positive HSV-1 serostatus, whereas Shangaan ethnicity was associated with HSV-2 seropositivity. Increasing age was associated with higher IgG titres to both EBV and CMV, whereas CD4 cell count was negatively associated with EBV and CMV IgG titres. Moreover, IgG titres of HSV-1 and 2, VZV and CMV, and CMV and EBV were positively correlated. The high HHV seroprevalence emphasises the importance of awareness of these viral infections in HIV-infected individuals in South Africa

Simian varicella virus infection of Chinese rhesus macaques produces ganglionic infection in the absence of rash

Varicella-zoster virus (VZV) causes varicella (chickenpox), becomes latent in ganglia along the entire neuraxis, and may reactivate to cause herpes zoster (shingles). VZV may infect ganglia via retrograde axonal transport from infected skin or through hematogenous spread. Simian varicella virus (SVV) infection of rhesus macaques provides a useful model system to study the pathogenesis of human VZV infection. To dissect the virus and host immune factors during acute SVV infection, we analyzed four SVV-seronegative Chinese rhesus macaques infected intratracheally with cell-associated 5 × 103 plaque-forming units (pfu) of SVV-expressing green fluorescent protein (n = 2) or 5 × 104 pfu of wild-type SVV (n = 2). All monkeys developed viremia and SVV-specific adaptive B- and T-cell immune responses, but none developed skin rash. At necropsy 21 days postinfection, SVV DNA was found in ganglia along the entire neuraxis and in viscera, and SVV RNA was found in ganglia, but not in viscera. The amount of SVV inoculum was associated with the extent of viremia and the immune response to virus. Our findings demonstrate that acute SVV infection of Chinese rhesus macaques leads to ganglionic infection by the hematogenous route and the induction of a virus-specific adaptive memory response in the absence of skin rash

Current Perspectives on the Management of Herpesvirus Infections in Solid Organ Transplant Recipients

Solid organ transplant recipients (SOTRs) are at high risk of human herpesvirus (HHV)-related morbidity and mortality due to the use of immunosuppressive therapy. We aim to increase awareness and understanding of HHV disease burden in SOTRs by providing an overview of current prevention and management strategies as described in the literature and guidelines. We discuss challenges in both prevention and treatment as well as future perspectives.</p

Reanalysis of single-cell RNA sequencing data does not support herpes simplex virus 1 latency in non-neuronal ganglionic cells in mice

Most individuals are latently infected with herpes simplex virus type 1 (HSV-1), and it is well-established that HSV-1 establishes latency in sensory neurons of peripheral ganglia. However, it was recently proposed that latent HSV-1 is also present in immune cells recovered from the ganglia of experimentally infected mice. Here, we reanalyzed the single-cell RNA sequencing (scRNA-Seq) data that formed the basis for that conclusion. Unexpectedly, off-target priming in 3’ scRNA-Seq experiments enabled the detection of non-polyadenylated HSV-1 latency-associated transcript (LAT) intronic RNAs. However, LAT reads were near-exclusively detected in mixed populations of cells undergoing cell death. Specific loss of HSV-1 LAT and neuronal transcripts during quality control filtering indicated widespread destruction of neurons, supporting the presence of contaminating cell-free RNA in other cells following tissue processing. In conclusion, the reported detection of latent HSV-1 in non-neuronal cells is best explained using compromised scRNA-Seq datasets. IMPORTANCE Most people are infected with herpes simplex virus type 1 (HSV-1) during their life. Once infected, the virus generally remains in a latent (silent) state, hiding within the neurons of peripheral ganglia. Periodic reactivation (reawakening) of the virus may cause fresh diseases such as cold sores. A recent study using single-cell RNA sequencing (scRNA-Seq) proposed that HSV-1 can also establish latency in the immune cells of mice, challenging existing dogma. We reanalyzed the data from that study and identified several flaws in the methodologies and analyses performed that invalidate the published conclusions. Specifically, we showed that the methodologies used resulted in widespread destruction of neurons which resulted in the presence of contaminants that confound the data analysis. We thus conclude that there remains little to no evidence for HSV-1 latency in immune cells.</p

Reanalysis of single-cell RNA sequencing data does not support herpes simplex virus 1 latency in non-neuronal ganglionic cells in mice

Most individuals are latently infected with herpes simplex virus type 1 (HSV-1), and it is well-established that HSV-1 establishes latency in sensory neurons of peripheral ganglia. However, it was recently proposed that latent HSV-1 is also present in immune cells recovered from the ganglia of experimentally infected mice. Here, we reanalyzed the single-cell RNA sequencing (scRNA-Seq) data that formed the basis for that conclusion. Unexpectedly, off-target priming in 3’ scRNA-Seq experiments enabled the detection of non-polyadenylated HSV-1 latency-associated transcript (LAT) intronic RNAs. However, LAT reads were near-exclusively detected in mixed populations of cells undergoing cell death. Specific loss of HSV-1 LAT and neuronal transcripts during quality control filtering indicated widespread destruction of neurons, supporting the presence of contaminating cell-free RNA in other cells following tissue processing. In conclusion, the reported detection of latent HSV-1 in non-neuronal cells is best explained using compromised scRNA-Seq datasets. IMPORTANCE Most people are infected with herpes simplex virus type 1 (HSV-1) during their life. Once infected, the virus generally remains in a latent (silent) state, hiding within the neurons of peripheral ganglia. Periodic reactivation (reawakening) of the virus may cause fresh diseases such as cold sores. A recent study using single-cell RNA sequencing (scRNA-Seq) proposed that HSV-1 can also establish latency in the immune cells of mice, challenging existing dogma. We reanalyzed the data from that study and identified several flaws in the methodologies and analyses performed that invalidate the published conclusions. Specifically, we showed that the methodologies used resulted in widespread destruction of neurons which resulted in the presence of contaminants that confound the data analysis. We thus conclude that there remains little to no evidence for HSV-1 latency in immune cells.</p

Whole exome sequencing of patients with varicella-zoster virus and herpes simplex virus induced acute retinal necrosis reveals rare disease-associated genetic variants

Purpose: Herpes simplex virus (HSV) and varicella-zoster virus (VZV) are neurotropic human alphaherpesviruses endemic worldwide. Upon primary infection, both viruses establish lifelong latency in neurons and reactivate intermittently to cause a variety of mild to severe diseases. Acute retinal necrosis (ARN) is a rare, sight-threatening eye disease induced by ocular VZV or HSV infection. The virus and host factors involved in ARN pathogenesis remain incompletely described. We hypothesize an underlying genetic defect in at least part of ARN cases. Methods: We collected blood from 17 patients with HSV-or VZV-induced ARN, isolated DNA and performed Whole Exome Sequencing by Illumina followed by analysis in Varseq with criteria of CADD score &gt; 15 and frequency in GnomAD &lt; 0.1% combined with biological filters. Gene modifications relative to healthy control genomes were filtered according to high quality and read-depth, low frequency, high deleteriousness predictions and biological relevance. Results: We identified a total of 50 potentially disease-causing genetic variants, including missense, frameshift and splice site variants and on in-frame deletion in 16 of the 17 patients. The vast majority of these genes are involved in innate immunity, followed by adaptive immunity, autophagy, and apoptosis; in several instances variants within a given gene or pathway was identified in several patients.Discussion: We propose that the identified variants may contribute to insufficient viral control and increased necrosis ocular disease presentation in the patients and serve as a knowledge base and starting point for the development of improved diagnostic, prophylactic, and therapeutic applications.</p

Local CD4 and CD8 T-Cell Reactivity to HSV-1 Antigens Documents Broad Viral Protein Expression and Immune Competence in Latently Infected Human Trigeminal Ganglia

Herpes simplex virus type 1 (HSV-1) infection results in lifelong chronic infection of trigeminal ganglion (TG) neurons, also referred to as neuronal HSV-1 latency, with periodic reactivation leading to recrudescent herpetic disease in some persons. HSV-1 proteins are expressed in a temporally coordinated fashion during lytic infection, but their expression pattern during latent infection is largely unknown. Selective retention of HSV-1 reactive T-cells in human TG suggests their role in controlling reactivation by recognizing locally expressed HSV-1 proteins. We characterized the HSV-1 proteins recognized by virus-specific CD4 and CD8 T-cells recovered from human HSV-1-infected TG. T-cell clusters, consisting of both CD4 and CD8 T-cells, surrounded neurons and expressed mRNAs and proteins consistent with in situ antigen recognition and antiviral function. HSV-1 proteome-wide scans revealed that intra-TG T-cell responses included both CD4 and CD8 T-cells directed to one to three HSV-1 proteins per person. HSV-1 protein ICP6 was targeted by CD8 T-cells in 4 of 8 HLA-discordant donors. In situ tetramer staining demonstrated HSV-1-specific CD8 T-cells juxtaposed to TG neurons. Intra-TG retention of virus-specific CD4 T-cells, validated to the HSV-1 peptide level, implies trafficking of viral proteins from neurons to HLA class II-expressing non-neuronal cells for antigen presentation. The diversity of viral proteins targeted by TG T-cells across all kinetic and functional classes of viral proteins suggests broad HSV-1 protein expression, and viral antigen processing and presentation, in latently infected human TG. Collectively, the human TG represents an immunocompetent environment for both CD4 and CD8 T-cell recognition of HSV-1 proteins expressed during latent infection. HSV-1 proteins recognized by TG-resident T-cells, particularly ICP6 and VP16, are potential HSV-1 vaccine candidates

Virus-specific T_RM cells of both donor and recipient origin reside in human kidney transplants

Tissue-resident lymphocytes (TRLs) are critical for local protection against viral pathogens in peripheral tissue. However, it is unclear if TRLs perform a similar role in transplanted organs under chronic immunosuppressed conditions. In this study, we aimed to characterize the TRL compartment in human kidney transplant nephrectomies and examine its potential role in antiviral immunity. The TRL compartment of kidney transplants contained diverse innate, innate-like, and adaptive TRL populations expressing the canonical residency markers CD69, CD103, and CD49a. Chimerism of donor and recipient cells was present in 43% of kidney transplants and occurred in all TRL subpopulations. Paired single-cell transcriptome and T cell receptor (TCR) sequencing showed that donor and recipient tissue–resident memory T (TRM) cells exhibit striking similarities in their transcriptomic profiles and share numerous TCR clonotypes predicted to target viral pathogens. Virus dextramer staining further confirmed that CD8 TRM cells of both donor and recipient origin express TCRs with specificities against common viruses, including CMV, EBV, BK polyomavirus, and influenza A. Overall, the study results demonstrate that a diverse population of TRLs resides in kidney transplants and offer compelling evidence that TRM cells of both donor and recipient origin reside within this TRL population and may contribute to local protection against viral pathogens.</p

Varicella zoster virus-induced autophagy in human neuronal and hematopoietic cells exerts antiviral activity

Autophagy is a degradational pathway with pivotal roles in cellular homeostasis and survival, including protection of neurons in the central nervous system (CNS). The significance of autophagy as antiviral defense mechanism is recognized and some viruses hijack and modulate this process to their advantage in certain cell types. Here, we present data demonstrating that the human neurotropic herpesvirus varicella zoster virus (VZV) induces autophagy in human SH-SY5Y neuronal cells, in which the pathway exerts antiviral activity. Productively VZV-infected SH-SY5Y cells showed increased LC3-I-LC3-II conversion as well as co-localization of the viral glycoprotein E and the autophagy receptor p62. The activation of autophagy was dependent on a functional viral genome. Interestingly, inducers of autophagy reduced viral transcription, whereas inhibition of autophagy increased viral transcript expression. Finally, the genotype of patients with severe ocular and brain VZV infection were analyzed to identify potential autophagy-associated inborn errors of immunity. Two patients expressing genetic variants in the autophagy genes ULK1 and MAP1LC3B2, respectively, were identified. Notably, cells of both patients showed reduced autophagy, alongside enhanced viral replication and death of VZV-infected cells. In conclusion, these results demonstrate a neuro-protective role for autophagy in the context of VZV infection and suggest that failure to mount an autophagy response is a potential predisposing factor for development of severe VZV disease.</p