Adeno-associated virus type 2 in US children with acute severe hepatitis.

As of August 2022, clusters of acute severe hepatitis of unknown aetiology in children have been reported from 35 countries, including the USA1,2. Previous studies have found human adenoviruses (HAdVs) in the blood from patients in Europe and the USA3-7, although it is unclear whether this virus is causative. Here we used PCR testing, viral enrichment-based sequencing and agnostic metagenomic sequencing to analyse samples from 16 HAdV-positive cases from 1 October 2021 to 22 May 2022, in parallel with 113 controls. In blood from 14 cases, adeno-associated virus type 2 (AAV2) sequences were detected in 93% (13 of 14), compared to 4 (3.5%) of 113 controls (P < 0.001) and to 0 of 30 patients with hepatitis of defined aetiology (P < 0.001). In controls, HAdV type 41 was detected in blood from 9 (39.1%) of the 23 patients with acute gastroenteritis (without hepatitis), including 8 of 9 patients with positive stool HAdV testing, but co-infection with AAV2 was observed in only 3 (13.0%) of these 23 patients versus 93% of cases (P < 0.001). Co-infections by Epstein-Barr virus, human herpesvirus 6 and/or enterovirus A71 were also detected in 12 (85.7%) of 14 cases, with higher herpesvirus detection in cases versus controls (P < 0.001). Our findings suggest that the severity of the disease is related to co-infections involving AAV2 and one or more helper viruses.

A Case Series of Children with Acute Hepatitis and Human Adenovirus Infection.

BACKGROUND: Human adenoviruses typically cause self-limited respiratory, gastrointestinal, and conjunctival infections in healthy children. In late 2021 and early 2022, several previously healthy children were identified with acute hepatitis and human adenovirus viremia. METHODS: We used International Classification of Diseases, 10th Revision, codes to identify all children (<18 years of age) with hepatitis who were admitted to Children's of Alabama hospital between October 1, 2021, and February 28, 2022; those with acute hepatitis who also tested positive for human adenovirus by whole-blood quantitative polymerase chain reaction (PCR) were included in our case series. Demographic, clinical, laboratory, and treatment data were obtained from medical records. Residual blood specimens were sent for diagnostic confirmation and human adenovirus typing. RESULTS: A total of 15 children were identified with acute hepatitis - 6 (40%) who had hepatitis with an identified cause and 9 (60%) who had hepatitis without a known cause. Eight (89%) of the patients with hepatitis of unknown cause tested positive for human adenovirus. These 8 patients plus 1 additional patient referred to this facility for follow-up were included in this case series (median age, 2 years 11 months; age range, 1 year 1 month to 6 years 5 months). Liver biopsies indicated mild-to-moderate active hepatitis in 6 children, some with and some without cholestasis, but did not show evidence of human adenovirus on immunohistochemical examination or electron microscopy. PCR testing of liver tissue for human adenovirus was positive in 3 children (50%). Sequencing of specimens from 5 children showed three distinct human adenovirus type 41 hexon variants. Two children underwent liver transplantation; all the others recovered with supportive care. CONCLUSIONS: Human adenovirus viremia was present in the majority of children with acute hepatitis of unknown cause admitted to Children's of Alabama from October 1, 2021, to February 28, 2022, but whether human adenovirus was causative remains unclear. Sequencing results suggest that if human adenovirus was causative, this was not an outbreak driven by a single strain. (Funded in part by the Centers for Disease Control and Prevention.).

Functional Fc receptors are crucial in antibody-mediated protection against cytomegalovirus.

Human cytomegalovirus is a medically important pathogen. Previously, using murine CMV (MCMV), we provided evidence that both neutralizing and nonneutralizing antibodies can confer protection from viral infection in vivo. In this study, we report that serum derived from infected animals had a greater protective capacity in MCMV-infected RAG-/- mice than serum from animals immunized with purified virus. The protective activity of immune serum was strictly dependent on functional Fcγ receptors (FcγR). Deletion of individual FcγRs or combined deletion of FcγRI and FcγRIV had little impact on the protection afforded by serum. Adoptive transfer of CD115-positive cells from noninfected donors demonstrated that monocytes represent important cellular mediators of the protective activity provided by immune serum. Our studies suggest that Fc-FcγR interactions and monocytic cells are critical for antibody-mediated protection against MCMV infection in vivo. These findings may provide new avenues for the development of novel strategies for more effective CMV vaccines or antiviral immunotherapies.

SOX2 downregulation of PML increases HCMV gene expression and growth of glioma cells.

The presence of human cytomegalovirus (HCMV) in glioblastoma (GBM) and improved outcomes of GBM patients receiving therapies targeting the virus have implicated HCMV in GBM progression. However, a unifying mechanism that accounts for the contribution of HCMV to the malignant phenotype of GBM remains incompletely defined. Here we have identified SOX2, a marker of glioma stem cells (GSCs), as a key determinant of HCMV gene expression in gliomas. Our studies demonstrated that SOX2 downregulated promyelocytic leukemia (PML) and Sp100 and consequently facilitated viral gene expression by decreasing the amount of PML nuclear bodies in HCMV-infected glioma cells. Conversely, the expression of PML antagonized the effects of SOX2 on HCMV gene expression. Furthermore, this regulation of SOX2 on HCMV infection was demonstrated in a neurosphere assay of GSCs and in a murine xenograft model utilizing xenografts from patient-derived glioma tissue. In both cases, SOX2 overexpression facilitated the growth of neurospheres and xenografts implanted in immunodeficient mice. Lastly, the expression of SOX2 and HCMV immediate early 1 (IE1) protein could be correlated in tissues from glioma patients, and interestingly, elevated levels of SOX2 and IE1 were predictive of a worse clinical outcome. These studies argue that HCMV gene expression in gliomas is regulated by SOX2 through its regulation of PML expression and that targeting molecules in this SOX2-PML pathway could identify therapies for glioma treatment.

Acquisition of Furin Cleavage Site and Further SARS-CoV-2 Evolution Change the Mechanisms of Viral Entry, Infection Spread, and Cell Signaling.

Circulation of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in the human population leads to further viral evolution. The new variants that arise during this evolution are more infectious. Our data suggest that newer variants have shifted from utilizing both cathepsin/endosome- and TMPRSS2-mediated entry mechanisms to rely on a TMPRSS2-dependent entry pathway. Accordingly, only the early lineages of SARS-CoV-2 are capable of infecting and forming syncytia in Vero/ACE2 cells which lack TMPRSS2 expression. The presence of an intact multibasic furin cleavage site (FCS) in the S protein was a key requirement for cell-to-cell fusion. Deletion of FCS makes SARS-CoV-2 more infectious in vitro but renders it incapable of syncytium formation. Cell-to-cell fusion likely represents an alternative means of virus spread and is resistant to the presence of high levels of neutralizing monoclonal antibodies (MAbs) and immune sera in the media. In this study, we also noted that cells infected with SARS-CoV-2 with an intact FCS or alphavirus replicon expressing S protein (VEErep/S) released high levels of free S1 subunit. The released S1 is capable of activating the TLR4 receptor and inducing a pro-inflammatory response. Thus, S1 activation of TLR4 may be an important contributor to SARS-CoV-2-induced COVID-19 disease and needs to be considered in the design of COVID mRNA vaccines. Lastly, a VEErep/S-replicon was shown to produce large amounts of infectious, syncytium-forming pseudoviruses and thus could represent alternative experimental system for screening inhibitors of virus entry and syncytium formation. IMPORTANCE The results of this study demonstrate that the late lineages of SARS-CoV-2 evolved to more efficient use of the TMPRSS2-mediated entry pathway and gradually lost an ability to employ the cathepsins/endosome-mediated entry. The acquisition of a furin cleavage site (FCS) by SARS-CoV-2-specific S protein made the virus a potent producer of syncytia. Their formation is also determined by expression of ACE2 and TMPRSS2 and is resistant to neutralizing human MAbs and immune sera. Syncytium formation appears to be an alternative means of infection spread following the development of an adaptive immune response. Cells infected with SARS-CoV-2 with an intact FCS secrete high levels of the S1 subunit. The released S1 demonstrates an ability to activate the TLR4 receptor and induce pro-inflammatory cytokines, which represent a hallmark of SARS-CoV-2 pathogenesis. Alphavirus replicons encoding SARS-CoV-2 S protein cause spreading, syncytium-forming infection, and they can be applied as an experimental tool for studying the mechanism of syncytium formation.

Human Cytomegalovirus Hijacks WD Repeat Domain 11 for Virion Assembly Compartment Formation and Virion Morphogenesis.

Human cytomegalovirus (HCMV) has a large (∼235 kb) genome with more than 200 predicted open reading frames that exploits numerous cellular factors to facilitate its replication. A key feature of HCMV-infected cells is the emergence of a distinctive membranous cytoplasmic compartment termed the virion assembly compartment (vAC). Here, we report that host protein WD repeat domain 11 (WDR11) plays a key role in vAC formation and virion morphogenesis. We found that WDR11 was upregulated at both mRNA and protein levels during HCMV infection. At the late stage of HCMV replication, WDR11 relocated to the vAC and colocalized with markers of the trans-Golgi network (TGN) and vAC. Depletion of WDR11 hindered HCMV-induced membrane reorganization of the Golgi and TGN, altered vAC formation, and impaired HCMV secondary envelopment and virion morphogenesis. Further, motifs critical for the localization of WDR11 in TGN were identified by alanine-scanning mutagenesis. Mutation of these motifs led to WDR11 mislocation outside the TGN and loss of vAC formation. Taken together, these data indicate that host protein WDR11 is required for efficient viral replication at the stage of virion assembly, possibly by facilitating the remodeling of the endomembrane system for vAC formation and virion morphogenesis. IMPORTANCE During the late phase of human cytomegalovirus (HCMV) infection, the endomembrane system is dramatically reorganized, resulting in the formation of a unique structure termed the virion assembly compartment (vAC), which is critical for the assembly of infectious virions. The mechanism of HCMV-induced vAC formation is still not fully understood. In this report, we identified a host factor, WDR11, that plays an important role in vAC formation. Our findings argue that WDR11 contributes to the relocation of the Golgi and trans-Golgi network to the vAC, a membrane reorganization process that appears to be required for efficient virion maturation. The present work provides new insights into the vAC formation and HCMV virion morphogenesis and a potential novel target for antiviral treatment.

Receptor-Independent Therapies for Forensic Detainees with Schizophrenia-Dementia Comorbidity.

Forensic institutions throughout the world house patients with severe psychiatric illness and history of criminal violations. Improved medical care, hygiene, psychiatric treatment, and nutrition led to an unmatched longevity in this population, which previously lived, on average, 15 to 20 years shorter than the public at large. On the other hand, longevity has contributed to increased prevalence of age-related diseases, including neurodegenerative disorders, which complicate clinical management, increasing healthcare expenditures. Forensic institutions, originally intended for the treatment of younger individuals, are ill-equipped for the growing number of older offenders. Moreover, as antipsychotic drugs became available in 1950s and 1960s, we are observing the first generation of forensic detainees who have aged on dopamine-blocking agents. Although the consequences of long-term treatment with these agents are unclear, schizophrenia-associated gray matter loss may contribute to the development of early dementia. Taken together, increased lifespan and the subsequent cognitive deficit observed in long-term forensic institutions raise questions and dilemmas unencountered by the previous generations of clinicians. These include: does the presence of neurocognitive dysfunction justify antipsychotic dose reduction or discontinuation despite a lifelong history of schizophrenia and violent behavior? Should neurolipidomic interventions become the standard of care in elderly individuals with lifelong schizophrenia and dementia? Can patients with schizophrenia and dementia meet the Dusky standard to stand trial? Should neurocognitive disorders in the elderly with lifelong schizophrenia be treated differently than age-related neurodegeneration? In this article, we hypothesize that gray matter loss is the core symptom of schizophrenia which leads to dementia. We hypothesize further that strategies to delay or stop gray matter depletion would not only improve the schizophrenia sustained recovery, but also avert the development of major neurocognitive disorders in people living with schizophrenia. Based on this hypothesis, we suggest utilization of both receptor-dependent and independent therapeutics for chronic psychosis.

A Novel Strain-Specific Neutralizing Epitope on Glycoprotein H of Human Cytomegalovirus.

Human cytomegalovirus (HCMV) is a ubiquitous pathogen that causes severe clinical disease in immunosuppressed patients and congenitally infected newborn infants. Viral envelope glycoproteins represent attractive targets for vaccination or passive immunotherapy. To extend the knowledge of mechanisms of virus neutralization, monoclonal antibodies (MAbs) were generated following immunization of mice with HCMV virions. Hybridoma supernatants were screened for in vitro neutralization activity, yielding three potent MAbs, 6E3, 3C11, and 2B10. MAbs 6E3 and 3C11 blocked infection of all viral strains that were tested, while MAb 2B10 neutralized only 50% of the HCMV strains analyzed. Characterization of the MAbs using indirect immunofluorescence analyses demonstrated their reactivity with recombinantly derived gH. While MAbs 6E3 and 3C11 reacted with gH when expressed alone, 2B10 detected gH only when it was coexpressed with gB and gL. Recognition of gH by 3C11 was dependent on the expression of the entire ectodomain of gH, whereas 6E3 required residues 1 to 629 of gH. The strain-specific determinant for neutralization by Mab 2B10 was identified as a single Met→Ile amino acid polymorphism within gH, located within the central part of the protein. The polymorphism is evenly distributed among described HCMV strains. The 2B10 epitope thus represents a novel strain-specific antibody target site on gH of HCMV. The dependence of the reactivity of 2B10 on the simultaneous presence of gB/gH/gL will be of value in the structural definition of this tripartite complex. The 2B10 epitope may also represent a valuable tool for diagnostics to monitor infections/reinfections with different HCMV strains during pregnancy or after transplantation. IMPORTANCE HCMV infections are life threatening to people with compromised or immature immune systems. Understanding the antiviral antibody repertoire induced during HCMV infection is a necessary prerequisite to define protective antibody responses. Here, we report three novel anti-gH MAbs that potently neutralized HCMV infectivity. One of these MAbs (2B10) targets a novel strain-specific conformational epitope on gH that only becomes accessible upon coexpression of the minimal fusion machinery gB/gH/gL. Strain specificity is dependent on a single amino acid polymorphism within gH. Our data highlight the importance of strain-specific neutralizing antibody responses against HCMV. The 2B10 epitope may also represent a valuable tool for diagnostics to monitor infections/reinfections with different HCMV strains during pregnancy or after transplantation. In addition, the dependence of the reactivity of 2B10 on the simultaneous presence of gB/gH/gL will be of value in the structural definition of this tripartite complex.

Localization of the WD repeat-containing protein 5 to the Virion Assembly Compartment Facilitates Human Cytomegalovirus Assembly.

We previously reported that human cytomegalovirus (HCMV) utilizes the cellular protein WD repeat-containing protein 5 (WDR5) to facilitate capsid nuclear egress. Here, we further show that HCMV infection results in WDR5 localization in a juxtanuclear region, and that its localization to this cellular site is associated with viral replication and late viral gene expression. Furthermore, WDR5 accumulated in the virion assembly compartment (vAC) and co-localized with vAC markers of gamma-tubulin (γ-tubulin), early endosomes, and viral vAC marker proteins pp65, pp28, and glycoprotein B (gB). WDR5 co-immunoprecipitated with multiple virion proteins, including MCP, pp150, pp65, pIRS1, and pTRS1, which may explain WDR5 accumulation in the vAC during infection. WDR5 fractionated with virions either in the presence or absence of Triton X-100 and was present in purified viral particles, suggesting that WDR5 was incorporated into HCMV virions. Thus, WDR5 localized to the vAC and was incorporated into virions, raising the possibility that in addition to capsid nuclear egress, WDR5 could also participate in cytoplasmic HCMV virion morphogenesis.Importance Human cytomegalovirus (HCMV) has a large (∼235-kb) genome that contains over 170 ORFs and exploits numerous cellular factors to facilitate its replication. In the late phase of HCMV infection cytoplasmic membranes are reorganized to establish the virion assembly compartment (vAC), which has been shown to necessary for efficient assembly of progeny virions. We previously reported that WDR5 facilitates HCMV nuclear egress. Here, we show that WDR5 is localized to the vAC and incorporated into virions, perhaps contributing to efficient virion maturation. Thus, findings in this study identified a potential role for WDR5 in HCMV assembly in the cytoplasmic phase of virion morphogenesis.

Acute Hepatitis and Adenovirus Infection Among Children - Alabama, October 2021-February 2022.

During October-November 2021, clinicians at a children's hospital in Alabama identified five pediatric patients with severe hepatitis and adenovirus viremia upon admission. In November 2021, hospital clinicians, the Alabama Department of Public Health, the Jefferson County Department of Health, and CDC began an investigation. This activity was reviewed by CDC and conducted consistent with applicable federal law and CDC policy.

HCMV trimer- and pentamer-specific antibodies synergize for virus neutralization but do not correlate with congenital transmission.

Human cytomegalovirus (HCMV) causes substantial disease in transplant patients and harms the development of the nervous system in babies infected in utero. Thus, there is a major focus on developing safe and effective HCMV vaccines. Evidence has been presented that a major target of neutralizing antibodies (NAbs) is the HCMV pentamer glycoprotein gH/gL/UL128-131. In some studies, most of the NAbs in animal or human sera were found to recognize the pentamer, which mediates HCMV entry into endothelial and epithelial cells. It was also reported that pentamer-specific antibodies correlate with protection against transmission from mothers to babies. One problem with the studies on pentamer-specific NAbs to date has been that the studies did not compare the pentamer to the other major form of gH/gL, the gH/gL/gO trimer, which is essential for entry into all cell types. Here, we demonstrate that both trimer and pentamer NAbs are frequently found in human transplant patients' and pregnant mothers' sera. Depletion of human sera with trimer caused reductions in NAbs similar to that observed following depletion with the pentamer. The trimer- and pentamer-specific antibodies acted in a synergistic fashion to neutralize HCMV and also to prevent virus cell-to-cell spread. Importantly, there was no correlation between the titers of trimer- and pentamer-specific NAbs and transmission of HCMV from mothers to babies. Therefore, both the trimer and pentamer are important targets of NAbs. Nevertheless, these antibodies do not protect against transmission of HCMV from mothers to babies.

Comparing Nasopharyngeal and Midturbinate Nasal Swab Testing for the Identification of Severe Acute Respiratory Syndrome Coronavirus 2.

Testing of paired midturbinate (MT) nasal and nasopharyngeal (NP) swabs, collected by trained personnel from 40 patients with coronavirus disease 2019 (COVID-19), showed that more NP (76/95 [80%]) than MT swabs tested positive (61/95 [64%]) (P = .02). Among samples collected a week after study enrollment, fewer MT than NP samples were positive (45% vs 76%; P = .001).

Predictors of Nonseroconversion after SARS-CoV-2 Infection.

Not all persons recovering from severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection develop SARS-CoV-2-specific antibodies. We show that nonseroconversion is associated with younger age and higher reverse transcription PCR cycle threshold values and identify SARS-CoV-2 viral loads in the nasopharynx as a major correlate of the systemic antibody response.

Cell Fusion Induced by a Fusion-Active Form of Human Cytomegalovirus Glycoprotein B (gB) Is Inhibited by Antibodies Directed at Antigenic Domain 5 in the Ectodomain of gB.

Human cytomegalovirus (HCMV) is a ubiquitous pathogen that can cause severe clinical disease in allograft recipients and infants infected in utero Virus-neutralizing antibodies defined in vitro have been proposed to confer protection against HCMV infection, and the virion envelope glycoprotein B (gB) serves as a major target of neutralizing antibodies. The viral fusion protein gB is nonfusogenic on its own and requires glycoproteins H (gH) and L (gL) for membrane fusion, which is in contrast to requirements of related class III fusion proteins, including vesicular stomatitis virus glycoprotein G (VSV-G) or baculovirus gp64. To explore requirements for gB's fusion activity, we generated a set of chimeras composed of gB and VSV-G or gp64, respectively. These gB chimeras were intrinsically fusion active and led to the formation of multinucleated cell syncytia when expressed in the absence of other viral proteins. Utilizing a panel of virus-neutralizing gB-specific monoclonal antibodies (MAbs), we could demonstrate that syncytium formation of the fusogenic gB/VSV-G chimera can be significantly inhibited by only a subset of neutralizing MAbs which target antigenic domain 5 (AD-5) of gB. This observation argues for differential modes of action of neutralizing anti-gB MAbs and suggests that blocking the membrane fusion function of gB could be one mechanism of antibody-mediated virus neutralization. In addition, our data have important implications for the further understanding of the conformation of gB that promotes membrane fusion as well as the identification of structures in AD-5 that could be targeted by antibodies to block this early step in HCMV infection.IMPORTANCE HCMV is a major global health concern, and antiviral chemotherapy remains problematic due to toxicity of available compounds and the emergence of drug-resistant viruses. Thus, an HCMV vaccine represents a priority for both governmental and pharmaceutical research programs. A major obstacle for the development of a vaccine is a lack of knowledge of the nature and specificities of protective immune responses that should be induced by such a vaccine. Glycoprotein B of HCMV is an important target for neutralizing antibodies and, hence, is often included as a component of intervention strategies. By generation of fusion-active gB chimeras, we were able to identify target structures of neutralizing antibodies that potently block gB-induced membrane fusion. This experimental system provides an approach to screen for antibodies that interfere with gB's fusogenic activity. In summary, our data will likely contribute to both rational vaccine design and the development of antibody-based therapies against HCMV.

Human Cytomegalovirus Infection in Women With Preexisting Immunity: Sources of Infection and Mechanisms of Infection in the Presence of Antiviral Immunity.

Human cytomegalovirus (HCMV) infection remains an important cause of neurodevelopmental sequelae in infants infected in utero. Unique to the natural history of perinatal HCMV infections is the occurrence of congenital HCMV infections (cCMV) in women with existing immunity to HCMV, infections that have been designated as nonprimary maternal infection. In maternal populations with a high HCMV seroprevalence, cCMV that follows nonprimary maternal infections accounts for 75%-90% of all cases of cCMV infections as well as a large proportion of infected infants with neurodevelopmental sequelae. Although considerable effort has been directed toward understanding immune correlates that can modify maternal infections and intrauterine transmission, the source of virus leading to nonprimary maternal infections and intrauterine transmission is not well defined. Previous paradigms that included reactivation of latent virus as the source of infection in immune women have been challenged by studies demonstrating acquisition and transmission of antigenically distinct viruses, a finding suggesting that reinfection through exposure to an exogenous virus is responsible for some cases of nonprimary maternal infection. Additional understanding of the source(s) of virus that leads to nonprimary maternal infection will be of considerable value in the development and testing of interventions such as vaccines designed to limit the incidence of cCMV in populations with high HCMV seroprevalence.

Contribution of Congenital Cytomegalovirus Infection to Permanent Hearing Loss in a Highly Seropositive Population: The Brazilian Cytomegalovirus Hearing and Maternal Secondary Infection Study.

BACKGROUND: The exact contribution of congenital cytomegalovirus infection (cCMVI) to permanent hearing loss (HL) in highly seropositive populations is unknown. We determined the contribution of cCMVI to HL and estimated the effectiveness of newborn hearing screening (HS) in identifying neonates with CMV-related HL. METHODS: A total of 11 900 neonates born from a population with ≥97% maternal seroprevalence were screened for cCMVI and HL. cCMVI was confirmed by detection of CMV-DNA in saliva and urine at age <3 weeks. RESULTS: Overall, 68 (0.6%; 95% confidence interval [CI], 0.4-0.7) neonates were identified with cCMVI. Of the 91 (0.8%) newborns who failed the HS, 24 (26.4%) were confirmed with HL, including 7 (29.2%; 95% CI, 17.2-59.3) with cCMVI. Another newborn with cCMVI passed the HS but was confirmed with HL at age 21 days. Of the 62 neonates with cCMVI who underwent a complete hearing evaluation, 8 (12.9%; 95% CI, 6.7-23.4) had HL and most (7/8; 87.5%; 95% CI, 46.6-99.7) were identified by HS. The rate of CMV-related HL was 8 per 11 887 neonates (0.7 per 1000 live births). The prevalence ratio of HL among neonates with cCMVI compared to CMV-uninfected neonates was 89.5 (95% CI, 39.7-202.0). No late-onset cCMVI-related HL was detected during a median follow-up of 36 months. CONCLUSIONS: cCMVI is an important cause of HL in childhood in all settings. Integrating targeted cCMVI screening among neonates who fail a HS could be a reasonable, cost-effective strategy to identify newborns with early-onset cCMVI-related HL.

Correction to: Association of CMV genomic mutations with symptomatic infection and hearing loss in congenital CMV infection.

After publication of the original article [1], we were notified that Fig. 3 has "Fig. 1" posted on the top of it and Figs. 4 and 5 have "Genomic Position" in a different spot.

Brain-resident memory CD8+ T cells induced by congenital CMV infection prevent brain pathology and virus reactivation.

Congenital HCMV infection is a leading infectious cause of long-term neurodevelopmental sequelae. Infection of newborn mice with mouse cytomegalovirus (MCMV) intraperitoneally is a well-established model of congenital human cytomegalovirus infection, which best recapitulates the hematogenous route of virus spread to brain and subsequent pathology. Here, we used this model to investigate the role, dynamics, and phenotype of CD8+ T cells in the brain following infection of newborn mice. We show that CD8+ T cells infiltrate the brain and form a pool of tissue-resident memory T cells (TRM cells) that persist for lifetime. Adoptively transferred virus-specific CD8+ T cells provide protection against primary MCMV infection in newborn mice, reduce brain pathology, and remain in the brain as TRM cells. Brain CD8+ TRM cells were long-lived, slowly proliferating cells able to respond to local challenge infection. Importantly, brain CD8+ TRM cells controlled latent MCMV and their depletion resulted in virus reactivation and enhanced inflammation in brain.

Human Cytomegalovirus Immediate Early 1 Protein Causes Loss of SOX2 from Neural Progenitor Cells by Trapping Unphosphorylated STAT3 in the Nucleus.

The mechanisms underlying neurodevelopmental damage caused by virus infections remain poorly defined. Congenital human cytomegalovirus (HCMV) infection is the leading cause of fetal brain development disorders. Previous work has linked HCMV infection to perturbations of neural cell fate, including premature differentiation of neural progenitor cells (NPCs). Here, we show that HCMV infection of NPCs results in loss of the SOX2 protein, a key pluripotency-associated transcription factor. SOX2 depletion maps to the HCMV major immediate early (IE) transcription unit and is individually mediated by the IE1 and IE2 proteins. IE1 causes SOX2 downregulation by promoting the nuclear accumulation and inhibiting the phosphorylation of STAT3, a transcriptional activator of SOX2 expression. Deranged signaling resulting in depletion of a critical stem cell protein is an unanticipated mechanism by which the viral major IE proteins may contribute to brain development disorders caused by congenital HCMV infection.IMPORTANCE Human cytomegalovirus (HCMV) infections are a leading cause of brain damage, hearing loss, and other neurological disabilities in children. We report that the HCMV proteins known as IE1 and IE2 target expression of human SOX2, a central pluripotency-associated transcription factor that governs neural progenitor cell (NPC) fate and is required for normal brain development. Both during HCMV infection and when expressed alone, IE1 causes the loss of SOX2 from NPCs. IE1 mediates SOX2 depletion by targeting STAT3, a critical upstream regulator of SOX2 expression. Our findings reveal an unanticipated mechanism by which a common virus may cause damage to the developing nervous system and suggest novel targets for medical intervention.

WDR5 Facilitates Human Cytomegalovirus Replication by Promoting Capsid Nuclear Egress.

WD repeat-containing protein 5 (WDR5) is essential for assembling the VISA-associated complex to induce a type I interferon antiviral response to Sendai virus infection. However, the roles of WDR5 in DNA virus infections are not well described. Here, we report that human cytomegalovirus exploits WDR5 to facilitate capsid nuclear egress. Overexpression of WDR5 in fibroblasts slightly enhanced the infectious virus yield. However, WDR5 knockdown dramatically reduced infectious virus titers with only a small decrease in viral genome replication or gene expression. Further investigation of late steps of viral replication found that WDR5 knockdown significantly impaired formation of the viral nuclear egress complex and induced substantially fewer infoldings of the inner nuclear membrane. In addition, fewer capsids were associated with these infoldings, and there were fewer capsids in the cytoplasm. Restoration of WDR5 partially reversed these effects. These results suggest that WDR5 knockdown impairs the nuclear egress of capsids, which in turn decreases virus titers. These findings reveal an important role for a host factor whose function(s) is usurped by a viral pathogen to promote efficient replication. Thus, WDR5 represents an interesting regulatory mechanism and a potential antiviral target.IMPORTANCE Human cytomegalovirus (HCMV) has a large (∼235-kb) genome with over 170 open reading frames and exploits numerous cellular factors to facilitate its replication. HCMV infection increases protein levels of WD repeat-containing protein 5 (WDR5) during infection, overexpression of WDR5 enhances viral replication, and knockdown of WDR5 dramatically attenuates viral replication. Our results indicate that WDR5 promotes the nuclear egress of viral capsids, the depletion of WDR5 resulting in a significant decrease in production of infectious virions. This is the first report that WDR5 favors HCMV, a DNA virus, replication and highlights a novel target for antiviral therapy.