Search

Intranasal infection is commonly used to establish a SARS-CoV-2 mouse model due to its non-invasive procedures and a minimal effect from the operation itself. However, mice intranasally infected with SARS-CoV-2 have a high mortality rate, which limits the utility of this model for exploring therapeutic strategies and the sequelae of non-fatal COVID-19 cases. To resolve these limitations, an aerosolised viral administration method has been suggested. However, an in-depth pathological analysis comparing the two models is lacking. Here, we show that inhalation and intranasal SARS-CoV-2 (10 6 PFU) infection models established in K18-hACE2 mice develop unique pathological features in both the respiratory and central nervous systems, which could be directly attributed to the infection method. While the inhalation-infection model exhibited relatively milder pathological parameters, it closely mimicked the prevalent chest CT pattern observed in COVID-19 patients with focal, peripheral lesions and fibrotic scarring in the recuperating lung. We also found the evidence of direct neuron-invasion from the olfactory receptor neurons to the olfactory bulb in the intranasal model and showed the trigeminal nerve as an alternative route of transmission to the brain in inhalation infected mice. Even after viral clearance confirmed at 14 days post-infection, mild lesions were still found in the brain of inhalation-infected mice. These findings suggest that the inhalation-infection model has advantages over the intranasal-infection model in closely mimicking the pathological features of non-fatal symptoms of COVID-19, demonstrating its potential to study the sequelae and possible interventions for long COVID., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper, (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Viruses, such as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), use respiratory epithelial cells as an entry point for infection. Within the nasal cavity, the olfactory epithelium (OE) is particularly sensitive to infections which may lead to olfactory dysfunction. In patients suffering from coronavirus disease 2019, deficits in olfaction have been characterized as a distinctive symptom. Here, we used the K18hACE2 mice to study the spread of SARS-CoV-2 infection and inflammation in the olfactory system (OS) after 7 d of infection. In the OE, we found that SARS-CoV-2 selectively targeted the supporting/sustentacular cells (SCs) and macrophages from the lamina propria. In the brain, SARS-CoV-2 infected some microglial cells in the olfactory bulb (OB), and there was a widespread infection of projection neurons in the OB, piriform cortex (PC), and tubular striatum (TuS). Inflammation, indicated by both elevated numbers and morphologically activated IBA1 + cells (monocyte/macrophage lineages), was preferentially increased in the OE septum, while it was homogeneously distributed throughout the layers of the OB, PC, and TuS. Myelinated OS axonal tracts, the lateral olfactory tract, and the anterior commissure, exhibited decreased levels of 2',3'-cyclic-nucleotide 3'-phosphodiesterase, indicative of myelin defects. Collectively, our work supports the hypothesis that SARS-CoV-2 infected SC and macrophages in the OE and, centrally, microglia and subpopulations of OS neurons. The observed inflammation throughout the OS areas and central myelin defects may account for the long-lasting olfactory deficit., Competing Interests: E.M-L., T.M., B.B., N.S., S.J.M., K.H., N.Y., C.W., and C.A.G. authors declare no competing financial interests. A.I. co-founded RIGImmune, Xanadu Bio, and PanV and is a member of the Board of Directors of Roche Holding and Genentech., (Copyright © 2024 Martin-Lopez et al.)

Parkinson's disease (PD) is a prevalent neurodegenerative disorder that affects 7-10 million individuals worldwide. A common early symptom of PD is olfactory dysfunction (OD), and more than 90% of PD patients suffer from OD. Recent studies have highlighted a high incidence of OD in patients with SARS-CoV-2 infection. This review investigates the potential convergence of OD in PD and COVID-19, particularly focusing on the mechanisms by which neuroinflammation contributes to OD and neurological events. Starting from our fundamental understanding of the olfactory bulb, we summarize the clinical features of OD and pathological features of the olfactory bulb from clinical cases and autopsy reports in PD patients. We then examine SARS-CoV-2-induced olfactory bulb neuropathology and OD and emphasize the SARS-CoV-2-induced neuroinflammatory cascades potentially leading to PD manifestations. By activating microglia and astrocytes, as well as facilitating the aggregation of α-synuclein, SARS-CoV-2 could contribute to the onset or exacerbation of PD. We also discuss the possible contributions of NF-κB, the NLRP3 inflammasome, and the JAK/STAT, p38 MAPK, TLR4, IL-6/JAK2/STAT3 and cGAS-STING signaling pathways. Although olfactory dysfunction in patients with COVID-19 may be reversible, it is challenging to restore OD in patients with PD. With the emergence of new SARS-CoV-2 variants and the recurrence of infections, we call for continued attention to the intersection between PD and SARS-CoV-2 infection, especially from the perspective of OD., Competing Interests: Declaration of Competing Interest Authors declare no conflict of interest., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Anosmia was identified as a hallmark of COVID-19 early in the pandemic, however, with the emergence of variants of concern, the clinical profile induced by SARS-CoV-2 infection has changed, with anosmia being less frequent. Here, we assessed the clinical, olfactory and neuroinflammatory conditions of golden hamsters infected with the original Wuhan SARS-CoV-2 strain, its isogenic ORF7-deletion mutant and three variants: Gamma, Delta, and Omicron/BA.1. We show that infected animals develop a variant-dependent clinical disease including anosmia, and that the ORF7 of SARS-CoV-2 contributes to the induction of olfactory dysfunction. Conversely, all SARS-CoV-2 variants are neuroinvasive, regardless of the clinical presentation they induce. Taken together, this confirms that neuroinvasion and anosmia are independent phenomena upon SARS-CoV-2 infection. Using newly generated nanoluciferase-expressing SARS-CoV-2, we validate the olfactory pathway as a major entry point into the brain in vivo and demonstrate in vitro that SARS-CoV-2 travels retrogradely and anterogradely along axons in microfluidic neuron-epithelial networks., (© 2023. The Author(s).)

Current understanding of coronavirus disease 2019 (COVID-19) pathophysiology is limited by disease heterogeneity, complexity, and a paucity of studies assessing patient tissues with advanced molecular tools. Rapid autopsy tissues were evaluated using multiscale, next-generation RNA-sequencing methods (bulk, single-nuclei, and spatial transcriptomics) to provide unprecedented molecular resolution of COVID-19-induced damage. Comparison of infected/uninfected tissues revealed four major regulatory pathways. Effectors within these pathways could constitute novel therapeutic targets, including the complement receptor C3AR1, calcitonin receptor-like receptor, or decorin. Single-nuclei RNA sequencing of olfactory bulb and prefrontal cortex highlighted remarkable diversity of coronavirus receptors. Angiotensin-converting enzyme 2 was rarely expressed, whereas basigin showed diffuse expression, and alanyl aminopeptidase, membrane, was associated with vascular/mesenchymal cell types. Comparison of lung and lymph node tissues from patients with different symptoms (one had died after a month-long hospitalization with multiorgan involvement, and the other had died after a few days of respiratory symptoms) with digital spatial profiling resulted in distinct molecular phenotypes. Evaluation of COVID-19 rapid autopsy tissues with advanced molecular techniques can identify pathways and effectors, map diverse receptors at the single-cell level, and help dissect differences driving diverging clinical courses among individual patients. Extension of this approach to larger data sets will substantially advance the understanding of the mechanisms behind COVID-19 pathophysiology., (Copyright © 2021 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.)

Anosmia, the loss of smell, is a common and often the sole symptom of COVID-19. The onset of the sequence of pathobiological events leading to olfactory dysfunction remains obscure. Here, we have developed a postmortem bedside surgical procedure to harvest endoscopically samples of respiratory and olfactory mucosae and whole olfactory bulbs. Our cohort of 85 cases included COVID-19 patients who died a few days after infection with SARS-CoV-2, enabling us to catch the virus while it was still replicating. We found that sustentacular cells are the major target cell type in the olfactory mucosa. We failed to find evidence for infection of olfactory sensory neurons, and the parenchyma of the olfactory bulb is spared as well. Thus, SARS-CoV-2 does not appear to be a neurotropic virus. We postulate that transient insufficient support from sustentacular cells triggers transient olfactory dysfunction in COVID-19. Olfactory sensory neurons would become affected without getting infected., Competing Interests: Declaration of interests T.D.H., A.N., L.P., and J.W.R. are employees and stockholders at NanoString Technologies, Inc., (Copyright © 2021 The Author(s). Published by Elsevier Inc. All rights reserved.)

Background: Citomegalovirus (CMV) infects approximately 1% of live newborns. About 10% of the infants affected by congenital CMV infection are symptomatic at birth and up to 60% of these infants will develop permanent neurological disabilities. Depending on gestational age (GA) at the time of infection, the involvement of central nervous system (CNS) can lead to malformations of cortical development, calcifications, periventricular white matter lesions and cysts, ventriculomegaly and cerebellar hypoplasia., Case Presentation: We report the MRI findings in a Caucasian female born at 32 weeks of post-menstrual age with post-birth diagnosis of congenital CMV infection showing an unusual and peculiar marked T2 hyperintensity of the inner part of olfactory bulbs in addition to the CMV related diffuse brain involvement. Despite the known extensively described fetal and neonatal Magnetic Resonance Imaging (MRI) findings in CMV infected fetuses and newborns, any in vivo MRI depiction of olfactory system damage have never been reported so far. Nevertheless, in murine studies CMV is known to infect the placenta during pregnancy showing particular tropism for neural stem cells of the olfactory system and previous neuropathologic study on CMV infected human fetal brains from 23 to 28 weeks of GA reported damage in the olfactory bulbs (OB) consisting in disseminated cytomegalic cells, inflammation, necrosis and neuronal and radial glial cell loss. Therefore, we assume an OB involvement and damage in congenital CMV infection., Conclusion: To our knowledge this is the first in vivo MRI evidence of OB damage in a newborn with congenital CMV infection that may give new insights on CMV infection., (© 2021. The Author(s).)

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative pathogen of coronavirus disease 2019 (COVID-19). It is known as a respiratory virus, but SARS-CoV-2 appears equally, or even more, infectious for the olfactory epithelium (OE) than for the respiratory epithelium in the nasal cavity. In light of the small area of the OE relative to the respiratory epithelium, the high prevalence of olfactory dysfunctions (ODs) in COVID-19 has been bewildering and has attracted much attention. This review aims to first examine the cytological and molecular biological characteristics of the OE, especially the microvillous apical surfaces of sustentacular cells and the abundant SARS-CoV-2 receptor molecules thereof, that may underlie the high susceptibility of this neuroepithelium to SARS-CoV-2 infection and damages. The possibility of SARS-CoV-2 neurotropism, or the lack of it, is then analyzed with regard to the expression of the receptor (angiotensin-converting enzyme 2) or priming protease (transmembrane serine protease 2), and cellular targets of infection. Neuropathology of COVID-19 in the OE, olfactory bulb, and other related neural structures are also reviewed. Toward the end, we present our perspectives regarding possible mechanisms of SARS-CoV-2 neuropathogenesis and ODs, in the absence of substantial viral infection of neurons. Plausible causes for persistent ODs in some COVID-19 convalescents are also examined.

The Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), the causative agent of human COVID-19, not only causes flu-like symptoms and gut microbiome complications but a large number of infected individuals also experience a host of neurological symptoms including loss of smell and taste, seizures, difficulty concentrating, decreased alertness, and brain inflammation. Although SARS-CoV-2 infections are not more prevalent in Parkinson's disease patients, a higher mortality rate has been reported not only associated with older age and longer disease duration, but also through several mechanisms, such as interactions with the brain dopaminergic system and through systemic inflammatory responses. Indeed, a number of the neurological symptoms seen in COVID-19 patients, as well as the alterations in the gut microbiome, are also prevalent in patients with Parkinson's disease. Furthermore, biochemical pathways such as oxidative stress, inflammation, and protein aggregation have shared commonalities between Parkinson's disease and COVID-19 disease progression. In this review, we describe and compare the numerous similarities and intersections between neurodegeneration in Parkinson's disease and RNA viral infections, emphasizing the current SARS-CoV-2 global health crisis., (© 2021. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Coronavirus disease 2019 (COVID-19) pandemic is caused by the novel coronavirus that has spread rapidly around the world, leading to high mortality because of multiple organ dysfunction; however, its underlying molecular mechanism is unknown. To determine the molecular mechanism of multiple organ dysfunction, a bioinformatics analysis method based on a time-order gene co-expression network (TO-GCN) was performed. First, gene expression profiles were downloaded from the gene expression omnibus database (GSE161200), and a TO-GCN was constructed using the breadth-first search (BFS) algorithm to infer the pattern of changes in the different organs over time. Second, Gene Ontology enrichment analysis was used to analyze the main biological processes related to COVID-19. The initial gene modules for the immune response of different organs were defined as the research object. The STRING database was used to construct a protein-protein interaction network of immune genes in different organs. The PageRank algorithm was used to identify five hub genes in each organ. Finally, the Comparative Toxicogenomics Database played an important role in exploring the potential compounds that target the hub genes. The results showed that there were two types of biological processes: the body's stress response and cell-mediated immune response involving the lung, trachea, and olfactory bulb (olf) after being infected by COVID-19. However, a unique biological process related to the stress response is the regulation of neuronal signals in the brain. The stress response was heterogeneous among different organs. In the lung, the regulation of DNA morphology, angiogenesis, and mitochondrial-related energy metabolism are specific biological processes related to the stress response. In particular, an effect on tracheal stress response was made by the regulation of protein metabolism and rRNA metabolism-related biological processes, as biological processes. In the olf, the distinctive stress responses consist of neural signal transmission and brain behavior. In addition, myeloid leukocyte activation and myeloid leukocyte-mediated immunity in response to COVID-19 can lead to a cytokine storm. Immune genes such as SRC , RHOA , CD40LG , CSF1 , TNFRSF1A , FCER1G , ICAM1 , LAT , LCN2 , PLAU , CXCL10 , ICAM1 , CD40 , IRF7 , and B2M were predicted to be the hub genes in the cytokine storm. Furthermore, we inferred that resveratrol, acetaminophen, dexamethasone, estradiol, statins, curcumin, and other compounds are potential target drugs in the treatment of COVID-19., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Zou, Su, Wang, Yi, Qiu, Yin, Zhou, Niu, Wang and Su.)

To provide instructive clues for clinical practice and further research of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, we analyzed the existing literature on viral neuroinvasion of SARS-CoV-2 in coronavirus disease 2019 (COVID-19) patients. To date, SARS-CoV-2 has been detected in the cerebrospinal fluid (CSF) or brain parenchyma in quite a few patients, which provide undeniable evidence for the neuroinvasive potential of this novel coronavirus. In contrast with the cerebrum and cerebellum, the detection rate of SARS-CoV-2 was higher in the olfactory system and the brainstem, both of which also showed severe microgliosis and lymphocytic infiltrations. As compared with the number of patients who underwent viral testing in the central nervous system (CNS), the number of patients showing positive results seems very small. However, it seems too early to conclude that the neuroinvasion of SARS-CoV-2 is rare in COVID-19 patients because the detection methods or sampling procedures in some studies may not be suitable or sufficient to reveal the CNS infection induced by neurotropic viruses. Moreover, the primary symptoms and/or causes of death were distinctly different among examined patients, which probably caused more conspicuous pathological changes than those due to the direct infection that usually localized to specific brain areas. Unfortunately, most autopsy studies did not provide sufficient details about neurological symptoms or suspected diagnoses of the examined patients, and the documentation of neuropathological changes was often incomplete. Given the complex pathophysiology of COVID-19 and the characteristics of neurotropic viruses, it is understandable that any study of the CNS infection may inevitably have limitations., (© 2021 Wiley Periodicals LLC.)

Viral encephalitis initiates a series of immunological events in the brain that can lead to brain damage and death. Astrocytes express IFN-β in response to neurotropic infection, whereas activated microglia produce proinflammatory cytokines and accumulate at sites of infection. Here, we observed that neurotropic vesicular stomatitis virus (VSV) infection causes recruitment of leukocytes into the central nervous system (CNS), which requires MyD88, an adaptor of Toll-like receptor and interleukin-1 receptor signaling. Infiltrating leukocytes, and in particular CD8 + T cells, protected against lethal VSV infection of the CNS. Reconstitution of MyD88, specifically in neurons, restored chemokine production in the olfactory bulb as well as leukocyte recruitment into the infected CNS and enhanced survival. Comparative analysis of the translatome of neurons and astrocytes verified neurons as the critical source of chemokines, which regulated leukocyte infiltration of the infected brain and affected survival., (Copyright © 2021 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.)

Neurological manifestations are frequently reported in the COVID-19 patients. Neuromechanism of SARS-CoV-2 remains to be elucidated. In this study, we explored the mechanisms of SARS-CoV-2 neurotropism via our established non-human primate model of COVID-19. In rhesus monkey, SARS-CoV-2 invades the CNS primarily via the olfactory bulb. Thereafter, viruses rapidly spread to functional areas of the central nervous system, such as hippocampus, thalamus, and medulla oblongata. The infection of SARS-CoV-2 induces the inflammation possibly by targeting neurons, microglia, and astrocytes in the CNS. Consistently, SARS-CoV-2 infects neuro-derived SK-N-SH, glial-derived U251, and brain microvascular endothelial cells in vitro. To our knowledge, this is the first experimental evidence of SARS-CoV-2 neuroinvasion in the NHP model, which provides important insights into the CNS-related pathogenesis of SARS-CoV-2.

COVID-19 is a pandemic viral infection caused by a novel coronavirus, SARS-CoV2, which is a global concern of the twenty-first century for its rapid spreading in a short period. Apart from its known acute respiratory involvements, the CNS manifestations of COVID-19 are common. These neurological symptoms are diverse and could range from mild nonspecific or specific symptoms such as the loss of various sensory perceptions, the worrying autoimmune Guillain-Barré syndrome, to the life-threatening acute disseminated encephalomyelitis, and the CNS-mediated respiratory distress. An autopsy report documented the presence of SARS-CoV2 in brain tissues of a COVID-19 patient. However, there is no definite conclusion on the mechanisms of SARS-CoV2 neuroinvasion. These proposed mechanisms include the direct viral invasion, the systemic blood circulation, or the distribution of infected immune cells. Concerning these different neuropathophysiologies, COVID-19 patients who are presenting with either the early-onset, multiple, and severe CNS symptoms or rapid respiratory deterioration should be suspected for the direct viral neuroinvasion, and appropriate management options should be considered. This article reviews the neurological manifestations, the proposed neuroinvasive mechanisms, and the potential neurological sequelae of SARS-CoV2.

The complete features of the neurological complications of coronavirus disease 2019 (COVID-19) still need to be elucidated, including associated cranial nerve involvement. In the present study we describe cranial nerve lesions seen in magnetic resonance imaging (MRI) of six cases of confirmed COVID-19, involving the olfactory bulb, optic nerve, abducens nerve, and facial nerve. Cranial nerve involvement was associated with COVID-19, but whether by direct viral invasion or autoimmunity needs to be clarified. The development of neurological symptoms after initial respiratory symptoms and the absence of the virus in the cerebrospinal fluid (CSF) suggest the possibility of autoimmunity.

Neuropilin-1 (NRP-1), a member of a family of signaling proteins, was shown to serve as an entry factor and potentiate SARS Coronavirus 2 (SARS-CoV-2) infectivity in vitro. This cell surface receptor with its disseminated expression is important in angiogenesis, tumor progression, viral entry, axonal guidance, and immune function. NRP-1 is implicated in several aspects of a SARS-CoV-2 infection including possible spread through the olfactory bulb and into the central nervous system and increased NRP-1 RNA expression in lungs of severe Coronavirus Disease 2019 (COVID-19). Up-regulation of NRP-1 protein in diabetic kidney cells hint at its importance in a population at risk of severe COVID-19. Involvement of NRP-1 in immune function is compelling, given the role of an exaggerated immune response in disease severity and deaths due to COVID-19. NRP-1 has been suggested to be an immune checkpoint of T cell memory. It is unknown whether involvement and up-regulation of NRP-1 in COVID-19 may translate into disease outcome and long-term consequences, including possible immune dysfunction. It is prudent to further research NRP-1 and its possibility of serving as a therapeutic target in SARS-CoV-2 infections. We anticipate that widespread expression, abundance in the respiratory and olfactory epithelium, and the functionalities of NRP-1 factor into the multiple systemic effects of COVID-19 and challenges we face in management of disease and potential long-term sequelae., Competing Interests: The authors have declared that no competing interests exist.

Anosmia is one of the most prevalent symptoms of SARS-CoV-2 infection during the COVID-19 pandemic. However, the cellular mechanism behind the sudden loss of smell has not yet been investigated. The initial step of odour detection takes place in the pseudostratified olfactory epithelium (OE) mainly composed of olfactory sensory neurons surrounded by supporting cells known as sustentacular cells. The olfactory neurons project their axons to the olfactory bulb in the central nervous system offering a potential pathway for pathogens to enter the central nervous system by bypassing the blood brain barrier. In the present study, we explored the impact of SARS-CoV-2 infection on the olfactory system in golden Syrian hamsters. We observed massive damage of the OE as early as 2 days post nasal instillation of SARS-CoV-2, resulting in a major loss of cilia necessary for odour detection. These damages were associated with infection of a large proportion of sustentacular cells but not of olfactory neurons, and we did not detect any presence of the virus in the olfactory bulbs. We observed massive infiltration of immune cells in the OE and lamina propria of infected animals, which may contribute to the desquamation of the OE. The OE was partially restored 14 days post infection. Anosmia observed in COVID-19 patient is therefore likely to be linked to a massive and fast desquamation of the OE following sustentacular cells infection with SARS-CoV-2 and subsequent recruitment of immune cells in the OE and lamina propria., (Copyright © 2020 Elsevier Inc. All rights reserved.)

The olfactory mucosa, where the first step of odor detection occurs, is a privileged pathway for environmental toxicants and pathogens toward the central nervous system. Indeed, some pathogens can infect olfactory sensory neurons including their axons projecting to the olfactory bulb allowing them to bypass the blood-brain barrier and reach the central nervous system (CNS) through the so-called olfactory pathway. The respiratory syncytial virus (RSV) is a major respiratory tract pathogen but there is growing evidence that RSV may lead to CNS impairments. However, the mechanisms involved in RSV entering into the CNS have been poorly described. In this study, we wanted to explore the capacity of RSV to reach the CNS via the olfactory pathway and to better characterize RSV cellular tropism in the nasal cavity. We first explored the distribution of RSV infectious sites in the nasal cavity by in vivo bioluminescence imaging and a tissue clearing protocol combined with deep-tissue imaging and 3D image analyses. This whole tissue characterization was confirmed with immunohistochemistry and molecular biology approaches. Together, our results provide a novel 3D atlas of mouse nasal cavity anatomy and show that RSV can infect olfactory sensory neurons giving access to the central nervous system by entering the olfactory bulb. Cover Image for this issue: doi: 10.1111/jnc.14765., (© 2019 International Society for Neurochemistry.)

The main neurological manifestation of COVID-19 is loss of smell or taste. The high incidence of smell loss without significant rhinorrhea or nasal congestion suggests that SARS-CoV-2 targets the chemical senses through mechanisms distinct from those used by endemic coronaviruses or other common cold-causing agents. Here we review recently developed hypotheses about how SARS-CoV-2 might alter the cells and circuits involved in chemosensory processing and thereby change perception. Given our limited understanding of SARS-CoV-2 pathogenesis, we propose future experiments to elucidate disease mechanisms and highlight the relevance of this ongoing work to understanding how the virus might alter brain function more broadly., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Usually, pandemic COVID-19 disease, caused by SARS-CoV2, presents with mild respiratory symptoms such as fever, cough, but frequently also with anosmia and neurological symptoms. Virus-cell fusion is mediated by angiotensin-converting enzyme 2 (ACE2) and transmembrane serine protease 2 (TMPRSS2) with their organ expression pattern determining viral tropism. Clinical presentation suggests rapid viral dissemination to the central nervous system leading frequently to severe symptoms including viral meningitis. Here, we provide a comprehensive expression landscape of ACE2 and TMPRSS2 proteins across human postmortem nasal and olfactory tissue. Sagittal sections through the human nose complemented with immunolabelling of respective cell types represent different anatomically defined regions including olfactory epithelium, respiratory epithelium of the nasal conchae and the paranasal sinuses along with the hardly accessible human olfactory bulb. ACE2 can be detected in the olfactory epithelium as well as in the respiratory epithelium of the nasal septum, the nasal conchae, and the paranasal sinuses. ACE2 is located in the sustentacular cells and in the glandular cells in the olfactory epithelium as well as in the basal cells, glandular cells, and epithelial cells of the respiratory epithelium. Intriguingly, ACE2 is not expressed in mature or immature olfactory receptor neurons and basal cells in the olfactory epithelium. Similarly, ACE2 is not localized in the olfactory receptor neurons albeit the olfactory bulb is positive. Vice versa, TMPRSS2 can also be detected in the sustentacular cells and the glandular cells of the olfactory epithelium. Our findings provide the basic anatomical evidence for the expression of ACE2 and TMPRSS2 in the human nose, olfactory epithelium, and olfactory bulb. Thus, they are substantial for future studies that aim to elucidate the symptom of SARS-CoV2 induced anosmia via the olfactory pathway., (© 2021 S. Karger AG, Basel.)

Background: Pseudorabies virus (PRV, or suid herpesvirus, SuHV-1), a member of the herpesvirus family, has an extremely broad host range and threatens the pig industry in China. PRV can evade host innate immunity and infect the kidney, lung, brain and other tissues. At the same time, many studies have reported that microRNA (miRNA) can affect the replication of viruses by regulating gene expression levels., Results: Here, to identify changes in miRNA expression and post-transcriptional regulation associated with PRV infection in the lung, spleen, and olfactory bulb, we sequenced small RNAs in tissues of rats infected or uninfected with PRV strain XJ (PRV-XJ). Sixty-one, 199 and 29 differentially-expressed miRNAs were identified in the lung, spleen, and olfactory bulb, respectively, of infected compared with uninfected rats. Among the miRNAs differentially-expressed in PRV-infected rats, 36, 171, and 15 miRNAs showed tissue-selective expression in the olfactory bulb, lung and spleen, respectively. All differentially-expressed miRNAs were analyzed for their GO functional annotations and KEGG pathway associations ., Conclusions: In PRV-XJ-infected rats, miRNAs were differentially expressed in the lung, spleen and olfactory bulb. These miRNAs were involved in regulating various pathways of the nervous, respiratory and immune systems, and may affect the tissue tropism of the virus and play pivotal roles in viral infection and proliferation.

Olfaction guides goal-directed behaviours including feeding. To investigate how central olfactory neural circuits control feeding behaviour in mice, we performed retrograde tracing from the lateral hypothalamus (LH), an important feeding centre. We observed a cluster of retrogradely labelled cells distributed in the posteroventral region of the olfactory peduncle. Histochemical analyses revealed that the majority of these retrogradely labelled projection neurons expressed glutamic acid decarboxylase 65/67 (GAD65/67), but not vesicular glutamate transporter 1 (VGluT1). We named this region containing GABAergic projection neurons the ventral olfactory nucleus (VON) to differentiate it from the conventional olfactory peduncle. VON neurons were less immunoreactive for DARPP-32, a striatal neuron marker, compared to neurons in the olfactory tubercle and nucleus accumbens, which distinguished the VON from the ventral striatum. Fluorescent labelling confirmed putative synaptic contacts between VON neurons and olfactory bulb projection neurons. Rabies-virus-mediated trans-synaptic labelling revealed that VON neurons received synaptic inputs from the olfactory bulb, other olfactory cortices, horizontal limb of the diagonal band, and prefrontal cortex. Collectively, these results identify novel GABAergic projection neurons in the olfactory cortex that may integrate olfactory sensory and top-down inputs and send inhibitory output to the LH, which may modulate odour-guided LH-related behaviours.

Mammalian Bornavirus (BoDV-1) typically causes a fatal neurologic disorder in horses and sheep, and was recently shown to cause fatal encephalitis in humans with and without transplant reception. It has been suggested that BoDV-1 enters the central nervous system (CNS) via the olfactory pathway. However, (I) susceptible cell types that replicate the virus for successful spread, and (II) the role of olfactory ensheathing cells (OECs), remained unclear. To address this, we studied the intranasal infection of adult rats with BoDV-1 in vivo and in vitro, using olfactory mucosal (OM) cell cultures and the cultures of purified OECs. Strikingly, in vitro and in vivo, viral antigen and mRNA were present from four days post infection (dpi) onwards in the olfactory receptor neurons (ORNs), but also in all other cell types of the OM, and constantly in the OECs. In contrast, in vivo, BoDV-1 genomic RNA was only detectable in adult and juvenile ORNs, nerve fibers, and in OECs from 7 dpi on. In vitro, the rate of infection of OECs was significantly higher than that of the OM cells, pointing to a crucial role of OECs for infection via the olfactory pathway. Thus, this study provides important insights into the transmission of neurotropic viral infections with a zoonotic potential.

Human coronaviruses (HCoVs) are recognized respiratory pathogens for which accumulating evidence indicates that in vulnerable patients the infection can cause more severe pathologies. HCoVs are not always confined to the upper respiratory tract and can invade the central nervous system (CNS) under still unclear circumstances. HCoV-induced neuropathologies in humans are difficult to diagnose early enough to allow therapeutic interventions. Making use of our already described animal model of HCoV neuropathogenesis, we describe the route of neuropropagation from the nasal cavity to the olfactory bulb and piriform cortex and then the brain stem. We identified neuron-to-neuron propagation as one underlying mode of virus spreading in cell culture. Our data demonstrate that both passive diffusion of released viral particles and axonal transport are valid propagation strategies used by the virus. We describe for the first time the presence along axons of viral platforms whose static dynamism is reminiscent of viral assembly sites. We further reveal that HCoV OC43 modes of propagation can be modulated by selected HCoV OC43 proteins and axonal transport. Our work, therefore, identifies processes that may govern the severity and nature of HCoV OC43 neuropathogenesis and will make possible the development of therapeutic strategies to prevent occurrences. IMPORTANCE Coronaviruses may invade the CNS, disseminate, and participate in the induction of neurological diseases. Their neuropathogenicity is being increasingly recognized in humans, and the presence and persistence of human coronaviruses (HCoV) in human brains have been proposed to cause long-term sequelae. Using our mouse model relying on natural susceptibility to HCoV OC43 and neuronal cell cultures, we have defined the most relevant path taken by HCoV OC43 to access and spread to and within the CNS toward the brain stem and spinal cord and studied in cell culture the underlying modes of intercellular propagation to better understand its neuropathogenesis. Our data suggest that axonal transport governs HCoV OC43 egress in the CNS, leading to the exacerbation of neuropathogenesis. Exploiting knowledge on neuroinvasion and dissemination will enhance our ability to control viral infection within the CNS, as it will shed light on underlying mechanisms of neuropathogenesis and uncover potential druggable molecular virus-host interfaces., (Copyright © 2018 American Society for Microbiology.)

Background: Flaviviruses are a group of diverse and emerging arboviruses and an immense global health problem. A number of flaviviruses are neurotropic, causing severe encephalitis and even death. Type I interferons (IFNs) are the first line of defense of the innate immune system against flavivirus infection. IFNs elicit the concerted action of numerous interferon-stimulated genes (ISGs) to restrict both virus infection and replication. Viperin (virus-inhibitory protein, endoplasmic reticulum-associated, IFN-inducible) is an ISG with broad-spectrum antiviral activity against multiple flaviviruses in vitro. Its activity in vivo restricts neurotropic infections to specific regions of the central nervous system (CNS). However, the cell types in which viperin activity is required are unknown. Here we have examined both the regional and cell-type specificity of viperin in the defense against infection by several model neurotropic flaviviruses., Methods: Viral burden and IFN induction were analyzed in vivo in wild-type and viperin -/- mice infected with Langat virus (LGTV). The effects of IFN pretreatment were tested in vitro in primary neural cultures from different brain regions in response to infection with tick-borne encephalitis virus (TBEV), West Nile virus (WNV), and Zika virus (ZIKV)., Results: Viperin activity restricted nonlethal LGTV infection in the spleen and the olfactory bulb following infection via a peripheral route. Viperin activity was also necessary to restrict LGTV replication in the olfactory bulb and the cerebrum following CNS infection, but not in the cerebellum. In vitro, viperin could restrict TBEV replication in primary cortical neurons, but not in the cerebellar granule cell neurons. Interferon-induced viperin was also very important in primary cortical neurons to control TBEV, WNV, and ZIKV., Conclusions: Our findings show that viperin restricts replication of neurotropic flaviviruses in the CNS in a region- and cell-type-specific manner. The most important sites of activity are the olfactory bulb and cerebrum. Activity within the cerebrum is required in the cortical neurons in order to restrict spread. This study exemplifies cell type and regional diversity of the IFN response within the CNS and shows the importance of a potent broad-spectrum antiviral ISG.

Vesicular stomatitis virus (VSV) and rabies and Chandipura viruses belong to the Rhabdovirus family. VSV is a common laboratory virus to study viral evolution and host immune responses to viral infection, and recombinant VSV-based vectors have been widely used for viral oncolysis, vaccination, and gene therapy. Although the tropism of VSV is broad, and its envelope glycoprotein G is often used for pseudotyping other viruses, the host cellular components involved in VSV infection remain unclear. Here, we demonstrate that the host protein leucine-rich repeat-containing G protein-coupled receptor 4 (Lgr4) is essential for VSV and VSV-G pseudotyped lentivirus (VSVG-LV) to infect susceptible cells. Accordingly, Lgr4-deficient mice had dramatically decreased VSV levels in the olfactory bulb. Furthermore, Lgr4 knockdown in RAW 264.7 cells also significantly suppressed VSV infection, and Lgr4 overexpression in RAW 264.7 cells enhanced VSV infection. Interestingly, only VSV infection relied on Lgr4, whereas infections with Newcastle disease virus, influenza A virus (A/WSN/33), and herpes simplex virus were unaffected by Lgr4 status. Of note, assays of virus entry, cell ELISA, immunoprecipitation, and surface plasmon resonance indicated that VSV bound susceptible cells via the Lgr4 extracellular domain. Pretreating cells with an Lgr4 antibody, soluble LGR4 extracellular domain, or R-spondin 1 blocked VSV infection by competitively inhibiting VSV binding to Lgr4. Taken together, the identification of Lgr4 as a VSV-specific host factor provides important insights into understanding VSV entry and its pathogenesis and lays the foundation for VSV-based gene therapy and viral oncolytic therapeutics., (© 2017 by The American Society for Biochemistry and Molecular Biology, Inc.)

Background: Herpes simplex virus 1 (HSV-1) infection can result in a life-threatening condition known as herpes simplex encephalitis (HSE). Trafficking patterns by which the virus reaches the central nervous system (CNS) following ocular infection are unresolved. We evaluated early viral dissemination pathways following ocular infection that involve trafficking to the olfactory bulb (OB). Additionally, we have characterized the capacity of HSV-1 to establish latency within OB tissue and profiled the local T lymphocyte response over the course of the acute infection into latency., Methods: Scarified corneas of C57BL/6 or reporter-inducible Rosa mice (Rosa Td/Tm ) were inoculated with HSV-1 and assessed for viral dissemination into the peripheral nervous system (PNS) and CNS by RT-PCR and confocal microscopy. T cells and the resident microglia activation signatures were analyzed by flow cytometry. T cell effector function in the form of IFN-γ secretion was measured by T cells isolated from OB in comparison to T cells from other nervous system sites known to harbor HSV-1-specific memory T cells., Results: Following ocular infection, HSV-1 viral titers from nasal secretions were detected as early as 48 h through 8 days post infection (8 DPI). HSV-1 gene expression was expressed as early as 2 days following ocular infection in the OB and was consistent with an enhanced expression in the ophthalmic, maxillary, and mandibular branch of the trigeminal nerve ganglia (TG). Rosa fluorescence protein expression (RFP + ) representing HSV-1-infected cells from Rosa Td/Tm mice was detected in the OB before other areas of the CNS (2 DPI). Additionally, during acute infection, most infected cells appeared to be anatomically distributed within the OB rather than other regions of the CNS. During latency (i.e., 30 DPI and beyond) despite no detectable infectious virus or lytic gene expression and low levels of latency associated transcripts, total effector (CD44 + CD62 - ) CD4 + T, CD8 + T, HSV-1-specific CD8 + T cells, and MHC class II positive resident microglia numbers continued to increase. CD4 + and CD8 + T cell populations isolated from the OB during latency were capable of responding to PMA/ionomycin in the production of IFN-γ similar to T cells from other tissue that possess latent virus including the TG and brain stem., Conclusions: It is currently understood that HSV-1 traffics to the TG following ocular infection. We have identified a second conduit by which HSV-1 can directly access the CNS bypassing the brain stem. We have also recognized that the OB is unique in that during HSV-1 latency, latency-associated transcripts levels were marginally above uninfected controls. Despite these findings, the local immune response mimicked the phenotype of an active infection during latency.

Viruses have long been implicated in the pathogenesis of classical encephalitis lethargica, which was first described by Constantin von Economo in 1917. In this article, I propose the hypothesis that an airborne virus travels along the olfactory conduit to infect the olfactory bulb; this local infection or induced neuroinflammation, in turn, retrogradely targets certain neuronal populations with sleep-wake regulatory functions in the hypothalamus and midbrain, leading to the development of wakeful inactivity, a hallmark clinical feature of the disease. Furthermore, the olfactory vector hypothesis may also explain the pathomechanism of the debilitating complication of the disease, i.e., postencephalitic parkinsonism, in terms of a recently discovered nigro-olfactory projection., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Structural and ultrastructural alterations in human olfactory pathways and putative associations with human herpesvirus 6 (HHV-6) infection were studied. The olfactory bulb/tract samples from 20 subjects with an unspecified encephalopathy determined by pathomorphological examination of the brain autopsy, 17 healthy age-matched and 16 younger controls were used. HHV-6 DNA was detected in 60, 29, and 19% of cases in these groups, respectively. In the whole encephalopathy group, significantly more HHV-6 positive neurons and oligodendrocytes were found in the gray matter, whereas, significantly more HHV-6 positive astrocytes, oligodendrocytes, microglia/macrophages and endothelial cells were found in the white matter. Additionally, significantly more HHV-6 positive astrocytes and, in particular, oligodendrocytes were found in the white matter when compared to the gray matter. Furthermore, when only HHV-6 PCR+ encephalopathy cases were studied, we observed similar but stronger associations between HHV-6 positive oligodendrocytes and CD68 positive cells in the white matter. Cellular alterations were additionally evidenced by anti-S100 immunostaining, demonstrating a significantly higher number of S100 positive cells in the gray matter of the whole encephalopathy group when compared to the young controls, and in the white matter when compared to both control groups. In spite the decreased S100 expression in the PCR+ encephalopathy group when compared to PCR- cases and controls, groups demonstrated significantly higher number of S100 positive cells in the white compared to the gray matter. Ultrastructural changes confirming the damage of myelin included irregularity of membranes and ballooning of paranodal loops. This study shows that among the cellular targets of the nervous system, HHV-6 most severely affects oligodendrocytes and the myelin made by them., Competing Interests: The authors have declared that no competing interests exist.

The severity of clinical symptoms induced by pseudorabies virus (PRV) infection of its natural host is inversely related to the age of the pig. During this study, 2- and 15-week-old pigs were inoculated with PRV strain NIA3. This resulted in important clinical disease, although the associated morbidity and mortality were lower in older pigs. Quantitative PCR analysis of viral DNA in different organs confirmed the general knowledge on PRV pathogenesis. Several new findings and potential explanations for the observed age-dependent differences in virulence, however, were determined from the study of viral and cytokine mRNA expression at important sites of neuropathogenesis. First, only limited viral and cytokine mRNA expression was detected in the nasal mucosa, suggesting that other sites may serve as the primary replication site. Second, PRV reached the trigeminal ganglion (TG) and brain stem rapidly upon infection but, compared to 2-week-old pigs, viral replication was less pronounced in 15-week-old pigs, and the decrease in viral mRNA expression was not preceded by or associated with an increased cytokine expression. Third, extensive viral replication associated with a robust expression of cytokine mRNA was detected in the olfactory bulbs of pigs from both age categories and correlated with the observed neurological disease. Our results suggest that age-dependent differences in PRV-induced clinical signs are probably due to enhanced viral replication and associated immunopathology in immature TG and the central nervous system neurons of 2-week-old pigs and that neurological disease is related with extensive viral replication and an associated immune response in the olfactory bulb., Importance: It is well known that alphaherpesvirus infections of humans and animals result in more severe clinical disease in newborns than in older individuals and that this is probably related to differences in neuropathogenesis. The underlying mechanisms, however, remain unclear. Pseudorabies virus infection of its natural host, the pig, provides a suitable infection model to study this more profoundly. We show here that the severe neurological disease observed in 2-week-old pigs does not appear to be related to a hampered innate immune response but is more likely to reflect the immature development state of the trigeminal ganglia (TG) and central nervous system (CNS) neurons, resulting in an inefficient suppression of viral replication. In 15-week-old pigs, viral replication was efficiently suppressed in the TG and CNS without induction of an extensive immune response. Furthermore, our results provide evidence that neurological disease could, at least in part, be related to viral replication and associated immunopathology in the olfactory bulb., (Copyright © 2017 American Society for Microbiology.)

The vesicular stomatitis virus (VSV) causes encephalitis in mice when inoculated intranasally. The deer mouse (Peromyscus maniculatus), a native New World rodent, is also susceptible to VSV infection and develops similar central nervous system (CNS) lesions to those observed in other rodent species. Chemokines, such as regulated on activation, normal T-cell expressed and secreted (RANTES; CCL-5) and monocyte chemoattractant protein (MCP)-1 (CCL-2), which are important for chemotaxis and activation of inflammatory cells, are expressed during the course of VSV encephalitis. However, the role of CNS resident cells in chemokine expression is poorly characterized. Here, we show that during vesicular stomatitis New Jersey virus (VSNJV) encephalitis in deer mice, RANTES and MCP-1 are expressed only in the olfactory bulb (OB), where the virus was localized. This chemokine expression was followed by the influx of inflammatory cells to the OB later in the course of acute disease. Neurons, astrocytes and microglia expressed RANTES, while MCP-1 was expressed by neurons and astrocytes. Although astrocytes and microglia responded to VSNJV infection by expressing chemokines, neurons were the cell type that was predominantly infected. Therefore, infected neurons may have a critical role in initiating an immune response in the OB. The signalling between neurons and other CNS resident cells is most likely the mechanism by which astrocytes and microglia are activated during the course of VSV encephalitis., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

In this autopsy-based study, human herpesvirus-6 (HHV-6) and -7 (HHV-7) genomic sequence frequency, HHV-6 variants, HHV-6 load and the expression of HHV-6 antigens in brain samples from the individuals, with and without unspecified encephalopathy (controls), using nested and real-time polymerase chain reactions, restriction endonuclease, and immunohistochemical analysis were examined. GraphPad Prism 6.0 Mann-Whitney nonparametric and chi-square test and Fisher's exact test were used for statistical analysis. The encephalopathy diagnoses were shown by magnetic resonance imaging made during their lifetime and macro- and microscopically studied autopsy tissue materials. Widespread HHV-6 and/or HHV-7 positivity was detected in the brain tissue of various individuals with encephalopathy, as well as in controls (51/57, 89.4 % and 35/51, 68.6 %, respectively; p = 0.009). Significantly higher detection frequency of single HHV-6 and concurrent HHV-6 + HHV-7 DNA was found in pia mater meninges, frontal lobe, temporal lobe, and olfactory tract DNAs in individuals with encephalopathy compared to the control group. HHV-6 load and higher frequency of the viral load >10 copies/10(6) cells significantly differed in samples from individuals with and without encephalopathy. The expression of HHV-6 antigens was revealed in different neural cell types with strong predominance in the encephalopathy group. In all HHV-6-positive autopsy samples of individuals with and without encephalopathy, HHV-6B was revealed. Significantly higher detection frequency of beta-herpesvirus DNA, more often detected HHV-6 load >10 copies/10(6) cells, as well as the expression of HHV-6 antigens in different brain tissue samples from individuals with encephalopathy in comparison with control group indicate on potential involvement of these viruses in encephalopathy development.

Herpes Simplex Virus type I (HSV-1) latently infects peripheral nervous system (PNS) sensory neurons, and its reactivation leads to recurring cold sores. The reactivated HSV-1 can travel retrograde from the PNS into the central nervous system (CNS) and is known to be causative of Herpes Simplex viral encephalitis. HSV-1 infection in the PNS is well documented, but little is known on the fate of HSV-1 once it enters the CNS. In the murine model, HSV-1 genome persists in the CNS once infected through an ocular route. To gain more details of HSV-1 infection in the CNS, we characterized HSV-1 infection of the tree shrew (Tupaia belangeri chinensis) brain following ocular inoculation. Here, we report that HSV-1 enters the tree shrew brain following ocular inoculation and HSV-1 transcripts, ICP0, ICP4, and LAT can be detected at 5 days post-infection (p.i.), peaking at 10 days p.i. After 2 weeks, ICP4 and ICP0 transcripts are reduced to a basal level, but the LAT intron region continues to be expressed. Live virus could be recovered from the olfactory bulb and brain stem tissue. Viral proteins could be detected using anti-HSV-1 antibodies and anti-ICP4 antibody, during the acute stage but not beyond. In situ hybridization could detect LAT during acute infection in most brain regions and in olfactory bulb and brain stem tissue well beyond the acute stage. Using a homogenate from these tissues' post-acute infection, we did not recover live HSV-1 virus, supporting a latent infection, but using a modified explant cocultivation technique, we were able to recover reactivated virus from these tissues, suggesting that the HSV-1 virus latently infects the tree shrew CNS. Compared to mouse, the CNS acute infection of the tree shrew is delayed and the olfactory bulb contains most latent virus. During the acute stage, a portion of the infected tree shrews exhibit symptoms similar to human viral encephalitis. These findings, together with the fact that tree shrews are closely related to primates, provided a valuable alternative model to study HSV-1 infection and pathogenesis in the CNS.

In 1935, the olfactory route was hypothesized to be a portal for virus entry into the central nervous system (CNS). This hypothesis was based on experiments in which nasophayngeal infection with poliovirus in monkeys was prevented from spreading to their CNS via transection of olfactory tracts between the olfactory neuroepithelium (ONE) of the nasal cavity and the olfactory bulb (OB). Since then, numerous neurotropic viruses have been observed to enter the CNS via retrograde transport along axons of olfactory sensory neurons whose cell bodies reside in the ONE. Importantly, this route of infection can occur even after subcutaneous inoculation of arboviruses that can cause encephalitis in humans. While the olfactory route is now accepted as an important pathway for viral entry into the CNS, it is unclear whether it provides a way for infection to spread to other brain regions. More recently, studies of antiviral innate and adaptive immune responses within the olfactory bulb suggest it provides early virologic control. Here we will review the data demonstrating that neurotropic viruses gain access to the CNS initially via the olfactory route with emphasis on findings that suggest the OB is a critical immunosensory effector organ that effectively clears virus.

Background: Although type I interferons (IFNs)-key effectors of antiviral innate immunity are known to be induced via different pattern recognition receptors (PRRs), the cellular source and the relative contribution of different PRRs in host protection against viral infection is often unclear. IPS-1 is a downstream adaptor for retinoid-inducible gene I (RIG-I)-like receptor signaling. In this study, we investigate the relative contribution of IPS-1 in the innate immune response in the different brain regions during infection with tick-borne encephalitis virus (TBEV), a flavivirus that causes a variety of severe symptoms like hemorrhagic fevers, encephalitis, and meningitis in the human host., Methods: IPS-1 knockout mice were infected with TBEV/Langat virus (LGTV), and viral burden in the peripheral and the central nervous systems, type I IFN induction, brain infiltrating cells, and inflammatory response was analyzed., Results: We show that IPS-1 is indispensable for controlling TBEV and LGTV infections in the peripheral and central nervous system. Our data indicate that IPS-1 regulates neuropathogenicity in mice. IFN response is differentially regulated in distinct regions of the central nervous system (CNS) influencing viral tropism, as LGTV replication was mainly restricted to olfactory bulb in wild-type (WT) mice. In contrast to the other brain regions, IFN upregulation in the olfactory bulb was dependent on IPS-1 signaling. IPS-1 regulates basal levels of antiviral interferon-stimulated genes (ISGs) like viperin and IRF-1 which contributes to the establishment of early viral replication which inhibits STAT1 activation. This diminishes the antiviral response even in the presence of high IFN-β levels. Consequently, the absence of IPS-1 causes uncontrolled virus replication, in turn resulting in apoptosis, activation of microglia and astrocytes, elevated proinflammatory response, and recruitment of inflammatory cells into the CNS., Conclusions: We show that LGTV replication is restricted to the olfactory bulb and that IPS-1 is a very important player in the olfactory bulb in shaping the innate immune response by inhibiting early viral replication and viral spread throughout the central nervous system. In the absence of IPS-1, higher viral replication leads to the evasion of antiviral response by inhibiting interferon signaling. Our data suggest that the local microenvironment of distinct brain regions is critical to determine virus permissiveness.

An increased incidence in the sleep-disorder narcolepsy has been associated with the 2009-2010 pandemic of H1N1 influenza virus in China and with mass vaccination campaigns against influenza during the pandemic in Finland and Sweden. Pathogenetic mechanisms of narcolepsy have so far mainly focused on autoimmunity. We here tested an alternative working hypothesis involving a direct role of influenza virus infection in the pathogenesis of narcolepsy in susceptible subjects. We show that infection with H1N1 influenza virus in mice that lack B and T cells (Recombinant activating gene 1-deficient mice) can lead to narcoleptic-like sleep-wake fragmentation and sleep structure alterations. Interestingly, the infection targeted brainstem and hypothalamic neurons, including orexin/hypocretin-producing neurons that regulate sleep-wake stability and are affected in narcolepsy. Because changes occurred in the absence of adaptive autoimmune responses, the findings show that brain infections with H1N1 virus have the potential to cause per se narcoleptic-like sleep disruption.

Equid herpesvirus type 1 (EHV-1) is an important pathogen of horses worldwide, associated with respiratory, reproductive and/or neurological disease. A mouse model for EHV-1 infection has been established but fails to reproduce some important aspects of the viral pathogenesis. Then, we investigated the susceptibility of rabbits to EHV-1 aiming at proposing this species as an alternative model for EHV-1 infection. Weanling rabbits inoculated intranasal with EHV-1 Kentucky D (10(7) TCID50/animal) shed virus in nasal secretions up to day 8-10 post-inoculation (pi), presented viremia up to day 14 pi and seroconverted to EHV-1 (virus neutralizing titers 4 to 64). Most rabbits (75%) developed respiratory disease, characterized by serous to hemorrhagic nasal discharge and mild to severe dyspnea. Some animals (20%) presented neurological signs as circling, bruxism and opisthotonus. Six animals died during acute disease (days 3-6); infectious virus and/or viral DNA were detected in the lungs, trigeminal ganglia (TG), olfactory bulbs (OBs) and cerebral cortex/brain (CC). Histological examination showed necrohemorrhagic, multifocal to coalescent bronchointerstitial pneumonia and diffuse alveolar edema. In two rabbits euthanized at day 50 pi, latent EHV-1 DNA was detected in the OBs. Dexamethasone administration at day 50 pi resulted in virus reactivation, demonstrated by virus shedding, viremia, clinical signs, and increase in VN titers and/or by detection of virus DNA in lungs, OBs, TGs and/or CC. These results demonstrate that rabbits are susceptible to EHV-1 infection and develop respiratory and neurological signs upon experimental inoculation. Thus, rabbits may be used to study selected aspects of EHV-1 biology and pathogenesis, extending and complementing the mouse model., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Viral neuroinvasion is a critical step in the pathogenesis of viral encephalitis. Multiple mechanisms of neuroinvasion have been identified, but their relative contribution to central nervous system (CNS) infection remains unclear for many viruses. In this study, we examined neuroinvasion of the mosquito-borne bunyavirus La Crosse (LACV), the leading cause of pediatric viral encephalitis in the USA. We found that the olfactory bulb (OB) and tract were the initial areas of CNS virus infection in mice. Removal of the OB reduced the incidence of LACV-induced disease demonstrating the importance of this area to neuroinvasion. However, we determined that infection of the OB was not due to axonal transport of virus from olfactory sensory neurons as ablation of these cells did not affect viral pathogenesis. Instead, we found that OB capillaries were compromised allowing leakage of virus-sized particles into the brain. Analysis of OB capillaries demonstrated specific alterations in cytoskeletal and Rho GTPase protein expression not observed in capillaries from other brain areas such as the cortex where leakage did not occur. Collectively, these findings indicate that LACV neuroinvasion occurs through hematogenous spread in specific brain regions where capillaries are prone to virus-induced activation such as the OB. Capillaries in these areas may be "hot spots" that are more susceptible to neuroinvasion not only for LACV, but other neurovirulent viruses as well.

Unlabelled: Previously we found that following intranasal (i.n.) infection with neurotropic vesicular stomatitis virus (VSV) type I interferon receptor (IFNAR) triggering of neuroectodermal cells was critically required to constrain intracerebral virus spread. To address whether locally active IFN-β was induced proximally, we studied spatiotemporal conditions of VSV-mediated IFN-β induction. To this end, we performed infection studies with IFN-β reporter mice. One day after intravenous (i.v.) VSV infection, luciferase induction was detected in lymph nodes. Upon i.n. infection, luciferase induction was discovered at similar sites with delayed kinetics, whereas on days 3 and 4 postinfection enhanced luciferase expression additionally was detected in the foreheads of reporter mice. A detailed analysis of cell type-specific IFN-β reporter mice revealed that within the olfactory bulb IFN-β was expressed by neuroectodermal cells, primarily by astrocytes and to a lesser extent by neurons. Importantly, locally induced type I IFN triggered distal parts of the brain as indicated by the analysis of ISRE-eGFP mice which after i.n. VSV infection showed enhanced green fluorescent protein (eGFP) expression throughout the brain. Compared to wild-type mice, IFN-β(-/-) mice showed increased mortality to i.n. VSV infection, whereas upon i.v. infection no such differences were detected highlighting the biological significance of intracerebrally expressed IFN-β. In conclusion, upon i.n. VSV instillation, IFN-β responses mounted by astrocytes within the olfactory bulb critically contribute to the antiviral defense by stimulating distal IFN-β-negative brain areas and thus arresting virus spread., Importance: The central nervous system has long been considered an immune privileged site. More recently, it became evident that specialized immune mechanisms are active within the brain to control pathogens. Previously, we showed that virus, which entered the brain via the olfactory route, was arrested within the olfactory bulb by a type I IFN-dependent mechanism. Since peripheral type I IFN would not readily cross the blood-brain barrier and within the brain thus far no abundant type I IFN responses have been detected, here we addressed from where locally active IFN originated from. We found that upon intranasal VSV instillation, primarily astrocytes, and to a lesser extent neurons, were stimulated within the olfactory bulb to mount IFN-β responses that also activated and protected distal brain areas. Our results are surprising because in other infection models astrocytes have not yet been identified as major type I IFN producers., (Copyright © 2015, American Society for Microbiology. All Rights Reserved.)

Central nervous system (CNS) disease is the most common extrarespiratory complication of influenza in humans. However, the pathogenesis, including the route of virus entry, is largely unknown. Here we present, for the first time, evidence of influenza virus entry into the CNS via the olfactory route in an immune-compromised infant. Since the nasal cavity is a primary site of influenza virus replication and is directly connected to the CNS via the olfactory nerve, these results imply that influenza virus invasion of the CNS may occur more often than previously believed., (© The Author 2014. Published by Oxford University Press on behalf of the Infectious Diseases Society of America. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.)

Unlabelled: Serious permanent neurological or psychiatric dysfunction may result from virus infections in the central nervous system (CNS). Olfactory sensory neurons are in direct contact with the external environment, making them susceptible to infection by viruses that can enter the brain via the olfactory nerve. The rarity of full brain viral infections raises the important question of whether unique immune defense mechanisms protect the brain. Here we show that both RNA (vesicular stomatitis virus [VSV]) and DNA (cytomegalovirus [CMV]) virus inoculations of the nasal mucosa leading to olfactory bulb (OB) infection activate long-distance signaling that upregulates antiviral interferon (IFN)-stimulated gene (ISG) expression in uninfected remote regions of the brain. This signaling mechanism is dependent on IFN-α/β receptors deep within the brain, leading to the activation of a distant antiviral state that prevents infection of the caudal brain. In normal mice, VSV replication is limited to the OB, and these animals typically survive the infection. In contrast, mice lacking the IFN-α/β receptor succumbed to the infection, with VSV spreading throughout the brain. Chemical destruction of the olfactory sensory neurons blocked both virus trafficking into the OB and the IFN response in the caudal brain, indicating a direct signaling within the brain after intranasal infection. Most signaling within the brain occurs across the 20-nm synaptic cleft. The unique long-distance IFN signaling described here occurs across many millimeters within the brain and is critical for survival and normal brain function., Importance: The olfactory mucosa can serve as a conduit for a number of viruses to enter the brain. Yet infections in the CNS rarely occur. The mechanism responsible for protecting the brain from viruses that successfully invade the OB, the first site of infection subsequent to infection of the nasal mucosa, remains elusive. Here we demonstrate that the protection is mediated by a long-distance interferon signaling, particularly IFN-β released by infected neurons in the OB. Strikingly, in the absence of neurotropic virus infection, ISGs are induced in the posterior regions of the brain, activating an antiviral state and preventing further virus invasion.

Porcine production is a primary market in the world economy. Controlling swine diseases in the farm is essential in order to achieve the sector necessities. Aujeszky's disease is a viral condition affecting pigs and is endemic in many countries of the world, causing important economic losses in the swine industry. microRNAs (miRNAs) are non-coding RNAs which modulates gene expression in animals, plants and viruses. With the aim of understanding miRNA roles during the Aujeszky's disease virus [ADV] (also known as suid herpesvirus type 1 [SuHV-1]) infection, the expression profiles of host and viral miRNAs were determined through deep sequencing in SuHV-1 infected porcine cell line (PK-15) and in an animal experimental SuHV-1 infection with virulent (NIA-3) and attenuated (Begonia) strains. In the in vivo approach miR-206, miR-133a, miR-133b and miR-378 presented differential expression between virus strains infection. In the in vitro approach, most miRNAs were down-regulated in infected groups. miR-92a and miR-92b-3p were up-regulated in Begonia infected samples. Functional analysis of all this over expressed miRNAs during the infection revealed their association in pathways related to viral infection processes and immune response. Furthermore, 8 viral miRNAs were detected by stem loop RT-qPCR in both in vitro and in vivo approaches, presenting a gene regulatory network affecting 59 viral genes. Most described viral miRNAs were related to Large Latency Transcript (LLT) and to viral transcription activators EP0 and IE180, and also to regulatory genes regarding their important roles in the host-pathogen interaction during viral infection.

Herpes simplex virus 1 (HSV-1) is a ubiquitous and important human pathogen. It is known to persist in trigeminal ganglia (TG), but how it reaches this site has been difficult to determine, as viral transmission is sporadic, pathogenesis is complicated, and early infection is largely asymptomatic. We used mice to compare the most likely natural HSV-1 host entry routes: oral and nasal. Intranasal infection was 100-fold more efficient than oral and targeted predominantly the olfactory neuroepithelium. Live imaging of HSV-1-expressed luciferase showed infection progressing from the nose to the TG and then reemerging in the facial skin. The brain remained largely luciferase negative throughout. Infected cell tagging by viral Cre recombinase expression in floxed reporter gene mice showed nasal virus routinely reaching the TG and only rarely reaching the olfactory bulbs. Thus, HSV-1 spread from the olfactory neuroepithelium to the TG and reemerged peripherally without causing significant neurological disease. This recapitulation of typical clinical infection suggests that HSV-1 might sometimes also enter humans via the respiratory tract.

Western equine encephalitis virus (WEEV; Alphavirus) is a mosquito-borne virus that can cause severe encephalitis in humans and equids. Previous studies have shown that intranasal infection of outbred CD-1 mice with the WEEV McMillan (McM) strain result in high mortality within 4 days of infection. Here in vivo and ex vivo bioluminescence (BLM) imaging was applied on mice intranasally infected with a recombinant McM virus expressing firefly luciferase (FLUC) to track viral neuroinvasion by FLUC detection and determine any correlation between BLM and viral titer. Immunological markers of disease (MCP-1 and IP-10) were measured and compared to wild type virus infection. Histopathology was guided by corresponding BLM images, and showed that neuroinvasion occurred primarily through cranial nerves, mainly in the olfactory tract. Olfactory bulb neurons were initially infected with subsequent spread of the infection into different regions of the brain. WEEV distribution was confirmed by immunohistochemistry as having marked neuronal infection but very few infected glial cells. Axons displayed infection patterns consistent with viral dissemination along the neuronal axis. The trigeminal nerve served as an additional route of neuroinvasion showing significant FLUC expression within the brainstem. The recombinant virus WEEV.McM.FLUC had attenuated replication kinetics and induced a weaker immunological response than WEEV.McM but produced comparable pathologies. Immunohistochemistry staining for FLUC and WEEV antigen showed that transgene expression was present in all areas of the CNS where virus was observed. BLM provides a quantifiable measure of alphaviral neural disease progression and a method for evaluating antiviral strategies.

Gliogenesis under pathophysiological conditions is of particular clinical relevance since it may provide regeneration-promoting cells recruitable for therapeutic purposes. There is accumulating evidence that aldynoglial cells with Schwann cell-like growth-promoting properties emerge in the lesioned CNS. However, the characterization of these cells and the signals triggering their in situ generation have remained enigmatic. In the present study, we used the p75 neurotrophin receptor (p75(NTR) ) as a marker for Schwann cells to study gliogenesis in the well-defined canine distemper virus (CDV)-induced demyelination model. White matter lesions of CDV-infected dogs contained bi- to multipolar, p75(NTR) -expressing cells that neither expressed MBP, GFAP, BS-1, or P0 identifying oligodendroglia, astrocytes, microglia, and myelinating Schwann cells nor CDV antigen. Interestingly, p75(NTR) -expression became apparent prior to the onset of demyelination in parallel to the expression of β-amyloid precursor protein (β-APP), nonphosphorylated neurofilament (n-NF), BS-1, and CD3, and peaked in subacute lesions with inflammation. To study the role of infiltrating immune cells during differentiation of Schwann cell-like glia, organotypic slice cultures from the normal olfactory bulb were established. Despite the absence of infiltrating lymphocytes and macrophages, a massive appearance of p75(NTR) -positive Schwann-like cells and BS-1-positive microglia was noticed at 10 days in vitro. It is concluded that axonal damage as an early signal triggers the differentiation of tissue-resident precursor cells into p75(NTR) -expressing aldynoglial Schwann cells that retain an immature pre-myelin state. Further studies have to address the role of microglia during this process and the regenerative potential of aldynoglial cells in CDV infection and other demyelinating diseases., (Copyright © 2011 Wiley Periodicals, Inc.)

Parkinson's disease, as well as many other parkinsonisms, including most toxic, neurodegenerative and familial types are typically asymmetric. No explanation for this phenomenon exists. A summary of the frequency of asymmetry in a spectrum of parkinsonian disorders is provided. Evidence against asymmetry being the result of normal asymmetries of the substantia nigrais reviewed. Asymmetry either results from a greater susceptibility on one side or a spreading pathology entering or starting on one side of the CNS. With the increasing evidence for spreading pathologies (toxins, viruses, α-synuclein), knowledge of neuroanatomical connections, and literature implicating spreading pathogens from the enteric and olfactory nerves, potential explanations can be theorized and explored, including the possibility of a pathogen preferentially entering or originating in the olfactory bulb on one side, with subsequent involvement of the other side., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

Herpesviruses are ubiquitous pathogens that cause much disease. The difficulty of clearing their established infections makes host entry an important target for control. However, while herpesviruses have been studied extensively in vitro, how they cross differentiated mucus-covered epithelia in vivo is unclear. To establish general principles we tracked host entry by Murid Herpesvirus-4 (MuHV-4), a lymphotropic rhadinovirus related to the Kaposi's Sarcoma-associated Herpesvirus. Spontaneously acquired virions targeted the olfactory neuroepithelium. Like many herpesviruses, MuHV-4 binds to heparan sulfate (HS), and virions unable to bind HS showed poor host entry. While the respiratory epithelium expressed only basolateral HS and was bound poorly by incoming virions, the neuroepithelium also displayed HS on its apical neuronal cilia and was bound strongly. Incoming virions tracked down the neuronal cilia, and either infected neurons or reached the underlying microvilli of the adjacent glial (sustentacular) cells and infected them. Thus the olfactory neuroepithelium provides an important and complex site of HS-dependent herpesvirus uptake.