Unconventional Use of LC3 by Coronaviruses through the Alleged Subversion of the ERAD Tuning Pathway

Pathogens of bacterial and viral origin hijack pathways operating in eukaryotic cells in many ways in order to gain access into the host, to establish themselves and to eventually produce their progeny. The detailed molecular characterization of the subversion mechanisms devised by pathogens to infect host cells is crucial to generate targets for therapeutic intervention. Here we review recent data indicating that coronaviruses probably co-opt membranous carriers derived from the endoplasmic reticulum, which contain proteins that regulate disposal of misfolded polypeptides, for their replication. In addition, we also present models describing potential mechanisms that coronaviruses could employ for this hijacking

Unconventional use of LC3 by coronaviruses through the alleged subversion of the ERAD tuning pathway

Pathogens of bacterial and viral origin hijack pathways operating in eukaryotic cells in many ways in order to gain access into the host, to establish themselves and to eventually produce their progeny. The detailed molecular characterization of the subversion mechanisms devised by pathogens to infect host cells is crucial to generate targets for therapeutic intervention. Here we review recent data indicating that coronaviruses probably co-opt membranous carriers derived from the endoplasmic reticulum, which contain proteins that regulate disposal of misfolded polypeptides, for their replication. In addition, we also present models describing potential mechanisms that coronaviruses could employ for this hijacking

SynGEM: An intranasal prefusion-like RSV F subunit vaccine

Respiratory syncytial virus (RSV) is an important cause of respiratory tract disease in (naive) young infants, older infants, the elderly and immune-compromised. Despite the medical need and the market potential, no licensed vaccine is available. Mucosis B.V. is a Dutch biotech company developing innovative mucosal vaccines, based on the Bacterium-Like Particles (BLP) technology. Such vaccines can be administered needle-free, e.g. through the nasal mucosa. In support of the mucosal approach, there is accumulating evidence that RSV F-specific local S-IgA antibodies secreted in the upper respiratory tract of humans correlate well with protection. Because of its ability to induce broadly neutralizing antibodies the RSV F protein is the most attractive antigen. The current view is that in particular serum antibodies directed against the prefusion form of RSV F belong to the most potent neutralizing antibodies and the ability to elicit these is a pivotal attribute for a successful RSV vaccine. We studied different variants of F with respect to their conformation using neutralizing monoclonal antibodies (mAbs), following the view that F proteins mimicking the meta-stable prefusion form of F expose a more extensive and relevant epitope repertoire than F proteins corresponding to the postfusion F structure. Both addition of a trimerization motif and mutation of the furin cleavage sites increased the reactivity of F with the prefusion-specific mAb D25, with the highest reactivity being observed for F proteins in which both these features were combined. This candidate antigen, called Flys-GCN, is suitable in the development of mucosal as well as intramuscular RSV vaccines. Here we describe the development of our intranasal candidate RSV vaccine, SynGEM, which is based on the validated BLP technology. The non-living BLPs allow for presentation of stable, trimeric prefusion-like RSV F proteins bound to the particle surface. Intranasal vaccination of naïve and convalescent mice with SynGEM induced long-lasting virus neutralizing RSV-specific serum IgG and robust levels of local IgA. Cotton rats immunized intranasally with SynGEM were protected upon RSV challenge, as represented by a low viral load in the lungs. Enhanced levels of pre- versus postfusion specific antibodies were observed in individual animals, confirming the ability of the SynGEM vaccine to induce potent neutralizing antibodies. A clinical Phase I study with intranasal SynGEM is planned for 2016

Low gH/gL (sub)species-specific antibody levels indicate elephants at risk of fatal elephant endotheliotropic herpesvirus hemorrhagic disease

DATA AVAILABILITY STATEMENT : The data supporting the findings of this study are available from the corresponding authors upon reasonable request.Elephant endotheliotropic herpesviruses (EEHVs), of which eleven (sub)species are currently distinguished, infect either Asian (Elephas maximus) or African elephants (Loxodonta species). While all adult elephants are latently infected with at least one EEHV (sub)species, young elephants, specifically those with low to non-detectable EEHV-specific antibody levels, may develop fatal hemorrhagic disease (EEHV-HD) upon infection. However, animals with high antibody levels against EEHV(1A) gB, an immunodominant antigen recognized by antibodies elicited against multiple (sub)species, may also occasionally succumb to EEHV-HD. To better define which animals are at risk of EEHV-HD, gB and gH/gL ELISAs were developed for each of the Asian elephant EEHV subspecies and assessed using 396 sera from 164 Asian elephants from European zoos. Antibody levels measured against gB of different (sub)species correlated strongly with one another, suggesting high cross-reactivity. Antibody levels against gH/gL of different subspecies were far less correlated and allowed differentiation between these (sub)species. Importantly, while high gB-specific antibody levels were detected in the sera of several EEHV-HD fatalities, all fatalities (n = 23) had low antibody levels against gH/gL of the subspecies causing disease. Overall, our data indicate that (sub)species-specific gH/gL ELISAs can be used to identify animals at risk of EEHV-HD when infected with a particular EEHV (sub)species.Named Fund Friends of VetMed.https://www.mdpi.com/journal/viruseshj2024Veterinary Tropical DiseasesSDG-03:Good heatlh and well-bein

EEHV1A glycoprotein B subunit vaccine elicits humoral and cell-mediated immune responses in mice

DATA AVAILABILITY : Data will be made available on request.Asian elephants are an endangered species facing many threats, including severe hemorrhagic disease (HD) caused by the elephant endotheliotropic herpesvirus (EEHV). EEHV-HD is the leading cause of death in captive juvenile Asian elephants in North America and Europe, and also affects elephants in their natural range countries. Significant challenges exist for successful treatment of EEHV-HD, which include timely recognition of disease onset and limited availability of highly effective treatment options. To address this problem, our goal is to prevent lethal disease in young elephants by developing a vaccine that elicits robust and durable humoral and cell-mediated immunity against EEHV. EEHV glycoprotein B (gB) is a major target for cellular and humoral immunity in elephants previously exposed to EEHV. Therefore, we generated a vaccine containing recombinant EEHV1A gB together with a liposome formulated TLR-4 and saponin combination adjuvant (SLA-LSQ). CD-1 mice that received one or two vaccinations with the vaccine elicited significant anti-gB antibody and polyfunctional CD4+ and CD8+ T cell responses, while no adverse effects of vaccination were observed. Overall, our findings demonstrate that an adjuvanted gB protein subunit vaccine stimulates robust humoral and cell-mediated immune responses and supports its potential use in elephants.The Cytometry and Cell Sorting Core at Baylor College of Medicine with funding from the CPRIT Core Facility Support Award, the NIH, the International Elephant Foundation (IEF) and Houston Zoo and funds acquired via Named Fund Friends of VetMed to the Utrecht University EEHV research group.http://www.elsevier.com/locate/vaccineam2023Veterinary Tropical Disease

Young elephants in a large herd maintain high levels of elephant endotheliotropic herpesvirus-specific antibodies and do not succumb to fatal haemorrhagic disease

Elephant endotheliotropic herpesviruses (EEHVs) have co-existed with elephants for millions of years, yet may cause fatal haemorrhagic disease (EEHV-HD), typically in elephants between 1 and 10 years of age. EEHV is omnipresent in (sub)adult elephants, and young elephants with low EEHV-specific antibody levels are at risk for EEHV-HD, suggesting that fatal disease may occur due to an insufficiently controlled primary infection. To further address this hypothesis, sera of three large elephant cohorts were subjected to a multiple EEHV species ELISA: (I) 96 Asian elephants between 0 and 57 years, including 13 EEHV-HD fatalities, from European zoo herds typically sized five to six elephants, (II) a herd of 64 orphaned elephants aged 0–15 years at the Elephant Transit Home in Sri Lanka and (III) 31 elephants aged 8–63 years, part of a large herd of 93 elephants at Pinnawala Elephant Orphanage, Sri Lanka. All Sri Lankan elephants showed high EEHV-specific antibody levels regardless of their age. While antibody levels of most European zoo elephants were comparable to those of Sri Lankan elephants, the average antibody level of the European juveniles (1–5 years of age) was significantly lower than those of age-matched Sri Lankan individuals. Moreover, the European juveniles showed a gradual decrease between 1 and 4 years of age, to be attributed to waning maternal antibodies. Maintenance of high levels of antibodies in spite of waning maternal antibodies in young Sri Lankan elephants is likely due to the larger herd size that increases the likelihood of contact with EEHV-shedding elephants. Together with the observation that low levels of EEHV-specific antibodies correlate with increased numbers of EEHV-HD fatalities, these results suggest that infection in presence of high maternal antibody levels may protect calves from developing EEHV-HD, while at the same time activating an immune response protective in future encounters with this virus.http://wileyonlinelibrary.com/journal/tbedhj2022Veterinary Tropical Disease

Associations of autozygosity with a broad range of human phenotypes

In many species, the offspring of related parents suffer reduced reproductive success, a phenomenon known as inbreeding depression. In humans, the importance of this effect has remained unclear, partly because reproduction between close relatives is both rare and frequently associated with confounding social factors. Here, using genomic inbreeding coefficients (FROH) for >1.4 million individuals, we show that FROH is significantly associated (p < 0.0005) with apparently deleterious changes in 32 out of 100 traits analysed. These changes are associated with runs of homozygosity (ROH), but not with common variant homozygosity, suggesting that genetic variants associated with inbreeding depression are predominantly rare. The effect on fertility is striking: FROH equivalent to the offspring of first cousins is associated with a 55% decrease [95% CI 44–66%] in the odds of having children. Finally, the effects of FROH are confirmed within full-sibling pairs, where the variation in FROH is independent of all environmental confounding

Studying the dynamics of coronavirus replicative structures.

Coronaviruses (CoVs) generate specialized membrane compartments, which consist of double membrane vesicles connected to convoluted membranes, the so-called replicative structures, where viral RNA synthesis takes place. These sites harbor the CoV replication-transcription complexes (RTCs): multi-protein complexes consisting of 16 nonstructural proteins (nsps), the CoV nucleocapsid protein (N) and presumably host proteins. To successfully establish functional membrane-bound RTCs all of the viral and host constituents need to be correctly spatiotemporally organized during viral infection. Few studies, however, have investigated the dynamic processes involved in the formation and functioning of the (subunits of) CoV RTCs and the replicative structures in living cells. In this chapter we describe several protocols to perform time-lapse imaging of CoV-infected cells and to study the kinetics of (subunits of) the CoV replicative structures. The approaches described are not limited to CoV-infected cells; they can also be applied to other virus-infected or non-infected cells

Studying the dynamics of coronavirus replicative structures.

Coronaviruses (CoVs) generate specialized membrane compartments, which consist of double membrane vesicles connected to convoluted membranes, the so-called replicative structures, where viral RNA synthesis takes place. These sites harbor the CoV replication-transcription complexes (RTCs): multi-protein complexes consisting of 16 nonstructural proteins (nsps), the CoV nucleocapsid protein (N) and presumably host proteins. To successfully establish functional membrane-bound RTCs all of the viral and host constituents need to be correctly spatiotemporally organized during viral infection. Few studies, however, have investigated the dynamic processes involved in the formation and functioning of the (subunits of) CoV RTCs and the replicative structures in living cells. In this chapter we describe several protocols to perform time-lapse imaging of CoV-infected cells and to study the kinetics of (subunits of) the CoV replicative structures. The approaches described are not limited to CoV-infected cells; they can also be applied to other virus-infected or non-infected cells