Biology subjects

Noval, M. G.

Publications and source records attributed to Noval, M. G..

12 recordsLinked to original sources

bioRxiv2026

GM-CSF and M-CSF Driven Differentiation Differentially Regulates Chikungunya Virus Infection and Antiviral Responses in Human Monocyte-Derived Macrophages

Chikungunya virus is an arthritogenic alphavirus causing debilitating joint pain in infected individuals. The mechanisms driving CHIKV-associated arthralgia is poorly understood, however, macrophages have been implicated as potential reservoirs of persistent viral material and mediators of immunopathology. Granulocyte Macrophage-Colony Stimulating Factor (GM-CSF) and Macrophage-Colony Stimulating Factor (M-CSF) are cytokines that serve as myeloid growth factors that bias macrophages toward pro-inflammatory and anti-inflammatory phenotypes, respectively. In this study, we examined how cytokine-driven macrophage differentiation via GM-CSF and M-CSF influences susceptibility to and responses against CHIKV infection in vitro. Using parallel donor-matched cultures of primary macrophages, we show that GM-CSF-differentiated macrophages are highly permissive to CHIKV and Mayaro virus (MAYV) infection and support robust viral replication, whereas M-CSF-differentiated macrophages are resistant to CHIKV replication and lack detectable levels of viral protein expression. Despite these differences, we observed pro-inflammatory, M1-skewing of CHIKV-infected macrophages, regardless of differentiation state. Interestingly, we observe higher production of IFN and IP10/CXCL10 in M-CSF differentiated macrophages, suggesting that M-CSF promotes an antiviral state that restricts CHIKV infection. Stimulation of macrophages with double-stranded RNA (polyinosinic:polycytidylic acid; poly(I:C)), but not with single-stranded RNA (resiquimod, R848), recapitulated the antiviral cytokine and chemokine response induced by CHIKV infection. These findings suggest that dsRNA sensing plays a more prominent role than ssRNA sensing in driving the macrophage antiviral response to CHIKV. Together, these findings highlight macrophage differentiation as a critical determinant of CHIKV susceptibility and antiviral immunity in humans, with implications for understanding inflammatory pathogenesis during infection.

microbiology↗

bioRxiv2024

Differential restriction of chikungunya virus in primary human cardiac endothelial cells occurs at multiple steps in the viral life cycle

Arthropod-borne viruses (arboviruses) constitute a significant ongoing public health threat, as the mechanisms of pathogenesis remain incompletely understood. Cardiovascular symptomatology is emerging as an important manifestation of arboviral infection. We have recently studied the cardiac tropism and mechanisms implicated in cardiac damage in mice for the alphavirus chikungunya virus (CHIKV), and we therefore sought to evaluate the cardiac tropism of other emerging alphaviruses and arboviruses. Using human primary cardiac cells, we found that arboviruses from diverse viral families were able to replicate within these cells. Interestingly, we noted that while the closely related alphavirus Mayaro virus (MAYV) could replicate to high titers in primary human cardiac microvascular endothelial cells, pulmonary, and brain endothelial cells, the Indian Ocean Lineage of CHIKV (CHIKV-IOL) was completely restricted in all endothelial cells tested. Upon further investigation, we discovered that this restriction occurs at both entry and egress stages. Additionally, we observed that compared to CHIKV, MAYV may antagonize or evade the innate immune response more efficiently in human cardiac endothelial cells to increase infection. Overall, this study explores the tropism of arboviruses in human primary cardiac cells and characterizes the strain-specific restriction of CHIKV-IOL in human endothelial cells. Further work is needed to understand how the differential restriction of alphaviruses in human endothelial cells impacts pathogenesis in a living model, as well as the specific host factors responsible. Author SummaryMosquito-borne viruses, such as those within the alphavirus genus, are an ongoing concern to human health globally. While we have recently begun to explore the mechanisms of CHIKV-induced pathology in the heart, little is known about other arboviruses and alphaviruses related to CHIKV. Here, we identified that multiple cardiac cell types are susceptible to infection by several arboviruses important to public health. Specifically, we noted differences in how two related viruses, CHIKV and MAYV, infect endothelial cells from multiple origins. This work highlights the potential for other emerging arboviruses, in addition to CHIKV, to directly infect cardiac tissue. Moreover, it emphasizes the ability for MAYV, an emerging virus that is less studied, to infect various endothelial cell types to high titers, suggesting the need for further research on MAYV pathogenesis. Finally, this research identifies differences in infection between individual strains of CHIKV, suggesting a finely tuned mechanism of restriction in endothelial cells that can be further explored, as well as the need to study use different viral strains for future alphavirus research.

microbiology↗

bioRxiv2023

The chikungunya virus E1 glycoprotein fusion loop and hinge alter glycoprotein dynamics leading to cell and host specific changes in infectivity.

Alphaviruses infect both mammals and insects, yet the distinct mechanisms that alphaviruses use to infect different hosts are not well defined. In this study, we characterize CHIKV E1 variants in the fusion loop (E1-M88L) and hinge region (E1-N20Y) in vitro and in vivo to understand how these regions of the E1 glycoprotein contribute to host-specific infection. Through cell culture assays, we found that CHIKV E1-N20Y enhanced infectivity in mosquito cells while the CHIKV E1-M88L variant enhanced virus binding and infectivity in both BHK-21 and C6/36 cells, and led to changes in the virus cholesterol-dependence in BHK-21 cells. Given these in vitro results and that residue E1-M88L is in a defined Mxra8 interacting domain, we hypothesized that this residue may be important for receptor usage. However, while the CHIKV E1-M88L variant increased replication in Mxra8-deficient mice compared to WT CHIKV, it was attenuated in vitro in mouse fibroblasts, suggesting that residue E1-M88 may function in a cell-type dependent manner to alter entry. Finally, using molecular dynamics to understand how potential changes in the E1 glycoprotein may impact the CHIKV glycoprotein E1-E2 complex, we found that E1-M88L and other E1 domain II variants lead to changes in both E1 and E2 dynamics. Taken together, these studies show that key residues in the CHIKV E1 fusion loop and hinge region function through changes in E1-E2 dynamics to facilitate cell- and host-dependent entry. ImportanceArthropod-borne viruses (arboviruses) are significant global public health threats, and their continued emergence around the world highlights the need to understand how these viruses replicate at the molecular level. The alphavirus class II glycoproteins are critical for virus entry in mosquitoes and mammals, yet how these proteins function is not completely understood. Therefore, to address these gaps in our knowledge, it is critical to dissect how distinct glycoprotein domains function in vitro and in vivo. Here, we show that changes in the CHIKV E1 fusion loop and hinge contribute to host-specific entry and E1-E2 dynamics, furthering our knowledge of how alphaviruses infect mammals and insects.

microbiology↗

bioRxiv2023

SARS-CoV-2 infection triggers pro-atherogenic inflammatory 1 responses in human coronary vessels

COVID-19 patients present higher risk for myocardial infarction (MI), acute coronary syndrome, and stroke for up to 1 year after SARS-CoV-2 infection. While the systemic inflammatory response to SARS-CoV-2 infection likely contributes to this increased cardiovascular risk, whether SARS-CoV-2 directly infects the coronary vasculature and attendant atherosclerotic plaques to locally promote inflammation remains unknown. Here, we report that SARS-CoV-2 viral RNA (vRNA) is detectable and replicates in coronary atherosclerotic lesions taken at autopsy from patients with severe COVID-19. SARS-CoV-2 localizes to plaque macrophages and shows a stronger tropism for arterial lesions compared to corresponding perivascular fat, correlating with the degree of macrophage infiltration. In vitro infection of human primary macrophages highlights that SARS-CoV-2 entry is increased in cholesterol-loaded macrophages (foam cells) and is dependent, in part, on neuropilin-1 (NRP-1). Furthermore, although viral replication is abortive, SARS-CoV-2 induces a robust inflammatory response that includes interleukins IL-6 and IL-1{beta}, key cytokines known to trigger ischemic cardiovascular events. SARS-CoV-2 infection of human atherosclerotic vascular explants recapitulates the immune response seen in cultured macrophages, including pro-atherogenic cytokine secretion. Collectively, our data establish that SARS-CoV-2 infects macrophages in coronary atherosclerotic lesions, resulting in plaque inflammation that may promote acute CV complications and long-term risk for CV events.

immunology↗

bioRxiv2022

Type I interferon and MAVS signaling restricts chikungunya virus heart infection and cardiac tissue damage

Chikungunya virus (CHIKV) infection has been associated with severe cardiac manifestations, yet, how CHIKV infection leads to heart disease remains unknown. Here, we leveraged both mouse models and human primary cells to define the mechanisms of CHIKV heart infection. We found that CHIKV actively replicates in cardiac fibroblasts and is cleared without significant tissue damage through the induction of a local type-I interferon response from both infected and non-infected cardiac cells. Importantly, signaling through the mitochondrial antiviral-signaling protein (MAVS) is required for viral clearance from the heart. In the absence of MAVS, persistent infection leads to focal myocarditis and major vessel vasculitis persisting for up to 60 days post-infection, suggesting CHIKV can lead to vascular inflammation and potential long-lasting cardiovascular complications. This study provides a model of CHIKV cardiac infection and mechanistic insight into CHIKV-induced heart disease, underscoring the importance of monitoring cardiac function in patients with CHIKV infections.

microbiology↗

bioRxiv2021

Emerging chikungunya virus variants at the E1-E1 inter-glycoprotein spike interface impact virus attachment and Inflammation

Chikungunya virus (CHIKV) is a re-emerging arthropod-borne alphavirus and a serious threat to human health. Therefore, efforts toward elucidating how this virus causes disease and the molecular mechanisms underlying steps of the viral replication cycle are crucial. Using an in vivo transmission system that allows intra-host evolution, we identified an emerging CHIKV variant carrying a mutation in the E1 glycoprotein (V156A) in the serum of mice and saliva of mosquitoes. E1 V156A has since emerged in humans during an outbreak in Brazil, co-occurring with a second mutation, E1 K211T, suggesting an important role for these residues in CHIKV biology. Given the emergence of these variants, we hypothesized that they function to promote CHIKV infectivity and subsequent disease. Here, we show that E1 V156A and E1 K211T modulate virus attachment and fusion and impact binding to heparin, a homolog of heparan sulfate, a key entry factor on host cells. These variants also exhibit differential neutralization by anti-glycoprotein monoclonal antibodies, suggesting structural impacts on the particle that may be responsible for altered interactions at the host membrane. Finally, E1 V156A and E1 K211T exhibit increased titers in an adult arthritic mouse model and induce increased foot-swelling at the site of injection. Taken together, this work has revealed new roles for E1 where discrete regions of the glycoprotein are able to modulate cell attachment and swelling within the host. IMPORTANCEAlphaviruses represent a growing threat to human health worldwide. Chikungunya virus (CHIKV) has rapidly spread to new geographic regions in the last several decades, causing overwhelming outbreaks of disease, yet there are no approved therapeutics. The CHIKV glycoproteins are key determinants of CHIKV adaptation and virulence. In this study, we characterize the naturally emerging E1 glycoprotein variants, V156A and K211T. We demonstrate that E1 V156A and K211T function in virus attachment to cells, a role that until now has been only attributed to the CHIKV E2 glycoprotein. We also demonstrate E1 V156A and K211T to increase foot-swelling in mice. Observing that these variants and other pathogenic variants occur at the E1-E1 inter-spike interface, we highlight this structurally important region as critical for multiple steps during CHIKV infection. Together, these studies further defines the function of E1 in CHIKV infection and can inform the development of therapeutic or preventative strategies.

microbiology↗

bioRxiv2021

Gut microbiome dysbiosis during COVID-19 is associated with increased risk for bacteremia and microbial translocation.

The microbial populations in the gut microbiome have recently been associated with COVID-19 disease severity. However, a causal impact of the gut microbiome on COVID-19 patient health has not been established. Here we provide evidence that gut microbiome dysbiosis is associated with translocation of bacteria into the blood during COVID-19, causing life-threatening secondary infections. Antibiotics and other treatments during COVID-19 can potentially confound microbiome associations. We therefore first demonstrate in a mouse model that SARS-CoV-2 infection can induce gut microbiome dysbiosis, which correlated with alterations to Paneth cells and goblet cells, and markers of barrier permeability. Comparison with stool samples collected from 96 COVID-19 patients at two different clinical sites also revealed substantial gut microbiome dysbiosis, paralleling our observations in the animal model. Specifically, we observed blooms of opportunistic pathogenic bacterial genera known to include antimicrobial-resistant species in hospitalized COVID-19 patients. Analysis of blood culture results testing for secondary microbial bloodstream infections with paired microbiome data obtained from these patients indicates that bacteria may translocate from the gut into the systemic circulation of COVID-19 patients. These results are consistent with a direct role for gut microbiome dysbiosis in enabling dangerous secondary infections during COVID-19.

microbiology↗

bioRxiv2021

Structurally conserved domains between flavivirus and alphavirus fusion glycoproteins contribute to replication in mammals and infectious virion production.

Alphaviruses and flaviviruses have class II fusion glycoproteins that are essential for virion assembly and infectivity. Importantly, the tip of domain II is structurally conserved between the alphavirus and flavivirus fusion proteins, yet whether these structural similarities between virus families translate to functional similarities is unclear. Using in vivo evolution of Zika virus (ZIKV), we identified several novel emerging variants including an envelope glycoprotein variant in {beta}-strand c (V114M) of domain II. We have previously shown that the analogous {beta}-strand c and the ij loop, located in the tip of domain II of the alphavirus E1 glycoprotein, are important for infectivity. This led us to hypothesize that flavivirus E {beta}-strand c also contributes to flavivirus infection. We generated this ZIKV glycoprotein variant and found that while it had little impact on infection in mosquitoes, it reduced replication in human cells and mice, and increased virus sensitivity to ammonium chloride, as seen for alphaviruses. In light of these results and given our alphavirus ij loop studies, we mutated a conserved alanine at the tip of the flavivirus ij loop to valine to test its effect on ZIKV infectivity. Interestingly, this mutation inhibited infectious virion production of ZIKV and yellow fever virus, but not West Nile virus. Together, these studies show that structurally analogous residues in the alphavirus and flavivirus class II fusion proteins contribute to virus infection in vivo and highlight these shared domains as targets for broad-spectrum arbovirus therapies. Author SummaryArboviruses are a significant global public health threat, yet there are no antivirals targeting these viruses. This problem is in part due to our lack of knowledge on the molecular mechanisms involved in the arbovirus life cycle. In particular, virus entry and assembly are essential processes in the virus life cycle and steps that can be targeted for the development of antiviral therapies. Therefore, understanding common, fundamental mechanisms used by different arboviruses for entry and assembly is essential. In this study, we show that flavivirus and alphavirus residues located in structurally conserved and analogous regions of the class II fusion proteins contribute to common mechanisms of entry, dissemination, and infectious virion production. These studies highlight how class II fusion proteins function and provide novel targets for development of antivirals.

microbiology↗

bioRxiv2021

Combination of a Sindbis-SARS-CoV-2 spike vaccine and αOX40 antibody elicits protective immunity against SARS-CoV-2 induced disease and potentiates long-term SARS-CoV-2-specific humoral and T-cell immunity

The COVID-19 pandemic caused by the coronavirus SARS-CoV-2 is a major global public threat. Currently, a worldwide effort has been mounted to generate billions of effective SARS-CoV-2 vaccine doses to immunize the worlds population at record speeds. However, there is still demand for alternative effective vaccines that rapidly confer long-term protection and rely upon cost-effective, easily scaled-up manufacturing. Here, we present a Sindbis alphavirus vector (SV), transiently expressing the SARS-CoV-2 spike protein (SV.Spike), combined with the OX40 immunostimulatory antibody (OX40) as a novel, highly effective vaccine approach. We show that SV.Spike plus OX40 elicits long-lasting neutralizing antibodies and a vigorous T-cell response in mice. Protein binding, immunohistochemical and cellular infection assays all show that vaccinated mice sera inhibits spike functions. Immunophenotyping, RNA Seq transcriptome profiles and metabolic analysis indicate a reprogramming of T-cells in vaccinated mice. Activated T-cells were found to mobilize to lung tissue. Most importantly, SV.Spike plus OX40 provided robust immune protection against infection with authentic coronavirus in transgenic mice expressing the human ACE2 receptor (hACE2-Tg). Finally, our immunization strategy induced strong effector memory response, potentiating protective immunity against re-exposure to SARS-CoV-2 spike protein. Our results show the potential of a new Sindbis virus-based vaccine platform to counteract waning immune response that can be used as a new candidate to combat SARS-CoV-2. Given the strong T-cell responses elicited, our vaccine is likely to be effective against variants that are proving challenging, as well as, serve as a platform to develop a broader spectrum pancoronavirus vaccine. Similarly, the vaccine approach is likely to be applicable to other pathogens.

immunology↗

bioRxiv2020

High titers of multiple antibody isotypes against the SARS-CoV-2 spike receptor-binding domain and nucleoprotein associate with better neutralization.

Understanding antibody responses to SARS-CoV-2 is indispensable for the development of containment measures to overcome the current COVID-19 pandemic. Here, we determine the ability of sera from 101 recovered healthcare workers to neutralize both authentic SARS-CoV-2 and SARS-CoV-2 pseudotyped virus and address their antibody titers against SARS-CoV-2 nucleoprotein and spike receptor-binding domain. Interestingly, the majority of individuals have low neutralization capacity and only 6% of the healthcare workers showed high neutralizing titers against both authentic SARS-CoV-2 virus and the pseudotyped virus. We found the antibody response to SARS-CoV-2 infection generates antigen-specific isotypes as well as a diverse combination of antibody isotypes, with high titers of IgG, IgM and IgA against both antigens correlating with neutralization capacity. Importantly, we found that neutralization correlated with antibody titers as quantified by ELISA. This suggests that an ELISA assay can be used to determine seroneutralization potential. Altogether, our work provides a snapshot of the SARS-CoV-2 neutralizing antibody response in recovered healthcare workers and provides evidence that possessing multiple antibody isotypes may play an important role in SARS-CoV-2 neutralization.

microbiology↗

bioRxiv2019

Mapping the evolutionary landscape of Zika virus infection in immunocompromised mice.

The fundamental basis of how arboviruses evolve in nature and what regulates the adaptive process remain unclear. To address this problem, we established a Zika virus (ZIKV) vector-borne transmission system in immunocompromised mice that mimics evolutionary characteristics of ZIKV infection in humans. Using this system, we defined factors that influence the evolutionary landscape of ZIKV infection and show that transmission route and specific organ microenvironments impact viral diversity and defective viral genome (DVG) production. In addition, we identified in mice the emergence of ZIKV mutants previously seen in natural infections, including variants present in currently circulating strains, as well as mutations unique to the mouse infections. With these studies, we have established an insect-to-mouse transmission model to study ZIKV evolution. We also defined how organ microenvironments and infection route impact the ZIKV evolutionary landscape, providing a deeper understanding of the factors that regulate arbovirus evolution and emergence.

microbiology↗

bioRxiv2019

Evolution-driven attenuation of alphaviruses highlights key glycoprotein determinants regulating viral infectivity and dissemination

Understanding the fundamental mechanisms of arbovirus transmission and pathogenesis is essential to develop new strategies for treatment and prevention. We previously took an in vivo evolution-based approach and identified the chikungunya virus E1 glycoprotein residue 80 to play a critical role in viral transmission and pathogenesis. In this study, we address the genetic robustness and function of position 80 and demonstrate that this highly conserved residue is a key determinant in alphavirus infectivity and dissemination through modulation of viral fusion and cholesterol dependence. In addition, in studying the evolution of position 80, we identified a network of glycoprotein residues, including epidemic determinants, that regulate virus dissemination and infectivity. These studies underscore the importance of taking evolution-based approaches to not only identify key viral determinants driving arbovirus transmission and pathogenesis but also to uncover fundamental aspects of arbovirus biology.

microbiology↗