Free Virus Research Articles

Viral involvement in microbial anaerobic methane oxidation-mediated arsenic mobilization in paddy soil

Anaerobic oxidation of methane (AOM) facilitates As mobilization, posing a significant environmental risk. Soil viruses potentially participate in the microbial AOM process, yet their roles in methane-mediated As mobilization of paddy soil remain elusive. Here, an anaerobic microcosm study was conducted by inoculating microbial suspension with extracellular free virus and mitomycin C (MC)-induced virus, along with 13CH4 injection. The results showed that extracellular free virus enhanced while MC-induced virus suppressed 13CH4-mediated As mobilization. During the AOM process, both viruses inhibited 13CH4 oxidation to 13CO2. However, the extracellular free virus suppressed whereas the MC-induced virus enhanced 13CH4 consumption, likely attributed to the viral influence on the ANME-2d abundance. The methane consumption differences were inferred to influence As reduction, as evidenced by a strong correlation between As(III) and 13CH4 consumption concentrations. Moreover, virus-mediated methane assimilation into microbial biomass carbon influenced the overall microbial population. An increased abundance of Geobacter in the extracellular free virus treatment elevated net As(III) concentrations (up to 260%) relative to treatment without virus in the presence of 13CH4. In contrast, MC-induced virus led to a net 122% reduction in As(III) concentration due to decreased Geobacter abundance. These findings provide new insights into soil viruses in microbial AOM-driven As mobilization, highlighting their crucial functions in soil ecosystems.

Coastal bacteria and protists assimilate viral carbon and nitrogen.

Free viruses are the most abundant type of biological particles in the biosphere, but the lack of quantitative knowledge about their consumption by heterotrophic protists and bacterial degradation has hindered the inclusion of virovory in biogeochemical models. Using isotope-labeled viruses added to three independent microcosm experiments with natural microbial communities followed by isotope measurements with single-cell resolution and flow cytometry, we quantified the flux of viral C and N into virovorous protists and bacteria and compared the loss of viruses due to abiotic vs biotic factors. We found that some protists can obtain most of their C and N requirements from viral particles and that viral C and N get incorporated into bacterial biomass. We found that bacteria and protists were responsible for increasing the daily removal rate of viruses by 33% to 85%, respectively, compared to abiotic processes alone. Our laboratory incubation experiments showed that abiotic processes removed roughly 50% of the viruses within a week, and adding biotic processes led to a removal of 83% to 91%. Our data provide direct evidence for the transfer of viral C and N back into the microbial loop through protist grazing and bacterial breakdown, representing a globally significant flux that needs to be investigated further to better understand and predictably model the C and N cycles of the hydrosphere.

Targeted recruitment of immune effector cells for rapid eradication of influenza virus infections

Despite much research, considerable data suggest that influenza virus remains a serious health problem because i) the effectiveness of current vaccines ranges only from 19% to 60%, ii) available therapies remain ineffective in advanced stages of disease, iii) death rates vary between 25,000 and 72,000/year in the United States, and iv) avian influenza strains are now being transmitted to dairy cattle that in turn are infecting humans. To address these concerns, we have developed zanDR, a bispecific small molecule that binds and inhibits viral neuraminidase expressed on both free virus and virus-infected cells and recruits naturally occurring anti-rhamnose and anti-dinitrophenyl (DNP) antibodies with rhamnose and DNP haptens. Because the neuraminidase inhibition replicates the chemotherapeutic mechanism of zanamivir and oseltamivir, while rhamnose and DNP recruit endogenous antibodies much like an anti-influenza vaccine, zanDR reproduces most of the functions of current methods of protection against influenza virus infections. Importantly, studies on cells in culture demonstrate that both of the above protective mechanisms remain highly functional in the zanDR conjugate, while studies in lethally infected mice with advanced-stage disease establish that a single intranasal dose of zanDR not only yields 100% protection but also reduces lung viral loads faster and ~1,000× more thoroughly than current antiviral therapies. Since zanDR also lowers secretion of proinflammatory cytokines and protects against virus-induced damage to the lungs better than current therapies, we suggest that combining an immunotherapy with a chemotherapy in single pharmacological agent constitutes a promising approach for treating the more challenging forms of influenza.

Nectin1 is a pivotal host factor involved in attachment and entry of red-spotted grouper nervous necrosis virus in the early stages of the viral life cycle.

Nervous necrosis virus (NNV) is a highly neurotropic virus that poses a persistent threat to the survival of multiple fish species. However, its inimitable neuropathogenesis remains largely elusive. To rummage potential partners germane to the nervous system, we investigated the interaction between red-spotted grouper NNV (RGNNV) and grouper brain by immunoprecipitation coupled with mass spectrometry and discerned Nectin1 as a novel host factor subtly involved in viral early invasion events. Nectin1 was abundant in neural tissues and implicated in the inception of tunnel nanotubes triggered by RGNNV. Its overexpression not only dramatically potentiated the replication dynamics of RGNNV in susceptible cells, but also empowered non-sensitive cells to expeditiously capture free virions within 2 min. This potency was impervious to low temperatures but was dose-dependently suppressed by soluble protein or specific antibody of Nectin1 ectodomain, indicating Nectin1 as an attachment receptor for RGNNV. Mechanistically, efficient hijacking of virions by Nectin1 strictly depended on intricate linkages to different modules of viral capsid protein, especially the direct binding between the IgC1 loop and P-domain. More strikingly, despite abortive proliferation in Nectin1-reconstructed CHSE-214 cells, a non-sensitive cell, RGNNV could gain access to the intracellular compartment by capitalizing on Nectin1, thereby inducing canonical cytoplasmic vacuolation. Altogether, our findings delineate a candidate entrance for RGNNV infiltration into the nervous system, which may shed unprecedented insights into the exploration and elucidation of RGNNV pathogenesis.IMPORTANCENervous necrosis virus (NNV) is one of the most virulent pathogens in the aquaculture industry, which inflicts catastrophic damage to ecology, environment, and economy annually around the world. Nevertheless, its idiosyncratic invasion and latency mechanisms pose enormous hardships to epidemic prevention and control. In this study, deploying grouper brain as a natural screening library, a single-transmembrane glycoprotein, Nectin1, was first identified as an emergent functional receptor for red-spotted grouper NNV (RGNNV) that widely allocated in nervous tissues and directly interacted with viral capsid protein through distinct Ig-like loops to bridge virus-host crosstalk, apprehend free virions, and concomitantly propel viral entry. Our findings illuminate the critical role of Nectin1 in RGNNV attachment and entry and provide a potential target for future clinical intervention strategies in the therapeutic race against RGNNV.

Modeling virus-stimulated proliferation of CD4[formula omitted] T-cell, cell-to-cell transmission and viral loss in HIV infection dynamics

Human immunodeficiency virus (HIV) can persist in infected individuals despite prolonged antiretroviral therapy and it may spread through two modes: virus-to-cell and cell-to-cell transmissions. Understanding viral infection dynamics is pivotal for elucidating HIV pathogenesis. In this study, we incorporate the loss term of virions, and both virus-to-cell and cell-to-cell infection modes into a within-host HIV model, which also takes into consideration the proliferation of healthy target cells stimulated by free viruses. By constructing suitable Lyapunov function and applying geometric methods, we establish global stability results of the infection free equilibrium and the infection persistent equilibrium, respectively. Our findings highlight the crucial role of the basic reproduction number in the threshold dynamics. Moreover, we use the loss rate of virions as the bifurcation parameter to investigate stability switches of the positive equilibrium, local Hopf bifurcation, and its global continuation. Numerical simulations validate our theoretical results, revealing rich viral dynamics including backward bifurcation, saddle–node bifurcation, and bistability phenomenon in the sense that the infection free equilibrium and a limit cycle are both locally asymptotically stable. These insights contribute to a deeper understanding of HIV dynamics and inform the development of effective therapeutic strategies.

A Review of the Therapeutical Perspective of Phytochemicals in the Treatment of HIV

Abstract: HIV/AIDS is one of the most widespread global diseases. It is caused by the human immunodeficiency virus (HIV), which leads to the development of acquired immunodeficiency syndrome (AIDS). In AIDS patients, the immune system progressively weakens, making them vulnerable to various infections and cancers. The virus can be transmitted through the exchange of bodily fluids such as blood, semen, vaginal fluid, and breast milk, where HIV can be found in both free virus particles and infected immune cells. Although drug resistance remains a challenge for HIV/AIDS treatment, researchers are exploring the use of natural substances in the development of anti-HIV drugs. Clinical trials based on plant-derived compounds are currently being conducted. Experimental research suggests that plants may have the potential to treat HIV. However, the identification of active compounds, testing of extracts, and creation of suitable bioassays are all complex processes. Natural product scientists face the challenge of separating these complicated extracts into their active components. Various phytochemicals found in Astragalus membranaceus Bunge, Chelidonium majus L, Combretum molle (R. Br. ex. G. Don.) Engl& Diels, Dittrichiaviscosa (L.) Greuter, Helichrysum populifolium, Hoodia gordonii (Masson) Sweet ex Decne, and Hypericumperf, have been investigated pharmacologically and pharmacokinetically. Further research is needed to optimize formulations and production in accordance with preclinical safety and efficacy studies.

Dynamics of Activation and Regulation of the Immune Response to Attack by Viral Pathogens Using Mathematical Modeling

In this paper, a mathematical model is developed to simulate the activation of regulatory T lymphocytes dynamics. The model considers the adaptive immune response and consists of epithelial cells, infected cells, free virus particles, helper and cytotoxic T lymphocytes, B lymphocytes, and regulatory T lymphocytes. A mathematical analysis was carried out to discuss the conditions of existence and stability of equilibrium solutions in terms of the basic reproductive number. In addition, the definitions and properties necessary to preserve the positivity and stability of the model are shown. The precision of these mathematical models can be affected by numerous sources of uncertainty, partly due to the balance between the complexity of the model and its predictive capacity to depict the biological process accurately. Nevertheless, these models can provide remarkably perspectives on the dynamics of infection and assist in identification specific immunological traits that improve our comprehension of immune mechanisms. The theoretical results are validated by numerical simulations using data reported in the literature. The construction, analysis, and simulation of the developed models demonstrate that the increased induced regulatory T lymphocytes effectively suppress the inflammatory response in contrast to similar cells at lower contents, playing a key role in maintaining self-tolerance and immune homeostasis.

Accounting for cellular-level variation in lysis: implications for virus-host dynamics.

Viral impacts on microbial populations depend on interaction phenotypes-including viral traits spanning the adsorption rate, latent period, and burst size. The latent period is a key viral trait in lytic infections. Defined as the time from viral adsorption to viral progeny release, the latent period of bacteriophage is conventionally inferred via one-step growth curves in which the accumulation of free virus is measured over time in a population of infected cells. Developed more than 80 years ago, one-step growth curves do not account for cellular-level variability in the timing of lysis, potentially biasing inference of viral traits. Here, we use nonlinear dynamical models to understand how individual-level variation of the latent period impacts virus-host dynamics. Our modeling approach shows that inference of the latent period via one-step growth curves is systematically biased-generating estimates of shorter latent periods than the underlying population-level mean. The bias arises because variability in lysis timing at the cellular level leads to a fraction of early burst events, which are interpreted, artefactually, as an earlier mean time of viral release. We develop a computational framework to estimate latent period variability from joint measurements of host and free virus populations. Our computational framework recovers both the mean and variance of the latent period within simulated infections including realistic measurement noise. This work suggests that reframing the latent period as a distribution to account for variability in the population will improve the study of viral traits and their role in shaping microbial populations.IMPORTANCEQuantifying viral traits-including the adsorption rate, burst size, and latent period-is critical to characterize viral infection dynamics and develop predictive models of viral impacts across scales from cells to ecosystems. Here, we revisit the gold standard of viral trait estimation-the one-step growth curve-to assess the extent to which assumptions at the core of viral infection dynamics lead to ongoing and systematic biases in inferences of viral traits. We show that latent period estimates obtained via one-step growth curves systematically underestimate the mean latent period and, in turn, overestimate the rate of viral killing at population scales. By explicitly incorporating trait variability into a dynamical inference framework that leverages both virus and host time series, we provide a practical route to improve estimates of the mean and variance of viral traits across diverse virus-microbe systems.

Genomic and induction evidence for bacteriophage contributions to sargassum-bacteria symbioses

BackgroundSymbioses between primary producers and bacteria are crucial for nutrient exchange that fosters host growth and niche adaptation. Yet, how viruses that infect bacteria (phages) influence these bacteria-eukaryote interactions is still largely unknown. Here, we investigate the role of viruses on the genomic diversity and functional adaptations of bacteria associated with pelagic sargassum. This brown alga has dramatically increased its distribution range in the Atlantic in the past decade and is predicted to continue expanding, imposing severe impacts on coastal ecosystems, economies, and human health.ResultsWe reconstructed 73 bacterial and 3963 viral metagenome-assembled genomes (bMAGs and vMAGs, respectively) from coastal Sargassum natans VIII and surrounding seawater. S. natans VIII bMAGs were enriched in prophages compared to seawater (28% and 0.02%, respectively). Rhodobacterales and Synechococcus bMAGs, abundant members of the S. natans VIII microbiome, were shared between the algae and seawater but were associated with distinct phages in each environment. Genes related to biofilm formation and quorum sensing were enriched in S. natans VIII phages, indicating their potential to influence algal association in their bacterial hosts. In-vitro assays with a bacterial community harvested from sargassum surface biofilms and depleted of free viruses demonstrated that these bacteria are protected from lytic infection by seawater viruses but contain intact and inducible prophages. These bacteria form thicker biofilms when growing on sargassum-supplemented seawater compared to seawater controls, and phage induction using mitomycin C was associated with a significant decrease in biofilm formation. The induced metagenomes were enriched in genomic sequences classified as temperate viruses compared to uninduced controls.ConclusionsOur data shows that prophages contribute to the flexible genomes of S. natans VIII-associated bacteria. These prophages encode genes with symbiotic functions, and their induction decreases biofilm formation, an essential capacity for flexible symbioses between bacteria and the alga. These results indicate that prophage acquisition and induction contribute to genomic and functional diversification during sargassum-bacteria symbioses, with potential implications for algae growth.1vVC6YGWLCm_DZc419o4RCVideo

An optimal solution for tumor growth model using generalized Bessel polynomials

In this paper, a mathematical model is given that depicts the interactions between cancer cells and viruses in the setting of oncolytic virotherapy. The model is separated into three classes, namely, concentrations of uninfected tumor cells in the population “ ”, free virus “ ”, and cancerous cells infected “ ”. Applying Caputo fractional derivative, the model is fractionalized, and using generalized Bessel polynomials, an optimal problem is solved utilizing Lagrange multipliers method. The results show that the presented method has high accuracy and is suitable for solving the nonlinear systems based on partial differential equations especially tumors models.

Highly restrictive and directional penetration of the blood cerebral spinal fluid barrier by JCPyV.

The human polyomavirus JCPyV is an opportunistic pathogen that infects greater than 60% of the world's population. The virus establishes a persistent and asymptomatic infection in the urogenital system but can cause a fatal demyelinating disease in immunosuppressed or immunomodulated patients following invasion of the CNS. The mechanisms responsible for JCPyV invasion into CNS tissues are not known but direct invasion from the blood to the cerebral spinal fluid via the choroid plexus has been hypothesized. To study the potential of the choroid plexus as a site of neuroinvasion, we used an adult human choroid plexus epithelial cell line to model the blood-cerebrospinal fluid (B-CSF) barrier in a transwell system. We found that these cells formed a highly restrictive barrier to virus penetration either as free virus or as virus associated with extracellular vesicles (EVJC+). The restriction was not absolute and small amounts of virus or EVJC+ penetrated and were able to establish foci of infection in primary astrocytes. Disruption of the barrier with capsaicin did not increase virus or EVJC+ penetration leading us to hypothesize that virus and EVJC+ were highly cell-associated and crossed the barrier by an active process. An inhibitor of macropinocytosis increased virus penetration from the basolateral (blood side) to the apical side (CSF side). In contrast, inhibitors of clathrin and raft dependent transcytosis reduced virus transport from the basolateral to the apical side of the barrier. None of the drugs inhibited apical to basolateral transport suggesting directionality. Pretreatment with cyclosporin A, an inhibitor of P-gp, MRP2 and BCRP multidrug resistance transporters, restored viral penetration in cells treated with raft and clathrin dependent transcytosis inhibitors. Because choroid plexus epithelial cells are known to be susceptible to JCPyV infection both in vitro and in vivo we also examined the release of infectious virus from the barrier. We found that virus was preferentially released from the cells into the apical (CSF) chamber. These data show clearly that there are two mechanisms of penetration, direct transcytosis which is capable of seeding the CSF with small amounts of virus, and infection followed by directional release of infectious virions into the CSF compartment.

Spatial information allows inference of the prevalence of direct cell-to-cell viral infection.

The role of direct cell-to-cell spread in viral infections-where virions spread between host and susceptible cells without needing to be secreted into the extracellular environment-has come to be understood as essential to the dynamics of medically significant viruses like hepatitis C and influenza. Recent work in both the experimental and mathematical modelling literature has attempted to quantify the prevalence of cell-to-cell infection compared to the conventional free virus route using a variety of methods and experimental data. However, estimates are subject to significant uncertainty and moreover rely on data collected by inhibiting one mode of infection by either chemical or physical factors, which may influence the other mode of infection to an extent which is difficult to quantify. In this work, we conduct a simulation-estimation study to probe the practical identifiability of the proportion of cell-to-cell infection, using two standard mathematical models and synthetic data that would likely be realistic to obtain in the laboratory. We show that this quantity cannot be estimated using non-spatial data alone, and that the collection of data which describes the spatial structure of the infection is necessary to infer the proportion of cell-to-cell infection. Our results provide guidance for the design of relevant experiments and mathematical tools for accurately inferring the prevalence of cell-to-cell infection in in vitro and in vivo contexts.

Herpes simplex virus spreads rapidly in human foreskin, partly driven by chemokine-induced redistribution of Nectin-1 on keratinocytes.

HSV infects keratinocytes in the epidermis of skin via nectin-1. We established a human foreskin explant infection model to investigate HSV entry and spread. HSV1 entry could only be achieved by the topical application of virus via high density microarray projections (HD-MAPs) to the epidermis, which penetrated beyond one third of its thickness, simulating in vivo microtrauma. Rapid lateral spread of HSV1 to a mean of 13 keratinocytes wide occurred after 24 hours and free virus particles were observed between keratinocytes, consistent with an intercellular route of spread. Nectin-1 staining was markedly decreased in foci of infection in the epidermis and in the human keratinocyte HaCaT cell line. Nectin-1 was redistributed, at the protein level, in adjacent uninfected cells surrounding infection, inducible by CCL3, IL-8 (or CXCL8), and possibly CXCL10 and IL-6, thus facilitating spread. These findings provide the first insights into HSV1 entry and spread in human inner foreskin in situ.

Enterovirus A71 crosses a human blood-brain barrier model through infected immune cells.

Enterovirus A71 (EV-A71) is associated with neurological conditions such as acute meningitis and encephalitis. The virus is detected in the bloodstream, and high blood viral loads are associated with central nervous system (CNS) manifestations. We used an in vitro blood-brain barrier (BBB) model made up of human brain-like endothelial cells (hBLECs) and brain pericytes grown in transwell systems to investigate whether three genetically distinct EV-A71 strains (subgenogroups C1, C1-like, and C4) can cross the human BBB. EV-A71 poorly replicated in hBLECs, which released moderate amounts of infectious viruses from their luminal side and trace amounts of infectious viruses from their basolateral side. The barrier properties of hBLECs were not impaired by EV-A71 infection. We investigated the passage through hBLECs of EV-A71-infected white blood cells. EV-A71 strains efficiently replicated in immune cells, including monocytes, neutrophils, and NK/T cells. Attachment to hBLECs of immune cells infected with the C1-like virus was higher than attachment of cells infected with C1-06. EV-A71 infection did not impair the transmigration of immune cells through hBLECs. Overall, EV-A71 targets different white blood cell populations that have the potential to be used as a Trojan horse to cross hBLECs more efficiently than cell-free EV-A71 particles.IMPORTANCEEnterovirus A71 (EV-A71) was first reported in the USA, and numerous outbreaks have since occurred in Asia and Europe. EV-A71 re-emerged as a new multirecombinant strain in 2015 in Europe and is now widespread. The virus causes hand-foot-and-mouth disease in young children and is involved in nervous system infections. How the virus spreads to the nervous system is unclear. We investigated whether white blood cells could be infected by EV-A71 and transmit it across human endothelial cells mimicking the blood-brain barrier protecting the brain from adverse effects. We found that endothelial cells provide a strong roadblock to prevent the passage of free virus particles but allow the migration of infected immune cells, including monocytes, neutrophils, and NK/T cells. Our data are consistent with the potential role of immune cells in the pathogenesis of EV-A71 infections by spreading the virus in the blood and across the human blood-brain barrier.

Widespread occurrence and diverse origins of polintoviruses influence lineage-specific genome dynamics in stony corals.

Stony corals (Order: Scleractinia) are central to vital marine habitats known as coral reefs. Numerous stressors in the Anthropocene are contributing to the ongoing decline in coral reef health and coverage. While viruses are established modulators of marine microbial dynamics, their interactions within the coral holobiont and impact on coral health and physiology remain unclear. To address this key knowledge gap, we investigated diverse stony coral genomes for 'endogenous' viruses. Our study uncovered a remarkable number of integrated viral elements recognized as 'Polintoviruses' (Class Polintoviricetes) in thirty Scleractinia genomes; with several species harboring hundreds to thousands of polintoviruses. We reveal massive paralogous expansion of polintoviruses in stony coral genomes, alongside the presence of integrated elements closely related to Polinton-like viruses (PLVs), a group of viruses that exist as free virions. These results suggest multiple integrations of polintoviruses and PLV-relatives, along with paralogous expansions, shaped stony coral genomes. Re-analysis of existing gene expression data reveals all polintovirus structural and non-structural hallmark genes are expressed, providing support for free virion production from polintoviruses. Our results, revealing a significant diversity of polintovirus across the Scleractinia order, open a new research avenue into polintovirus and their possible roles in disease, genomic plasticity, and environmental adaptation in this key group of organisms.

Dynamical analysis of a general delayed HBV infection model with capsids and adaptive immune response in presence of exposed infected hepatocytes.

The aim of this paper is to develop and investigate a novel mathematical model of the dynamical behaviors of chronic hepatitis B virus infection. The model includes exposed infected hepatocytes, intracellular HBV DNA-containing capsids, uses a general incidence function for viral infection covering a variety of special cases available in the literature, and describes the interaction of cytotoxic T lymphocytes that kill the infected hepatocytes and the magnitude of B-cells that send antibody immune defense to neutralize free virions. Further, one time delay is incorporated to account for actual capsids production. The other time delays are used to account for maturation of capsids and free viruses. We start with the analysis of the proposed model by establishing the local and global existence, uniqueness, non-negativity and boundedness of solutions. After defined the threshold parameters, we discuss the stability properties of all possible steady state constants by using the crafty Lyapunov functionals, the LaSalle's invariance principle and linearization methods. The impacts of the three time delays on the HBV infection transmission are discussed through local and global sensitivity analysis of the basic reproduction number and of the classes of infected states. Finally, an application is provided and numerical simulations are performed to illustrate and interpret the theoretical results obtained. It is suggested that, a good strategy to eradicate or to control HBV infection within a host should concentrate on any drugs that may prolong the values of the three delays.

Co-infection dynamics between HIV-HTLV-I disease with the effects of Cytotoxic T-lymphocytes, saturated incidence rate and study of optimal control

The spreading of HIV or HTLV-I among the cells has received the great attention in recent modelling study to explore the virus infection dynamics. The co-infection of HIV and HTLV-I with the effect of Cytotoxic T-lymphocytes (CTLs) immune response is also important from epidemiological point of view. To identify the co-infection scenario of HIV and HTLV-I with the CTLs effect we proposed in this paper a six compartmental ODE-model with uninfected, HIV-infected, HTLV-I infected CD4+T cells and free HIV virus particles with HIV specific CTLs and HTLV-I specific CTLs. The rates of infection of the cases are considered here saturated type and proliferation rate of uninfected and HIV infected CD4+ T-cells are of logistic terms. To establish the well-posedness of the model we have shown that the solution of the proposed model is non-negative and bounded. We obtain the basic reproduction number which is the maximum of the HIV-related reproduction and the HTLV-I related reproduction number. Along with the disease free equilibrium point the system contains other seven endemic equilibrium points containing infection by single disease or both. Analytically, we establish the local and global stability conditions of the equilibrium points and also we establish that the system experiences transcritical bifurcation by the generation of only HIV or HTLV-I infected endemic equilibrium point. Using numerical simulations, we validate the theoretical results and found two infection paths, one initiating with HIV and other with HTLV-I, both cases ultimately become co-infected. Finally, using the optimal control analysis we found the optimal policy for treatment using AVR, RTI & PI for HIV or AZT for HTLV-I control and lastly concluded by some recommendations.

Delay induced stability switch in a mathematical model of CD8 T-cell response to SARS-CoV-2 mediated by receptor ACE2.

The pathogen SARS-CoV-2 binds to the receptor angiotensin-converting enzyme 2 (ACE2) of the target cells and then replicates itself through the host, eventually releasing free virus particles. After infection, the CD8 T-cell response is triggered and appears to play a critical role in the defense against virus infections. Infected cells and their activated CD8 T-cells can cause tissue damage. Here, we established a mathematical model of within-host SARS-CoV-2 infection that incorporates the receptor ACE2, the CD8 T-cell response, and the damaged tissues. According to this model, we can get the basic reproduction number R0 and the immune reproduction number R1. We provide the theoretical proof for the stability of the disease-free equilibrium, immune-inactivated equilibrium, and immune-activated equilibrium. Finally, our numerical simulations show that the time delay in CD8 T-cell production can induce complex dynamics such as stability switching. These results provide insights into the mechanisms of SARS-CoV-2 infection and may help in the development of effective drugs against COVID-19.

Abstract 6331: A novel automated platform for rapid manufacturing of CAR T cells

Abstract Autologous cell therapy using genetically modified T cells to express chimeric antigen receptor (CAR) has yielded durable responses in cancers patients. However, currently only a small portion of patients are privileged to receive the treatment. High cost and long manufacturing time contribute to the key limitations prohibiting cell therapy broad adoption. Researchers discovered that reducing ex vivo culture improves anti-tumor activity of CAR T cells1 and recently demonstrated the process of generating functional CAR T cells within 24 hours2. This rapid process provides great promise in cost reduction of CAR T therapy and opportunity to have cells manufactured in close-to-patient settings. CART cell manufacturing process involves: (1) collection of peripheral blood; (2) isolation of T cells; (3) performing transduction via viral vector; (4) washing CAR T cells to remove viral vector, and (5) harvesting CAR T cells in IV fluid. Conventional process uses magnetic separation and serial centrifugation technologies on multiple platforms with manual operations. Here we demonstrate the feasibility of rapid manufacturing CAR T cells on an automated platform with closed fluidics without centrifugation steps. The platform, MARS® Atlas, is built with innovative technologies in immune-magnetic cell isolation and acoustic cell washing and implemented as a modular system. In this feasibility test, T cells were isolated from peripheral blood from a healthy donoron MARS® magnetic module. eGFP expressing lentivirus were added to the enriched T cells. After short incubation, cells and virus were washed with fresh medium on the MARS® acoustic module and cells were incubated with virus overnight without activation. Infected cells were washed on the acoustic module to remove free virus. Product CAR T cells were harvested in saline solution and ready for QC analysis. Starting from 20mL of peripheral blood, magnetic T cell separation was done 2mL/min achieving 90% T cell purity, 90% recovery. In the first acoustic wash, medium exchange of the mixture of cells and virus took place at 0.9mL/min flow rate. As a result, 90% removal of contaminating molecule and 99.6% recovery of T cells were achieved. The second acoustic wash completed medium exchange of transfected cells at 0.9mL/min with 96.7% removal of virus and 90% CAR T cell recovery. As a result, 18 million GFP expressing T cells were produced. In this study we have demonstrated the feasibility of rapidly manufacturing CAR-T cells on an automated platform MARS® Atlas. This result indicates the MARS® Atlas being a potential CAR T manufacturing platform to be used in close-to-patient settings to enable low-cost cell therapy. 1 Saba Ghassemi, Reducing Ex Vivo Culture Improves the Antileukemic Activity of Chimeric Antigen Receptor (CAR) T Cells, Cancer Immunol Res; 6(9) September 2018 2 Saba Ghassemi, Rapid manufacturing of non-activated potent CAR T cells, Nature Biomedical Engineering | VOL 6 | February 2022 | 118-128 Citation Format: Silin Sa, Ethan Nguyen, Jie Pan, Liping Yu. A novel automated platform for rapid manufacturing of CAR T cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6331.

Mammalian orthoreovirus can exit cells in extracellular vesicles.

Several egress pathways have been defined for many viruses. Among these pathways, extracellular vesicles (EVs) have been shown to function as vehicles of non-lytic viral egress. EVs are heterogenous populations of membrane-bound structures released from cells as a form of intercellular communication. EV-mediated viral egress may enable immune evasion and collective viral transport. Strains of nonenveloped mammalian orthoreovirus (reovirus) differ in cell lysis phenotypes, with T3D disrupting cell membranes more efficiently than T1L. However, mechanisms of reovirus egress and the influence of transport strategy on infection are only partially understood. To elucidate reovirus egress mechanisms, we infected murine fibroblasts (L cells) and non-polarized human colon epithelial (Caco-2) cells with T1L or T3D reovirus and enriched cell culture supernatants for large EVs, medium EVs, small EVs, and free reovirus. We found that both reovirus strains exit cells in association with large and medium EVs and as free virus particles, and that EV-enriched fractions are infectious. While reovirus visually associates with large and medium EVs, only medium EVs offer protection from antibody-mediated neutralization. EV-mediated protection from neutralization is virus strain- and cell type-specific, as medium EVs enriched from L cell supernatants protect T1L and T3D, while medium EVs enriched from Caco-2 cell supernatants largely fail to protect T3D and only protect T1L efficiently. Using genetically barcoded reovirus, we provide evidence that large and medium EVs can convey multiple particles to recipient cells. Finally, T1L or T3D infection increases the release of all EV sizes from L cells. Together, these findings suggest that in addition to exiting cells as free particles, reovirus promotes egress from distinct cell types in association with large and medium EVs during lytic or non-lytic infection, a mode of exit that can mediate multiparticle infection and, in some cases, protection from antibody neutralization.