HSV-1 Replication Research Articles

Role of IL-27 in HSV-1-Induced Herpetic Stromal Keratitis.

Herpetic stromal keratitis (HSK) is a painful and vision-impairing disease caused by recurrent HSV-1 infection of the cornea. The virus replication in the corneal epithelium and associated inflammation play a dominant role in HSK progression. Current HSK treatments targeting inflammation or virus replication are partially effective and promote HSV-1 latency, and long-term use can cause side effects. Thus, understanding molecular and cellular events that control HSV-1 replication and inflammation is crucial for developing novel HSK therapies. In this study, we report that ocular HSV-1 infection induces the expression of IL-27, a pleiotropic immunoregulatory cytokine. Our data indicate that HSV-1 infection stimulates IL-27 production by macrophages. Using a primary corneal HSV-1 infection mouse model and IL-27 receptor knockout mice, we show that IL-27 plays a critical role in controlling HSV-1 shedding from the cornea, the optimum induction of effector CD4+ T cell responses, and limiting HSK progression. Using invitro bone marrow-derived macrophages, we show that IL-27 plays an antiviral role by regulating macrophage-mediated HSV-1 killing, IFN-β production, and IFN-stimulated gene expression after HSV-1 infection. Furthermore, we report that IL-27 is critical for macrophage survival, Ag uptake, and the expression of costimulatory molecules involved in the optimum induction of effector T cell responses. Our results indicate that IL-27 promotes endogenous antiviral and anti-inflammatory responses and represents a promising target for suppressing HSK progression.

CLEAR Strategy Inhibited HSV Proliferation Using Viral Vectors Delivered CRISPR-Cas9.

Herpes simplex virus type 1 (HSV-1) is a leading cause of encephalitis and infectious blindness. The commonly used clinical therapeutic drugs are nucleoside analogues such as acyclovir. However, current drugs for HSV cannot eliminate the latent virus or viral reactivation. Therefore, the development of new treatment strategies against latent HSV has become an urgent need. To comprehensively suppress the proliferation of HSV, we designed the CLEAR strategy (coordinated lifecycle elimination against viral replication). VP16, ICP27, ICP4, and gD-which are crucial genes that perform significant functions in different stages of the HSV infection lifecycle-were selected as targeting sites based on CRISPR-Cas9 editing system. In vitro and in vivo investigations revealed that genome editing by VP16, ICP27, ICP4 or gD single gene targeting could effectively inhibit HSV replication. Moreover, the combined administration method (termed "Cocktail") showed superior effects compared to single gene editing, which resulted in the greatest decrease in viral proliferation. Lentivirus-delivered CRISPR-Cas9/gRNA editing could effectively block HSV replication. The CLEAR strategy may provide new insights into the potential treatment of refractory HSV-1-associated diseases, particularly when conventional approaches have encountered resistance.

Oncolytic herpes simplex virus armed with a bacterial GBP1 degrader improves antitumor activity

Oncolytic viruses (OVs) encoding various transgenes are being evaluated for cancer immunotherapy. Diverse factors such as cytokines,immune checkpoint inhibitors, tumor-associated antigens, and Tcell engagers have been exploited as transgenes. These modifications are primarily aimed to reverse the immunosuppressive tumor microenvironment. By contrast, antiviral restriction factors that inhibit the replication of OVs and result in suboptimal oncolytic activity have received far less attention. Here, we report that guanylate-binding protein 1 (GBP1) is potently induced duringHSV-1 infection and restricts HSV-1 replication. Mechanistically, GBP1 remodels cytoskeletal organization to impede nuclear entry of HSV-1 genome. Previous studies have established that IpaH9.8, a bacterial E3 ubiquitin ligase, targets GBPs for proteasomal degradation. We therefore engineered an oncolytic HSV-1 to express IpaH9.8 and found that the modified OV effectively antagonized GBP1, replicated to a higher titer invitro and showed superior antitumor activity invivo. Our study features a strategy for improving the replication of OVs via targeting a restriction factor and achieving promising therapeutic efficacy.

A supramolecular gel with unique rheological properties for treating corneal virus infection

Herpes simplex keratitis (HSK) caused by herpes simplex virus-1 infection of the cornea has taken a significant toll on the visual health of people worldwide. Anti-viral hydrogels are preferred in the treatment of HSK due to enhanced topical drug retention and reduced side effects, however, current polymer-based gelling systems usually generate clumpy or uneven gel layers on ocular surface, and are easily cleared by eyelid blinking. This study proposes a supramolecular gel co-assembled by all-small-molecule building blocks including ganciclovir and 2′-deoxyguanosine in the presence of potassium ions. The developed gel performs fascinating rheological and texture properties including high viscoelastic, good spread ability, and deformation recovery behavior, enabling the formation of a uniform and thin gel layer on ocular surface to resist the clearance by eyelid blinking and tear washing. The gel shows similar ocular convenience, but much longer drug retention in ocular tissues and higher therapeutic efficacy compared to the clinical ganciclovir gel on the primary and recurrent mouse HSK models, rabbit HSK models, and also efficiently inhibited the HSV-1 replication in ex vivo human corneal tissues. The results suggest that this all-small-molecule supramolecular gel could be a promising drug formulation for treating HSK. Data and materials availabilityThe main data supporting the results in this study are available within the paper and its Supplementary Information. The associated raw data and analyzed datasets are too large to be readily shared publicly, but they are available from the corresponding author on reasonable request.

A potential anti-HIV-1 compound, Q308, inhibits HSV-2 infection and replication in vitro and in vivo

HSV-2 is a common human pathogen worldwide that causes genital herpes. Due to the lack of an effective HSV-2 vaccine in the foreseeable future, there is an urgent need to develop effective, safe and affordable anti-HSV-2 agents. Our previous studies confirmed that a small-molecule compound, Q308, effectively inhibits the reactivation of latent HIV and might be developed as an anti-HIV-1 agent. Patients infected with HSV-2 are generally more susceptible to HIV-1 infection than normal humans. In this study, we found that Q308 treatment had strong inhibitory activity against both HSV-2 and acyclovir-resistant HSV-2 strains in vitro and reduced the viral titers in tissue. And this treatment effectively ameliorated the cytokine storm and pathohistological changes caused by HSV-2 infection in HSV-2-infected mice. Unlike nucleoside analogs such as acyclovir, Q308 inhibited post-viral entry events by attenuating the synthesis of viral proteins. Furthermore, Q308 treatment blocked HSV-2-induced PI3K/AKT phosphorylation due to its inhibition on viral infection and replication. Overall, Q308 treatment exhibits potent anti-HSV-2 activity by inhibiting viral replication both in vitro and in vivo. Q308 is a promising lead compound for the development of new anti-HSV-2/HIV-1 therapies, particularly against acyclovir-resistant HSV-2 strains.

Myristic acid as a checkpoint to regulate STING-dependent autophagy and interferon responses by promoting N-myristoylation

Stimulator of interferon gene (STING)-triggered autophagy is crucial for the host to eliminate invading pathogens and serves as a self-limiting mechanism of STING-induced interferon (IFN) responses. Thus, the mechanisms that ensure the beneficial effects of STING activation are of particular importance. Herein, we show that myristic acid, a type of long-chain saturated fatty acid (SFA), specifically attenuates cGAS-STING-induced IFN responses in macrophages, while enhancing STING-dependent autophagy. Myristic acid inhibits HSV-1 infection-induced innate antiviral immune responses and promotes HSV-1 replication in mice in vivo. Mechanistically, myristic acid enhances N-myristoylation of ARF1, a master regulator that controls STING membrane trafficking. Consequently, myristic acid facilitates STING activation-triggered autophagy degradation of the STING complex. Thus, our work identifies myristic acid as a metabolic checkpoint that contributes to immune homeostasis by balancing STING-dependent autophagy and IFN responses. This suggests that myristic acid and N-myristoylation are promising targets for the treatment of diseases caused by aberrant STING activation.

Chloroquinolone Carboxamide Derivatives as New Anti-HSV-1 Promising Drugs.

Since the emergence of HSV resistant strains, new antiviral agents have emerged and still are urgently needed, especially those with alternative targets. In this work, we evaluated new quinolone derivatives as anti-HSV. For this study, cells were infected and treated with different components to evaluate the profile of HSV replication in vitro. In addition, studies were performed to determine the pharmacokinetic toxicity and profile of the compound. Indeed the EC50 values of these promising molecules ranged between 8 μM and 32 μM. We have also showed that all compounds inhibited the expression of ICP27 viral proteins, which gives new insights in the search for new target for antiherpetic therapy. Chlorine in positions C6 and phosphonate in position C1 have shown to be important for viral inhibition. The chloroquinolone carboxamide derivatives fulfilled "Lipinsky Rule of Five" for good oral bioavailability and showed higher intestinal absorption and blood brain barrier penetration, as well as lower toxicity profile. Although the inhibition activities of chloroquinolone carboxamide derivatives were lower than acyclovir, they showed different modes of action in comparison to the drugs currently available. These findings encourage us to continue pre-clinical studies for the development of new anti-HSV-1 agents.

Analysis of Antioxidant and Antiviral Effects of Olive (Olea europaea L.) Leaf Extracts and Pure Compound Using Cancer Cell Model.

The present study aims to assess the antioxidant and antiviral effectiveness of leaf extracts obtained from Olea europaea L. var. sativa and Olea europaea L. var. sylvestris. The total antioxidant activity was determined via both an ammonium phosphomolybdate assay and a nitric oxide radical inhibition assay. Both extracts showed reducing abilities in an in vitro system and in human HeLa cells. Indeed, after oxidative stress induction, we found that exposition to olive leaf extracts protects human HeLa cells from lipid peroxidation and increases the concentration of enzyme antioxidants such as catalase (CAT), superoxide dismutase (SOD), and glutathione peroxidase. Additionally, OESA treatment affects viral DNA accumulation more than OESY, probably due to the exclusive oleuropein content. In fact, subtoxic concentrations of oleuropein inhibit HSV-1 replication, stimulating the phosphorylation of PKR, c-FOS, and c-JUN proteins. These results provide new knowledge about the potential health benefits and mechanisms of action of oleuropein and oleuropein-rich extracts.

Osteopontin contributes to virus resistance associated with type I IFN expression, activation of downstream ifn-inducible effector genes, and CCR2+CD115+CD206+ macrophage infiltration following ocular HSV-1 infection of mice.

Ocular pathology is often associated with acute herpes simplex virus (HSV)-1 infection of the cornea in mice. The present study was undertaken to determine the role of early T lymphocyte activation 1 protein or osteopontin (OPN) in corneal inflammation and host resistance to ocular HSV-1 infection. C57BL/6 wild type (WT) and osteopontin deficient (OPN KO) mice infected in the cornea with HSV-1 were evaluated for susceptibility to infection and cornea pathology. OPN KO mice were found to possess significantly more infectious virus in the cornea at day 3 and day 7 post infection compared to infected WT mice. Coupled with these findings, HSV-1-infected OPN KO mouse corneas were found to express less interferon (IFN)-α1, double-stranded RNA-dependent protein kinase, and RNase L compared to infected WT animals early post infection that likely contributed to decreased resistance. Notably, OPN KO mice displayed significantly less corneal opacity and neovascularization compared to WT mice that paralleled a decrease in expression of vascular endothelial growth factor (VEGF) A within 12 hr post infection. The change in corneal pathology of the OPN KO mice aligned with a decrease in total leukocyte infiltration into the cornea and specifically, in neutrophils at day 3 post infection and in macrophage subpopulations including CCR2+CD115+CD206+ and CD115+CD183+CD206+ -expressing cells. The infiltration of CD4+ and CD8+ T cells into the cornea was unaltered comparing infected WT to OPN KO mice. Likewise, there was no difference in the total number of HSV-1-specific CD4+ or CD8+ T cells found in the draining lymph node with both sets functionally competent in response to virus antigen comparing WT to OPN KO mice. Collectively, these results demonstrate OPN deficiency directly influences the host innate immune response to ocular HSV-1 infection reducing some aspects of inflammation but at a cost with an increase in local HSV-1 replication.

Antiviral Potential of Melissa officinalis L.: A Literature Review.

The use of synthetic drugs has increased in recent years; however, herbal medicine is yet more trusted among a huge population worldwide; This could be due to minimal side effects, affordable prices, and traditional beliefs. Lemongrass (Melissa officinalis) has been widely used for reducing stress and anxiety, increasing appetite and sleep, reducing pain, healing wounds, and treating poisonous insect bites and bee stings for a long time. Today, research has shown that this plant can also fight viruses including Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), Herpes Simplex Virus (HSV), and Human Immunodeficiency Virus (HIV) through various mechanisms such as inhibiting HSV-1 from binding to host cell, inhibiting HSV-1 replication during the post-adsorption or inhibiting main protease and spike protein of SARS-CoV-2, furthermore, be effective in treating related diseases. This Review investigated the antiviral properties of Melissa officinalis and its effect on viral diseases. More in vitro and in vivo studies are needed to determine Melissa officinaliss underlying mechanism, and more randomized controlled trials should be done to identify its effect in humans. Also, due to the usefulness and lack of side effects, it can be used more as a complementary medicine.

Garlic essential oil shows no antiviral effects on the replication of HSV-1 in the virus-infected VERO cells

Garlic essential oil shows no antiviral effects on the replication of HSV-1 in the virus-infected VERO cells

Punica granatum Peel and Leaf Extracts as Promising Strategies for HSV-1 Treatment.

Punica granatum is a rich source of bioactive compounds which exhibit various biological effects. In this study, pomegranate peel and leaf ethanolic crude extracts (PPE and PLE, respectively) were phytochemically characterized and screened for antioxidant, antimicrobial and antiviral activity. LC-PDA-ESI-MS analysis led to the identification of different compounds, including ellagitannins, flavonoids and phenolic acids. The low IC50 values, obtained by DPPH and FRAP assays, showed a noticeable antioxidant effect of PPE and PLE comparable to the reference standards. Both crude extracts and their main compounds (gallic acid, ellagic acid and punicalagin) were not toxic on Vero cells and exhibited a remarkable inhibitory effect on herpes simplex type 1 (HSV-1) viral plaques formation. Specifically, PPE inhibited HSV-1 adsorption to the cell surface more than PLE. Indeed, the viral DNA accumulation, the transcription of viral genes and the expression of viral proteins were significantly affected by PPE treatment. Amongst the compounds, punicalagin, which is abundant in PPE crude extract, inhibited HSV-1 replication, reducing viral DNA and transcripts accumulation, as well as proteins of all three phases of the viral replication cascade. In contrast, no antibacterial activity was detected. In conclusion, our findings indicate that Punica granatum peel and leaf extracts, especially punicalagin, could be a promising therapeutic candidate against HSV-1.

3D-Printed EVA Devices for Antiviral Delivery and Herpes Virus Control in Genital Infection.

Herpes viruses are widespread in the human population and can cause many different diseases. Genital herpes is common and can increase the risk of HIV infection and neonatal herpes. Acyclovir is the most used drug for herpes treatment; however, it presents some disadvantages due to its poor oral bioavailability. In this study, some ethylene vinyl acetate devices with different acyclovir amounts (0, 10, and 20 wt.%) were manufactured by fused filament fabrication in two different geometries, an intrauterine device, and an intravaginal ring. Thermal analyses suggested that the crystallinity of EVA decreased up to 8% for the sample loaded with 20 wt.% of acyclovir. DSC, SEM, and FTIR analyses confirmed that the drug was successfully incorporated into the EVA matrix. Moreover, the drug release tests suggested a burst release during the first 24 h followed by a slower release rate sustained up to 80 days. Biological assays showed the biocompatibility of the EVA/ACV device, as well as a 99% reduction in vitro replication of HSV-1. Finally, the EVA presented a suitable performance for 3D printing manufacturing that can contribute to developing personalized solutions for long-term herpes treatment.

Lipopolysaccharide enhances HSV-1 replication and inflammatory factor release in the ARPE-19 cells

PurposeDuring acute retinal necrosis (ARN), retinal pigment epithelial (RPE) cells could be stimulated by both herpes simplex virus (HSV) and lipopolysaccharide (LPS). We aim to investigate the impact of LPS on HSV-1 infection and inflammatory factors in human retinal pigment epithelial cell lines (ARPE-19 cells). MethodsARPE-19 cells were infected by HSV-1F strain and HSVg4 strain, a modified HSV strain with GFP genes cloned in, for 1 h. Different concentrations of LPS were added. Green fluorescence protein (GFP) of HSVg4 and the infected cell protein 4 (ICP4) expression were observed. Cell culture supernatants were collected to detect 34 kinds of related cytokines and chemokines by multiplex immunoassay assay. ResultsUnder LPS treatment, the cytopathic effect displayed as enlarged multinucleated cells, and the GFP fluorescence intensity and ICP4 expression increased in the HSV-1-infected ARPE-19 cells. HSV-1 infection stimulated cytokines IL-1α, IL-1β, IL-1RA, IL-2, IL-4, IL-6, IL-9, IL-12P70, IL-15, IL-18, IL-21, IL-27, TNF-α, IFN-γ and chemokines CXCL1, CXCL8, CXCL10, CXCL12, CCL2, CCL3, CCL4, CCL5, CCL11 while LPS further enhanced their expression. ConclusionLPS promoted HSV-1 infection and inflammatory factor release in ARPE-19 cells, indicating that ARN could deteriorate when complicated with endotoxemia.

PUL36 Deubiquitinase Activity Augments Both the Initiation and the Progression of Lytic Herpes Simplex Virus Infection in IFN-Primed Cells.

The evolutionarily conserved, structural HSV-1 tegument protein pUL36 is essential for both virus entry and assembly. While its N-terminal deubiquitinase (DUB) activity is dispensable for infection in cell culture, it is required for efficient virus spread in vivo, as it acts as a potent viral immune evasin. Interferon (IFN) induces the expression of hundreds of antiviral factors, including many ubiquitin modulators, which HSV-1 needs to neutralize to efficiently initiate a productive infection. Herein, we discover two functions of the conserved pUL36 DUB during lytic replication in cell culture in an understudied but equally important scenario of HSV-1 infection in IFN-treated cells. Our data indicate that the pUL36 DUB contributes to overcoming the IFN-mediated suppression of productive infection in both the early and late phases of HSV-1 infection. We show that incoming tegument-derived pUL36 DUB activity contributes to the IFN resistance of HSV-1 in IFN-primed cells to efficiently initiate lytic virus replication. Subsequently, the de novo expressed DUB augmented the efficiency of virus replication and increased the output of infectious virus. Notably, the DUB defect was only apparent when IFN was applied prior to infection. Our data indicate that IFN-induced defense mechanisms exist and that they work to both neutralize infectivity early on and slow the progression of HSV-1 replication in the late stages of infection. Also, our data indicate that pUL36 DUB activity contributes to the disarming of these host responses. IMPORTANCE HSV-1 is a ubiquitous human pathogen that is responsible for common cold sores and may also cause life-threatening disease. pUL36 is an essential, conserved herpesvirus protein with N-terminal deubiquitinating (DUB) activity. The DUB is dispensable for HSV-1 replication in cell culture but represents an important viral immune evasin in vivo. IFN plays a pivotal role in HSV-1 infection and suppresses viral replication both in vitro and in vivo. Here, we show that DUB activity contributes to overcoming IFN-induced cellular resistance in order to more efficiently initiate lytic replication and produce infectious virions. As such, DUB activity in the incoming virions increases their infectivity, while the de novo synthesized DUB augments productive infection. Thus, the HSV-1 DUB antagonizes the activity of IFN-inducible effector proteins to facilitate productive infection at multiple levels. Our findings underscore the importance of using more challenging cell culture systems to fully understand virus protein functions.

RIG-I contributes to dsDNA-induced innate immune activation in human brain microvascular endothelial cells

Human brain microvascular epithelial cells (HBMECs) play a pivotal role in maintaining the stability of the blood-brain barrier (BBB), a potent physiological barrier to prevent the invasion of exotic pathogens. Our previous study indicated that polyI:C, an analog of double-stranded RNA (dsRNA), could initiate the TLR3/IFNs antiviral signaling pathway in HBMECs. However, the response of HBMECs to dsDNA remains unclear. In this study, we demonstrated that a dsDNA mimic, poly(dA:dT), could induce the production of a series of antiviral factors, including IFN-β, IFN-λ1, and ISGs. Furthermore, the poly(dA:dT)-activated HBMECs could effectively restrain HSV-1 replication. In addition, we found that RIG-I rather than cGAS and IFI16 had a more crucial role in sensing poly(dA:dT). These observations indicate that HBMECs possess a dsDNA sensing system, and RIG-I may partly contribute to the dsDNA-induced antiviral innate immunity.

Knockout of signal peptide peptidase in the eye reduces HSV-1 replication and eye disease in ocularly infected mice.

We previously reported that knocking out signal peptide peptidase (SPP), a glycoprotein K (gK) binding partner, in mouse peripheral sensory neurons reduced latency-reactivation in infected mice without affecting primary virus replication or eye disease. Since virus replication in the eye plays an essential role in eye disease, we generated a conditional knockout mouse lacking SPP expression in the eye by crossing Pax6 (paired box 6)-Cre mice that have intact Pax6 expression with SPPflox/flox mice. Significantly less SPP protein expression was detected in the eyes of Pax6-SPP-/- mice than in WT control mice. HSV-1 replication in the eyes of Pax6-SPP-/- mice was significantly lower than in WT control mice. Levels of gB, gK, and ICP0 transcripts in corneas, but not trigeminal ganglia (TG), of Pax6-SPP-/- infected mice were also significantly lower than in WT mice. Corneal scarring and angiogenesis were significantly lower in Pax6-SPP-/- mice than in WT control mice, while corneal sensitivity was significantly higher in Pax6-SPP-/- mice compared with WT control mice. During acute viral infection, absence of SPP in the eye did not affect CD4 expression but did affect CD8α and IFNγ expression in the eye. However, in the absence of SPP, latency-reactivation was similar in Pax6-SPP-/- and WT control groups. Overall, our results showed that deleting SPP expression in the eyes reduced primary virus replication in the eyes, reduced CD8α and IFNγ mRNA expression, reduced eye disease and reduced angiogenesis but did not alter corneal sensitivity or latency reactivation to HSV-1 infection. Thus, blocking gK binding to SPP in the eye may have therapeutic potential by reducing both virus replication in the eye and eye disease associated with virus replication.

Direct cleavage of caspase-8 by herpes simplex virus 1 tegument protein US11

The HSV-1 tegument protein Us11 counteracts the antiviral defense mechanisms by precluding the host protein shutoff. Previous works demonstrated that Us11 prevents heat-and staurosporine-induced apoptosis and inhibits autophagy. Therefore, in the present study, we investigated the hypothesis that HSV-1, through Us11, could recruit caspase-8, a key enzyme regulating programmed cell death. We first show that HSV-1 promotes the accumulation of caspase-8-p18 active fragments in both semi permissive THP-1 cells and fully permissive HEp-2 cells to HSV-1 replication. Using a recombinant virus R3630 (ΔUs11/ΔUs12) and a plasmid encoding Us11-recombinant protein we have proven that Us11 promotes p18 accumulation, which does not trigger the apoptotic signaling. Additional, in an in vitro model, we demonstrated that Us11-recombinant protein induces caspase-8-p18 cleavage by physically interacting with the caspase-8 recombinant protein. Finally, we found that, during HSV-1 replication, activated-caspase-8 cleaves Atg3 protein to potentially block autophagy and support its replication.

Swarms of chemically modified antiviral siRNA targeting herpes simplex virus infection in human corneal epithelial cells.

Herpes simplex virus type 1 (HSV-1) is a common virus of mankind and HSV-1 infections are a significant cause of blindness. The current antiviral treatment of herpes infection relies on acyclovir and related compounds. However, acyclovir resistance emerges especially in the long term prophylactic treatment that is required for prevention of recurrent herpes keratitis. Earlier we have established antiviral siRNA swarms, targeting sequences of essential genes of HSV, as effective means of silencing the replication of HSV in vitro or in vivo. In this study, we show the antiviral efficacy of 2´-fluoro modified antiviral siRNA swarms against HSV-1 in human corneal epithelial cells (HCE). We studied HCE for innate immunity responses to HSV-1, to immunostimulatory cytotoxic double stranded RNA, and to the antiviral siRNA swarms, with or without a viral challenge. The panel of studied innate responses included interferon beta, lambda 1, interferon stimulated gene 54, human myxovirus resistance protein A, human myxovirus resistance protein B, toll-like receptor 3 and interferon kappa. Our results demonstrated that HCE cells are a suitable model to study antiviral RNAi efficacy and safety in vitro. In HCE cells, the antiviral siRNA swarms targeting the HSV UL29 gene and harboring 2´-fluoro modifications, were well tolerated, induced only modest innate immunity responses, and were highly antiviral with more than 99% inhibition of viral release. The antiviral effect of the 2’-fluoro modified swarm was more apparent than that of the unmodified antiviral siRNA swarm. Our results encourage further research in vitro and in vivo on antiviral siRNA swarm therapy of corneal HSV infection, especially with modified siRNA swarms.

Bulgarian Medicinal Extracts as Natural Inhibitors with Antiviral and Antibacterial Activity.

Background: Bulgaria is a country with a wide range of medicinal plants, with uses in traditional medicine dating back for centuries. Methods: Disc diffusion assay was used to evaluate the antimicrobial activity of the plant extracts. A cytopathic effect inhibition test was used for the assessment of the antiviral activity of the extracts. The virucidal activity of the extracts, their influence on the stage of viral adsorption, and their protective effect on uninfected cells were reported using the end-point dilution method, and Δlgs was determined as compared to the untreated controls. Results: The results of the study reveal that the antibacterial potential of G. glabra and H. perforatum extracts in Gram-positive bacteria is more effective than in Gram-negative bacteria. When applied during the replication of HSV-1 and HCov-OC-43, only some of the extracts showed weak activity, with SI between 2 to 8.5. Almost all tested extracts inhibited the extracellular virions of the studied enveloped viruses (HSV-1 and HCov-OC-43) to a greater extent than of the non-enveloped viruses (PV-1 and HAdV-5). They inhibited the stage of viral adsorption (HSV-1) in the host cell (MDBK) to varying degrees and showed a protective effect on healthy cells (MDBK) before they were subjected to viral invasion (HSV-1). Conclusion: The antipathogenic potential of extracts of H. perforatum and G. glabra suggests their effectiveness as antimicrobial agents. All 13 extracts of the Bulgarian medicinal plants studied can be used to reduce viral yield in a wide range of viral infections.