Anti-herpesviral property and mode of action of a polysaccharide from brown seaweed (<Emphasis Type="Italic">Hydroclathrus clathratus</Emphasis>)

A sulfated polysaccharide, designated HC-b1, was isolated from the brown seaweed Hydroclathrus clathratus. It was found to be a strong inhibitor of herpes simplex virus type 1 (HSV-1), including acyclovir-resistant strain and clinical strain. HC-b1 inhibited the plaque formation of HSV-1 in a dose-dependent manner. It could protect Vero cells from infection by HSV-1 if the cells were incubated with HC-b1 before exposure to the virus. It also had inactivating effect against HSV-1 since the pretreatment of the virus with HC-b1 caused significant reduction of viral infectivity. Time of addition studies demonstrated that HC-b1 exerted its antiviral action at the early stage of virus replication cycle. The presence of HC-b1 could not effectively inhibit the replication of HSV-1 about 45 min after the penetration period started. The antiviral action of HC-b1 appeared to inhibit the attachment of herpes simplex virus on host cell membrane through interfering with the processes of adsorption and penetration.     

Cytotoxicity and potential antiviral evaluation of violacein produced by Chromobacterium violaceum

Natural products are an inexhaustible source of compounds with promising pharmacological activities including antiviral action. Violacein, the major pigment produced by Chromobacterium violaceum, has been shown to have antibiotic, antitumoral and anti-Trypanosoma cruzi activities. The goal of the present work was to evaluate the cytotoxicity of violacein and also its potential antiviral properties.The cytotoxicity of violacein was investigated by three methods: cell morphology evaluation by inverted light microscopy and cell viability tests using the Trypan blue dye exclusion method and the MTT assay. The cytotoxic concentration values which cause destruction in 50% of the monolayer cells (CC50) were different depending on the sensitivity of the method. CC50 values were > or =2.07 +/- 0.08 microM for FRhK-4 cells: > or =2.23 +/- 0.11 microM for Vero cells; > or =2.54 +/- 0.18 microM for MA104 cells; and > or =2.70 +/- 0.20 microM for HEp-2 cells. Violacein showed no cytopathic inhibition of the following viruses: herpes simplex virus type 1 (HSV-1) strain 29-R/acyclovir resistant, hepatitis A virus (strains HM175 and HAF-203) and adenovirus type 5 nor did it show any antiviral activity in the MTT assay. However violacein did show a weak inhibition of viral replication: 1.42 +/- 0.68%, 14.48 +/- 5.06% and 21.47 +/- 3.74% for HSV-1 (strain KOS); 5.96 +/- 2.51%, 8.75 +/- 3.08% and 17.75 +/- 5.19% for HSV-1 (strain ATCC/VR-733); 5.13 +/- 2.38 %, 8.18 +/- 1.11% and 8.51 +/- 1.94% for poliovirus type 2; 8.30 +/- 4.24%; 13.33 +/- 4.66% and 24.27 +/- 2.18% for simian rotavirus SA11, at 0.312, 0.625 and 1.250 mM, respectively, when measured by the MTT assay.     

Different antiviral potencies of BV-araU and related nucleoside analogues against herpes simplex virus type 1 in human cell lines and Vero cells.

Antiviral potencies against herpes simplex virus type 1 (HSV-1) of 1-beta-D-arabinofuranosyl-E-5-(2-bromovinyl)uracil (BV-araU) and ten other nucleoside analogues in human embryonic lung fibroblast (HEL) cells were compared with those in Vero cells. 5-Halogenovinylarabinosyluracils, highly active in HEL cells, were inactive against all three laboratory-stocked strains of HSV-1 but exerted moderate antiviral effects on three clinical isolates in Vero cells. The reduction in anti-HSV-1 potencies of other representative nucleoside analogues in Vero cells was much less than those of 5-halogenovinylarabinosyluracils. However, significant antiviral potencies of BV-araU against laboratory strains were observed in other human and monkey fibroblast cells including an immortalized cell line. Significant enhancement of antiviral activity of BV-araU against a laboratory strain and a clinical isolate was demonstrated in Vero cells by the addition of 0.1 microM aminopterin or FUdR, an inhibitor of thymidylate synthesis. The potentiated anti-HSV-1 activity in Vero cells was comparable to the potency in HEL cells without the inhibitor. These results suggested that high activity of thymidylate synthesis and a large thymidylate pool size in Vero cells seem to be related to loss of anti-HSV-1 potency of BV-araU. Original tissue type, species, and the immortality may not be responsible for the reduced antiviral activity of BV-araU in Vero cells.     

Anti-herpes simplex virus activity of sulfated galactans from the red seaweeds Gymnogongrus griffithsiae and Cryptonemia crenulata

This study presents the chemical composition and antiviral activity against herpes simplex virus type 1 (HSV-1) and 2 (HSV-2) of sulfated galactan crude extracts and main fractions obtained from two red seaweeds collected in Brazil, Gymnogongrus griffithsiae and Cryptonemia crenulata. Most of the eighteen tested products, including homogeneous kappa/iota/nu carrageenan and DL-galactan hybrid, exhibited antiherpetic activity with inhibitory concentration 50% (IC50) values in the range 0.5-5.6 microg/ml, as determined in a virus plaque reduction assay in Vero cells. The galactans lacked cytotoxic effects and showed a broad spectrum of antiviral activity against HSV-1 and HSV-2. No direct virus inactivation was observed after virion treatment with the galactans. The mode of action of these compounds could be mainly ascribed to an inhibitory effect on virus adsorption. Most importantly, a significant protection against a murine vaginal infection with HSV-2 was afforded by topical treatment with the sulfated galactans.     

In vitro bactericidal action of a complex of natural cytokins on Streptococcus pyogenes

The data on the antimicrobial in vitro action of a complex of natural cytokins (CNC)--the substance and the preparation "Superlymph"--on virulent museum and clinical macrolid-resistant group A beta-hemolytic streptococci, are presented. CNC suppressed the growth of museum test culture of streptococci in the concentration dependent manner. Moreover, the preparation "Superlymph" proved to be an effective antistreptococcal remedy in its action on erythromycin-resistant group A streptococcal (GAS) strains. The substance of the preparation induced the lysis of GAS cells, grown on lawn plates and in the system of spectrophotometric analysis. The effect thus established could be caused by cationic antimicrobial peptides (protegrins), contained in "Superlymph".     

The relationship between incorporation of E-5-(2-Bromovinyl)-2'-deoxyuridine into herpes simplex virus type 1 DNA with virus infectivity and DNA integrity

E-5-(2-Bromovinyl)-2'-deoxyuridine (BrvdUrd) produced a dose-dependent shift in the density of herpes simplex virus type 1 (HSV-1) DNA at concentrations which yielded potent inhibition of virus replication in cultured Vero cells. Although the density of cellular DNA was not altered by these concentrations of BrvdUrd, incorporation of this analogue into cellular DNA of HSV-1-infected cells has been previously observed in this laboratory. The degree of inhibition correlated with the amount of BrvdUrd substituted for thymidine in HSV-1 DNA. BrvdUrd-substituted DNA was more labile as determined by a dose-dependent increase in single strand breaks when examined by centrifugation in alkaline sucrose gradients. Thus, the potent antiviral action of BrvdUrd observed in cell culture correlates not only with its incorporation into HSV-1 DNA but also with an altered stability of this DNA.     

Specific phosphorylation of 5-ethyl-2'-deoxyuridine by herpes simplex virus-infected cells and incorporation into viral DNA

5-Ethyl-2'-deoxyuridine (EDU) is a potent and selective inhibitor of the replication of herpes simplex virus type 1 (HSV-1) and 2 (HSV-2), which is currently being pursued for the topical treatment of HSV-1 and HSV-2 infections in humans. Using [4-14C]EDU as the radiolabeled analogue of EDU, it was ascertained that, at antivirally active doses, EDU is phosphorylated to a much greater extent by HSV-infected Vero cells than by mock-infected cells. Within the HSV-1-infected cells, EDU was incorporated to a much greater extent into viral DNA than cellular DNA. Using varying doses of EDU, a close correlation was found between the incorporation of EDU into viral DNA, the inhibition of viral DNA synthesis, and the inhibition of virus yield. It is postulated that the selectivity of EDU as an antiviral agent depends on both its preferential phosphorylation by the virus-infected cell and its preferential incorporation into viral DNA. The latter than results in a suppression of viral DNA synthesis and, hence, shutoff of viral progeny formation.     

Mechanism of action of the suppression of herpes simplex virus type 2 replication by pterocarnin A

The purpose of this study was to investigate the in vitro antiviral properties of pterocarnin A, extracted from the bark of Pterocarya stenoptera (Juylandaceae). Results showed that pterocarnin A exhibited anti-herpes simplex virus (HSV) activity. It had a low selectivity index (SI) value and only possessed some level of cell cytotoxic effect at high antiviral concentrations. Mechanism studies demonstrated that pterocarnin A inhibited herpes simplex virus type 2 (HSV-2) from attaching and penetrating into cells. It also actively suppressed HSV-2 multiplication in Vero cells even when added 12 h after infection. This observation indicated that pterocarnin A affected the late stage(s) of HSV-2 infection cycle. Pterocarnin A also significantly reduced viral infectivity at high concentrations. From these observations, it was concluded that pterocarnin A suppressed both early and late in the replication cycle of HSV-2. The various modes of action of pterocarnin A in interfering with certain steps of viral infection thus merit further investigation.     

Antiviral activity of fattiviracin FV-8 against human immunodeficiency virus type 1 (HIV-1)

A novel antiviral agent, fattiviracin FV-8, purified from the culture broth of Streptomyces microflavus strain No. 2445, showed potent antiviral activities against human immunodeficiency virus type 1 (HIV-1), herpes simplex virus type 1 (HSV-1), varicella-zoster virus (VZV), and influenza A and B viruses. The action mechanism of fattiviracin FV-8 against HIV-1 was examined. As a result, the agent was thought to act on HIV-1 particles directly without lysis of the particles, and it affords the inhibition of viral entry into the host cells.     

Antiviral activity of recombinant cyanovirin-N against HSV-1

In this study, a standard strain of HSV-1 (strain SM₄₄) was used to investigate the antiviral activity of the recombinant Cyanovirin-N (CV-N) against Herpes simplex virus type 1 (HSV-1) in vitro and in vivo. Cytopathic effect (CPE) and MTT assays were used to evaluate the effect of CV-N on HSV-1 in Vero cells. The number of copies of HSV-DNA was detected by real-time fluorescence quantitative PCR (FQ-PCR). The results showed that CV-N had a low cytotoxicity on Vero cells with a CC₅₀ of 359.03±0.56 μg/mL, and that it could not directly inactivate HSV-1 infectivity. CV-N not only reduced the CPE of HSV-1 when added before or after viral infection, with a 50% inhibitory concentration (IC₅₀) with 2.26 and 30.16μg/mL respectively, but it also decreased the copies of HSV-1 DNA in infected host cells. The encephalitis model for HSV-1 infection was conducted in Kunming mice, and treated with three dosages of CV-N (0.5, 5 &; 10 mg/kg) which was administered intraperitoneally at 2h, 3d, 5d, 7d post infection. The duration for the appearance of symptoms of encephalitis and the survival days were recorded and brain tissue samples were obtained for pathological examination (HE staining). Compared with the untreated control group, in the 5mg/kg CV-N and 10mg/kg CV-N treated groups, the mice suffered light symptoms and the number of survival days were more than 9d and 14d respectively. HE staining also showed that in 5mg/kg CV-N and 10mg/kg CV-N treated groups, the brain cells did not show visible changes, except for a slight inflammation. Our results demonstrated that CV-N has pronounced antiviral activity against HSV-1 both in vitro and in vivo, and it would be a promising new candidate for anti-HSV therapeutics.     

Comparison of disc diffusion, Etest and broth microdilution for testing susceptibility of carbapenem-resistant P. aeruginosa to polymyxins

Background

New prophylactic and therapeutic tools are needed for the treatment of herpes simplex virus infections. Several essential oils have shown to possess antiviral activity in vitro against a wide spectrum of viruses.

Aim

The present study was assess to investigate the activities of the essential oil obtained from leaves of Artemisia arborescens against HSV-1 and HSV-2

Methods

The cytotoxicity in Vero cells was evaluated by the MTT reduction method. The IC₅₀ values were determined by plaque reduction assay. In order to characterize the mechanism of action, yield reduction assay, inhibition of plaque development assay, attachment assay, penetration assay and post-attachment virus neutralization assay were also performed.

Results

The IC₅₀ values, determined by plaque reduction assay, were 2.4 and 4.1 μg/ml for HSV-1 and HSV-2, respectively, while the cytotoxicity assay against Vero cells, as determined by the MTT reduction method, showed a CC₅₀ value of 132 μg/ml, indicating a CC₅₀/IC₅₀ ratio of 55 for HSV-1 and 32.2 for HSV-2. The antiviral activity of A. arborescens essential oil is principally due to direct virucidal effects. A poor activity determined by yield reduction assay was observed against HSV-1 at higher concentrations when added to cultures of infected cells. No inhibition was observed by attachment assay, penetration assay and post-attachment virus neutralization assay. Furthermore, inhibition of plaque development assay showed that A. arborescens essential oil inhibits the lateral diffusion of both HSV-1 and HSV-2.

Conclusion

This study demonstrates the antiviral activity of the essential oil in toto obtained from A. arborescens against HSV-1 and HSV-2. The mode of action of the essential oil as antiherpesvirus agent seems to be particularly interesting in consideration of its ability to inactivate the virus and to inhibit the cell-to-cell virus diffusion.     

Efficacy of therapeutic and prophylactic actions of ultralow doses of antibodies to gamma-interferon in experimental murine model of herpes virus]

The process of the disease due to herpes simplex virus types 1 and 2 (HSV-1 and 2) was studied on white uninbred mice weighing 10 to 12 g. The animals were infected intracerebrally or intraperitoneally. Intraperitoneal contamination of the animals with MS strain of HSV-2 was used for the experimental model of the herpes simplex infection. The prophylactic antiherpes action of ultralow doses of the human gamma-interferon antibodies (ULD of anti-IFN-gamma) at a course of its intragastral administration was evaluated. The preparation was shown to have a significant (p < 0.05) protective effect in a dose of 10 LD50, evident from a 10-fold decrease of the HSV-2 accumulation in the brain, a lower percentage of the animal deaths and an increase of the average lifespan of the animals by 3.3 days. The study of the therapeutic action of ULD of anti-IFN-gamma at a course of its intragastral administration showed that the preparation had no significant positive effect on the disease process in the animals infected with HSV-2 in a dose of 10 LD50. However, a positive effect associated with delayed virus replication in the brain was observed in the study on the therapeutic effect of ULD of anti-IFN-gamma after its intragastral administration to the mice infected with a sublethal dose of the virus.     

The inhibitory effect of essential oils on herpes simplex virus type-1 replication in vitro

The antiviral effect of 12 essential oils on herpes simplex virus type-1 (HSV-1) replication was examined in vitro. The replication ability of HSV-1 was suppressed by incubation of HSV-1 with 1% essential oils at 4 C for 24 hr. Especially, lemongrass completely inhibited the viral replication even at a concentration of 0.1%, and its antiviral activity was dependent on the concentrations of the essential oil. When Vero cells were treated with the essential oil before or after viral adsorption, no antiviral activity was found, which suggests that the antiviral activity of essential oils including lemongrass may be due to the direct interaction with virions.     

Antiviral property and mechanisms of a sulphated polysaccharide from the brown alga Sargassum patens against Herpes simplex virus type 1

A sulphated polysaccharide (SP-2a) from the brown alga Sargassum patens (Kütz.) Agardh (Sargassaceae) was found to significantly inhibit the in vitro replication of both the acyclovir (ACV)-sensitive and -resistant strains of Herpes simplex virus type 1 (HSV-1), in dose-dependent manners, with 50% inhibitions occurring with 1.5–5.3 μg/ml of the polysaccharide. SP-2a exhibited extracellular virucidal activity only against the ACV-sensitive strains, but not the resistant strain, at the concentration of 100 μg/ml. The strongest antiviral activities against the different strains of HSV-1 were observed when this polysaccharide was present during and after adsorption of the virus to host cells. The inhibitory effect of SP-2a on virus adsorption occurred dose-dependently in all the HSV-1 strains tested, and the adsorption of the ACV-resistant DM2.1 strain was reduced by 81.9% (relative to control) with 4 μg/ml of the polysaccharide. This study clearly demonstrated that the antiviral mode of action of SP-2a is mediated mainly by inhibiting virus attachment to host cells, and this sulphated polysaccharide might have different modes of action against the ACV-sensitive and -resistant strains of HSV-1.     

Trichosanthin suppresses the elevation of p38 MAPK, and Bcl-2 induced by HSV-1 infection in Vero cells

Trichosanthin (TCS) is a type 1 ribosome-inactivating protein (RIP) effective against HIV-1 and HSV-1 replication. The mechanism of its antiviral activity is not clear. Many believe that it is related to ribosome inactivation. Some RIPs and viral infection affect the phosphorylation of MAPK and Bcl-2 and these proteins may be the common element linking RIP and viral infection. This study investigated the effect of HSV-1 infection on p38 MAPK and Bcl-2 as well as possible interference by TCS. Results showed that HSV-1 infection induced an elevation of phosphorylated p38 and Bcl-2 in Vero cells, which could be partially blocked by TCS. At the same time, both viral replication and host cells viability were lowered. Viral replication, Vero cell viability, p38 MAPK and Bcl-2 were further reduced with the addition of a p38 MAPK inhibitor (SB203580). This suggested that TCS may interfere with MAPK and Bcl-2 signals generated by infection leading to inhibition of viral replication. In summary, our results demonstrated that HSV-1 infection in Vero cells induced an elevation of p38 MAPK and Bcl-2. TCS suppressed this rise and reduced viral replication. The MAPK family may play a role in the antiviral mechanism of TCS.     

Optimization of virus yield as a strategy to improve rabies vaccine production by Vero cells in a bioreactor

To improve rabies vaccine production by Vero cells, we have developed a strategy based on high cell density culture and optimization of virus yield. We have first optimized cell growth in spinner flask using a Taguchi's L8 experimental design. We analyzed the effects of the following factors: initial glucose and glutamine concentrations, Cytodex 1 concentration and the regulation of glucose level at 1 g l(-1). We have also investigated the effect of the following factor interactions: Cytodex 1 concentration/glutamine concentration, Cytodex 1 concentration/glucose concentration and glucose concentration/glutamine concentration. Statistical analysis of the collected data pointed to the initial glucose concentration, the regulation of glucose level at 1 g l(-1) and the interactions between Cytodex 1 concentration/initial glucose concentration and Cytodex 1 concentration/initial glutamine concentration as the parameters that affected cell growth. Using the optimal conditions determined earlier, we have studied Vero cell growth in a 7-l bioreactor and in batch culture, and obtained a cell density level equal to 3.6 +/- 0.2 x 10(6) cells ml-1. Cell infection with rabies virus (LP 2061/Vero strain) at a multiplicity of infection (MOI) of 0.3 using M199 medium supplemented with 0.2% bovine serum albumin (BSA), yielded a maximal virus titer equal to 8 +/- 1.6 x 10(7) Fluorescent Focus Units (FFU) ml-1. We have also studied Vero cell growth in a 7-l bioreactor using recirculation as a perfusion culture mode during cell proliferation step and perfusion for virus multiplication phase. In comparison to batch culture, we reached a higher cell density level that was equal to 10.1 +/- 0.5 x 10(6) cells ml-1. Cell infection under the conditions previously indicated, yielded 14l of virus harvest that had a virus titer equal to 2.6 +/- 0.5 x 10(7) FFU ml-1. The activity of the inactivated virus harvest showed a protective activity that meets WHO requirements.     

6-Azathymidine-4'-thionucleosides: synthesis and antiviral evaluation

The synthesis of dideoxy-6-azathymidine 4'-thionucleoside 1-(2,3-dideoxy-4-thio-beta-D-erythro-pentofuranosyl)-(6-azathymidine) (2), and the L-nucleoside, 1-(4-thio-beta-L-erythro-pentofuranosyl)-(6-azathymidine) (3) and their evaluation against a wide panel of antiviral assays are described. The L-thionucleoside (3) was devoid of antiviral activity. The dideoxy-thionucleoside (2) was moderately active against vaccinia virus (VV) and the herpes simplex virus strains HSV-1 (strain KOS) and HSV-2 (strain G) (MIC 12 microM) and retained inhibitory activity vs a thymidine kinase-deficient strain HSV-1/TK(-), suggesting that (2) is not dependent on viral TK-catalysed phosphorylation for antiviral activity and/or may use an alternative metabolic activation pathway.     

In vitro anti-herpetic activity of sulfated polysaccharide fractions from Caulerpa racemosa

A sulfated polysaccharide fraction was isolated from the hot water extract of the green alga Caulerpa racemosa and designated HWE. This polymer, which contained galactose, glucose, arabinose and xylose as the major component sugars, had [alpha](D)(30) + 46.2 degrees in water and contained 9% sulfate hemiester groups. Sugar linkage analysis indicates that HWE was branched and mainly contained 1,3- and 1,3,6-linked galactose, 1,3,4-linked arabinose, 1,4-linked glucose and terminal- and 1,4-linked xylose residues. Sulfation was deduced from infrared spectroscopy and methylation analysis to occur on O-6 of galactose and O-3 of arabinose. The native polysaccharide could be fractionated by size exclusion chromatography into two overlapping fractions and the major fraction has a hydrodynamic volume similar to that of 70 kDa dextran. HWE was a selective inhibitor of reference strains and TK(-) acyclovir-resistant strains of herpes simplex virus type 1 (HSV-1) and type 2 (HSV-2) in Vero cells, with antiviral effective concentration 50% (EC(50)) values in the range of 2.2-4.2 microg/ml and lacking cytotoxic effects. Furthermore, HWE did not exhibit anticoagulant properties at concentrations near the EC(50).     

人集落刺激因子-1(CSF-1)对常见呼吸道病毒的抑制作用

本文观察集落刺激因子-1(CSF-1)及其诱生剂:内毒素(LPS)、胞壁二肽(MDP)和干扰素(IFN-α)对下列病毒所致细胞病变的抑制,包括不同型别腺病毒5株,单疱病毒Ⅰ型和Ⅱ型,流感病毒A_3型,鼻病毒,ECHO11型各1株,在人胚肺传代细胞株上病变抑制的结果如下:对HSV-1、HSV-2、腺病毒6、11、22型,流感A_3型,鼻病毒。(CSF-1)和IFN-α,一样有明显抑毒效果,LPS MDP联合使用对以上病毒有明显增强抑毒作用。CSF-1和IFN-α的抑毒作用能分别被CSF-1和IFN-α的抗体所解除。 CSF-1在人胚肺传代细胞和人胚皮肤肌肉传代细胞上对VSV的抑毒效果在人胚肺细胞中效果比人胚皮肤肌肉细胞更明显。LPS10ng/ml作用48小时比作用24小时效果更强。LPS MDP和IFN-α对二种细胞都有同样高效的抑毒作用。     

Synthesis and biological evaluation of novel homochiral carbocyclic nucleosides from 1-amino-2-indanols

New chiral purinyl and 8-azapurinyl carbanucleoside derivatives based on indanol were synthesized from commercial available (1S,2S)-trans-1-amino-2-indanol and (1R,2R)-trans-1-amino-2-indanol using a linear methodology. The antiviral activity and cytotoxicity of these compounds were evaluated against herpes simplex virus type 1 (HSV-1) in Vero cells, bovine viral diarrhea virus (BVDV) in Mardin-Darby bovine kidney (MDBK) cells and hepatitis B virus (HBV) in HepG2 2.2.15 cell line. Three compounds, showed an inhibition of the HBsAg levels similar to reference drug lamivudine. One chloropurinyl nucleoside, derived from the cis-1-amino-2-indanol, was cytotoxic on MDBK cells and it could be a lead for developing anticancer agents.