Virology & Mycology
Open Access

Antiviral Activity of a Marine Seaweed Tubinaria ornata (turner) J. Agardh (phaeophyceae) Against Herpes Simplex Virus ii (HSV-II)

Poonam Sethi^{* *}Correspondence: Poonam Sethi, Department of Plant Biology and Plant Biotechnology, Guru Nanak College (Autonomous), Chennai, India, Email: ,

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Introduction

Plants yield many bio medically useful substances. Apart from land plants, the lotic and lentic sources have also served as an outburst of a large group of structurally unique natural products of significance. Uniformly accepted standards for in vitro susceptibility testing are not available for antiviral drugs. Antiviral resistance is another critical aspect of clinical importance. [1].This paper also describes the inhibitory activity of dextran sulphate a standard agent against the viruses. Over the past three decades, the importance of marine-derived physiologically active substances effective on mammalian tissues and on pathogenic microbes affecting the mammalian system has received much attention leading to the development of ‘marine biomedical research [2].

Materials and Methods

Turbinaria ornata (Turner) J. Agardh a genus of Phaeophyceae, Phaeophyta, was selected as the experimental algae, collected from the rocky shores of Rameswaram, Tamil Nadu. The collection was made in the month of November 2018, authenticated by Dr. R.Thevanathan Dean of Sciences, Presidency College, Chennai.

Botanical description of the experimental plant

Plants erect and stiff, 20-30 cm long when reproductive, usually isolated or in small groups, often rusty brown to dark brown; holdfast bearing one (or more) terete erect portion, basally a conical or irregular holdfast with several unbranched or dichotomously branched stolons, these often remaining when erect portion torn off, or appearing before erect portion formed.

Macroscopical characters of the experimental plant

The plant body consists of branched cylindrical axis and terminal clusters of funnel shaped expanded bodies and the surface of the plant body is smooth and even (Figure1).

Figure 1: (A-G) Test alga and vero cell line cultures.

A- Turbinaria ornata, J. Agardh; B- Vero cell line in culture with HSV-2 showing 25% cytopathic effect; C- Vero cell line in culture with HSV-2 showing 50% cyto pathic effect; D- Vero cell line in culture with HSV-2 showing 75% cyto pathic effect; E- Vero cell line in culture with HSV-2 showing 100%cyto pathic effect; F-Vero cell line at cytotoxic concentrations of aqueous extract residue; G-Vero cell line at cytotoxic concentrations of ethanolic extract residue

Extract preparation

The extract was prepared by dissolving ten grams of the finely chopped experimental plant in 100.0 mL of double distilled, Millipore filtered water and kept in a shaker. After 48 hrs, it was filtered through cheese cloth and the filtrate was freeze dried- Aqueous extract. Similarly the ethanolic extracts were prepared by dissolving ten grams of the finely chopped experimental plant in 100.0 mL of 90% ethanol. From the stock solution having the residue with distilled water of varying concentrations of the extract residue was used for antiviral studies.

Cell line used for in vitro assay

VERO cell lines and the virus were, obtained from KIPM, Kings Institute of Preventive Medicine, Guindy, Chennai and maintained in MEM containing 5% fetal calf serum, kept in incubator at 37°C.

Virus

Herpes Simplex Virus HSV-2 [HSV-2/P6/Hep. 2 dt 10/12/01] Standard strains of HSV-2 P6/ Hep2 dt 10/12/01 of TCID50 107.1 were obtained from the Department of Virology, King Institute of Preventive Medicine, Chennai. Vero cell lines were used for the culture and routine maintenance of this human pathogenic virus.

Cytotoxicity assay

As a prerequisite for the studies on the antiviral potential of these extract residues their cytotoxicity was studied on the Vero cell line used for the culture of the (HSV-2) .The non-cytotoxic levels of the different extracts were used for the antiviral assay. Antiviral assay

Vero cells were seeded onto 96 well plate at a concentration of 1.0 × 105 cells per ml and a volume of 90 μl per well. The minimal active concentrations which had activity against the viruses was selected and subjected to 100 μl of 100 TCID50 of the virus which was added to each well.

The plates were incubated at 37ºC in a 5% CO2 atmosphere and observed. In the Pretreatment assay the extract was added to the cell line before the subjection of the virus [3]. while in post treatment assay the extracts were subjected to the cell line after the addition of the virus. Dextran sulphate, a sulfated polysaccharide was simultaneously used as a positive control in all the experiments because of its known broad spectrum antiviral properties [4].

Results

The various concentrations of the extracts ranging from 1 μg, 10 μg, 100 μg and 500 μg upto 1000 μg were subjected to cytotoxicity studies on the vero cell line which acts as a host for the virus HSV-2 as shown in Tables1-4.

Table 1: Cytotoxicity of the preparations of turbinaria ornata on vero cell line.

Table 2: Cytotoxicity of extracts and dextran sulphate.

Table 3: IC50 Values for the extract residues of Turbinaria ornata against HSV-2.

Table 4: Cytotoxicity of extracts and dextran sulphate.

The aqueous extract residues of the algae exhibited good activity against HSV-2 virus. The IC50 values for the aqueous extract residue of Turbinaria ornata were 100 μg/mL that of ethanolic was 250 μg/mL. Nevertheless, the two fractions of the experimental algae appeared to be effective. The observations made against HSV-2 indicated that the alga was effective against the virus, irrespective of the nature of the extract used. Dextran sulphate, a sulfated polysaccharide had IC50 value of 50 μg/mL. After the cytotoxicity testing the Optical Density value were measured in an ELISA plate.

Discussion

Both the aqueous and ethanolic extracts were found to be active against HSV-2. Further for combating drug resistant viral strain, it has been suggested that the new antiherpes drugs should be non-nucleosides. Many efforts have been carried out to screen for antiviral agents from natural sources and Marine algae are promising as important sources of antiviral agents [5,6].

Conclusion

Earlier works also showed that the aqueous extracts from five out of eight species of Hong Kong seaweeds exhibited some potential antiviral activities and very low cytotoxicity. Thus Turbinaria could be a promising source of natural products with anti-herpetic effect at non-toxic concentrations, just like seaweeds from other geographical regions.

References

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