In vitro evaluation of marine-microorganism extracts for anti-viral activity

Abstract

Viral-induced infectious diseases represent a major health threat and their control remains an unachieved goal, due in part to the limited availability of effective anti-viral drugs and measures. The use of natural products in drug manufacturing is an ancient and well-established practice. Marine organisms are known producers of pharmacological and anti-viral agents. In this study, a total of 20 extracts from marine microorganisms were evaluated for their antiviral activity. These extracts were tested against two mammalian viruses, herpes simplex virus (HSV-1) and vesicular stomatitis virus (VSV), using Vero cells as the cell culture system, and two marine virus counterparts, channel catfish virus (CCV) and snakehead rhabdovirus (SHRV), in their respective cell cultures (CCO and EPC). Evaluation of these extracts demonstrated that some possess antiviral potential. In sum, extracts 162M(4), 258M(1), 298M(4), 313(2), 331M(2), 367M(1) and 397(1) appear to be effective broad-spectrum antivirals with potential uses as prophylactic agents to prevent infection, as evident by their highly inhibitive effects against both virus types. Extract 313(2) shows the most potential in that it showed significantly high inhibition across all tested viruses. The samples tested in this study were crude extracts; therefore the development of antiviral application of the few potential extracts is dependent on future studies focused on the isolation of the active elements contained in these extracts.

Figure 1

Representation of viral attachment/entry inhibition by marine extracts. Viruses (VSV) were pre-incubated with test extract (100 μg/ml). Plates (Vero cells) were infected for one hour, after which plates were allowed to incubate for 24-36 hrs, until adequate plaques were observed. Plates were stained with crystal violet staining and pictures were taken. Plaques were counted and inhibition was determined relative to controls. Row 1: Extract 397(1), showing marked plaque reduction (≥ 80%) relative to the controls; Row 2: Extract 312(2), showing marked plaque reduction (≥ 90%) relative to the controls; Row 3: 338M(1), showing no marked plaque reduction (< 20%) relative to the controls; Row 4: Control of 0.1% DMSO.

Figure 2

Representation of viral replication inhibition by marine extracts. Cells (CCO) were seeded into TC-12.5 cm² flasks and then infected with virus (CCV) at an MOI of 0.1. Following a 1-hr incubation, media was completely removed and infected cultures were subsequently incubated for approximately 3 days with 2.5 ml/flask of media containing extracts (100 μg/ml). Pictures were taken to track the progression of viral-induced CPE. As shown, pictures were taken at 72 and 84 hours post-infection. Extracts 397(1) and 397M(1) show > 90% viral inhibition, under the parameters of the experiment, relative to the control. Extract 162M(4) shows no inhibition relative to the control.