Development of a Plaque Reduction Assay as an Antiphage Activity Evaluation Method

Background: Antiviral screening of newly isolated or synthesized compounds is an important matter which requires a reliable antiviral test. In order to address this issue, the development of a rapid antiphage test has been conducted. To achieve this goal, the antiphage activity of three antiviral drugs (Acyclovir, Lamivudine and Trifluridin) against phage CP51 which infects Bacillus cereus (ATCC 10876) was investigated. Materials and Methods: Phage lysate was prepared by inoculation of bacterial culture with few phage plaques. The number of phage has to be about one million units per milliliter. The antiviral drugs were dissolved in suitable solvents and different concentrations of each drug were prepared. Phage lysate (0.1ml) mixed with appropriate amount of each drug. After 30 minutes incubation at ambient temperature or without any incubation, 500 μl inoculum of 5 hours old liquid culture of B.cereus added to the mixture. Then, melted top agar (1.4-1.9 ml) was subjoined at the end and the final admixture was immediately seeded on the solid PA agar. After 24 h, plaques were counted. Results: Out of three drugs, only trifluridine significantly decreased the plaque forming unit ratio (p<0.05) at higher concentration (133 μg/ml). The results of preincubation method and non-incubation method did not show any significant differences (p>0.05). Conclusion: Briefly, the scientific evidences from this study supported the development of one rapid qualitative and quantitative antiphage assay.