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. 2017 Jul 19;7:5816. doi: 10.1038/s41598-017-04449-3

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© The Author(s) 2017

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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66E2 inhibits hepatitis E virus replication. (A) P6Luc expressing Huh7 cells were treated 66E2 and CMC. At 48 h post-infection guassia luciferase secreted into the medium was monitored and plotted as % RLU. DMSO treated Huh7 cells were used as control. (B) Cytotoxicity of the compounds was tested and the values are depicted as percentages with the DMSO treated cells taken as 100%. (C) Reverse-transcriptase quantitative PCR analysis was performed to determine the levels of HEV replication in the p6luc transfected Huh7 cells in the presence of 66E2 and CMC. Results are expressed as the percentage relative to DMSO treated control. (D and E) S10-3 cells were infected with p6 virus in the presence of 66E2 and CMC along with DMSO as vehicle control. After 48 h, total RNA was extracted and reverse-transcriptase quantitative PCR analysis was performed to determine the levels of positive (D) and negative sense (E) p6 RNA. Results are expressed as % relative to DMSO treated control and are representative of two independent assays. All the assays in this figure were performed in triplicates and results presented are representative of three independent assays. (F) RdRp assay using purified HEV RdRp. RNA harboring 130 bases from 5′ end and 210 bases from 3′ end of HEV genome was used as a template. The amount of product formed is quantified and given below as percent synthesis relative to the sample treated with DMSO. The data presented are representative of three independent experiments.