We developed a cell culture system and characterized HEV particles; we identified 3 ORF2 capsid proteins (ORF2i, ORF2g, and ORFc). These findings will advance our understanding of the HEV life cycle and improve diagnosis.

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Hepatitis C virus (HCV) envelope glycoprotein complex is composed of E1 and E2 subunits. E2 is the receptor-binding protein as well as the major target of neutralizing antibodies, whereas the functions of E1 remain poorly defined. Here, we took advantage of the recently published structure of the N-terminal region of the E1 ectodomain to interrogate the functions of this glycoprotein by mutating residues within this 79-amino-acid region in the context of an infectious clone. The phenotypes of the mutants were characterized to determine the effects of the mutations on virus entry, replication, and assembly. Furthermore, biochemical approaches were also used to characterize the folding and assembly of E1E2 heterodimers. Thirteen out of 19 mutations led to viral attenuation or inactivation. Interestingly, two attenuated mutants, T213A and I262A, were less dependent on claudin-1 for cellular entry in Huh-7 cells. Instead, these viruses relied on claudin-6, indicating a shift in receptor dependence for these two mutants in the target cell line. An unexpected phenotype was also observed for mutant D263A which was no longer infectious but still showed a good level of core protein secretion. Furthermore, genomic RNA was absent from these noninfectious viral particles, indicating that the D263A mutation leads to the assembly and release of viral particles devoid of genomic RNA. Finally, a change in subcellular colocalization between HCV RNA and E1 was observed for the D263A mutant. This unique observation highlights for the first time cross talk between HCV glycoprotein E1 and the genomic RNA during HCV morphogenesis. IMPORTANCE Hepatitis C virus (HCV) infection is a major public health problem worldwide. It encodes two envelope proteins, E1 and E2, which play a major role in the life cycle of this virus. E2 has been extensively characterized, whereas E1 remains poorly understood. Here, we investigated E1 functions by using site-directed mutagenesis in the context of the viral life cycle. Our results identify unique phenotypes. Unexpectedly, two mutants clearly showed a shift in receptor dependence for cell entry, highlighting a role for E1 in modulating HCV particle interaction with a cellular receptor(s). More importantly, another mutant led to the assembly and release of viral particles devoid of genomic RNA. This unique phenotype was further characterized, and we observed a change in subcellular colocalization between HCV RNA and E1. This unique observation highlights for the first time cross talk between a viral envelope protein and genomic RNA during morphogenesis.

The recent emergence and re-emergence of viral infections transmitted by vectors, such as the Zika virus (ZIKV) and Dengue virus (DENV), is a cause for international concern. These highly pathogenic arboviruses represent a serious health burden in tropical and subtropical areas of the world. Despite the high morbidity and mortality associated with these viral infections, antiviral therapies are missing. Medicinal plants have been widely used to treat various infectious diseases since millenaries. Several compounds extracted from plants exhibit potent effects against viruses in vitro, calling for further investigations regarding their efficacy as antiviral drugs. Here, we demonstrate that an extract from Psiloxylon mauritianum, an endemic medicinal plant from Reunion Island, inhibits the infection of ZIKV in vitro without exhibiting cytotoxic effects. The extract was active against different ZIKV African and Asian strains, including an epidemic one. Time-of-drug-addition assays revealed that the P. mauritianum extract interfered with the attachment of the viral particles to the host cells. Importantly, the P. mauritianum extract was also able to prevent the infection of human cells by four dengue virus serotypes. Due to its potency and ability to target ZIKV and DENV particles, P. mauritianum may be of value for identifying and characterizing antiviral compounds to fight medically-important flaviviruses.

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Zika virus (ZIKV) and Dengue virus (DENV) are mosquito-borne viruses of the Flavivirus genus that could cause congenital microcephaly and hemorrhage, respectively, in humans, and thus present a risk to global public health. A preventive vaccine against ZIKV remains unavailable, and no specific antiviral drugs against ZIKV and DENV are licensed. Medicinal plants may be a source of natural antiviral drugs which mostly target viral entry. In this study, we evaluate the antiviral activity of Doratoxylum apetalum, an indigenous medicinal plant from the Mascarene Islands, against ZIKV and DENV infection. Our data indicated that D. apetalum exhibited potent antiviral activity against a contemporary epidemic strain of ZIKV and clinical isolates of four DENV serotypes at non-cytotoxic concentrations in human cells. Time-of-drug-addition assays revealed that D. apetalum extract acts on ZIKV entry by preventing the internalisation of virus particles into the host cells. Our data suggest that D. apetalum-mediated ZIKV inhibition relates to virus particle inactivation. We suggest that D. apetalum could be a promising natural source for the development of potential antivirals against medically important flaviviruses.

Zika virus (ZIKV) is an emerging mosquito-borne virus of medical concern. ZIKV infection may represent a serious disease, causing neonatal microcephaly and neurological disorders. Nowadays, there is no approved antiviral against ZIKV. Several indigenous or endemic medicinal plants from Mascarene archipelago in Indian Ocean have been found able to inhibit ZIKV infection. The purpose of our study was to determine whether essential oil (EO) from Reunion Island medicinal plant Ayapana triplinervis, whose thymohydroquinone dimethyl ether (THQ) is the main component has the potential to prevent ZIKV infection in human cells. Virological assays were performed on human epithelial A549 cells infected with either GFP reporter ZIKV or epidemic viral strain. Zebrafish assay was employed to evaluate the acute toxicity of THQ in vivo. We showed that both EO and THQ inhibit ZIKV infection in human cells with IC50 values of 38 and 45 µg/mL, respectively. At the noncytotoxic concentrations, EO and THQ reduced virus progeny production by 3-log. Time-of-drug-addition assays revealed that THQ could act as viral entry inhibitor. At the antiviral effective concentration, THQ injection in zebrafish does not lead to any signs of stress and does not impact fish survival, demonstrating the absence of acute toxicity for THQ. From our data, we propose that THQ is a new potent antiviral phytocompound against ZIKV, supporting the potential use of medicinal plants from Reunion Island as a source of natural and safe antiviral substances against medically important mosquito-borne viruses.

The mosquito-borne viruses dengue (DENV) and Zika (ZIKV) viruses are two medically important pathogens in tropical and subtropical regions of the world. There is an urgent need of therapeutics against DENV and ZIKV, and medicinal plants are considered as a promising source of antiviral bioactive metabolites. In the present study, we evaluated the ability of Phyllanthus phillyreifolius, an endemic medicinal plant from Reunion Island, to prevent DENV and ZIKV infection in human cells. At non-cytotoxic concentration in vitro, incubation of infected A549 cells with a P. phillyreifolius extract or its major active phytochemical geraniin resulted in a dramatic reduction of virus progeny production for ZIKV as well as four serotypes of DENV. Virological assays showed that P. phillyreifolius extract-mediated virus inhibition relates to a blockade in internalization of virus particles into the host cell. Infectivity studies on ZIKV showed that both P. phillyreifolius and geraniin cause a loss of infectivity of the viral particles. Using a zebrafish model, we demonstrated that administration of P. phillyreifolius and geraniin has no effect on zebrafish locomotor activity while no morbidity nor mortality was observed up to 5 days post-inoculation. Thus, P. phillyreifolius could act as an important source of plant metabolite geraniin which is a promising antiviral compound in the fight against DENV and ZIKV.