No abstract

BackgroundThe mosquito transmitted Dengue virus (DENV) remains a significant public health problem in many tropical and subtropical countries. Increasing evidence has suggested that during the infection process cellular lipids play important roles at several stages of the replication cycle. This study sought to characterize the changes in lipid metabolism gene expression and investigated the role of one enzyme, fatty acid synthase, in DENV infection.MethodsTranscriptional profiles of genes associated with lipid metabolism were evaluated by real-time PCR after infection of different cell lines (HepG2 and HEK293T/17) and with different DENVs (laboratory adapted and low passage). Expression profiles of genes were evaluated by western blotting. A critical lipid metabolism protein, fatty acid synthase was down-regulated through siRNA and inhibited with orlistat and the effect on DENV infection determined by flow cytometry, plaque assay, western blotting and confocal microscopy.ResultsThe results showed alterations of gene transcription and expression were seen in genes variously associated with lipogenesis, lipolysis and fatty acid β-oxidation during DENV infection. Interference of fatty acid synthase with either siRNA or orlistat had marked effects on virus production, with orlistat having an EC50 value of 10.07 μM at 24 h post infection. However, non-structural protein expression was largely unaffected.ConclusionsWhile drug treatment reduced virus titer by up to 3Log10, no significant effect on DENV non-structural protein expression was observed, suggesting that fatty acid synthase acts through an effect on virion formation.Electronic supplementary materialThe online version of this article (doi:10.1186/s12985-017-0685-9) contains supplementary material, which is available to authorized users.

Many mosquito transmitted viruses of the genera Alphavirus and Flavivirus are human pathogens of significant concern, and there is currently no specific antiviral for any member of these two genera. This study sought to investigate the broad utility of orlistat (tetrahydrolipstatin) in reducing virus infection for several mosquito borne viruses including flaviviruses (dengue virus (DENV; nine isolates analyzed), Japanese encephalitis virus (JEV; one isolate analyzed) and Zika virus (ZIKV; 2 isolates analyzed)) as well as an alphavirus (chikungunya virus; CHIKV; 2 isolates analyzed). Three different treatment regimens were evaluated, namely pre-treatment (only), post-treatment (only) and pre-and post-treatment, and three factors were evaluated, namely level of infection, virus titer and genome copy number. Results showed that all three treatment modalities were able to significantly reduce virus titer for all viruses investigated, with the exception of three isolates of DENV in the pre-treatment only regimen. Pre-and post-treatment was more effective in reducing the level of infection and genome copy number of all viruses investigated than either pre-treatment or post-treatment alone. collectively, these results suggest orlistat has potential as a broad-spectrum agent against multiple mosquito transmitted viruses.Mosquito transmitted viruses are a significant public health problem in many tropical and sub-tropical countries, and some of the most significant human pathogens belong to the genera Flavivirus and Alphavirus 1 . The genus Flavivirus consists of 53 virus species 2 of which more than half are transmitted by mosquitoes and the majority of these have the potential to infect humans 3 . Medically important mosquito transmitted viruses in the genus Flavivirus include dengue virus (DENV), Japanese encephalitis virus (JEV), Zika virus (ZIKV) and yellow fever virus (YFV). The genus Alphavirus consists of 31 virus species 2 the majority of which are spread by mosquitoes, and medically important alphaviruses include chikungunya virus (CHIKV), Ross River virus, Semliki Forest Virus and Sindbis virus 4 .Viruses in the genera Flavivirus and Alphavirus have a number of similarities. Viruses in both genera are classified in group IV in the Baltimore classification system 5 as they possess a positive sense single stranded RNA genome. The genome sizes are approximately equivalent (flaviviruses approximately 9.2-11 kb 6 , alphaviruses approximately 9.7-12 kb 7 , but while the ten flavivirus proteins (capsid (C), pre-membrane (prM), envelope (E), NS1, NS2A, NS2B, NS3, NS4A, NS4B and NS5) are encoded by a single open reading frame, the nine alphavirus proteins are encoded by two open reading frames, the first of which encodes the non-structural proteins (nsP1, nsP2, nsP3 and nsP4), while the second open reading frame encodes the structural proteins (C, E1, E2, E3), as well as a protein (6 K) of uncertain function 4 . Viruses in both genera encode a protein with RNA-dependent RNA polymerase (RdRP) activity that undertakes gen...

Between 2018 and 2019, the incidence of chikungunya was approximately 15,000 cases across 60 provinces in Thailand. Here, the clinical presentations in chikungunya, emergent pattern, and genomic diversity of the chikungunya virus (CHIKV) causing this massive outbreak were demonstrated. A total of 1,806 sera samples from suspected cases of chikungunya were collected from 13 provinces in Thailand, and samples were tested for the presence of CHIKV RNA, IgG, and IgM using real-time PCR, enzyme-linked immunoassay (ELISA), commercial immunoassay (rapid test). The phylogenetic tree of CHIKV whole-genome and CHIKV E1 were constructed using the maximum-likelihood method. CHIKV infection was confirmed in 547 (42.2%) male and 748 (57.8%) female patients by positive real-time PCR results and/or CHIKV IgM antibody titers. Unsurprisingly, CHIKV RNA was detected in >80% of confirmed cases between 1 and 5 days after symptom onset, whereas anti-CHIKV IgM was detectable in >90% of cases after day 6. Older age was clearly one of the risk factors for the development of arthralgia in infected patients. Although phylogenetic analysis revealed that the present CHIKV Thailand strain of 2018–2020 belongs to the East, Central, and Southern African (ECSA) genotype similar to the CHIKV strains that caused outbreaks during 2008–2009 and 2013, all present CHIKV Thailand strains were clustered within the recent CHIKV strain that caused an outbreak in South Asia. Interestingly, all present CHIKV Thailand strains possess two mutations, E1-K211E, and E2-V264A, in the background of E1-226A. These mutations are reported to be associated with virus-adapted Aedes aegypti. Taken together, it was likely that the present CHIKV outbreak in Thailand occurred as a result of the importation of the CHIKV strain from South Asia. Understanding with viral genetic diversity is essential for epidemiological study and may contribute to better disease management and preventive measures.

BackgroundA number of studies have implicated the direct involvement of the liver in dengue virus (DENV) infection, and it has been widely shown that liver cells subsequently undergo apoptosis. The mechanism by which liver cells undergo apoptosis in response to DENV infection remains unclear. To provide further information on the mechanism of apoptosis in DENV infected liver cells, HepG2 cells were infected with DENV 2 and analyzed for the induction of ER stress, apoptosis and autophagy.ResultsIn response to DENV infection, HepG2 cells showed the induction of both the ER resident unfolded protein response as well as the Noxa/PUMA stress response pathways. Proteolytic activation of caspases 4, 7, 8 and 9 was observed as well as changes in mitochondrial transmembrane potential. Increased monodansylcadaverine staining was observed in DENV infected cells, consistent with the previously reported induction of autophagy.ConclusionsThese results are consistent with a model in which the induction of multiple ER stress pathways is coupled with the induction of multiple cell death pathways as a mechanism to ensure the removal of infected liver cells from the system.