NS1’ protein expression in the JaOArS982 strain of Japanese encephalitis virus does not enhance virulence in mice

Using a mouse model,we previously demonstrated that subcutaneous infection with the JaTH160 strainof Japaneseencephalitis virus (JEV) causes significantly higher virulence and strongervirus propagation in the brain compared with that of the JaOArS982strain. We also showed that the JaTH160 strain,but not JaOArS982, expresses the NS1’ protein and that NS1’ enhances JEVproduction in avian cells and embryonated chicken eggs. In this study, weexamined whether NS1’ expression affects virulence in mice infected with theJaOArS982 and JaTH160 strains using the corresponding recombinant viruses S982-ICand JaTH-IC.Expression of the NS1’ protein in S982-IC diminishedthe mortality in mice, whereas S982-IC viruses without NS1’ caused 40% mortality.However, the viral loads in the brains of these mice were not significantlydifferent despite the difference in NS1’ expression. JaTH-IC viruses depletedof the NS1’ protein exhibited high mortality levels, similar to those of thevirus expressing NS1’.Previousstudies showed that the NS1’ protein plays a role in the enhanced virulenceof the JEV SA14 strain in mice. However, ourcurrent data suggest that NS1’ protein expression in S982-IC reduces,rather than enhances, the mortality in mice. Thus, the effect of NS1’ on pathogenicity <i>invivo</i> may vary among virus strains. Our data also suggest that the reducedmortality resulting from NS1’ expression in S982-IC is not simply due to viralreplication in the brains. Furtherinvestigation is needed to uncover the mechanism by which NS1’ affectspathogenicity in JEV-infected animals.

Epidemiological survey of severe fever with thrombocytopenia syndrome virus in ticks in Nagasaki, Japan

Severe fever with thrombocytopenia syndrome (SFTS) is an emerging disease that is endemic in East Asia. The SFTS virus (SFTSV) is transmitted to other organisms by infected ticks and is endemic to Nagasaki in western Japan. However, epidemiological information regarding SFTSV in Nagasaki ticks has not been elucidated. In this study, we first examined the sensitivities of SFTSV gene detection by real-time RT-PCR and virus isolation in cultured cells and mice. These methods could detect SFTSV in the samples containing more than 4 × 10⁰ ffu. Next, we attempted to isolate SFTSV and to detect viral gene in 2,222 nymph and adult ticks collected from May to August 2013 among seven regions of Nagasaki. However, neither virus isolation nor viral gene detection were confirmed in those tick pools. SFTSV positivity rates are considered very low in ticks and viral loads in ticks are also very limited. Further investigation by increasing the number of ticks and including larval samples in the investigation, as well as improved detection methods, may be required to find SFTSV-positive ticks in this region.

NS1’ Protein Expression in the JaOArS982 Strain of Japanese Encephalitis Virus Does Not Enhance Virulence in Mice

Using a mouse model, we previously demonstrated that subcutaneous infection with the JaTH160 strain of Japanese encephalitis virus (JEV) causes significantly higher virulence and stronger virus propagation in the brain compared with that of the JaOArS982 strain. We also showed that the JaTH160 strain, but not JaOArS982, expresses the NS1’ protein and that NS1’ enhances JEV production in avian cells and embryonated chicken eggs. In this study, we examined whether NS1’ expression affects virulence in mice infected with the JaOArS982 and JaTH160 strains using the corresponding recombinant viruses S982-IC and JaTH-IC. Expression of the NS1’ protein in S982-IC diminished the mortality in mice, whereas S982-IC viruses without NS1’ caused 40–60% mortality. However, the viral loads in the brains of these mice were not significantly different despite the dvariation in NS1’ expression. JaTH-IC viruses depleted of the NS1’ protein exhibited high mortality levels, similar to those of the virus expressing NS1’. Previous studies showed that the NS1’ protein plays a role in the enhanced virulence of the JEV SA14 strain in mice. However, our current data suggest that NS1’ protein expression in S982-IC reduces, rather than enhances, the mortality in mice. Thus, the effect of NS1’ on pathogenicity <i>in vivo</i> may vary among virus strains. Our data also suggest that the reduced mortality resulting from NS1’ expression in S982-IC is not simply due to viral replication in the brains. Further investigation is needed to uncover the mechanism by which NS1’ affects pathogenicity in JEV-infected animals.

Epidemiological Survey of Severe Fever with Thrombocytopenia Syndrome Virus in Ticks in Nagasaki, Japan

Severe fever with thrombocytopenia syndrome (SFTS) is an emerging disease endemic in East Asia. Transmitted to other organisms by infected ticks, the SFTS virus (SFTSV) and is endemic to Nagasaki in western Japan. However, epidemiological information regarding SFTSV in Nagasaki ticks has not been available to date. In this study, we began by examining the sensitivities of SFTSV gene detection by real-time RT-PCR and virus isolation in cultured cells and mice. These methods could detect SFTSV in the samples containing more than 4 × 10⁰ ffu. Next, we attempted to isolate SFTSV and to detect viral gene in 2,222 nymph and adult ticks collected from May to August 2013 among seven regions of Nagasaki. However, neither virus isolation nor viral gene detection were confirmed in the tick pools. SFTSV positivity rates are considered to be very low in ticks, and viral loads are also very limited. Further investigations increasing the number of ticks and including larval samples as well as improved detection methods, may be required to find SFTSV-positive ticks in this region.

First Isolation of Dengue Virus from the 2010 Epidemic in Nepal

Dengue is an emerging disease in Nepal and was first observed as an outbreak in nine lowland districts in 2006. In 2010, however, a large epidemic of dengue occurred with 4,529 suspected and 917 serologically-confirmed cases and five deaths reported in government hospitals in Nepal. The collection of demographic information was performed along with an entomological survey and clinical evaluation of the patients. A total of 280 serum samples were collected from suspected dengue patients. These samples were subjected to routine laboratory investigations and IgM-capture ELISA for dengue serological identification, and 160 acute serum samples were used for virus isolation, RT-PCR, sequencing and phylogenetic analysis. The results showed that affected patients were predominately adults, and that 10% of the cases were classified as dengue haemorrhagic fever/ dengue shock syndrome. The genetic characterization of dengue viruses isolated from patients in four major outbreak areas of Nepal suggests that the DENV-1 strain was responsible for the 2010 epidemic. Entomological studies identified <i>Aedes aegypti</i> in all epidemic areas. All viruses belonged to a monophyletic single clade which is phylogenetically close to Indian viruses. The dengue epidemic started in the lowlands and expanded to the highland areas. To our knowledge, this is the first dengue isolation and genetic characterization reported from Nepal.