Interleukin 12-containing influenza virus-like-particle vaccine elevate its protective activity against heterotypic influenza virus infection.

To produce monovalent and bivalent influenza vaccines composed of virus-like particles (VLPs) containing hemagglutinin (HA), we generated four recombinant Baculoviruses derived from nuclear polyhedrosis virus (BmNPV) and nuclear polyhedrosis virus (AcNPV). Monovalent Fukushima (A/tufted duck/Fukushima/16/2011 [H5N1]) (FkH5) and Anhui (A/Anhui/1/2013 [H7N9]) (AnH7) VLP influenza vaccines were produced in silkworm pupae infected with FkH5-BmNPV or AnH7-BmNPV. To produce a bivalent FkH5 and AnH7 vaccine, the pupae were simultaneously inoculated with FkH5-BmNPV and AnH7-BmNPV.

Overexpression of a virus-like particle influenza vaccine in Eri silkworm pupae, using Autographa californica nuclear polyhedrosis virus and host-range expansion.

Ecological investigations of silkworms have revealed that Eri silkworms (Samia cynthia ricini) possess useful morphological and ecological characteristics for virus-like particle (VLP) production, namely non-seasonal breeding, longer lengths, and heavier weights than Bombyx mori silkworms. Furthermore, when vector DNA from Bombyx mori nuclear polyhedrosis virus (BmNPV), which is unable to replicate in Sf9 cells from Eri silkworms, was replaced with the Autographa californica nuclear polyhedrosis virus (AcNPV) vector, three improved AcNPV influenza virus recombinants capable of replication in Sf9 cells were obtained. Although VLP antigens produced previously in silkworms were not evaluated individually, the present recombinant Fukushima (FkH5) and Anhui (AnH7) VLP antigens were detected in tissue fluids and fat bodies of Eri silkworms.

Functional growth inhibition of influenza A and B viruses by liquid and powder components of leaves from the subtropical plant Melia azedarach L.

We evaluated the anti-influenza-virus effects of Melia components and discuss the utility of these components. The effects of leaf components of Melia azedarach L. on viruses were examined, and plaque inhibition tests were performed.

Development of a Japanese encephalitis virus-like particle vaccine in silkworms using codon-optimised prM and envelope genes.

We have successfully prepared a Japanese encephalitis virus (JEV) - Nakayama virus like particle (NVLP) vaccine using synthetic codon-optimized prM and E genes. The expression of the recombinant JEV Nakayama-BmNPV (JEV-NNPV) virus was determined in infected silkworm Bm-N cells by fluorescence and Western blot analysis. The recombinant was inoculated into silkworm pupae and the yield of Nakayama VLP (NVLP) reached a peak in the homogenates after 3 days.

Quantitative analysis of the yield of avian H7 influenza virus haemagglutinin protein produced in silkworm pupae with the use of the codon-optimized DNA: A possible oral vaccine.

In this study, we aimed to quantitatively compare the increased production of three H7 influenza virus-like particle (VLP) haemagglutinin (HA) with the use of a codon-optimized single HA gene in silkworm pupae. Recombinant baculovirus (Korea H7-BmNPV) could produce 0.40 million HA units per pupa, corresponding to 1832μg protein.

The large-scale production of an artificial influenza virus-like particle vaccine in silkworm pupae.

We successfully established a mass production system for an influenza virus-like particle (VLP) vaccine using a synthetic H5 hemagglutinin (HA) gene codon-optimized for the silkworm. A recombinant baculovirus containing the synthetic gene was inoculated into silkworm pupae. Four days after inoculation, the hemagglutination titer in homogenates from infected pupae reached a mean value of 0.

An amino acid substitution (V3I) in the Japanese encephalitis virus NS4A protein increases its virulence in mice, but not its growth rate in vitro.

Our previous studies have shown that the Japanese encephalitis virus (JEV) strain Mie/40/2004 is the most virulent of the strains isolated by us in Japan from 2002 to 2004. Comparison of the amino acid sequence of Mie/40/2004 with those of low-virulence strains revealed that an isoleucine residue at position 3 of the Mie/40/2004 NS4A protein may increase viral pathogenicity. A recombinant virus with a single valine-to-isoleucine substitution (V3I) at position 3 in the low-virulence Mie/41/2002 background (rJEV-Mie41-NS4A(V3I)) exhibited increased virulence in mice compared with the Mie/41/2002 parent strain.

Molecular and virological analyses of dengue virus responsible for dengue outbreak in East Timor in 2005.

A severe dengue outbreak occurred in East Timor in 2005. The dengue virus genome was detected by TaqMan RT-PCR in 40 serum samples, as follows: dengue virus type-3 (DENV-3) in 37 samples, DENV-2 in 2 samples, and DENV-1 in one sample. One DENV-1 genome, one DENV-2 genome, and 5 DENV-3 genomes were sequenced, and these specimens were aligned with the previously determined envelope (E) gene sequences.

A single mutation in the Japanese encephalitis virus E protein (S123R) increases its growth rate in mouse neuroblastoma cells and its pathogenicity in mice.

We previously reported that the Japanese encephalitis virus (JEV) strain Mie/41/2002 has weak pathogenicity compared with the laboratory strain Beijing-1. To identify the determinants of its growth nature and pathogenicity, we produced intertypic viruses, rJEV(EB1-M41), rJEV(nEB1-M41) and rJEV(cEB1-M41), which contained the entire, the N-terminal, and the C-terminal half, respectively, of the Beijing-1 E region in the Mie/41/2002 background. The growth of rJEV(EB1-M41) in mouse neuroblastoma N18 cells and virulence in mice were similar to those of Beijing-1.

Dengue virus type 2 isolated from an imported dengue patient in Japan: first isolation of dengue virus from Nepal.

We report the isolation of dengue virus type 2 from a dengue patient returning to Japan from Nepal in October, 2004. This is the first isolate of dengue virus in Nepal. According to nucleotide homology, the virus was closest to a dengue virus type 2 isolate from India.

Molecular epidemiological analyses of Japanese encephalitis virus isolates from swine in Japan from 2002 to 2004.

To characterize Japanese encephalitis virus (JEV) strains recently prevalent in Japan, JEV surveillance was performed in pigs from 2002 to 2004. Eleven new JEV isolates were obtained and compared with previous isolates from Japan and other Asian countries. All of the isolates were classified into genotype 1 by nucleotide sequence analysis of the E gene.

Phylogenetic analysis of dengue viruses isolated from imported dengue patients: possible aid for determining the countries where infections occurred.

Background: Molecular epidemiology of dengue viruses in endemic countries have been reported, but few were reported on the imported dengue cases among travelers. We analyzed dengue viruses isolated from imported dengue cases in Japan who were infected while traveling in endemic regions of the world.

Method: We sequenced the complete envelope (E) gene of 33 dengue virus strains isolated from patients returning from Asia, Oceania, South Pacific islands, and South America to Japan where no domestic dengue virus infection occurs.

Rapid genome sequencing of RNA viruses.

We developed a system for rapid determination of viral RNA sequences whereby genomic sequence is obtained from cultured virus isolates without subcloning into plasmid vectors. This method affords new opportunities to address the challenges of unknown or untypeable emerging viruses.

Development and evaluation of fluorogenic TaqMan reverse transcriptase PCR assays for detection of dengue virus types 1 to 4.

The fluorogenic TaqMan reverse transcriptase PCR (RT-PCR) assay was developed for detecting each of the dengue virus (DV) types 1 to 4. DV genome was detected in all the 35 serum samples from confirmed dengue cases by the TaqMan RT-PCR, although it was not detected in 13 and 21% by conventional type-specific and cross-reactive RT-PCR, respectively.

Partial protective effect of inactivated Japanese encephalitis vaccine on lethal West Nile virus infection in mice.

The effect of mouse brain-derived, inactivated Japanese encephalitis (JE) vaccine on West Nile virus (WNV) infection was examined using a murine model. Mice were immunized with JE vaccine twice and challenged with lethal doses of WNV. When mice were intracranially challenged with WNV, none of the immunized mice were protected.