Akira Kotaki

Two cases of Zika fever imported from French Polynesia to Japan, December 2013 to January 2014

We present two cases of imported Zika fever to Japan, in travellers returning from French Polynesia, where an outbreak due to Zika virus (ZIKV) is ongoing since week 41 of 2013. This report serves to raise awareness among healthcare professionals, that the differential diagnosis of febrile and subfebrile patients with rash should include ZIKV infection, especially in patients returning from areas affected by this virus.

Autochthonous dengue fever, Tokyo, Japan, 2014.

After 70 years with no confirmed autochthonous cases of dengue fever in Japan, 19 cases were reported during August–September 2014. Dengue virus serotype 1 was detected in 18 patients. Phylogenetic analysis of the envelope protein genome sequence from 3 patients revealed 100% identity with the strain from the first patient (2014) in Japan.

Zika fever imported from Thailand to Japan, and diagnosed by PCR in the urines.

In July 2014, a Japanese traveller returning from Thailand was investigated for fever, headache, rash and conjunctivitis. Zika virus RNA was detected in his urine sample by real-time reverse transcription polymerase chain reaction. Serological tests showed cross reactivity of IgM against the dengue virus. Zika fever could be misdiagnosed or missed and should be considered in febrile patients with a rash, especially those returning from Thailand.

Detection of Dengue Virus Genome in Urine by Real-Time Reverse Transcriptase PCR: a Laboratory Diagnostic Method Useful after Disappearance of the Genome in Serum

ABSTRACT The reemergence of dengue virus (DENV) infection has created a requirement for improved laboratory diagnostic procedures. In this study, DENV genome detection in urine was evaluated as a diagnostic method. The DENV genome was detected by real-time reverse transcriptase PCR (RT-PCR) in urine and serum of dengue patients. The detection rate of DENV genome in urine was 25% (2/8) on disease days 0 to 3 and 32% (7/22) on days 4 to 5. The rate was 50% or higher on days 6 to 16, 52% (11/21) on days 6 to 7, 78% (7/9) on days 8 to 9, 80% (4/5) on days 10 to 11, 50% (2/4) on days 12 to 13, and 60% (3/5) on days 14 to 16. The last positive urine sample was on day 16. The detection rates in serum were highest on days 0 to 3 and were greater than 50% on days 0 to 7. Detection rates decreased thereafter, and the last positive detection was on day 11. These results indicate that the time frames for positive detection differ between urine and serum samples, whereby detection rates of 50% or higher are evident between days 6 to 16 for urine samples and days 0 to 7 for serum samples. Nucleotide sequences of PCR products were identical between urine and serum samples. The detection of DENV genome in urine samples by real-time RT-PCR is useful to confirm DENV infection, particularly after viremia disappears.

Importation of dengue virus type 3 to Japan from Tanzania and Côte d'Ivoire.

Travelers can introduce viruses from disease-endemic to non–disease-endemic areas. Serologic and virologic tests confirmed dengue virus infections in 3 travelers returning to Japan: 2 from Tanzania and 1 from Côte d’Ivoire. Phylogenetic analysis of the envelope gene showed that 2 genetically related virus isolates belonged to dengue virus type 3 genotype III.

Detection of Higher Levels of Dengue Viremia Using FcγR-Expressing BHK-21 Cells Than FcγR-Negative Cells in Secondary Infection but Not in Primary Infection

It has been reported that levels of viremia reflect the severity of illness in dengue virus infection. We assessed the levels of viremia in patients with primary and secondary infections, using 2 cell lines: FcγR-expressing BHK cells and FcγR-negative cells. In primary infection, virus titers were at similar levels between FcγR-expressing and FcγR-negative cells. In secondary infection, however, virus titers were ∼10 times higher in FcγR-expressing cells on days 1-6 when compared with FcγR-negative cells, indicating discrepancy in viremia titers between FcγR-expressing and FcγR-negative cells. The results suggest that dengue virus-antibody complexes with infectious capacity exist in patients with secondary infection, and these immune complexes can be detected by using FcγR-expressing cells. As it has been reported that principal target cells of dengue virus infection are FcγR-positive, monocyte/macrophage lineage cells, virus titers determined using FcγR-expressing cells may better reflect the actual viremic conditions in vivo.

Vero cell-derived inactivated West Nile (WN) vaccine induces protective immunity against lethal WN virus infection in mice and shows a facilitated neutralizing antibody response in mice previously immunized with Japanese encephalitis vaccine.

A novel Vero cell-derived inactivated WN vaccine (WN-VAX) was prepared from virus strain NY99-35262. Two immunizations with WN-VAX induced high levels of neutralizing antibody to WN virus. All immunized mice were protected against challenge with a lethal dose of WN virus. No WN viremia was detected, and the level of WN virus-neutralizing antibody increased rapidly. WN-VAX was then examined for immunogenicity in mice previously immunized with Japanese encephalitis vaccine (JE-VAX). Immunization with WN-VAX induced WN virus-neutralizing antibody in all mice previously immunized with JE-VAX but in only half of the control mice at 10 weeks. These results indicate that WN-VAX induced complete protective immunity against lethal WN infection and that the WN-VAX-induced antibody response is facilitated in JE-VAX-immunized mice. This WN-VAX is thus a candidate WN vaccine for humans.

Development of an antibody-dependent enhancement assay for dengue virus using stable BHK-21 cell lines expressing FcγRIIA

Dengue virus (DENV) causes a wide range of symptoms, from mild febrile illness, dengue fever (DF), to severe life threatening illness, dengue hemorrhagic fever (DHF) and dengue shock syndrome (DSS). Subneutralizing concentrations of antibody to DENV enhance DENV infection in Fc gammaR positive cells. This phenomenon is known as antibody-dependent enhancement (ADE). ADE is considered to be a risk factor for DHF and DSS. To develop an ADE assay for DENV, two stable BHK-21 cell lines were established that express Fc gammaRIIA (BHK-Fc gammaRIIA). The BHK-Fc gammaRIIA cell lines were used in an ADE assay with monoclonal antibody (4G2) to DENV, and DENV antibody-positive human sera. Virus growth was quantified directly in BHK-Fc gammaRIIA cells with a standard plaque assay procedure. ADE was detected with monoclonal antibody (4G2) to DENV. ADE was also detected with DENV antibody-positive human sera, but not with DENV antibody-negative human sera. The new ADE assay using BHK-Fc gammaRIIA cells is simple and practical, and is useful for defining the role of ADE in the pathogenesis of DENV infection.

Novel Dengue Virus Type 1 from Travelers to Yap State, Micronesia

Dengue virus type 1 (DENV-1), which was responsible for the dengue fever outbreak in Yap State, Micronesia, in 2004, was isolated from serum samples of 4 dengue patients in Japan. Genome sequencing demonstrated that this virus belonged to genotype IV and had a 29-nucleotide deletion in the 3´ noncoding region.

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.