Construction of the safe neutralizing assay system using pseudotyped Nipah virus and G protein-specific monoclonal antibody.

Nipah virus (NiV) is a recently emerged paramyxovirus that causes acute respiratory illness and fatal encephalitis in a broad spectrum of vertebrates, including humans. Due to its high pathogenicity and mortality rates, NiV requires handling in biosafety level-4 (BSL-4) containment facilities and no effective vaccines or therapeutic agents are currently available. Since current diagnostic tests for detecting serum neutralizing antibodies against NiV mainly employ live viruses, establishment of more safe and robust alternative diagnostic methods is an essential medical requirement. Here, we have developed a pseudotyped NiV and closely related Hendra virus (HeV) expressing envelope attachment (G) and fusion (F) glycoproteins using the Moloney murine leukemia virus (MuLV) packaging system. We additionally generated polyclonal antibodies (pAbs) against NiV-G and HeV-G and assessed their neutralizing activities for potential utilization in the pseudovirus-based neutralization assay and further application in the serum diagnostic test. To enhance the specificity of neutralizing antibody and sensitivity of the serological diagnostic test, monoclonal antibodies (mAbs) against NiV-G were generated, and among which four out of six mAb clones showed significant reactivity. Specifically, the 7G9 clone displayed the highest sensitivity. The selected mAb clones showed no cross-reactivity with HeV-G and efficient neutralizing activities against pseudotyped NiV. These results validate the safety and specificity of neutralization assays against NiV and HeV and present a useful tool to design effective vaccines and serological diagnosis.

Development and evaluation of a semifield test for repellent efficacy testing.

Estimation of the efficacy of mosquito repellents requires both laboratory and field tests. The results of field tests are more meaningful, but the safety of volunteers in such tests may be a significant concern. In the current study, we compared tests of mosquito repellent efficacy under semifield conditions in an outdoor enclosure with those under laboratory and field conditions. In this study, we assessed the efficacy of N,N-diethyl-meta-toluamide under laboratory conditions with human volunteers and under semifield and field conditions with Centers for Disease Control and Prevention traps and experimental mice. A semifield test may be a suitable replacement for the more difficult field test for assessment of mosquito repellent efficacy. Semifield tests should be considered when developing new guidelines for testing.

Protective efficacy of a human endogenous retrovirus envelope-coated, nonreplicable, baculovirus-based hemagglutin vaccine against pandemic influenza H1N1 2009.

Despite the advantages of DNA vaccines, overcoming their lower efficacy relative to that of conventional vaccines remains a challenge. Here, we constructed a human endogenous retrovirus (HERV) envelope-coated, nonreplicable, baculovirus-based HA vaccine against swine influenza A/California/04/2009(H1N1) hemagglutin (HA) (AcHERV-sH1N1-HA) as an alternative to conventional vaccines and evaluated its efficacy in two strains of mice, BALB/c and C57BL/6. A commercially available, killed virus vaccine was used as a positive control. Mice were intramuscularly administered AcHERV-sH1N1-HA or the commercial vaccine and subsequently given two booster injections. Compared with the commercial vaccine, AcHERV-sH1N1-HA induced significantly higher levels of cellular immune responses in both BALB/c and C57BL/6 mice. Unlike cellular immune responses, humoral immune responses depended on the strain of mice. Following immunization with AcHERV-sH1N1-HA, C57BL/6 mice showed HA-specific IgG titers 10- to 100-fold lower than those of BALB/c mice. In line with the different levels of humoral immune responses, the survival of immunized mice after intranasal challenge with sH1N1 virus (A/California/04/2009) depended on the strain. After challenge with 10-times the median lethal dose (MLD50) of sH1N1 virus, 100% of BALB/c mice immunized with the commercial vaccine or AcHERV-sH1N1-HA survived. In contrast, C57BL/6 mice immunized with AcHERV-sH1N1-HA or the commercial vaccine showed 60% and 70% survival respectively, after challenge with sH1N1 virus. In all mice, virus titers and results of histological analyses of lung tissues were consistent with the survival data. Our results indicate the importance of humoral immune response as a major defense system against influenza viral infection. Moreover, the complete survival of BALB/c mice immunized with AcHERV-sH1N1-HA after challenge with sH1N1 virus suggests the potential of baculoviral vector-based vaccines to achieve an efficacy comparable to that of killed virus vaccines.