Chimeric Dengue 2 PDK-53/West Nile NY99 Viruses Retain the Phenotypic Attenuation Markers of the Candidate PDK-53 Vaccine Virus and Protect Mice against Lethal Challenge with West Nile Virus

Abstract

West Nile (WN) virus, a member of the Flavivirus genus, is a mosquito-borne virus of the Japanese encephalitis (JE) serocomplex. The JE serocomplex contains viruses that cause central nervous system infections, such as JE virus in Asia; St. Louis encephalitis virus in the Americas; Rocio virus in Brazil; Murray Valley encephalitis virus in Australia, New Guinea, and New Zealand; and Kunjin virus (reclassified as subtype WN recently) in Australia (44). Before the mid 1990s, WN virus caused sporadic outbreaks of illness, ranging from fever to occasional encephalitis, in Africa, the Middle East, and Western Asia. However, since 1996, WN encephalitis in humans has been reported more frequently in Europe, the Middle East, northern and western Africa, and Russia (33). In 1999, WN virus first emerged in the western hemisphere in New York City and surrounding areas, where the virus caused the deaths of seven humans and numerous birds and horses (15,33). Since then, WN virus has spread throughout most of the continental United States, with more than 4,156 reported human cases and 284 deaths in 2002 (37) and 9,862 cases with 264 deaths in 2003 (12). WN virus activity in humans, birds, and horses has been documented in Canada, the Caribbean, and Central America. The rapid spread of WN virus suggests it may pose a significant public health problem in future years (15). There is no licensed human WN vaccine available to protect at-risk populations from WN illness.Dengue (DEN) viruses are also human pathogens that are transmitted by mosquitoes. These viruses cause illness in millions of people every year throughout tropical regions of the world. Flaviviruses of the DEN serocomplex are classified into four serotypes, DEN 1 to DEN 4 (D1 to D4). Flaviviruses contain a single-stranded positive-sense genomic RNA of approximately 11 kb with the genomic organization 5ЈNCR-CprM-E-NS1-NS2A-NS2B-NS3-NS4A-NS4B-NS5-3ЈNCR (where NCR is noncoding region, C is capsid, prM is premembrane, E is envelope, and NS is nonstructural protein). One of the most promising D2 vaccine candidates, strain PDK-53, was derived by passage of the wild-type D2 16681 virus 53 times in primary dog kidney (PDK) cells (48). To study the attenuation loci of the candidate D2 PDK-53 vaccine virus, as well as to develop effective tetravalent DEN vaccines against all four DEN serotypes, we constructed infectious cDNA clones of the D2 viruses (9, 26) and used them to engineer chimeric DEN viruses containing the prM-E genes of D1, D3, or D4 virus in the D2 genetic backbones (18,19). The uncloned D2 PDK-53 vaccine virus contains a mixture of two genotypic variants (26), designated 19) in this report. The PDK53-V variant contains all nine PDK-53 virus vaccine-specific nucleotide mutations, including the Glu-to-Val mutation at amino acid position NS3-250 (26). The PDK53-E variant contains eight of the nine mutations of the PDK-53 vaccine, and the NS3-250-Glu of its parental D2 16681 virus. Our results showed that the phenotypic markers associated with the attenuation of PDK...

Immunization of Flavivirus West Nile Recombinant Envelope Domain III Protein Induced Specific Immune Response and Protection against West Nile Virus Infection

“…With no specific treatment against WNV infection, precedence is given to developing an effective vaccine against it. A number of WNV vaccine candidates had been evaluated in animal models and approved for usage on horses (17)(18)(19)(20)(21)(22). These include an inactivated WNV horse vaccine and a recombinant vaccine using the canarypox virus to express WNV Ags (18,19).…”

“…These include an inactivated WNV horse vaccine and a recombinant vaccine using the canarypox virus to express WNV Ags (18,19). Another study used a chimeric live virus incorporating the NY99 WNV prM (precursor membrane protein), and the E gene inserted into the infectious clone backbone of the yellow fever or dengue virus serotype 4 have been constructed and serotype 4 and is now pending evaluation in clinical trials (21)(22). However, the risk of immune enhancement of heterologous flavivirus infection with inactivated virus and the legitimate safety concerns with the chimeric flaviviruses are highly debatable.…”

Identification of Mutated Cyclization Sequences That Permit Efficient Replication of West Nile Virus Genomes: Use in Safer Propagation of a Novel Vaccine Candidate

“…Live-attenuated vaccines (LAV) for yellow fever and Japanese encephalitis are in widespread use, and LAV for other flavivirus diseases, including dengue and West Nile encephalitis, are in different stages of development (7,8,16,18,21,29,34,35,40). LAV offer considerable advantage over inactivated viral vaccines or subunit vaccines in terms of efficacy and cost.…”

Monoclonal Antibody-Based Competitive Enzyme-Linked Immunosorbent Assay for Detecting and Quantifying West Nile Virus-Neutralizing Antibodies in Horse Sera

“…Neutralizing antibodies induced by domain III of flaviviral E proteins have been reported elsewhere as a major factor responsible for the greatest protection in vivo (24,29), and all types of recently developed bioengineered vaccines target the E protein (1,9,10,12,15,20,27,28,33,35). In our assay, competition for binding of MAb 5E8 by serum antibodies was observed in most of the serum samples from vaccinated horses (96.1%) as well as in serum samples from early infected mice.…”

“…Various types of vaccines for WNV have been explored for their ability to protect susceptible hosts against pathogenic WNV infection: formalin-inactivated (18,22), live attenuated (37), and recombinant chimeric virus vaccines (1,10,15,20,27); recombinant PrM/E or E protein vaccines (28,34); and DNA-based vaccines (9,12,33). Currently, a formalin-inactivated WNV vaccine (West Nile-Innovator; Fort Dodge Animal Health, IA) and a recombinant canarypox virus vector-based vaccine expressing PrM/E proteins of WNV (Recombitek; Merial Limited, GA) are commercially available for veterinary use in the United States (23).…”