ABSTRACT
Among the flaviviruses, dengue, with its four serotypes, has spread throughout the tropics. The most advanced vaccines developed so far include live attenuated viruses, which have been tested in humans but none has been licensed. Preclinical testing of dengue vaccine candidates is performed initially in mice and in nonhuman primates. In the latter the main criteria used to assay protection are neutralizing antibodies elicited by the vaccine candidate and the magnitude and duration of peripheral viremia upon challenge of previously immunized animals. Towards the identification of wild-type viruses that could be used in challenge experiments a total of 31 rhesus monkeys were inoculated subcutaneously of wild dengue types 1, 2, and 3 viruses. The viremia caused by the different viruses was variable but it was possible to identify dengue viruses useful as challenge strains.
Subject(s)
Humans , Animals , Male , Female , Dengue Virus/classification , Dengue Virus/pathogenicity , Viremia/virology , Chlorocebus aethiops , Disease Models, Animal , Macaca mulatta/virology , Vero Cells/virologyABSTRACT
A chimeric yellow fever (YF)-dengue serotype 2 (dengue 2) virus was constructed by replacing the premembrane and envelope genes of the YF 17D virus with those from dengue 2 virus strains of Southeast Asian genotype. The virus grew to high titers in Vero cells and, after passage 2, was used for immunogenicity and attenuation studies in rhesus monkeys. Subcutaneous immunization of naive rhesus monkeys with the 17D-D2 chimeric virus induced a neutralizing antibody response associated with the protection of 6 of 7 monkeys against viremia by wild-type dengue 2 virus. Neutralizing antibody titers to dengue 2 were significantly lower in YF-immune animals than in YF-naive monkeys and protection against challenge with wild-type dengue 2 virus was observed in only 2 of 11 YF-immune monkeys. An anamnestic response to dengue 2, indicated by a sharp increase of neutralizing antibody titers, was observed in the majority of the monkeys after challenge with wild-type virus. Virus attenuation was demonstrated using the standard monkey neurovirulence test. The 17D-D2 chimera caused significantly fewer histological lesions than the YF 17DD virus. The attenuated phenotype could also be inferred from the limited viremias compared to the YF 17DD vaccine. Overall, these results provide further support for the use of chimeric viruses for the development of a new live tetravalent dengue vaccine.
Subject(s)
Animals , Male , Female , Antibodies, Viral/biosynthesis , Viremia/immunology , Dengue Virus/immunology , Yellow fever virus/immunology , Amino Acid Sequence , Antibodies, Viral/immunology , Chlorocebus aethiops , Macaca mulatta , Molecular Sequence Data , Neutralization Tests , Recombination, Genetic , Reverse Transcriptase Polymerase Chain Reaction , Vero Cells , Dengue Virus/genetics , Yellow fever virus/geneticsABSTRACT
Several specied of non-human primates have been used in studies on experimental infection with hepatitis A virus (HAV). Attempts to infect a South-American marmoset (Callithrix jacchus) with a Brazilian HAV isolate (HAF-203) are described here. Four seronegative animals were inoculated intragastrically and one was sacrificed on day 11,20,47 and 62 after infection. One uninfected animal was included as control. Liver, small intestine, lymph node, spleen and kidney samples were collected for histological diagnosis and immunocytochemistry studies. Alanine aminotransferase (ALT) and aspartate aminotransferase (AST) serum enzymes and anti-HAV antibodies were monitored by a colorimetric procedure (Abbott) and an enzyme immunoassay (ELISA), respectively. Feces were collected daily for HAV antigen (HAVAg) detection by ELISA. Increased levels of HAVAg were detected in hepatocytes 11 days after infection, with a gradual decrease during the course of infection. Shedding of HAVAg in feces was observed from the late incubation to the early acute phase (20th day to 47th day after infection). The end of the incubation period was indicated by the initial increases in serum ALT and AST. Severe hepatic lesions such as piecemeal necrosis and bridging necrosis were detected during the acute phase, coinciding with the maximum transaminase levels and the appearance of anti-HAV antibodies. On the 62nd day (convalescent phase), the hepatic tissue showed evidence of regeneration and the transaminase values had returned to baselines. The serological, biochemical, antigenic and histological evidence of hepatitis A was similar to that observed in several primate models inoculated with other HAV isolates. The data suggest that C. jacchus can be a valuable model for the study of hepatitis A and for the evaluation of HAV vaccines