Production of yellow fever 17DD vaccine virus in primary culture of chicken embryo fibroblasts: yields, thermo and genetic stability, attenuation and immunogenicity

Abstract

While a good vaccine against yellow fever (YF) virus has been available for decades, the basic technology for the production of YF vaccine in chicken embryos has remained substantially unchanged since the 1940s. Here we describe the highly efficient and economic production of the 17DD strain of YF virus in chicken embryo fibroblast (CEF) cell cultures with viral titers ranging from 6.3 to 6.7 log10PFU/mL. Thermostability of two different formulations (5 and 50-dose vials) of the CEF vaccine virus was found to be as high as the current vaccines retaining the minimal titer required for YF 17D vaccines. The production passage in CEF did not lead to the selection of genetic variants as shown by nucleotide sequence analyses of the CEF-derived vaccine lots or the sequence of viruses recovered from monkeys experimentally inoculated with the CEF virus. YF 17DD virus produced in CEF was also indistinguishable from its seed lot virus parent in terms of plaque size and immunogenicity in mice and monkeys. Comparison of the CEF virus and the seed lot virus made in chicken embryo in the internationally accepted monkey neurovirulence test (MNVT) revealed a higher clinical score for the former. The differences in central nervous system (CNS) histological scores for monkeys inoculated with the chicken embryo and experimental CEF vaccines were at the borderline level of statistical significance. These data warrant further studies on the monkey attenuation of other batches of CEF-derived vaccines.

Current status and future prospects of yellow fever vaccines

“…The adaptive capacity of YFV has been investigated since the early development of all YFV vaccines, with multiple studies recognizing that passage history could reverse the attenuation or immunogenicity of the vaccine [76, 77]. A 17DD substrain seed was adapted to chick embryo fibroblasts (CEF), to permit growth of the vaccine lot by microcarrier bead culture technology [78]. The CEF-prepared vaccine lot was tested using the standard WHO monkey neurovirulence test; the authors noted elevated neurotropism scores in the monkeys tested with respect to a conventionality prepared seed, although these results were not statistically significant.…”

Subdoses of 17DD yellow fever vaccine elicit equivalent virological/immunological kinetics timeline

“…Thus, the spreading of risk areas and the restricted group of YF vaccine manufacturers creates a shortage on YF vaccine supply worldwide, which urges for new strategies of vaccination protocols including validation of new seed lots, need and timing of booster doses to maintain long lasting protection as well as dose sparing studies [8]. In regards to dose, the minimal number of viral particles has been established by WHO as at least 5,000PFU or approximately 3,000 IU.…”

Study of the genetic stability of measles virus CAM-70 vaccine strain after serial passages in chicken embryo fibroblasts primary cultures

“…The remaining tissue was washed with PBS, extruded and treated with trypsin to give a monodisperse cell suspension [17]. Cells were then seeded in roller bottles in 199 Hank's salts medium with fetal bovine serum, as previously described [18]. The seed virus FMS-7 was inoculated following the procedure of Freire and co-workers [18] using an incubation temperature of 30°C.…”

“…Cells were then seeded in roller bottles in 199 Hank's salts medium with fetal bovine serum, as previously described [18]. The seed virus FMS-7 was inoculated following the procedure of Freire and co-workers [18] using an incubation temperature of 30°C. The seed virus was passaged 10 times in CEF and the viruses from the passage levels used in this study were termed VSF-1 to VSF-10.…”