Generation of Yellow Fever virus vaccine in skeletal muscle cells of chicken embryos

Abstract

BACKGROUND The Yellow Fever (YF) vaccine is produced by the inoculation of embryonated chicken eggs with YF17DD virus on the ninth day of development. Full embryos are collected on the twelfth day of development for vaccine formulation. Skeletal muscle tissue is the main site where biosynthesis of viral particles occurs.OBJECTIVES This study aimed to analyse the experimental infection of skeletal muscle cells of chicken embryos by the 17DD Yellow Fever virus (YFV) in vivo and in vitro.METHODS Chicken embryos infected with YF17DD virus were analysed by immunofluorescence using confocal and super-resolution microscopes. Primary cultures of skeletal muscle cells of non-infected chicken embryos were evaluated for susceptibility and permissiveness to YF17DD virus using different protocols. This evaluation was performed based on morphological, viral titration, molecular biology, and colorimetric techniques.FINDINGS The present work phenotypically characterises embryonic chicken skeletal muscle cells as myogenic precursors expressing the Pax7 transcription factor in some cases. We demonstrated that these cells are susceptible to in vitro infection at different multiplicities of infection (MOIs), reproducing the same infection pattern observed in vivo. Furthermore, myogenic precursors and myoblasts are preferred infection targets, but establishment of infection does not depend on the presence of these cells. The peak of viral production occurred at 48 hpi, with decay occurring 72 hpi, when the cytopathic effect can be observed.MAIN CONCLUSIONS In conclusion, the primary culture of chicken skeletal muscle cells is a good model for studying muscle cells infected with YF17DD virus. This culture system displays satisfactory emulation of the in vitro phenomenon observed, contributing to our understanding of virus infection dynamics and leading to the development of alternative methods of vaccine production.

Next generation live-attenuated yellow fever vaccine candidate: Safety and immuno-efficacy in small animal models

“…Other YF vaccine candidates produced using cell culture-based methods have included an inactivated YF-17D vaccine produced from Vero cells [6] , and two live-attenuated YF-17DD vaccines produced from chicken embryo fibroblasts and skeletal muscle cells [30] , [31] . The inactivated YF-17D vaccine amplified on Vero cells (though not from a biological clone as in our study) is expected to ameliorate safety concerns associated with the live-attenuated vaccine (i.e.…”

The Present and Future of Yellow Fever Vaccines

“…In the current embryonated chicken egg manufacturing protocol, 17D is primarily propagated in the skeletal muscle tissue [ 121 ]. To better understand this process and to investigate development of an in vitro vaccine manufacturing protocol, primary cultures of chicken embryo skeletal muscle cells were infected with substrain 17DD.…”

Establishment methods and research progress of livestock and poultry immortalized cell lines: A review

“…First, collect fresh muscle tissue samples from a slaughterhouse and transport them to a cell culture laboratory under sterile conditions (5). Small-sized experimental animals such as chicken embryos, whose muscle tissue can also be separated directly on the laboratory sterile bench (6). Then, wash the muscle tissue with 70% ethanol or 1 × phosphate buffered saline (PBS) containing 1% penicillin-streptomycin to remove surface dirt, and cut it into small pieces.…”