Microbiological Assessment and Evaluation of Virus Yield in Germ-free Chicken Eggs

Increasing yield will have major implications for vaccine manufacturers. Internal bacterial contamination of specific pathogen-free (SPF) eggs results in batches of vaccines being rejected and may negatively impact vaccine yield.

The objectives of this study were to analyze Germ-Free (GF) eggs microbiologically and evaluate the yield of H1N1 when compared with SPF eggs from 2 different suppliers.

The contents of GF eggs were analyzed using three different methods (sterility test, total viable count, and bioluminescence). Eggs from each source were inoculated with H1N1 virus. Chick Embryo Fibroblast (CEF) cell cultures were generated from GF and SPF eggs from both suppliers and infected with H1N1. The supernatant was titrated for viral Haemagglutinin activity and virus infectivity while cellular RNA extracts were assayed for Interferon (α and β) gene expression by RT-qPCR using gene-specific primers.

The analysis of GF eggs confirmed that the contents were sterile. The batch yield of GF eggs was ~1.2-fold and ~1.8-fold higher than SPF supplier 1 and SPF supplier 2 virus yields, respectively. Replication of H1N1 was 2-fold higher in CEF derived from GF eggs than CEF derived from SPF supplier 1 or SPF supplier 2. IFN-β antiviral response of GF-derived CEF cells was 2-fold lower than SPF supplier 1-derived and ~3-fold lower than SPF supplier 2-derived CEF cells.

The content of GF eggs was sterile. The yield of H1N1 was higher in GF eggs than in SPF eggs, and the higher yield was associated with a significant reduction in interferon Beta levels.