The interferons (IFNs) are a large family of multifunctional secreted protein involved in antiviral defense, cell growth regulation, and immune activation. The human IFNs are used worldwide as antiviral drugs. Here, we present cDNAs encoding 13 novel feline IFN-omega (FeIFN-omega) subtypes that share 95%-99% amino acid sequence identity. FeIFN-omega2 and FeIFN-omega4 have seven additional amino acids at position 109 that are not present in other subtypes. Sequence identity of the present FeIFN proteins encoded by the 13 subtypes is approximately 57% compared with human IFN-omega (HuIFN-omega). All 13 FeIFN-omega subtypes were expressed in Escherichia coli using a periplasmic expression system. The antiviral activity of each product was evaluated in vitro. In addition, subtype FeIFN-omega2 was cytoplasm expressed in E. coli and secretion expressed in Pichia pastoris. The purified mature recombinant protein demonstrated significant antiviral activity on both homologous and heterologous animal cells in vitro.