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Vaccination of chickens with recombinant Salmonella expressing M2e and CD154 epitopes increases protection and decreases viral shedding after low pathogenic avian influenza challenge

Layton, S.L., Kapczynski, D.R., Higgins, S., Higgins, J., Wolfenden, A.D., Liljebjelke, K.A., Bottje, W.G., Swayne, D., Berghman, L.R., Kwon, Y.M., Hargis, B.M., Cole, K.
Poultry science 2009 v.88 no.11 pp. 2244
chickens, Influenza A virus, avian influenza, vaccination, viral antigens, recombinant antigens, recombinant vaccines, Salmonella enteritidis, polymerase chain reaction, nucleotide sequences, membrane proteins, oral administration, microbial colonization, immune response, liver, spleen, tonsils, immunoglobulin G, neutralization tests, strains, pathogenicity
Avian influenza (AI) is a significant public health concern and serious economic threat to the commercial poultry industry worldwide. Previous research demonstrates that antibodies against M2e confer protection against influenza challenge. Using the Red recombinase system in combination with overlapping extension PCR, we recently developed several novel attenuated Salmonella Enteritidis strains that express a protective M2e epitope in combination with a potential immune-enhancing CD154 peptide sequence on the Salmonella outer membrane protein lamB. Commercial Leghorn chicks were orally immunized (immunization dose: 10⁶ to 10⁸ cfu/chick) with saline (negative control) or one of the recombinant Salmonella strains [ΔaroA M2e-CD154, ΔhtrA M2e-CD154, ΔaroA-ΔhtrA M2e(4)-CD154] on day of hatch and 21 d posthatch. These candidate vaccine strains were evaluated for their ability to invade, colonize, and persist in tissues and elicit an M2e-specific antibody response. The vaccine candidate strain ΔaroA M2e-CD154 exhibited significantly greater organ invasion in the liver and spleen at d 7 (P > 0.05); however, no marked differences in colonization of the cecal tonsils were observed. Vaccinated chickens exhibited significantly increased M2e-specific IgG responses, which were further enhanced by simultaneous expression of CD154 (P < 0.05). Virus neutralization assays gave neutralizing indices of 6.6, 6.3, and 6.3 for ΔaroA M2e-CD154, ΔhtrA M2e-CD154, and ΔaroA-ΔhtrA M2e(4)-CD154 seven days post booster immunization, respectively, indicating effective neutralization of AI by serum IgG of vaccinated chickens. In a subsequent direct challenge study, specific-pathogen-free Leghorn chicks immunized with ΔaroA-ΔhtrA M2e(4)-CD154 offered significant protection against direct challenge with low pathogenic AI H7N2, but not highly pathogenic H5N1 AI. Taken together, these data suggest that these Salmonella-vectored vaccines expressing M2e in association with CD154 are effective at protecting chickens against low pathogenic AI.