Evaluation of chickens infected with a recombinant virulent NDV clone expressing chicken IL4

Authors: MARCANO, VALERIE - University Of Georgia; SUSTA, LEORNARDO - University Of Guelph; DIEL, DIEGO - Cornell University; CARDENA-GARCIA, STIVALIS - University Of Georgia; MILLER, PATTI - Retired ARS Employee; AFONSO, CLAUDIO - Retired ARS Employee; BROWN, CORRIE - University Of Georgia

Submitted to: Microbial Pathogenesis
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 7/25/2021
Publication Date: 7/31/2021
Citation: Marcana, V., Susta, L., Diel, D.G., Cardena-Garcia, S., Miller, P.J., Afonso, C.L., Brown, C.C. 2021. Evaluation of chickens infected with a recombinant virulent NDV clone expressing chicken IL4. Microbial Pathogenesis. 159:105116. https://doi.org/10.1016/j.micpath.2021.105116.
DOI: https://doi.org/10.1016/j.micpath.2021.105116

Interpretive Summary: Virulent Newcastle disease virus is a serious concern for poultry health because the virus is highly infectious and causes high mortality in susceptible species. The improved understanding of host responses in the pathogenesis of the disease are needed as we look for improved ways to control the virus. In an effort to look at important cytokines, chemicals that the host produces that are important in the immune response, the chicken interleuken 4 (chIL4) gene was inserted into a virulent Newcastle disease virus to have the virus express the IL4 during replication. When this virus was used to infect chickens, the virus still caused a severe disease with high mortality. In depth studies showed some affect on the lymphoid tissues, but disease and mortality was similar to the birds infected with the virulent virus that didn't express the chIL4 gene. Further work needs to be done to understand the immune response with virulent Newcastle disease viruses.

Technical Abstract: There is evidence that chicken IL4 (chIL4) functions similarly to its mammalian analogue byenhancing type 2 T helper (Th2) humoral immunity and promoting protection against parasitic infections;however, no studies have been performed to assess the effect of chIL4 on the pathogenesis of Newcastle disease(ND). To assess the role of chIL4 in velogenic NDV pathogenesis we created a vNDV infectious clone expressingchIL4. We hypothesized that co-expression of chIL4 during virus replication would result in decreased inflam-mation caused by the Th1 response and thereby increasing survival to challenge with vNDV.Methods: To evaluate the effect of chIL4 during early infection with velogenic Newcastle disease virus (NDV) inchickens, recombinant NDV clones expressing either chIL4 (rZJ1-IL4) or a control expressing green fluorescentprotein (rZJ1-GFP) were created by inserting an expression cassette in an intergenic region of the NDV genome.The pathogenesis of rZJ1-IL4 was assessed in 4-week-old specific pathogen free chickens. The extent of virusreplication was evaluated by titration in mucosal secretions and immunohistochemistry in multiple tissues.Expression of chIL4 was confirmed in tissues using immunohistochemistry.Results: Infection of birds with the rZJ1-IL4 resulted in successful viral replication in vivo and in vitro andgeneration of the chIL4 in tissues. All birds were clinically normal 2 DPI, with one bird in each group showingconjunctival swelling and enlarged spleens grossly. At 5 DPI, moderate or severe depression was observed inbirds infected with rZJ1-GFP or rZJ1-IL4, respectively. Neurological signs and thymic atrophy were observed inone bird infected with rZJ1-IL4. Grossly, conjunctival swelling, mottled spleen and proventricular hemorrhageswere observed at 5 DPI in one bird from each group. At 5 DPI, severe necrosis in the spleen, bursa and cecaltonsils were observed in birds infected with rZJ1-GFP, along with minimal evidence of chIL4 expression. Incontrast, splenic atrophy, and moderate necrosis in the bursa and cecal tonsils were observed in birds infectedwith rZJ1-IL4. In addition, chIL4 signal was found in all tissues of rZJ1-IL4 birds at 5DPI.Conclusions: The production of chIL4 by a recombinant NDV strain resulted in the activation of the positivefeedback loop associated with IL4 production. Insertion of chIL4 into NDV may decrease necrosis to lymphoidorgans while increasing the severity of lymphoid atrophy and prolonged disease. However, with a low number ofbirds it is difficult to determine whether these results are significant to disease outcome.