Alteration of amino acids in VP2 of very virulent infectious bursal disease virus results in tissue culture adaptation and attenuation in chickens

Reverse genetics technology offers the possibility to study the influence of particular amino acids of infectious bursa[ disease virus (IBDV) on adaptation to tissue culture. Genomic segments A and B of the very virulent (vv) IBDV field strain UK661 were completely cloned and sequenced, and the strain was rescued from full-length cDNA copies of both segments (UK661 rev). Using site-directed mutagenesis, alteration of a single amino acid in the segment A-encoded VP2 (A284T) resulted in a limited capacity of UK661 to replicate in tissue culture. Additional alteration of a second amino acid (Q253H) increased replication efficiency in tissue culture. The second mutant (UK661-Q253H-A284T) was used to infect chickens and results were compared with UK661 and UK661 rev. Whereas UK661 and UK661 rev induced 100% morbidity and 50-80% mortality, UK661-Q253H-A284T proved to be strikingly attenuated, producing neither morbidity nor mortality. Moreover, UK661 -Q253H-A284T-infected animals were protected from challenge infection. Thus, alteration of two specific amino acids in the VP2 region of IBDV resulted in tissue culture adaptation and attenuation in chickens of vvIBDV. The data demonstrate that VP2 plays a decisive role in pathogenicity of IBDV