Delayed nuclear localization of CRISPR/Cas9-modified fiber of fowl adenovirus serotype 8b reduces pathogenicity in Specific pathogen-free chicken embryonic liver cells.

Fowl adenovirus (FAdV) poses incessant outbreaks to poultry production worldwide, and Inclusion body hepatitis (IBH) is a predominant FAdV infectious disease. Currently, limited vaccines are available in Malaysia to fight against the local predominant FAdV strain 8b isolate (FAdV-8b), posing a desperate demand for efficient vaccine development. The fiber protein of FAdV is one of the major constituents of the adenoviral capsid involved in the virulence of pathogens. Hence, the aim was to modify the fiber gene of FAdV-8b UPMT27 to develop a live attenuated FAdV vaccine via the gene-editing CRISPR/Cas9 technology. Primary specific pathogen-free (SPF) chicken embryonic liver cells (CELs) infected with the modified isolated (cfUPMT27) were reported with significantly reduced cytopathic effects, delayed viral localization into the nucleus, and low apoptotic rates. cfUPMT27 isolate also exhibited constant amino acid substitution of Y179D in subsequent passages. Meanwhile, the liver of cfUPMT27 inoculated-SPF chicken embryonic eggs (CEE) was observed with mild hydropericardium and reported with a delayed mortality at 6-days post-infection (dpi). This holistic, integrative study incorporating genetic, pathology, and immunology analysis proposed cfUPMT27 isolate as a candidate vaccine for FAdV infections, providing efficient future protection in chickens.