Characterization of Frog Virus 3 knockout mutants lacking putative virulence genes

Area of Science:

• Virology
• Molecular Biology
• Pathogenesis

Background:

• Ranaviruses, such as Frog Virus 3 (FV3), are significant pathogens affecting amphibians.
• Understanding the genetic basis of ranavirus virulence is crucial for developing control strategies.

Purpose of the Study:

• To identify specific genes within FV3 that contribute to its virulence.
• To investigate the roles of viral caspase activation and recruitment domain-containing (CARD) protein (64R) and β-hydroxysteroid dehydrogenase homolog (52L) in FV3 pathogenesis.

Main Methods:

• Engineered knockout (KO) mutants of FV3 lacking specific genes: Δ64R-FV3 (vCARD) and Δ52L-FV3 (vβ-HSD).
• Compared the virulence, replication, and host response of KO mutants (including FV3-Δ18K and FV3-ΔvIF-2α) against wild-type (WT) FV3 in Xenopus tadpoles and cell lines.
• Assessed viral replication in amphibian (Xenopus A6 kidney) and non-amphibian cell lines.
• Evaluated sensitivity to interferon and induction of apoptosis.

Main Results:

• Δ64R-FV3 and Δ52L-FV3 mutants showed significantly reduced mortality and viral replication in Xenopus tadpoles compared to WT FV3.
• Replication of ΔvCARD-, ΔvβHSD-, and ΔvIF-2α-FV3 was markedly reduced in Xenopus A6 kidney cells, but not in non-amphibian cell lines.
• Δ64R- and ΔvIF-2α-FV3 mutants were more sensitive to interferon than WT FV3.
• Δ64R-, Δ18K-, and ΔvIF-2α-FV3 mutants triggered more apoptosis than WT FV3.

Conclusions:

• The viral CARD protein (64R) and β-HSD homolog (52L) genes are important contributors to FV3 pathogenesis.
• These genes play roles in viral replication in amphibian cells, immune evasion (interferon sensitivity), and modulation of host cell apoptosis.