Differential transcription of fathead minnow immune-related genes following infection with frog virus 3, an emerging pathogen of ectothermic vertebrates

Abstract

Frog virus 3 (FV3) and other ranaviruses are responsible for die-offs involving wild, farmed, and captive amphibians, fish, and reptiles. To ascertain which elements of the immune system respond to infection, we explored transcriptional responses following infection of fathead minnow cells with either wild type (wt) FV3 or a knock out (KO) mutant targeting the 18 kDa immediate early gene (18K). At 8 hr post infection we observed marked upregulation of multiple transcripts encoding proteins affecting innate and acquired immunity. Sequences expressed 4-fold or higher in wt-infected cells included transcripts encoding interferon (IFN), IFN regulatory factors (IRFs), IFN stimulated genes (ISGs) such as Mx and MHC class I, and interleukins IL-1β, IL-8, IL-17C and IL-12. Cells infected with the 18K KO mutant (Δ18K) showed qualitative differences and lower levels of induction. Collectively, these results indicate that ranavirus infection induced expression of multiple cellular genes affecting both innate and acquired immunity.

Xenopus-FV3 host-pathogen interactions and immune evasion

“…Using the technique described above we generated FV3 knock-out (KO) recombinants for 64R and 52L (Andino Fde et al, 2015). Because of its robust immediate early expression pattern and dispensable role for productive infection in cell culture (Cheng et al, 2014; Sample et al, 2007), the FV3 ICP18 or 18K (82R) gene was also selected as a virulence gene candidate and knocked out (Chen et al, 2011). Evidence from initial characterization of two FV3 KO viruses (Δ64R-, and Δ52L-FV3) is consistent with their involvement in immune evasion, whereas results obtained with Δ18-FV3 suggest a more complex role of the 18K gene.…”

Ranavirus Replication: Molecular, Cellular, and Immunological Events

“…Consistent with this suggestion, Cheng et al ( 2014 ) recently examined the response of FHM cells to infection with either wild type FV3 or a knock out mutant lacking the ranavirus vIF-2α and observed the induction of multiple immune-response genes at the transcriptional level including IFN, IL-8, GILT, IRF-3. Likewise, vaccination of groupers with inactivated SGIV induced the expression of numerous immune-related genes including Mx1, ISG15, IL-8, IL-1β, and MHC I/II indicating that the immune response is conserved among different fi sh species and similar to that seen in mammals (Ou-yang et al 2012 ).…”

“…It is proposed that the JNK pathway is important for SGIV replication and modulates the infl ammatory responses during virus infection ). Interestingly, activation of similar genes was seen following infection of FHM cells with FV3 (Cheng et al 2014 ).…”

Rana grylio virus TK and DUT gene locus could be simultaneously used for foreign gene expression

“…Briefly, selection markers were inserted into the locus of a target gene through homologous recombination. With the help of selection markers, like resistance to drugs or fluorescence, knockout mutants were generated through successively purification (Chen et al, 2011;Cheng et al, 2014). Finally, gene functions were inferred by changes in viral replication and virulence.…”