Temporal expression profiling of novel Spodoptera litura nucleopolyhedrovirus-encoded microRNAs upon infection of Sf21 cells.

Abstract

Baculoviruses are arthropod-specific pathogens, and find extensive applications in pest control strategies and recombinant protein expression. Spodoptera litura nucleopolyhedrovirus (SpltNPV) infects the tropical armyworm Spodoptera litura, which is an important polyphagous crop pest widely distributed in regions of Asia and Oceania. Using next-generation sequencing, we report stage-specific profiling of SpltNPV-encoded microRNAs (miRNAs) at different time intervals post-infection (p.i.) of Sf21 cells. Sequence length distribution analysis of the small RNA libraries revealed a significant increase in 20 nt reads and a reduction of other size fractions during late phases of infection. In silico miRNA prediction tools identified 48 novel SpltNPV-encoded miRNAs, of which 10 were validated experimentally in Sf21 cells using Northern blot analysis and TaqMan quantitative real-time (qRT)-PCR. The viral miRNAs were also found to be expressed in fat-body and mid-gut tissues of infected fifth-instar S. litura larva. qRT-PCR analysis confirmed that expression of most viral miRNAs was triggered 12 h p.i. and continued thereafter. Gene Ontology and KEGG (Kyoto Encyclopedia of Genes and Genomes) pathway annotation of computationally predicted targets of the reported miRNAs suggested a major impact of these miRNAs on cell signalling, protein translation and metabolic processes.