Endogenous viral elements and PIWI gene evolution in Spodoptera: diversity, conservation, and piRNA production

Endogenous viral elements (EVEs) are viral sequences that have integrated into host genomes. While these elements can be considered molecular fossils of past infections, in some cases, they are co-opted and contribute to host fitness. For instance, in arthropods, some EVEs derived from non-retro-transcribing RNA viruses (nrEVEs) generate PIWI-interacting RNAs (piRNAs), and in Aedes mosquitoes at least, they act as immune effectors. Here, we investigated the diversity and function of nrEVEs across four Spodoptera species (Lepidoptera: Noctuidae), which are notorious agricultural pests. Comparative genomic analyses revealed 21-27 nrEVEs per genome, with a strong enrichment on the Z chromosome. Most nrEVEs originated from (-)ssRNA viruses, particularly Rhabdoviridae, Orthomyxoviridae, Nyamiviridae, and Phenuiviridae. We identified species-specific integrations as well as a conserved orthologous nrEVE likely dating back ∼15 million years. Small RNA sequencing showed that only a subset of nrEVEs produced piRNAs, primarily in ovarian tissues, with a minority displaying sequence complementarity compatible with antiviral activity. Specifically, in Spodoptera frugiperda, we detected abundant nrEVE-derived piRNAs with high similarity to a currently known circulating virus, the Spodoptera frugiperda rhabdovirus (SfRV). Among them, we observed a piRNA peak with seed complementarity to the SfRV genome that correlated with the absence of infection. In parallel, analysis of PIWI-subfamily Argonaute genes, which codify for the proteins involved in the piRNA biogenesis, revealed lineage-specific expansions and signatures of positive selection in Spodoptera, exhibiting a distinct evolutionary trajectory compared to other lepidopteran families. Our results broaden our understanding of insect-virus coevolution suggesting a dual role for nrEVEs in noctuid moths: while most likely persist as genomic fossils, a subset displays features compatible with potential antiviral response through piRNA production.