Lnc557 promotes Bombyx mori nucleopolyhedrovirus replication by interacting with BmELAVL1 to enhance its stability and expression.

Bombyx mori nucleopolyhedrovirus (BmNPV) is a major pathogen that threatens the growth and sustainability of the sericultural industry. Currently, accumulated studies showed that long non-coding RNAs (lncRNAs) play important roles in the genesis and progression of various viruses and host-pathogens interactions. However, the functions and regulatory mechanisms of lncRNAs in insect-virus interaction are still limited. In this study, transcriptome sequencing and ribosome profiling sequencing (Ribo-seq) were performed in the BmNPV-infected midgut and control tissue, and a total of 9 differentially expressed (DE) lncRNAs and 27 small ORFs (sORFs) with micropeptide coding potential were identified. Among them, lncRNA XR_001139971.3 (lnc557) is verified to be significantly up-regulated upon BmNPV infection and may have the potential to encode a small peptide (ORF-674). The subcellular localization experiment showed that lnc557 was expressed in the cytoplasm. Overexpression of lnc557 promotes BmNPV replication and vice versa. By combining RNA pull-down, mass spectrometry, protein truncation and RNA immunoprecipitation (RIP) assays, we confirmed that lnc557 can bind to the RRM-5 domain of BmELAVL1 protein. Subsequently, we found that lnc557 could promote the expression of BmELAVL1 by enhancing the stability of BmELAVL1. Further, enhancing the expression of BmELAVL1 can promote the proliferation of BmNPV, while knockdown shows the opposite effect. Our data suggest that lnc557-mediated BmELAVL1 expression enhancement could play a positive role in BmNPV replication, which will provide a new insight into the molecular mechanism of interaction between Bombyx mori and virus.

Development of a rapid visual detection technology for BmNPV based on CRISPR/Cas13a system.

Pathogenic microorganism of silkworm are important factors that threaten the high-quality development of sericulture. Among them, Bombyx mori nucleopolyhedrovirus (BmNPV) caused diseases often lead to frequent outbreaks and high mortality, resulting in huge losses to sericultural industry. Current molecular detection methods for BmNPV require expensive equipment and sikilled technical personnel. As a result, the most commonly detection method for silkworm egg production enterprises involves observing the presence of polyhedra under a microscope. However, this method has low accuracy and sensitivity. There is an urgent need to develop a new detection technology with high sensitivity, high specificity, and applicability for silkworm farms, silkworm egg production enterprises and quarantine departments. In this study, we successfully established the CRISPR/Cas13a BmNPV visualized detection technology by combining Recombinase Polymerase Amplification (RPA) technology and CRISPR/Cas13a system. This technology is based on microplate lateral, flow test strips and portable fluorescence detector. The detection sensitivity can reach up to 1 copies/µL for positive standard plasmid and 1 fg/µL for BmNPV genome in 30-45 min, demonstrating high sensitivity. By detecting silkworm tissues infected with different pathogens, we determined that CRISPR/Cas13a detection technology has good specificity. In summary, the newly established nucleic acid detection technology for BmNPV is characterized by high sensitivity, high specificity, low cost and convenience for visualization. It can be applied in field detection and silkworm egg quality monitory system.