Abstract
The present work presents an improvement of microalgal antiviral dsRNA production for controlling disease in shrimp aquaculture. 307 bp of sequence targeting the VP28 gene of white spot syndrome virus (WSSV) was inserted between two convergent rrnS promoters in the novel vector p2XTRBL, which was then subcloned into the transformation vector pSS116 using Golden Gate assembly. The recombinant plasmid was transformed into the Chlamydomonas reinhardtii chloroplast, and transformants selected by the restoration of photosynthesis. The presence of the cassette and homoplasmy of the algal transformants was confirmed by PCR analysis. Transcribed sense and antisense VP28-RNA were hypothesised to form an RNA duplex in the chloroplast stroma, and quantitative RT-PCR indicated that ∼100 µg dsRNA was obtained per litre of transgenic microalgae culture. This accumulation of dsRNA represents a 10,000-fold increase relative to previous reports using convergent psaA promoters. Recombinant C. reinhardtii was assessed for its ability to prevent WSSV infection in shrimp larvae by direct feeding. After WSSV challenge, the survival of shrimp treated with dsRNA-expressing C. reinhardtii was significantly enhanced (95.2%) relative to the negative control without dsRNA treatment. The study suggests that this new algal production platform for dsRNA is significantly more efficient than the previous report, and it merits further scale-up and downstream processing studies.
Competing Interest Statement
The authors have declared no competing interest.