# Control of Aedes albopictus populations by silencing of the vesicular GABA transporter (vgat) and the vesicular monoamine transporter (vmat) genes using recombinant Chlorella shRNA

## TL;DR

This study shows that using genetically modified algae to deliver RNA that silences specific genes in mosquitoes can effectively kill them while being safe for other species.

## Contribution

A novel biocidal method using recombinant Chlorella to silence mosquito genes via oral delivery is demonstrated.

## Key findings

- Recombinant Chlorella expressing shRNA targeting vgat and vmat genes showed high lethality against Aedes albopictus larvae and adults.
- The shRNA-based method had no lethal effects on non-target organisms like Drosophila and tadpoles.
- Gene silencing caused specific behavioral and physiological impairments in mosquitoes, such as reduced sleep and flight issues.

## Abstract

Aedes albopictus is a primary vector for the transmission of dengue fever. RNA interference (RNAi)-based biocidal technology represents an important alternative and complement to conventional chemically synthesized insecticides.

Chlorella vulgaris was used as a carrier organism for RNAi-mediated gene silencing of Ae. albopictus. Short hairpin RNA (shRNA) expression vectors targeting the vgat and vmat genes of Ae. albopictus were constructed and subsequently used to transform C. vulgaris. The shRNA-transformed Chlorella were then administered to Ae. albopictus larvae or incorporated into attractive toxic sugar baits (ATSB) for adult feeding. The effects of silencing the vgat and vmat genes on Ae. albopictus were then investigated.

Both vgat and vmat shRNA-recombinant Chlorella exhibited high lethality against Ae. albopictus larvae and adults. Conversely, shRNA-recombinant Chlorella ATSBs were found to have no lethal effects on non-target organisms, including Drosophila melanogaster, Megalurothrips usitatus, Messor structor, and Lithobates catesbeiana tadpoles. Together, these results confirm the specificity and safety of using shRNA-recombinant Chlorella as a mosquito-killing agent. The results of a semi-field trial demonstrate that recombinant Chlorella ATSB maintained high lethality against Ae. albopictus and significantly reduced the number of eggs laid by this species. Additional experiments revealed that knockdown of the vgat gene in Ae. albopictus resulted in reduced sleep duration, leg twitching, and impaired flight. Conversely, knockdown of the vmat gene increased sleep duration, impaired walking and flight, and induced insensitivity to light levels.

Recombinant Chlorella expressing vgat and vmat shRNAs demonstrated strong lethality, high specificity, and good safety against both larval and adult Ae. albopictus. Oral delivery of shRNAs effectively knocked down mosquito target genes, providing a convenient approach for functional studies of mosquito genetics. Notably, this study is an important addition to the use of recombinant microalgae as a mosquito biocide.

The online version contains supplementary material available at 10.1186/s13071-025-07053-x.