Integration of Insecticidal Protein Vip3Aa1 into Beauveria bassiana Enhances Fungal Virulence to Spodoptera litura Larvae by Cuticle and Per Os Infection

The entomopathogenic fungus Beauveria bassiana acts slowly on insect pests through cuticle infection. Vegetative insecticidal proteins (Vip3A) of Bacillus thuringiensis kill lepidopteran pests rapidly, via per os infection, but their use for pest control is restricted to integration into transgenic plants. A transgenic B. bassiana strain (BbV28) expressing Vip3Aa1 (a Vip3A toxin) was thus created to infect the larvae of the oriental leafworm moth Spodoptera litura through conidial ingestion and cuticle adhesion. Vip3Aa1 (similar to 88 kDa) was highly expressed in the conidial cytoplasm of BbV28 and was detected as a digested form (similar to 62 kDa) in the larval midgut 18 and 36 h after conidial ingestion. The median lethal concentration (LC(50)) of BbV28 against the second-instar larvae feeding on cabbage leaves sprayed with conidial suspensions was 26.2-fold lower than that of the wild-type strain on day 3 and 1.1-fold lower on day 7. The same sprays applied to both larvae and leaves for their feeding reduced the LC(50) of the transformant 17.2- and 1.3-fold on days 3 and 7, respectively. Median lethal times (LT(50)s) of BbV28 were shortened by 23 to 35%, declining with conidial concentrations. The larvae infected by ingestion of BbV28 conidia showed typical symptoms of Vip3A action, i.e., shrinkage and palsy. However, neither LC(50) nor LT(50) trends differed between BbV28 and its parental strain if the infection occurred through the cuticle only. Our findings indicate that fungal conidia can be used as vectors for spreading the highly insecticidal Vip3A protein for control of foliage feeders such as S. litura.