Synergy between Aedes aegypti trypsin modulating oostatic factor and δ-endotoxins

Studies that were conducted at the University of Florida, FMEL and at the Ben-Gurion University of the Negev showed that starved first instar Aedes aegypti larvae are 35-fold more sensitive to Bacillus thuringiensis subsp. israelensis (Bti) toxins than fed larvae. Feeding larvae Pichia pastoris yeast cells expressing tmfA (synthetic gene coding for the Trypsin Modulating Oostatic Factor of Ae. aegypti) together with Escherichia coli cells expressing Bti toxin genes (cry4Aa, cry11Aa, cyt1Aa and p20) indicate that TMOF and Cry toxins are synergisitic. Codon-optimized synthetic genes, cry11-tmfA, gst-cry11-tmfA, cry4Aa-tmfA, gst-cry4Aa-tmfA and tmfA that were cloned into P. pastoris and fed to Ae. aegypti larvae caused rapid larval mortality. Cell free extracts of recombinant P. pastoris cells killed 40% of tested 4th instar larvae within 24 h, and mass spectra analysis confirmed that the recombinants synthesize Cry11Aa and Cry4Aa. This report shows for the first time that Cry toxins and TMOF are synergists to Ae. aegypti larvae when jointly fed or expressed in recombinant P. pastoris.