Transgenic Sweet Orange Expressing the Sarcotoxin IA Gene Produces High-Quality Fruit and Shows Tolerance to ‘Candidatus Liberibacter asiaticus’

Abstract

Huanglongbing (otherwise known as HLB or greening) is currently the most devastating citrus disease worldwide. HLB is primarily associated with the phloem-inhabiting bacterium '<i>Candidatus</i> Liberibacter asiaticus' (<i>C</i>Las). Currently, there are no citrus species resistant to <i>C</i>Las. Genetic transformation is one of the most effective approaches used to induce resistance against plant diseases. Antimicrobial peptides (AMPs) have shown potential breakthroughs to improve resistance to bacterial diseases in plants. In this paper, we confirm the <i>Agrobacterium</i>-mediated transformation of Pera sweet orange expressing the AMP <i>sarcotoxin IA</i> (<i>stx IA</i>) gene isolated from the flesh fly <i>Sarcophaga peregrina</i> and its reaction to <i>C</i>Las, involving plant performance and fruit quality assessments. Four independent transgenic lines, STX-5, STX-11, STX-12, and STX-13, and a non-transgenic control, were graft-inoculated with <i>C</i>Las. Based on our findings, none of the transgenic plants were immune to <i>C</i>Las. However, the STX-5 and STX-11 lines showed reduced susceptibility to HLB with mild disease symptoms and low incidence of plants with the presence of <i>C</i>Las. Fruit and juice quality were not affected by the genetic transformation. Further, no residues of the sarcotoxin IA protein were found in the juice of the STX-11 and STX-12 fruits, though detected in the juice of the STX-5 and STX-13 lines, as revealed by the immunoblotting test. However, juices from all transgenic lines showed low traces of sarcotoxin IA peptide in its composition. The accumulation of this peptide did not cause any deleterious effects on plants or in fruit/juice. Our findings reinforce the challenges of identifying novel approaches to managing HLB.