Assessment of grapefruit expressing anti-NodT antibody for huanglongbing resistance

Growers rely on broad-spectrum agrochemicals to manage one of the most economically important fruit tree diseases, huanglongbing (HLB, citrus greening disease), presumptively caused by the gram-negative bacterium ‘Candidatus Liberibacter asiaticus’ (CLas). While genetic resistance would be an attractive alternative to chemical management, this option is not yet available for HLB. Here, we tested whether a single chain variable fragment (scFv) antibody targeting the predicted CLas outer membrane transporter NodT can inhibit CLas growth or HLB disease development when expressed in ‘Duncan’ grapefruit (Citrus x paradisi). CLas NodT is similar to TolC and could be involved in Type I secretion and virulence. The scFv antibody was expressed in grapefruit trees as a C-terminal translational fusion to the Flowering Locus T protein of Poncirus trifoliata (FT-scFv) to promote expression in the phloem. Wild type and FT-scFv-expressing transgenic lines were challenged with CLas using three inoculation approaches: psyllid-mediated inoculation, vegetative graft inoculation, and natural exposure in grove-like conditions. With the first two approaches, HLB symptom expression and CLas bacterial titers in FT-scFv grapefruit lines were not significantly different from wild-type controls. In the grove trial, one FT-scFv line exhibited a slight, but significant, reduction in canopy chlorosis compared to wild-type controls. Wild type and FT-scFv lines were similar in all other metrics. We conclude that the expression of anti-NodT FT-scFv antibody in grapefruit did not decrease HLB susceptibility. Although our approach was unsuccessful, we hope that documenting our results will be useful for those seeking to develop HLB-resistant citrus germplasm in the future.