Generation of polyclonal antibody for serological detection of Candidatus liberibacter asiaticus, the causal agent of citrus Huanglongbing

Citrus huanglongbing (HLB), caused by Candidatus liberibacter asiaticus (CLas), is a dangerous bacterial plant disease worldwide. Developing a sensitive and reliable detection method-based antibody would be an effective tactic to control HLB. In this work, the serological potential of the outer membrane protein (OMP gene) of CLas was assessed. The OMP gene (878 bp) was positively amplified, cloned, and expressed in E. coli and validated by western blot analysis using 6xHis-tag. To produce polyclonal antibody, purified OMP (34 kDa) was used for immunizing rabbits. The highest titer of antisera and its excellent detection capability were determined by indirect-ELISA. The sensitivity and specificity of prepared IgG antibody against OMP were analyzed by serological assays. Western blot analysis proved that AP conjugated Anti-OMP antibody can detect the presence of native OMP in HLB-infected citrus plants. Indirect-ELISA confirmed that Anti-OMP also reacted with high affinity and sensitivity against CLas. Besides, it does not show cross-reactivity with non-pathogenic and other pathogenic plant agents. The high affinity and no cross-reactivity of Anti-OMP also validated in immunoblotting. Our research approved that Anti-OMP-IgG antibody has high sensitivity and specificity to CLas. The results suggest that obtained polyclonal antibody has high potential for relatively easy and accurate detection of CLas.