Impedimetric immunosensor versus qPCR for Huanglongbing detection

• Published in: Talanta (Oxford) 2025-02, Vol.283, p.127132, Article 127132
• Main Authors: Silva, Lucas Moreira, Martins, Elaine Cristina, Ferreira, Antonio Aparecido Pupim, Wulff, Nelson Arno, Yamanaka, Hideko
• Format: Article
• Language: English
• Subjects: antibodies; Antibodies, Immobilized - chemistry; Antibodies, Immobilized - immunology; antigens; Bacterial Outer Membrane Proteins - chemistry; Bacterial Outer Membrane Proteins - genetics; Bacterial Outer Membrane Proteins - immunology; Biomolecular technique; Biosensing Techniques - methods; Candidatus Liberibacter; carbon; Citriculture; Citrus; Citrus - chemistry; Citrus - microbiology; Diagnoses; Dielectric Spectroscopy; Electrochemical biosensor; electrodes; greening disease; Huanglongbing; Immunoassay - methods; immunosensors; laboratory diagnosis; leaves; Liberibacter; magnetic materials; magnetism; Magnetite Nanoparticles - chemistry; nanoparticles; neodymium; outer membrane proteins; Plant Diseases - microbiology; Psyllidae; quantitative polymerase chain reaction; Real-Time Polymerase Chain Reaction - methods
• ISSN: 0039-9140; 1873-3573; 1873-3573
• DOI: 10.1016/j.talanta.2024.127132

Citriculture is facing challenges due to the spread of various diseases, among which the most threatening, with worldwide occurrence, is Huanglongbing (HLB), caused by the bacteria Candidatus Liberibacter spp., vectored by psyllids. In orchards, HLB-infected plants are identified by visual observation of symptoms. For laboratory diagnosis, the gold standard is the polymerase chain reaction (PCR), requiring expertise and specific equipment, and with high financial cost. In this work, a selective impedimetric immunosensor was developed for the detection and determination of Ca. L. asiaticus (CLas) in citrus samples. An anti-HLB antibody against the outer membrane protein (OMP) sequence of CLas was obtained and immobilized on previously synthesized and characterized magnetic nanoparticles. The immobilized antibody was presented to various citrus leaf sample extracts. After the affinity reaction with the antigen, a washing step was performed to minimize matrix effects. The affinity reaction was monitored by electrochemical impedance spectroscopy, using a glassy carbon working electrode containing a neodymium magnet. The developed device was able to distinguish HLB-positive samples from HLB-negative samples and those with other infections. The results obtained with the proposed methodology were in good agreement with quantitative PCR (qPCR). [Display omitted] •Diagnosis of citriculture diseases.•The immunosensor for CLas detection is presented for the first time in the literature.•Immobilization of antibodies on magnetic iron oxide nanoparticles.•Comparison of electrochemical immunosensor performance with gold standard qPCR for HLB diagnosis.