Evaluation of peptide-based polyclonal antibodies for a lateral flow immunoassay to rapidly detect tomato leaf curl begomoviruses.

Tomato leaf curl diseases caused by begomoviruses result in significant yield losses in solanaceous and cucurbit crops across East Africa. To effectively manage and monitor these diseases, rapid and field-deployable diagnostic tools are essential. However, traditional Polymerase Chain Reaction (PCR)-based detection methods require laboratory facilities, making them unsuitable for on-site diagnosis. In this study, we developed epitope-targeted peptide antibodies for the serological detection of two begomoviruses in Kenya: tomato leaf curl Uganda virus (ToLCUV) and tomato leaf curl Kunene virus (ToLCKunV). We identified conserved coat protein epitopes on the basis of bioinformatics-based predictions and structural modeling and then used them to generate two peptide-specific antibodies (ToL-asA and ToL-asB). Both antibodies showed strong reactivity with extracts from plants infected with ToLCUV and ToLCKunV in various assays, including indirect ELISA, dot immunobinding assay, and western blotting. Notably, cross-reactivity with unrelated plant viruses was undetectable. In a double-antibody sandwich ELISA, specific antibody pairs generated strong detection signals, indicating that the targeted epitopes were accessible and suitable for immunochromatographic assays. Sequence alignment and phylogenetic analyses confirmed that the targeted epitopes are conserved in several begomoviruses prevalent in East Africa. Furthermore, the use of these antibodies improved viral capture in immunocapture PCR, significantly increasing detection sensitivity for analyses of crude plant extracts. Finally, the peptide antibodies were incorporated into a lateral flow immunoassay platform, thereby enabling the rapid visual detection of begomovirus infections. Overall, these findings demonstrate that epitope-based peptide antibodies are effective reagents for developing rapid and field-deployable diagnostic tools for tomato leaf curl diseases.