Reverse transcription-duplex-polymerase chain reaction for simultaneous detection of Citrus tristeza virus and ‘Candidatus Liberibacter’ from citrus plants

Tristeza and Huanglongbing are the most important diseases of citrus caused by Citrus tristeza virus (CTV) and 'Candidatus Liberibacter' species, respectively. A reverse transcription duplex polymerase chain reaction (RT-d-PCR) was developed to detect these pathogens simultaneously from infected citrus plants. Consistence of the technique was validated and specificity of the primers was confirmed by sequence analysis. Concurrent detection of CTV and 'Ca. Liberibacter' spp. decreases the risk of contamination and saves time in comparison to standard techniques (i.e., nested PCR) and reduces the cost as compared to high-end techniques (i.e., quantitative PCR), especially for the selection and production of pathogen free citrus plants for certification programs.