Development of a specific assay using RISA for detection of the bacterial agent of 'basses richesses' syndrome of sugar beet and confirmation of a Pentastiridius sp (Fulgoromopha, Cixiidae) as the economic vector

A technique for the specific diagnosis in insects of SBRp (the gamma-3 proteobacterium associated with the syndrome 'basses richesses' (SBR) of sugar beet crops in eastern France), using the RISA (rDNA intergenic spacer analysis) technique, was developed. PCR using the Alb1/Oliv1 primer pair specifically amplified a 16S-ITS region of SBRp and produced a characteristic DNA fingerprint. This PCR assay did not detect other closely related organisms, including the Arsenophonus endosymbiont of Diaphorina citri, the secondary endosymbiont of Glycaspis brimblecombei, or 'Candidatus Phlomobacter fragariae', a related phytopathogenic gamma-3 proteobacterium. Six different ribosomal operons, differing in their ITS region or partial 16S sequence, were identified in SBRp. PCR amplification with Alb1/Oliv1 of DNA samples from Hemiptera species (suborders Fulgoromorpha and Cicadomorpha) collected in sugar beet fields confirmed Pentastiridius sp. as the economic vector of SBR disease. The high percentage of field-Pentastiridius sp. specimens which tested positive for SBRp reflected the importance of SBR disease in sugar beet crops. This is the first time that the RISA technique has been used as a diagnostic test for a plant pathogenic bacterium in insects.