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Association of ‘Candidatus Liberibacter solanacearum’ with a Vegetative Disorder of Celery in Spain and Development of a Real-Time PCR Method for Its Detection

First, second, fourth, tenth, and eleventh authors: Plant Protection and Biotechnology Center, Instituto Valenciano de Investigaciones Agrarias (IVIA), 46113 Moncada, Valencia, Spain; third and twelfth authors: Grupo Virología Vegetal, Instituto Agroforestal Mediterráneo, Universidad Politécnica de Valencia, 46022 Valencia, Spain; fifth and sixth authors: Escola Superior de Agricultura Luiz de Queiroz, Universidade de São Paulo, 13418-900 Piracicaba, Brazil; seventh author: Servicio de Análisis Agroalimentario, Conselleria de Presidencia y de Agricultura, Pesca, Alimentación y Agua, Generalitat Valenciana, 46460 Silla, Valencia, Spain; and eighth and ninth authors: Departamento Técnico, Agrícola Villena Coop, V. 03400 Villena, Alicante, Spain.

A new symptomatology was observed in celery (Apium graveolens) in Villena, Spain in 2008. Symptomatology included an abnormal amount of shoots per plant and curled stems. These vegetative disorders were associated with ‘Candidatus Liberibacter solanacearum’ and not with phytoplasmas. Samples from plant sap were immobilized on membranes based on the spot procedure and tested using a newly developed real-time polymerase chain reaction assay to detect ‘Ca. L. solanacearum’. Then, a test kit was developed and validated by intralaboratory assays with an accuracy of 100%. Bacterial-like cells with typical morphology of ‘Ca. Liberibacter’ were observed using electron microscopy in celery plant tissues. A fifth haplotype of ‘Ca. L. solanacearum’, named E, was identified in celery and in carrot after analyzing partial sequences of 16S and 50S ribosomal RNA genes. From our results, celery (family Apiaceae) can be listed as a new natural host of this emerging bacterium.