Fluorescence in situ hybridization as a tool for studying phytoplasma-endophytes interaction in plant

Bacteria residing in plant tissues without inducing symptoms of diseases are defined as endophytes. They can protect plants from pathogens through three different mechanisms: competition; production of allelochemic inhibitors and induction of systemic resistance (ISR). Recently statistical analysis, carried out on LH-PCR (Length Heterogeneity-PCR) profiles, highlighted a different composition of endophytic bacterial community associated with healthy, GY diseased, and recovered grapevine plants. In this study we experimented fluorescence in situ hybridization (FISH) to study the interaction between phytoplasmas and endophytic bacteria in the model plant Catharanthus roseus L.. Both phytoplasmas and bacteria were co-localized by confocal microscopy using different probes to target 16SrV phytoplasmal and bacterial 16S rDNA, respectively. To avoid interference with autofluorescence of tissue constituents, bacterial probes were labelled with fluorophores emitting in the far-red (i.e. CY5). Phytoplasmal probes were labelled with FAM and they prove to be able in discriminating genetically different phytoplasmas, localized only in the phloem tissues. Endophytic bacteria were instead detected in the phloem, xylem and leaf parenchyma. These results, though preliminaries, show the great potentiality of FISH in studying the interaction between phytoplasmas and endophytic bacteria.