The endosymbiotic bacteria that live within the body of insects are involved in many aspects of the host physiology, including reproduction and defence. Thus, the exploitation of these microorganisms may have practical applications for the management of vector-borne diseases. In the Mediterranean area, Philaenus spumarius was identified as the main vector of Xylella fastidiosa. This xylem-restricted bacterium is responsible for several diseases in a variety of agricultural crops of high importance, for which there aren´t any effective control method. Thus, in this work, we evaluated different culture media for the isolation and growth of bacteria living within P. spumarius adults, for their potential exploitation in the management X. fastidiosa. Specifically, was compared the effect of minimal (Luria Bertani - LB) and complex (Modified Melin-Norkrans - MMN) media, with and without fetal bovine serum or gelling agents, on the number and diversity of bacteria. The collection of isolates obtained was further genetically characterized by BOX-PCR and sequencing of the 16S ribosomal RNA (rRNA) gene. Results showed no significant differences in the abundance and diversity of bacteria among the two media tested (LB and MMN). The addition of fetal bovine serum to the media lead to a slight increase in bacterial abundance, in particular in MMN medium. While the liquid media lead to a significant increase in abundance, the solid media facilitated growth of more diverse bacterial taxa. Comparison between BOX-PCR and 16S rRNA gene sequencing for the analysis of 57 bacterial isolates, revealed a greater discriminatory power of the former, allowing the differentiation of the bacteria even at the intra-species level. Clustering of the isolates using BOX-PCR fingerprinting was different from that obtained from the 16S rRNA gene phylogenetic tree. 16S rRNA gene sequencing method proved to be more suitable in phylogenetic evaluations, generally grouping isolates belonging to the same genus.

Acknowledgement: This work has received funding from the European Union’s Horizon 2020 research and innovation programme under Grant Agreement N. 727987 “Xylella fastidiosa Active Containment Through a multidisciplinary-Oriented Research Strategy, XF-ACTORS”.