Azospirillum brasilense az39 and pantoea sp.: gfp- labelling as a method to study root colonization

DIAZ HERRERA S; FAVALE N; BENAVIDES MP; ZAWOZNIK MS; GROPPA MD

Congreso; Reunión Conjunta de Sociedades de Biociencias; 2017

Plant growth promoting rhizobacteria (PGPR) are associated withthe roots of a wide variety of plant species leading to an increase ofwater and nutrients uptake, through diverse mechanisms.To obtain a tool for the study of root colonization by microorganisms,Azospirillum brasilense Az39 (a PGPR widely used in commercialformulations) and a strain of Pantoea sp. isolated fromwheat were labelled with the green fluorescent protein (GFP). Todemonstrate that bacterial labelling does not affect other bacterialproperties, the dynamics of growth in liquid medium, biofilm formation(by violet crystal assay) and indole acetic acid (IAA) production(by Salkowski method) of wild type and GFP-labelled strains werestudied. Preliminary results would indicate that GFP labelling did notaffect IAA production of A. brasilense, whereas biofilm formationwas lower in the transformed strain in LB medium. Great variabilitywas observed in Nfb medium. The dynamics of in vitro multiplication,the growth in semisolid Nfb medium and the morphology of the coloniesremained similar.Additionally, an inoculation assay of Arabidopsis thaliana plantswas carried out. After 6 days of inoculation with A. brasilense, theplants showed an evident fluorescence in the roots denoting thepresence of the bacteria. Fluorescence microscopy allowed to observea great colonization of the surface root and a certain numberof fluorescent cells arranged in a row inside a root segment, apparentlycolonizing a vascular bundle. While plants inoculated with labelledAzospirillum promoted root growth, inoculation with labelledPantoea sp. negatively affected Arabidopsis plants and a significantchlorosis in the leaves was observed. Labelled strains are importanttools to advance in the knowledge of the interaction between rhizosphericmicroorganisms and diverse crops of high economic impactfor our country.Keywords: Azospirillum brasilense Az39, Pantoea sp., GFP, Arabidopsisthaliana.