5. Isolation and characterization of endophytic bacteria from Handroanthus impetiginosus

YARTE M.E.; GISMONDI MI; LLORENTE B.; LARRABURU E.E.

San Miguel de Tucumán

Congreso; XII Congreso Argentino de Microbiología General; 2017

Asociación Civil de Microbiología General

Handroanthus impetiginosus "pink lapacho" (Bignoniaceae) is a medicinal, ornamental and forestaltree from the northwestern of Argentina. Propagation by seeds or cuttings of the pink lapacho isdifficult so in vitro culture methodologies and biofertilization are recommended technologies. Plantsusually interact with endophytic and rhizospheric bacteria with mutual benefits. Plant growth promotionby biofertilizers is mainly attributed to improve the acquisition of essential nutrients like nitrogen andphosphorus, or modulate the level of vegetal hormones. Plant growth promoting bacteria (PGPB)isolated from pink lapacho are scarcely described.The aim of this work was to isolate and characterize native endophytic bacteria from leaves and rootsof pink lapacho trees grown in National University of Luján, and to analyze their potential as PGPR.Superficial disinfected tissues were placed on nutritive agar plates and on nitrogen-free agar (Nfb)supplemented with antifungal and incubated at 30ºC until bacterial growth was observed. Morphologyof colonies, Gram staining and catalase activity were determined. Indole acetic acid (IAA) productionof isolated strains was colorimetrically evaluated in Luria-Bertani (LB) medium supplied withtryptophan (2.5 mg/mL) using Salkowski?s reagent. Strains able to solubilize inorganic phosphate wereidentified using Pikovskaya agar supplemented with Ca3(PO4 )2 (5 g/L) and green-bromocresol(PVK). Phosphate solubilization index (PSI) was recorded. Strain salt tolerance was evaluated in LBmedium supplemented with 5% or 10% NaCl. Bacterial identification was carried out by partialamplification and sequencing of 16S rRNA gene (V1 and V3 regions). Sequences were aligned toreference genomes retrieved from leBIBI database. Phylogenetic inference was performed by themaximum likelihood method followed by bootstrap analysis (1000 pseudoreplicates) as implementedin MEGA v6.0 software.Bacteria isolation of leaves and roots allowed obtaining 13 and 6 strains, respectively. Seventeenstrains were able to grow in Nfb indicating biological nitrogen fixation. All strains were able to grow inPVK although 13 showed solubilization halo.From total isolated strains, 10 were positive for IAA production. Leaf isolates (n=8) produced IAAconcentrations between 4.37 and 20.80 mg/ml whereas root isolates (n=2) produced 19.4 and 41.6mg/ml. Of them, root isolates were able to grow in medium with 5% NaCl and only one also grew at10% NaCl after 48 h. Among leaves endophytes, four isolates grew at both concentrations of NaCl.Molecular identification allowed determining Bacillus, Paenibacillus, Pseudomonas andMethylobacterium as endophytes genera.In conclusion, the isolated strains present plant growth promoting properties that can contribute toincrease environmental sustainability in agriculture.