Involvement of specific traits of olive beneficial rhizobacteria to protect against biotic and abiotic stresses

Abstract: Verticillium wilt of olive (VWO), caused by the fungus Verticillium dahliae Kleb., together with drought and salinity are considered serious (a)biotic constraints affecting olive cultivation worldwide. Different microorganisms have been identified as effective biocontrol agents (BCA) against V. dahliae and/or as useful tools to alleviate these stresses. A collection of olive beneficial rhizobacteria was screened, including the effective BCA against VWO Pseudomonas simiae PICF7 and Pseudomonas sp. PICF6, to assess them as protectants towards abiotic stresses. In this study, we pursued a double goal: i) on the one hand, to evaluate the potential involvement of selected phenotypes of strain PICF7 in root colonization and VWO biocontrol, and ii) on the other hand, to assess whether olive beneficial rhizobacteria can be used as protective agents to alleviate drought and salt stresses. For the latter objective we aimed to identify 1-aminocyclopropane-1-carboxylic acid deaminase (ACD) producers and tested the hypothesis that ACD producers lessen the effects of these abiotic stresses. A random transposon-insertion mutant bank of PICF7 was screened for the loss of phenotypes likely involved in rhizosphere/soil persistence (copper resistance), root colonization (biofilm formation) and plant growth promotion (phytase activity). Transposon insertions in genes putatively coding for the transcriptional regulator CusR or the chemotaxis protein CheV were found to affect copper resistance, whereas an insertion in the fleQ gene putatively encoding a flagellar regulatory protein hampered the ability to form a biofilm. However, these mutants displayed the same antagonistic effect against V. dahliae as the parental strain. Endophytic colonization of olive roots was only altered in mutants impaired in biofilm formation since they were never found inside olive roots. This rhizobacteria collection was in vitro screened for ACD activity. Strain PICF6 displayed this phenotype while PICF7 was defective, even though an ACD-coding gene was earlier predicted in its genome. An unidentified deaminase was confirmed instead. Greenhouse experiments with olive plants inoculated either with PICF6 or PICF7, or co-inoculated with both strains, and subjected to drought or salt stress were carried out. Several physiological and biochemical parameters increased in stressed plants (i. e., stomatal conductance and flavonoids content), irrespective they were previously bacterized. Finally, results demonstrated that the PICF7 phenotypes studied were irrelevant for VWO biocontrol and that neither PICF6 (ACD+) nor PICF7 (ACD-) lessened effects caused by these abiotic stresses, under our experimental conditions.