Molecular identification, in vitro copper resistance and antibiotics susceptibility of the causal agent of the olive knot disease in Morocco

Abstract

Aims: This study aimed to i) identify Pseudomonas savastanoi pv. savastanoi (Pss) as a causal agent of the olive knot on the basis of biochemical, pathogenicity and PCR technique ii) investigate in vitro bacterial resistance toward copperbased compounds and efficiency of some antibiotics on pathogen suppression. Methodology and results: Biochemical, pathogenicity and molecular identification based on alkaline method for the DNA extraction were performed to identify possible causal agent of the olive knot. Copper resistance for Pss strains was evaluated by inoculation of bacterial suspensions into YPG medium, containing the cupric sulfate at 0, 100, 250 and 500 ppm. The efficiency of eight antibiotics on Pss strain was evaluated at different concentrations. Fifty-nine isolates caused typical knots at the site of inoculation with bacterial suspensions. All isolates have been identified as Pss using specific primers. No resistance to copper was detected with concentration of 500 ppm. In contrast, copper resistance was found during 48 h with lower concentration (100 or 250 ppm). The maximal inhibition of Pss 2102-4M was observed with the highest concentration (20 μg/mL) of the Aureomycin, Streptomycin and Novobiocin with inhibition diameters of 30, 24 and 10 mm, respectively. Whereas, Colchicine, Bacitracin, Cephalex, Ampicillin and Cycloserine have no inhibitory effect on the Pss 2102-4M strain. Conclusion, significance and impact of study: The alkaline method for the DNA extraction from pure culture was reliable and rapid and can be recommended for molecular detection the causal agent of the olive knot. This is the first report determined copper resistance levels of Moroccan strains of Pss and in vitro evaluated for the susceptibility towards the antibiotics.