Rapid and sensitive methods for detection of Allorhizobium vitis, causal agent of grapevine crown gall

A rapid method and sensitive methods for extraction of bacterial DNA from pure culture and directly from plant materiel were compared in polymerase chain reaction with specific primers VCF3/VCR3 to see the reliable method that can used in the detection of tumorigenic strain of Allorhizobium vitis causal agent of grapevine crown gall. From the three tested methods of DNA extraction from pure culture, the alkaline method is the most effective technique for the extraction presenting a high sensitivity with a detection threshold equal to 5.104 CFU/ml. Five different protocols for extracting bacterial DNA from plant tissues of infected tomato, based on the use of an extraction buffer, were tested to see its usefulness in detecting pathogenic strain of A. vitisS4. Two protocols based on the use of Triton X-100 and Tween 20 were efficient for detecting A. vitis S4 directly from tomato tumors with a sensitivity of 103 CFU/ml for the both protocols. Consequently, these protocols were proposed as specific protocols for the detection of tumorigenic strain of A. vitis from symptomatic and asymptomatic plants.