Development and approbation of a new identification method of Pseudomonas savastanoi pv. phaseolicola

The article describes the development and testing of a new molecular genetic method for improving the identification scheme of Pseudomonas savastanoi pv. phaseolicola, the pathogen responsible for halo blight of beans in oil and leguminous crops. The reason for improving the methods for detecting and identifying the bacteria was to reduce the cost and time required for testing due to increased soybean and pea exports to importing countries that require seed products to be pathogenfree. During the development of a new confirmatory test based on the realtime PCR method, four types of primer systems were selected (PspF1/PspR1/PspP1; PspF2/PspR2/PspP2; PspF3/ PspR3/PspP3 and PspF4/PspR4/PspP4) using sites of consensus sequences of P. savastanoi pv. phaseolicola NZ_JAANPZ010000001, NC_007274 plasmids. The approbation of the new method was carried out in the bacteriology and GMO analysis laboratory of the Test Laboratory Center of the AllRussian Plant Quarantine Center. Analytical sensitivities were determined using pathogenfree soybean plant extracts supplemented with a P. savastanoi pv. phaseolicola suspension. The presence of a tested pathogen DNA was confirmed in extracts containing phytopathogenic bacteria at a concentration of at least 2 × 10² CFU/ml. Analytical specificity was determined to be 99.4% by testing 153 strains using the recommended realtime PCR with PspF3 and PspR3 primers and a PspP3 probe. As a result of the study, the PCR test is recommended for implementation in the diagnostic scheme as a confirmatory molecular genetic method.