Испытание различных методов идентификации возбудителя бактериальной пятнистости листьев цветной капусты Рseudomonas siringae pv. maculicola (mcculloch) Young et al.

Abstract

Pseudomonas syringae pv. maculicola (McCulloch) Young et al. is a pathogen of cauliflower bacterial spot that affects many plants of the Cruciferae family. The need for quality diagnostics of this species arose due to the mandatory phytosanitary inspection of places of production of plant products intended for export. Our research aims at determining a set of methods for the diagnosis of the pathogen of cauliflower bacterial leaf spot and evaluating these methods’ applicability in laboratory practice. The object of the research is P. syringae pv. maculicola – the causative agent of cauliflower bacterial leaf spot.The article presents the test results of two methods for the identification of Pseudomonas syringae pv. maculicola (McCulloch) Young et al. carried out in 2020 at the All-Russian Plant Quarantine Center (VNIIKR). The first method is based on the determination of biochemical properties using the API 20E test kit produced by bioMérieux’s (France); the second one – is a conventional PCR. The type bacterial strain CFBP 1657 obtained by specialists of the All-Russian Plant Quarantine Center (VNIIKR) from French collection of plant-associated bacteria (Cirm-CFBP) was used in the studies. A comparison of the biochemical properties of 23 bacteria of the genus Pseudomonas showed that there are only two characteristics within this test that distinguish P. s. pv. maculicola from other species pathovars: acetoin products and gelatin hydrolysis. Two pairs of primers with different targets in the P. syringae genome were also tested. PCR with PsyF/PsyR primers demonstrated the highest similarity of the obtained fragments with the NCBI database (97.2 %). The analytical sensitivity of PCR with PsyF/PsyR primers in plant and seed extracts was 105 CFU/ml. Determination of analytical specificity with 33 bacterial strains of the genus Pseudomonas revealed cross-reactions with strains of the following species: P. congelans, P. savastanoi pv. phaseolicola, P. savastanoi pv. glycinea, P. syringae pv. coronafaciens, P. syringae pv. syringae. Thus, to differentiate the species by means of PCR with PsyF/PsyR primers, the nucleotide sequence of the obtained amplification products should be additionally determined by Sanger sequencing.