Application of new methods for detecting and identifying Acidovorax Citrulli and optimization of the laboratory diagnostic scheme

The phytopathogenic bacterium Acidovorax citrulli is included in the quarantine lists of many countries, including the unified list of quarantine objects of the Eurasian Economic Union. The control of the introduction and spread of the quarantine organism into the territory of Russia is carried out through laboratory diagnostics. Diagnostic methods should ensure quality and the high speed of product health testing. The study aimed to use new methods for the detection and identification of A. citrulli and to optimize the laboratory diagnostic scheme. In the course of the work, cucumber seeds with an infectious load were prepared and tested. Testing was carried out according to an optimized diagnostic scheme using methods not previously used to diagnose A. citrulli. At each stage of work with samples, the time spent was recorded. The researchers tested the prepared infected seeds at the first stage using the standard qPCR method and then the qPCR method PAS F/R, PAS P which had not previously been used to diagnose A. citrulli. The new qPCR test PAS F/R, PAS P showed stable results at testing each sample in a row of each replication. The sample enrichment method was successfully applied, not previously used in diagnosing A. citrulli. The use of the method made it possible to separate samples containing live target bacteria from those containing only A. citrulli DNA within 2.5 days. After enrichment, the isolation and identification of the A. citrulli culture from the samples were successfully carried out. Based on the data obtained as a result of this study, the scheme for detecting and identifying the causative agent of bacterial fruit blotch of cucurbits was optimized. The proposed scheme will reduce diagnosing a dangerous quarantine organism from four weeks to one week.

plant quarantine, Acidovorax citrulli, qPCR test, method of sample enrichment, diagnosis of bacteria

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