RE SEARCH New Erwinia variants require more knowledge and specific tests Pulverising plant material DNA purification Implementation DIP assay PCR 1 product Sample 1 Sample 2 Sample 3 An overview of the DIP assay, which is a quick method to determine Pectobacterium wasabiae up to strain level. The plant material is pulverised and the DNA is purified. After this, with the help of PCR, four DNA fragments are simultaneously generated per microbial strain. These products can differ in length. If, from two samples, products of the individual PCR reactions always have identical lengths, they are infected with the same microbial strain and the infections are related. In this example, in samples 1 and 2, an identical microbial strain of Pectobacterium wasabiae was found, sample 3 shows a different strain. In Wageningen, a new DIP assay test has been developed for Pectobacterium wasabiae. ic Dickeya species are present. A test for Pectobacterium carotovorum subsp. brasiliense has recently been developed by Plant Research International (PRI) in Wageningen. This test was evaluated last year by the NAK for usage in practice. Don’t export Erwinia In order to be able to recognise new variants and, if necessary, adjust current tests, researchers regularly carry out new surveys. Both in and outside Europe, there are still other Dickeya and Pectobacterium species that can cause Black leg. Knowledge about the various variants and their properties are also relevant for export. Seed lots infected with Dickeya solani should not be exported to countries with a hot climate. Knowledge of the diversity of the bacterium is also of great importance for research into the survival, spreading, disinfection and control of the bacterium. For example, it transpired that organic control methods were not always equally effective for all variants of the bacterium. New DIP Assay test In Wageningen, a new DIP assay test has been developed for In order to be able to recognise new variants and, if necessary, adjust current tests, researchers regularly carry out new surveys. Pectobacterium wasabiae. Not only does it determine the Pectobacterium wasabiae species, but also the strain of Pectobacterium wasabiae found in the extract. Strains of this bacterium in Dutch seed lots show small differences which can be simply recognised with the DIP assay. The DIP assay is very suitable for research into the origin of the bacterium. If the same strain of the bacterium is found on a machine as well as in the seed at a crop farm, it can reasonably be assumed that the infections originated from the machine. Bacteria in seed potato lots were also compared with each other to establish whether there is a connection between the infections. The method has been used mainly in research, but can also be applied by inspection services and trading companies. ● Jan van der Wolf, Plant Research International PCR 2 product PCR 3 product PCR 4 product Firstly, knowledge about the diversity of Erwinia bacteria is very important during the inspection of the seed. Potato World 2015 • number 4 25 Pagina 24
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