Submitted to: BARC Poster Day
Publication Type: Abstract Only
Publication Acceptance Date: March 26, 2003
Publication Date: May 1, 2003
Citation: Kong, H.N., Lydon, J., Patterson, C.D. 2003. A PCR protocol for the identification of Pseudomonas syringae pv. tagetis based on gene mutations that disrupt tagetitoxin production. BARC Poster Day. Abstract 16. Technical Abstract: A PCR protocol that can be used to distinguish Pseudomonas syringae pv. tagetis from other P. syrinage pathovars and other P. syringae species that induce apical chlorosis was developed based on DNA sequences of genes characterized from non-toxigenic mutants of P. syringae pv. tagetis. Following Tn5 mutatagenesis and cloning of the effected DNA from non-toxigenic isolates of P. syringae pv. tagetis, DNA sequences of two affected genes were obtained. A BLAST search revealed that one of the genes had homology to exbB, which codes for an auxilliary protein in the TonB/ExbD/ExbB membrane transport system. The other affected gene had homology to asnB, which codes for an asparagine synthatase. PCR primer sets designated TAGTOX-9 and TAGTOX-10 that were designed from the sequences of the mutated genes that, when in PCRs with DNA from most strains of P. syringae pv. tagetis, produced amplicons of 507 and 733 bp, respectively. The same size amplicons were produced in PCRs containing DNA from chlorotic leaf and stem tissue obtained from sunflower plants infected with P. syringae pv. tagetis. In PCRs with DNA from 16 other P. syringae pathotypes, only DNA from P. syringae pv. helianthi produced the same size amplicons with the respective primer sets. Of significance was the low level of the 507 bp amplicon produced in PCRs with the P. syringae pv. helianthi DNA and TAGTOX-9 primers; results similar to those obtained in PCRs with the same primer set and DNA from non-toxigenic strains of P. syringae pv. tagetis. Results from PCRs with the TAGTOX-9 and TAGTOX-10 primers provide strong evidence that the newly described Pseudomonas syringae species CT99B016C isolated from Cirsium arvense displaying apical chlorosis is not a P. syringae pv. tagetis.