DNA Markers, Pathogenicity Test, and Multilocus Sequence Analysis to Differentiate and Characterize Cereal-Specific...

Xanthomonas translucens contains a group of bacterial pathogens that are closely related and have been divided into several pathovars based on their host range. X. translucens pv. undulosa (Xtu) and X. translucens pv. translucens (Xtt) are two important pathovars that cause bacterial leaf streak disease on wheat and barley, respectively. In this study, DNA markers were developed to differentiate Xtu and Xtt and were then used to characterize a collection of X. translucens strains with diverse origins, followed by confirmation and characterization with pathogenicity tests and multilocus sequence analysis/typing (MLSA/MLST). We first developed cleaved amplified polymorphic sequence markers based on the single-nucleotide polymorphisms within a cereal pathovar-specific DNA sequence. In addition, two Xtt-specific markers, designated Xtt-XopM and Xtt-SP1, were developed from comparative genomics among the sequenced Xtt/Xtu genomes. Using the developed markers, a collection of X. translucens strains were successfully identified as Xtu or Xtt. Pathogenicity tests on wheat and barley plants and MLSA of four housekeeping genes validated the pathovar assignation of those strains. Furthermore, MLSA revealed distinct subclades within both Xtu and Xtt groups. Seven and three sequence types were identified from MLST for Xtu and Xtt strains, respectively. The establishment of efficient Xtt/Xtu differentiation methods and characterization of those strains will be useful in studying disease epidemiology and host-pathogen interactions and breeding programs when screening for sources of resistance for these two important bacterial pathogens.