The genetic characterization of Xanthomonas arboricola pv. juglandis, the causal agent of walnut blight in Poland

Abstract

Leaves and fruits of walnut trees exhibiting symptoms of bacterial blight were collected from six locations in Poland. Isolations on agar media resulted in 18 bacterial isolates with colony morphology resembling that of the Xanthomonas genus. PCR using X1 and X2 primers specific for Xanthomonas confirmed that all isolates belonged to this genus. In pathogenicity tests on unripe walnut fruits, all isolates caused typical black necrotic lesions covering almost the entire pericarp. Results of selected phenotypic tests indicated that characteristics of all isolates were the same as described for the type strain of Xanthomonas arboricola pv. juglandis. Genetic analyses (PCR MP, ERIC-, BOX-PCR and MLSA) showed similarities between the studied isolates and the reference strain of X. arboricola pv. juglandis CFBP 7179 originating from France. However, reference strains I-391 from Portugal and LMG 746 from the UK were different. MLSA analysis of partial sequences of the fyuA, gyrB and rpoD genes of studied isolates and respective sequences from GenBank of pathotype strains of other pathovars of X. arboricola showed that the X. arboricola pv. juglandis isolates consisted of different phylogenetic lineages. An incongruence among MLSA gene phylogenies and traces of intergenic recombination events were proved. These data suggest that the sequence analysis of several housekeeping genes is necessary for proper identification of X. arboricola pathovars.

Development of SCAR markers for rapid and specific detection of Pseudomonas syringae pv. morsprunorum races 1 and 2, using conventional and real-time PCR

“…Commonly used methods for designing SCAR primers include rep-PCR (repetitive PCR) (Sangdee et al 2013 ), randomly amplified polymorphic DNA (RAPD) (Liu et al 2012 ; Cheng et al 2015 ), amplified fragment length polymorphism (AFLP) (Zhang et al 2012 ), PCR with universal rice primers (URP-PCR) (Lim et al 2009 ) and inter-simple sequence repeat (ISSR) (Giaj Merlera et al 2015 ). Although the PCR MP method was described so far as helpful in the study of genetic diversity of bacteria and yeast (Leibner-Ciszak et al 2010 ; Kałużna et al 2010b , 2014 ; Zasada et al 2014 ), it has not been previously reported to be used for the selection of SCAR markers. In this work, the PCR MP is for the first time used for the design of SCAR primers specific for detection of plant pathogenic bacteria.…”

Comparative Genomic and Phenotypic Characterization of Pathogenic and Non-Pathogenic Strains of Xanthomonas arboricola Reveals Insights into the Infection Process of Bacterial Spot Disease of Stone Fruits

“…MLSA has been shown to be useful to characterize strains of X . arboricola [ 2 , 5 , 18 , 45 , 46 ]. In this work, we utilized a MLSA scheme proposed by Young and collaborators [ 18 ], which is based in the analysis of partial sequences of the genes dnaK , fyuA , gyrB and rpoD .…”

Genetic diversity and pathogenicity of Monilinia polystroma – the new pathogen of cherries

“…The evolutionary rate of change within microsatellites is considerably higher than most other types of DNA; thus, the likelihood of polymorphism in these sequences is greater in comparison with other molecular markers (Schl€ otterer, 2000). In the PCR MP assay, the restriction patterns are a representation of the whole genome and are dependent upon the enzyme cleavage sequences and the melting temperature of the amplified restriction products (Kału_ zna et al, 2014). In contrast, in the RAPD assay, the arbitrary primers, although designed to amplify repetitive sequences, aligned more randomly to the genome (Wilson & Sharp, 2006).…”