PCR amplification of the hrcV gene through specific primers for detecting Pseudomonas syringae pathovars

Abstract

Pseudomonas syringae pathovars are important pathogens among phytopathogenic bacteria causing a variety of diseases in plants. These pathogens can rapidly disseminate in a large area leading to infection and destruction of plants. To prevent the incidence of the bacteria, appropriate detection methods should be employed. Routinely serological tests, being time-consuming and costly, are exploited to detect these pathogens in plants, soil, water and other resources. Over the recent years, DNA-based detection approaches which are stable, rapid, specific and reliable have been developed and sequence analysis of various genes are widely utilized to identify different strains of P. syringe. However, the greatest limitation of these genes is inability to detect numerous pathovars of P. syringae. Herein, by using bioinformatic analysis, we found the hrcV gene located at pathogenicity islands of bacterial genome with the potential of being used as a new marker for phylogenetic detection of numerous pathovars of P. syringae. Following design of specific primers to hrcV, we amplified a 440 bp fragment. Of 13 assayed pathovars, 11 were detected. Also, through experimental procedures and bioinformatic analysis it was revealed that the designed primers have the capacity to detect 19 pathovars. Our findings suggest that hrcV could be used as a gene with the merit of detecting more pathovars of P. syringae in comparison with other genes used frequently for detection purposes.

Identification and genetic diversity analysis of strain Pseudomonas syringae S5: A pathogen responsible for bacteriosis in Avena sativa plants

“…Pseudomonas protegens Pf‐5 was used as a negative control, and it did not produce an amplicon. In agreement with what is suggested by Vaseghi et al (), detection of the hrcV gene in the PsS5 strain is another piece of evidence that allows us to consider it belongs to the phytopathogenic species P. syringae .…”

“…The hrcV gene, located on the pathogenicity islands of the bacterial genome, has been proposed as an effective marker for the rapid detection of P. syringae pathovars. This gene was selected for its high specificity, prevalence and high evolutionary conservation in different P. syringae strains (Vaseghi, Bakhshinejad, Safaie, Parchin, & Sadeghizadeh, ). We amplified this gene by PCR using genomic DNA as a template and specific primers SAF‐F and SAF‐R.…”

Isolation and identification ofPseudomonas syringaefacilitated by a PCR targeting the wholeP. syringaegroup

“…Moreover, strains from the P. syringae group have extensive diversity in virulence gene repertoires like those for effectors, toxins or plant hormone production (Baltrus et al, 2011) that cannot be used to characterize the whole P. syringae group. Some molecular tools have been proposed for the specific detection of single pathogenic varieties so-called pathovars (Kong et al, 2004, Gervasi & Scortichini 2009, Cho et al, 2010, Gallelli et al, 2011, a group of pathovars (Tegli et al, 2010, Popovic et al, 2014, Vaseghi et al, 2014 or one phylogroup (Clarke et al, 2010, Cottyn et al, 2011.…”

Genetic diversity and pathogenicity of Pseudomonas syringae pv. aptata isolated from sugar beet

“…() and Vaseghi et al . (). In the present study, the Papt2F/1R specific primers were used, developed by Cachatori () from the gyrB gene (which is part of the core genome), and amplified DNA from all 104 isolates.…”