Detection of Pseudomonas syringae pathovars by thiol-linked DNA-Gold nanoparticle probes

DNA-Gold Nanoparticle (DNA-Gold NP) probes as a new generation of biosensor-based detection tools hold promise in biological sciences. They are of enormous importance in the emergence of rapid and sensitive procedures for detecting pathogenic microorganisms. Pseudomonas syringae pathovars are among the most significant phytopathogenic bacteria causing a variety of diseases in numerous agricultural crops. In this study, Gold NP-bound probes were used for colorimetric detection of P. syringae pathovars. For this purpose, subsequent to amplification of the hrcV gene by using specific primers, thiol-functionalized-DNA attached to Gold-NPs was added to a PCR-amplified fragment containing the hrcV gene. Wavelength changes were monitored before and after adding probes into target DNA. To investigate the specificity of binding of probes, a mixture of genomes extracted from plants and their specific bacteria and as well DNA isolated from bacteria not belonging to P. syringae species were employed. Also, sensitivity determination experiments which were conducted through gradually diminishing the concentration of genomic template indicated the sensitivity of our method in detecting as little as 15 ng/mu L of genomic template. Taken together, our method could detect 24 of 26 isolates which were examined. The results of our assay proved to be highly specific with potential for early detection of P. syringae. (c) 2013 Elsevier B.V. All rights reserved.