Intra-Laboratory Evaluation of DNA Extraction Methods and Assessment of a Droplet Digital PCR for the Detection of Xanthomonas citri pv. citri on Different Citrus Species

Autor: Nicoletta Pucci, Valeria Scala, Giuseppe Tatulli, Alessia L’Aurora, Simone Lucchesi, Manuel Salustri, Stefania Loreti
Jazyk: angličtina
Rok vydání: 2022
Předmět:
Zdroj: International Journal of Molecular Sciences, Vol 23, Iss 9, p 4975 (2022)
Druh dokumentu: article
ISSN: 1422-0067
1661-6596
DOI: 10.3390/ijms23094975
Popis: Xanthomonas citri pv. citri (Xcc) and X. citri pv. aurantifolii (Xca), causal agents of citrus bacterial canker, are both regulated by the European Union to prevent their introduction. Xcc is responsible for severe outbreaks of citrus production worldwide, therefore, a prompt and reliable detection is advisable for the early detection of this bacterium either in symptomatic or asymptomatic plant material. The current EPPO (European and Mediterranean Plant Protection Organization) diagnostic protocol, PM 7/44(1), includes several diagnostic tests even if new assays have been developed in the latter years for which validation data are needed. Recently, a test performance study was organized within the Valitest EU Project to validate Xcc diagnostic methods and provide evidence on the most reliable assays; however, the influence of DNA extraction methods (DEM) on the reliability of the detection has never been assessed. In this study we evaluate four different DEM, by following two different approaches: (i) a comparison by real-time PCR standard curves of bacterial DNA versus bacterial DNA added to plant DNA (lemon, leaves and fruit; orange fruit); and (ii) the evaluation of performance criteria of spiked samples (plant extract added with ten-fold diluted bacterial suspensions at known concentrations). Droplet digital PCR is developed and compared with real-time PCR, as the detection method.
Databáze: Directory of Open Access Journals
Externí odkaz:
Nepřihlášeným uživatelům se plný text nezobrazuje