December 31, 2013 Report Open Access

Development and validation of innovative diagnostic tools for detection and identification of the quarantine nematode Meloidogyne enterolobii in support of integrated plant protection strategies in the EU member states

Kiewnick, Sebastian; Braun-Kiewnick, Andrea; Viaene, Nicole; Hallmann, Johannes; Niere, Bkörn; Gamel, Sylvie; Roversi, Pio Federico; den Nijs, Loes; van de Vossenberg, Bart; Toktay, Halil; Prior, Tom; Sapp, Melanie

The aim of this Euphresco project was to coordinate ongoing and future research in the field of Meloidogyne enterolobii detection and identification and to validate new methods in a collaborative way. It contained six main topics:

- Test performance study on nematode extraction methods to detect motile stages (second stage juveniles) of M. enterolobii in soil samples.

- Test performance study on fast and simple DNA extraction from target nematode in complex nematode suspension after extraction from soil samples (Lysis buffer; Holterman et al., 2009). DNA needs to be clean enough (without inhibitors) for further by molecular detection/identification tools. Labs were encouraged but not required to use additional extraction methods. The aim was to test a fast and simple DNA extraction protocol that can be harmonized between NPPO laboratories in the future.

- Methods development and in-house validation of qPCR methods to detect, identify and potentially quantify M. enterolobii in complex nematode suspensions obtained from soil samples (develop protocols for robust, specific and sensitive detection/identification that can be harmonized between NPPO labs, independent of lab equipment and chemistry).

- Test performance study on protocols developed under topic 3.

- Final meeting to communicate and discuss results. Optional workshop on Barcoding as new diagnostic tool for fast and reliable nematode identification was offered to interested groups.

- Publication of results in peer-reviewed research journals and discussion for inclusion of protocols into EPPO diagnostic standard protocol. Identification of future research priorities in the field of Meloidogyne enterolobii research.