Corrigendum PM 7/024 (5) Xylella fastidiosa

Background information

In the EPPO diagnostic protocol PM 7/024 (5) Xylella fastidiosa (EPPO, 2023), the test of Hodgetts et al. (2021) is recommended under Section 3.4 on Screening test in Section 3.4.2.2 and as an identification and subspecies determination test in Section 4. It is described in full in Appendix 11.

It should be noted that during a Proficiency Test (PT) organized by the European Union Reference Laboratory for bacteriology (EURL-BAC) on Xylella fastidiosa some issues concerning analytical sensitivity and analytical specificity have been encountered with this test (Vreeburg et al., 2024a). A lower analytical sensitivity for subsp. pauca ST74 has been observed compared to the other subspecies. Therefore, using Hodgetts et al. (2021) as a single test for screening is not recommended.

In terms of assignment of subspecies in Section 4.2, the results of the PT showed that for Hodgetts et al. (2021) some of the subsp. pauca strains from sequence type 74 (ST74) can cross react with the test for subsp. fastidiosa (Vreeburg et al., 2024a). It is therefore recommended to laboratories using this test to include ST74 strains in the set of strains selected for verification of this test.

In Section 4.2 Molecular tests for the identification of X. fastidiosa and assignment of X. fastidiosa subspecies it is stated: ‘In other cases, subspecies assignment may be performed by subspecies-specific molecular tests (Pooler & Hartung, 1995, see Appendix 18; Hernandez-Martinez et al., 2006, see Appendices 19 and 20) or Sanger sequencing.’

It should be noted that during the PT organized by the EURL-BAC on Xylella fastidiosa subspecies determination, several participants used the conventional PCR tests of Hernandez-Martinez et al. (2006). These participants found that these conventional PCR tests could not distinguish between subsp. sandyi and subsp. morus, since both subspecies produced a PCR product of 638 bp (Vreeburg et al., 2024b).

It is therefore recommended to laboratories performing Hernandez-Martinez et al. (2006), that if a band of 638 bp is obtained, then additional tests (Appendices 10, 11, 16 or 17) should be performed to distinguish between subsp. sandyi and subsp. morus.

List of changes:

Using Hodgetts et al. (2021) as a single test for screening on plant samples is not recommended.

Section 4 of Appendix 11 for exclusivity is modified as follows (new text in bold):

[ ]: 100%.

With Hodgetts et al. (2021) some of the subsp. pauca strains from sequence type 74 (ST74) can cross react with the test for subsp. fastidiosa (Vreeburg et al., 2024a). It is therefore recommended to laboratories using this test to include ST74 strains in the set of strains selected for this test.

Section 4 of Appendix 19 and 20 for Analytical specificity is modified as follows (new text in bold):

[ ]

Hernandez-Martinez et al. (2006) cannot distinguish between subsp. sandyi and subsp. morus, since both subspecies produce a PCR product of 638 bp (Vreeburg et al., 2024b).

It is therefore recommended to laboratories performing Hernandez-Martinez et al. (2006), that if a band of 638 bp is obtained, then additional tests (Appendices 10, 11, 16 or 17) should be performed to distinguish between subsp. sandyi and subsp. morus.