Emergency measures for Xylella

The emergency measures target Xylella fastidiosa, Xylella taiwanensis and all of their sub-species. The sub-species of X. fastidiosa include: fastidiosa, multiplex, pauca , sandyi, tashke and pear leaf scorch (PLS; now called ‘Xylella taiwanensis’).

In this document ‘Xylella’ refers to the species Xylella fastidiosa and all of its sub-species, and Xylella taiwanensis.

Notes

1. There are scheduled fees associated with the growth of nursery stock in an Australian Government post- entry quarantine facility, which must be met by the importer. The importer is responsible for contacting the facility to confirm all arrangements, including space availability and number of plants, prior to the plant material arriving in Australia. Importers must clearly nominate the facility that their material will be sent to on the import permit application.
2. Nursery stock and tissue cultures from high-risk countries/regions must be produced through an arrangement approved by the National Plant Protection Organisation of the exporting country to meet Australia’s requirements (Appendix 4). These arrangements have also been distributed to NPPOs through the IPPC notification.
3. We will reserve the right to undertake testing to verify that a consignment is free of Xylella.
4. PCR tests that will detect Xylella including recognised sub-species. PCR testing will require the following two tests:
   1. the rimM gene sequence real-time PCR test of Harper et al. (2010)¹1
   2. the conventional PCR of Minsavage et al. (1994)2 or an equivalent PCR that detects X. fastidiosa sub-species pear leaf scorch (PLS).

This section is about the overarching systems and processes to ensure that nursery stock produced for export to Australia is grown, tested and confirmed free from infection by X. fastidiosa.

The National Plant Protection Organisation (NPPO) of the exporting country will approve the arrangements and ensure that the requirements are met. Several other parties, including the grower and testing laboratory will need to work with the NPPO to do this. The supplier will contact the NPPO of the exporting country to establish the arrangements. Australian importers should contact their overseas suppliers to ensure that the work is initiated for the arrangements.

The arrangements

The arrangements require these elements:

• Propagation, growth, testing, certification and export under the authority of the NPPO of the exporting country
• The facility where plants are grown is insect-proof so that it excludes all insects of the suborder Auchenorrhyncha (leafhoppers, froghoppers, sharpshooters, spittlebugs and treehoppers)
• Plants for export to Australia are grown for their entire life in the facility regardless of the propagation technique used (e.g. plants grown from seed, grown vegetatively or grown in tissue culture)
• All the mother plants are tested for X. fastidiosa by the approved protocol  up to 8 weeks before export:
  • nursery stock– the mother plants that are the immediate source from which the nursery stock plant lot was propagated are tested. This includes the mother tissue cultures or the mother plants that directly provided the material used to propagate the nursery stock plant lot.
  • Mother plants or mother tissue cultures will have been grown and protected within the facility for 12 months before testing. (Diagram 1)
• Prior to export, an official sample is drawn from the plant lot and tested according to the approved protocol for X. fastidiosa detailed in the PCR testing requirements.
• Plant consignments are packed and packaged to prevent infection by X. fastidiosa
• Phytosanitary certificates issued by the NPPO with additional declarations including information that enables tracing of plant lines for export to Australia to test results and the facility in which the lines were grown.

Roles and responsibilities

The exporter and the testing laboratory should verify the requirements before the testing. We retain the right to monitor the arrangements by auditing and by sampling and testing consignments after they arrive in Australia.

The NPPO

The NPPO will provide oversight of the arrangements to ensure that they meet the requirements. The NPPO is responsible for approving facilities or authorising any independent entities that approve facilities on its behalf. We may request that the NPPO provide records relevant to the approval and management of the arrangements.

The NPPO or an entity acting under the authority of the NPPO will:

• Inspect the production facility to confirm that:
  • phytosanitary requirements are met
  • the facility is insect proof
  • the facility is constructed and equipped to achieve requirements
  • records are maintained by the producer or the facility.
• Approve the facility under the arrangement
  • audit the facility and records
• Approve sampling of mother plants by the grower for testing or take samples of mother plants
• Take official samples of the plant lot for testing
• Maintain records of:
  • approved facilities
  • audits of facilities
  • phytosanitary practices within facilities
  • laboratory tests for X. fastidiosa.
• Certify that plants exported to Australia are free from X. fastidiosa. This will be based on evidence of systems that confirm:
  • plants were kept in the registered facility throughout their life, from propagation to export
  • testing has been undertaken by a competent laboratory and laboratory reports have been sighted.

• Phytosanitary Certificates must include:

  • additional declarations that indicate the status of the plant lot
  • the facility approval code/number
  • the laboratory report code/number.

  The grower

  The grower will ensure that:

  • phytosanitary conditions for Australia are met
  • plants are appropriately tested by an approved testing laboratory
  • samples of mother plants for testing are provided, if approved by NPPO
  • the facility is free from X. fastidiosa
  • the facility is insect-proof.

  The grower will also ensure that

  • complete records are maintained of:
    • plant lot identifying numbers or codes
    • the parentage of the exported plant lots
    • dates that plants are introduced to the facility
    • mother plants, i.e. plants that are the immediate propagation source from which plant lots were propagated
    • pathogens detected in plants in the facility
    • arthropods found in the facility
    • any plant material destroyed and the reason for the destruction.
  • all plants for export to Australia are transported in insect-proof closed containers or packaging.

  The testing laboratory

  The testing laboratory is approved by the NPPO as competent to undertake the testing required by Australia, using the prescribed testing methods.

  It will:

  • use the approved test protocol
  • record the plant lots or mother plants that are tested and the number of samples tested
  • provide the evidence of tests, results and operating processes to the NPPO, as required.

  Diagram 1: How the management system works

PCR testing will require two tests:

• rimM gene sequence real-time PCR test from Harper et al. (2010)¹
• conventional PCR from Minsavage et al. (1994)² or an equivalent PCR that detects X. fastidiosa sub-species pear leaf scorch (PLS).

To ensure accurate testing:

• The sample size (number of units) will be set according to Table 1 in ISPM 31³

  Ensuring a 95% confidence level that X. Fastidiosa is not present in more than 0.5% (level of detection) of each lot. A unit defined as an individual plant.

• Two tissue samples per unit will be tested from tissue cultures, bulbs and corms.
• Three tissue samples per unit will be tested from nursery stock plants and cuttings.
• Mid-ribs of leaves must be included in the sample if the plant has leaves.
• Living tissue with vascular structures will be sampled if the material has no leaves.

Note: a unit is a tissue culture plantlet, nursery stock plant, a bulb or a corm.

For efficient PCR testing:

• DNA extracted from up to 10 samples may be tested in a single PCR as a pool or batch, where a sample is defined as a single piece of tissue.
• samples from different species should not be pooled.

For accurate results:

• House-keeping gene positive controls must be run for each batch of tests to confirm that the DNA was extracted successfully.
• House-keeping positive controls must be run for each different plant species.

For proper documentation and certification:

• The laboratory must record the plant lots and mother plants that are tested and the number of samples tested.
• If the testing is conducted prior to export (nursery stock only) the:
  • exporting NPPO must verify the laboratory report and retain a copy, and
  • identifying code or number of the laboratory report must be provided on the Phytosanitary Certificate.

Harper, S. J., Ward, L. I., and Clover, G. R. G. (2010). Development of LAMP and real-time PCR methods for the rapid detection of Xylella fastidiosa for quarantine and field applications. Phytopathology 100:1282-1288.

Minsavage, G.V., Thompson, C.M., Hopkins, D.L., Leite, M.V.B.C. and Stall, R.E. (1994) Development of a Polymerase Chain Reaction protocol for detection of Xylella fastidiosa in plant tissue. Phytopathology 84: 456-461.

ISPM 31 in International Standards for Phytosanitary Methods, No. 1 to 32 (2009), the Secretariat of the International Plant Protection Convention, Food and Agriculture Organization of the United Nations, Rome. pp. 401–420. https://www.ippc.int/en/publications/588/