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Seeds ToBRFV –A pilot for harmonization of diagnostic protocols for seed
pests focused on ToBRFV. The NAPPO Seeds Expert Group successfully concluded the
ring test on the pilot project on harmonizing protocols for the detection of Tomato Brown Rugose Fruit
Virus (ToBRFV) in tomato and pepper seeds. Nine laboratories from the NAPPO region tested five
ToBRFV detection protocols, three conventional PCR-based (endpoint) and two Real Time PCR-
based, generating a total of 3,680 data points. Analytical, diagnostic, and calibrator samples were
used in this comparative study to evaluate the precision (repeatability, intermediate precision, and
reproducibility), accuracy (percent recovery and amplification efficiency), and selectivity (sensitivity
and specificity) for all the protocols tested. Resulting data was uploaded to and organized using a web
-based portal specifically designed for this project. The data was analyzed and reviewed by EG
members.
Four diagnostic protocols are transferrable (protocols were used by all labs in the ring test
successfully, and they should, theoretically, produce similar results during routine use)
between NAPPO’s National Plant Protection Organizations (NPPO) and three of them demonstrated
optimal performance and reproducibility for the detection of ToBRFV in tomato and pepper seeds.
The collaboration of NPPOs, academia, and industry greatly contributed to the successful completion
of the experimental phase of this project. The Seeds EG will draft a final report that will be presented
to the NAPPO Executive Committee. A scientific publication will also be written by the group. The
results of the ring test will allow NAPPO member countries to select one or more of the tested
protocols that showed optimal results thereby harmonizing phytosanitary measures and facilitating
efficient and timely seed trade in our NAPPO region.

The ring test outcomes were presented to Seeds Canada by the NAPPO Secretariat on February 6
and a member of from the Seeds EG will present the ring test results at the International Congress of
Plant Pathology to be held in August 20-25 2023 in Lyon, France. A detailed presentation is also
planned for the upcoming NAPPO Annual Meeting in December 2023 in Merida, Yucatan, Mexico.
The NAPPO Secretariat will post all EG reports and publications to the NAPPO website
(www.nappo.org). Stay tuned for more information!

Guidance for improving the regional implementation of ISPM 25

(Consignment in-transit). A two-day face-to-face meeting (February 16 and 17) of the
members of the ISPM 25-guidance writing subgroup (WSG) took place at the APHIS PPQ
Headquarters in Riverdale, Maryland. Wendy Asbil (CFIA), Erin Stiers (APHIS PPQ), Brendon
Reardon (EG Chairperson, APHIS PPQ), Stephanie Dubon (NAPPO AMC member – APHIS PPQ)
and Alonso Suazo (NAPPO TD) participated in the meeting.
The WSG reviewed the outline and status of the draft document and prioritized the sections that need
further development. The WSG discussed the most important in-transit scenarios and non-compliance
issues related to the in-transit movement of consignments at the U. S. - Canada border. Important
communication aspects to prevent or address non-compliance issues were also discussed. The
discussions concluded with the development of a list of recommendations for industry and NPPOs.

The WSG will develop an updated draft to share with the entire EG for review and feedback.

March 2023 -Vol 43 (1) 3