Transferability of PCR-based diagnostic protocols: An international collaborative case study assessing protocols targeting the quarantine pine pathogen Fusarium circinatum

[EN] Fusarium circinatum is a harmful pathogenic fungus mostly attacking Pinus species and also Pseudotsuga menziesii, causing cankers in trees of all ages, damping-off in seedlings, and mortality in cuttings and mother plants for clonal production. This fungus is listed as a quarantine pest in several parts of the world and the trade of potentially contaminated pine material such as cuttings, seedlings or seeds is restricted in order to prevent its spread to disease-free areas. Inspection of plant material often relies on DNA testing and several conventional or real-time PCR based tests targeting F. circinatum are available in the literature. In this work, an international collaborative study joined 23 partners to assess the transferability and the performance of nine molecular protocols, using a wide panel of DNA from 71 representative strains of F. circinatum and related Fusarium species. Diagnostic sensitivity, specificity and accuracy of the nine protocols all reached values >80%, and the diagnostic specificity was the only parameter differing significantly between protocols. The rates of false positives and of false negatives were computed and only the false positive rates differed significantly, ranging from 3.0% to 17.3%. The difference between protocols for some of the performance values were mainly due to cross-reactions with DNA from non-target species, which were either not tested or documented in the original articles. Considering that participating laboratories were free to use their own reagents and equipment, this study demonstrated that the diagnostic protocols for F. circinatum were not easily transferable to end-users. More generally, our results suggest that the use of protocols using conventional or real-time PCR outside their initial development and validation conditions should require careful characterization of the performance data prior to use under modified conditions (i.e. reagents and equipment). Suggestions to improve the transfer are proposed.This work was supported by COST action FP1406 Pinestrength . The work of the Estonian team was supported by the Estonian Science Foundation grants PSG136 and IUT21-04. The work of Portuguese team from INIAV was financed by INIAV I.P. Institute. The work at U. Aveiro (Portugal) was financed by European Funds through COMPETE and National Funds through the Portuguese Foundation for Science and Technology (FCT) to CESAM (UID/AMB/50017/2013 POCI-01- 0145-FEDER-007638). The work of Slovenian team was financed through Slovenian Research Agency (P4-0107) and by the Slovenian Ministry of Agriculture, Forestry and Food (Public Forestry Service). 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Global Geographic Distribution and Host Range of Fusarium circinatum, the Causal Agent of Pine Pitch Canker

Fusarium circinatum, the causal agent of pine pitch canker (PPC), is currently one of the most important threats of Pinus spp. globally. This pathogen is known in many pine-growing regions, including natural and planted forests, and can affect all life stages of trees, from emerging seedlings to mature trees. Despite the importance of PPC, the global distribution of F. circinatum is poorly documented, and this problem is also true of the hosts within countries that are affected. The aim of this study was to review the global distribution of F. circinatum, with a particular focus on Europe. We considered (1) the current and historical pathogen records, both positive and negative, based on confirmed reports from Europe and globally; (2) the genetic diversity and population structure of the pathogen; (3) the current distribution of PPC in Europe, comparing published models of predicted disease distribution; and (4) host susceptibility by reviewing literature and generating a comprehensive list of known hosts for the fungus. These data were collated from 41 countries and used to compile a specially constructed geo-database. A review of 6297 observation records showed that F. circinatum and the symptoms it causes on conifers occurred in 14 countries, including four in Europe, and is absent in 28 countries. Field observations and experimental data from 138 host species revealed 106 susceptible host species including 85 Pinus species, 6 non-pine tree species and 15 grass and herb species. Our data confirm that susceptibility to F. circinatum varies between different host species, tree ages and environmental characteristics. Knowledge on the geographic distribution, host range and the relative susceptibility of different hosts is essential for disease management, mitigation and containment strategies. The findings reported in this review will support countries that are currently free of F. circinatum in implementing effective procedures and restrictions and prevent further spread of the pathogen

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Phytophthora ramorum infection of coast live oak leaves in Californian forests and its capacity to sporulate in vitro

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A New Report of Phytophthora ramorum on Rhamnus purshiana in Northern California

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First Report ofPhytophthora cactorumAssociated With Beech Decline in Italy

During 2006 and 2007, declining mature beech trees (Fagus sylvatica) were recorded in two stands in the Natural Park of Monti Cimini in central Italy

COST Action (FP0801): Established and emerging Phytophthora: Increasing threats to woodland and forest ecosystems in Europe

COST Action FP0801 Working Group 4 focusses on the management and control of Phytophthora diseases and comprises 63 experts from 19 European countries, Tunisia, Australia, New Zealand and the USA. The main targets of WG4 are: 1. Collate informations on established and novel control and management methods and assess their efficacy and practicability for nurseries, individual trees and forests. 2. Explore the feasibility of environmentally friendly control methods, including biological control (antagonists, suppressive plants), low-impact chemical methods (fungistatic products, surfactants, nutrition and soil ameloration), physical methods (host removal, solarisation, water management, water filtration), and host resistance. 3. Discuss the concepts and efficacies of integrated management protocols and the reasons for their success or failure. WG4 members will be reviewing their national literature, collect and assess information and experiences from their countries and submit these data and data from their own trials to a WG4 database which will be accessible for the scientific community, decision makers, practitioners and the general public through the COST Action website. At the end of this COST Action the state-of-the-art and future perspectives on control and management methods for Phytophthora diseases will be published as a book comprising key papers written by leading experts with large-scale and longterm experiences and articles on certain aspects written by qualified WG4 members

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