Performance of diagnostic tests for the detection and identification of Pseudomonas syringae pv. actinidiae (Psa) from woody samples

Loreti, Stefania; Cunty, Amandine; Pucci, Nicoletta; Chabirand, Aude; Stefani, Emilio; Abelleira, A.; Balestra, Giorgio Mariano; Cornish, D. A.; Gaffuri, Francesca; Giovanardi, Davide; Gottsberger, Richard; Holeva, Maria C.; Karahan, Aynur Gül; Karafla, C. D.; Mazzaglia, A.; Taylor, Robert K.; Cruz, Leonor; López, Marı́a M.; Vanneste, J. L.; Poliakoff, F.

doi:10.1007/s10658-018-1509-5

Cited by 13 publications

(11 citation statements)

References 22 publications

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“…47 and Loreti, et al . 48 . The first scenario considered that an indeterminate result would be further assessed by the laboratory, and would always be rated “as expected” (i.e.…”

Section: Methodsmentioning

confidence: 99%

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

Ioos¹,

Aloi

Piškur

et al. 2019

Sci Rep

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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.

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“…47 and Loreti, et al . 48 . The first scenario considered that an indeterminate result would be further assessed by the laboratory, and would always be rated “as expected” (i.e.…”

Section: Methodsmentioning

confidence: 99%

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

Ioos¹,

Aloi

Piškur

et al. 2019

Sci Rep

View full text Add to dashboard Cite

show abstract

“…The molecular tests for screening recommended below have been selected based on test performance study results reported in Loreti et al (2018) except for the LAMP Ruinelli et al (2017). DNA extraction is described in Appendix .…”

Section: Detectionmentioning

confidence: 99%

“…Two test performance studies showed that isolation has a lower analytical and diagnostic sensitivity than the molecular tests (Loreti et al, 2014, 2018).…”

Section: Detectionmentioning

confidence: 99%

PM 7/120 (2) Pseudomonas syringae pv. actinidiae

2021

EPPO Bulletin

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“…In recent decades, rapid, specific and sensitive DNAbased methods have been made available for the detection of pathogens in plant material (Vincelli and Tisserat 2008). PCR and its variations, such as BIO-PCR and real-time PCR (qPCR), have become in many cases the standard methods for the detection and identification of phytopathogenic bacteria (Palacio-Bielsa et al 2009;Bull and Koike 2015;Loreti et al 2018).…”

Section: Introductionmentioning

confidence: 99%

Detection of Xanthomonas citri pv. viticola on grapevine by real-time PCR and BIO-PCR using primers designed from pathogenicity and xanthomonadin gene sequences

et al. 2019

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Grapevine bacterial canker caused byXanthomonas citri pv. viticola (X. campestris pv. viticola) (Xcv), was detected in Brazil in 1998 and is currently regarded as a quarantine disease with limited distribution in the country. To improve sensitivity and speed in the detection of Xcv in asymptomatic grapevines, two pairs of primers were designed, targeting sequences of a pathogenicity gene (hrpB) and the xanthomonadin coding cluster. Both pairs were tested in conventional PCR (cPCR) and real-time PCR (qPCR) formats. Primers targeting the hrpB gene showed cross reactions with other Xanthomonas spp. but were effective for use in both cPCR and qPCR, whereas primers for the xanthomonadin gene were highly specific for Xcv but showed low efficiency in qPCR. Enrichment of plant extracts in semi-selective medium before qPCR allowed a significant increase in sensitivity when compared to total DNA extraction, making it possible to detect as low as 10 1 CFU ml -1 . Under natural infection conditions, symptomatic and asymptomatic grapevines were tested by qPCR with hrpB primers and cPCR with xanthomonadin primers. In both cases, plant extracts were enriched for 36-72h. Xcv was detected in all symptomatic samples by qPCR and the result was confirmed by cPCR. For the asymptomatic samples, Xcv was detected in 93.4% with qPCR and in 89.5% with cPCR. These two methods offer advantages in terms of sensitivity and specificity, and they could be useful in quarantine programs, certification of grapevine propagating material and detection of inoculum sources in alternative hosts, contributing to the prevention of pathogen spread to disease-free areas.Keywords Grapevine bacterial canker . Vitis vinifera . Xanthomonas campestris pv. viticola. PCR-based diagnosis . qPCR

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Performance of diagnostic tests for the detection and identification of Pseudomonas syringae pv. actinidiae (Psa) from woody samples

Cited by 13 publications

References 22 publications

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

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

PM 7/120 (2) Pseudomonas syringae pv. actinidiae

Detection of Xanthomonas citri pv. viticola on grapevine by real-time PCR and BIO-PCR using primers designed from pathogenicity and xanthomonadin gene sequences

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