Assessment of Dickeya and Pectobacterium spp. on vegetables and ornamentals (Soft rot)
Wolf, J.M. van der; Bergsma-Vlami, M. ; Saddler, G. ; Hélias, V. ; Tsror, L. ; Yedida, Iris ; Pirhonen, M. ; Degefu, Yeshtila ; Tuomisto, J.T. ; Lojkowska, Ewa ; Li, Sean - \ 2020
Zenodo - 109 p.
Euphresco - Plant health - Dickeya - Pectobacterium - soft rot - vegetables - ornamentals - diagnostics - surveys - epidemiology - management
Dickeya and Pectobacterium belonging to the group of soft rot Pectobacteriaceae (SRP) are causing emerging problems in a wide range of vegetable and ornamental crops in Europe, including potato, carrot, cabbage, Chinese cabbage, celery, leek, pepper, parsley, Zantedeschia, hyacinth, Dahlia, Chrysanthemum, Philodendron, Freesia, Saintpaulia, Iris, Aglaonema, Crocus, Campanula and Phalaenopsis. The phytopathogens in both genera are genetically and phenotypically highly diverse. Disease problems in the different hosts are associated with the introduction of new variants or by spread of groups already present in Europe. Within this Euphresco project we aimed to identify and assess the risks of these new variants, and to develop management strategies, including reliable diagnostic methods to prevent introductions and further spread of SRP. To reach our goals, meetings were organized and collaborations were established with specialists worldwide. All information on meetings, protocols and activities of the Euphresco group are published on the Dickeya/Pectobacterium website, conveyed by the James Hutton Institute in Invergowrie (Scotland) (https://engage.hutton.ac.uk, contact person Dr I. Toth/Dr J. Fairly).
During the project, 1.5 days meetings were held in 2015 in Gdansk (Poland), in 2016 in Helsinki (Finland), in 2017 in Edinburgh (Scotland) and in 2018 in Emmeloord (The Netherlands). Meetings were attended by an average of 30 participants from organizations in EU member states, North- and Latin America, Africa, Asia and Australia.
One project’s objective was to develop methods for the detection and identification of Pectobacterium and Dickeya species in different matrices. For this, a panel of reference strains has been compiled for Dickeya and Pectobacterium species. Most strains have been deposited in international collections. For most strains also whole genome sequence data are available. During the course of this project, several diagnostic tests were developed and evaluated, often based on the TaqMan technology.
In several countries, surveys in potato and ornamental crops were conducted, but also in other matrices of the potato ecosystem, including water used for irrigation. In addition, new taxonomic groups that have been identified, are now new species including: P. versatile, P. aquaticum, P. fontis and P. polonicum. In potato, P. brasiliense became dominant as blackleg causing organism and has largely outcompeted D. solani in the last five years. In surface water in Europe, D. zeae was found to be the dominant SRP. In other continents, serious outbreaks of potato blackleg with other SRP has been reported, such as D. dianthicola in the USA and Australia. Various new SRP have been described, namely P. punjabense, P. peruviense, P. polaris, D. lacustris and D. zantedeschia. For the first time, D. fangzhongdai was described in Phalaenopsis. Not all species can cause potato blackleg.
Studies on various virulence factors were conducted for SRP, such as on chemoreceptors, small phenolic plant compounds interacting with signal molecules of Pectobacterium involved in the quorum sensing mechanism of the pathogen. A Tn-seq approach was developed and used to identify new virulence factors.
Information was exchanged on disease management strategies which include cultivation practices, resistance breeding, hygiene and the use of (bio-) control agents. A strict hygiene and an intensive monitoring of seed lots was found to be associated with a significant reduction of infections with blackleg causing SRP. A phage therapy has been developed to protect (seed) potato tubers against soft rot during storage. Various bacteriophages and bacterial antagonists were characterized and some evaluated for control of potato soft rot and/or blackleg. Steam treatments for seed tubers were found to decrease the blackleg incidence. Cold plasma treatment was found to kill SRP grown in vitro. Similarly, stabilized silver nanostructures killed SRP. It was found that seed potato lots can differ in suppressiveness against D. solani. Indications were found that the microbiome in tuber tissue plays a role in this.
First report of potato blackleg caused by Pectobacterium carotovorum subsp. brasiliensis in the Netherlands
Nunes Leite, L. ; Haan, E.G. de; Krijger, M.C. ; Kastelein, P. ; Zouwen, P.S. van der; Bovenkamp, G.W. van den; Tebaldi, N.D. ; Wolf, J.M. van der - \ 2014
New Disease Reports 29 (2014)2014. - ISSN 2044-0588 - p. 24 - 24.
Double-layer crystal violet pectate (DL-CVP) medium (Helias et al., 2012) was used to isolate pectinolytic bacteria from the stems of blackleg affected plants. Eleven isolates derived from ten seed lots, positive in the Pcb-specific PCR (Duarte et al., 2004), were characterised with multilocus sequence analysis (MLSA) using sequences of acnA, gapA, icdA, mdh, mtlD, pgi and proA (Panda et al., 2012) (GenBank Accession Nos. KJ576801-KJ576810, KJ596350-KJ596399, KJ608062-KJ608071). The phylogenetic analysis was done by constructing a maximum likelihood tree using sequences of reference strains P. atrosepticum, P. wasabiae, P.c. subsp. carotovorum, P.c. subsp. odororiferum, P. betavasculorum and Dickeya paradisiaca, derived from GenBank, for comparison (Fig. 1). The MLSA confirmed the identity of the Pcb strains. The Dutch strains clustered together in a homogeneous branch, which also showed that these strains were highly similar. Strains were motile, able to reduce sucrose and to grow at 39°C, resistant to erythromycin, and unable to ferment lactose according to the procedures described by Hayman et al. (1998). Two Pcb isolates were tested for virulence in a field experiment in the Netherlands in 2013. Two months after planting vacuum-infiltrated seed tubers, a high percentage of the plants (more than 30%) showed typical symptoms, whereas water-treated plants remained disease-free (Fig. 2). The pathogen was re-isolated from the diseased plants and identified as Pcb using MLSA. This is the first time Pcb is described as a potato blackleg-causing agent in the Netherlands, although a strain of Pcb isolated from potato in the Netherlands has been found in a culture collection before (De Boer et al., 2012).
Dickeya solani sp. nov., a pectinolytic plant pathogenic bacterium isolated from potato (Solanum tuberosum)
Wolf, J.M. van der; Nijhuis, E.H. ; Kowalewska, M.J. ; Saddler, G.S. ; Parkinson, N. ; Elphinstone, J.G. ; Pritchard, L. ; Toth, I.K. ; Lojkowska, E. ; Potrykus, M. ; Waleron, M. ; Vos, P. de; Cleenwerck, I. ; Pirhonen, M. ; Garlant, L. ; Hélias, V. ; Pothier, J.F. ; Pflüger, V. ; Duffy, B. ; Tsror, L. ; Manulis, S. - \ 2014
International Journal of Systematic and Evolutionary Microbiology 64 (2014)Pt. 3. - ISSN 1466-5026 - p. 768 - 774.
strains - identification - pseudomallei - chrysanthemi - relatedness - clonality - mallei - clade - genus
Pectinolytic bacteria were recently isolated from diseased potato plants exhibiting blackleg and slow wilt symptoms found in a number of European countries and Israel. These Gram-negative, motile, rods were identified as belonging to the genus Dickeya, previously the Pectobacterium chrysanthemi complex (Erwinia chrysanthemi), on the basis of a positive PCR reaction with pelADE primers, 16S rDNA sequence analyis, fatty acid methyl esterase analysis, the production of phosphatases and the ability to produce indole and acids from a-methyl glucoside. Differential physiological assays used previously to differentiate between strains of E. chrysanthemi, showed that these isolates belonged to biovar 3. Eight of the isolates, seven from potato and one from hyacinth, were analysed together with 21 reference strains representing all currently recognised taxa within the genus Dickeya. The new isolates formed a distinct genetic clade in multilocus sequence analysis (MLSA) using concatenated sequences of the intergenic spacer (IGS), as well as genes dnaX, recA, dnaN, fusA, gapA, purA, rplB, rpoS and gyrA. Characterization by whole-cell MALDI-TOF mass spectrometry, pulsed field gel electrophoresis after digestion of whole genome DNA with rare cutting restriction enzymes, average nucleotide identity analysis and DNA-DNA hybridization studies, showed that although related to D. dadantii, these isolates represent a novel species within the genus Dickeya, for which the name Dickeya solani sp. nov. (type strain IPO 2222T =LMG25993T =NCPPB4479T) is proposed
Two new effective semiselective crystal violet pectate media for isolation of Pectobacterium and Dickeya
Hélias, V. ; Hamon, P. ; Huchet, E. ; Wolf, J.M. van der; Andrivon, D. - \ 2012
Plant Pathology 61 (2012)2. - ISSN 0032-0862 - p. 339 - 345.
soft-rot erwinias - carotovora subsp atroseptica - selective medium - sp-nov. - potato - chrysanthemi - enrichment - soil
Pectolytic bacteria, including Pectobacterium spp. and Dickeya spp., are best isolated on crystal violet pectate (CVP), a semiselective medium containing pectin. The source of pectin is essential, because pectolytic bacteria are not able to degrade all of them. The aims of this study were to identify a new pectin source and to perfect formulations of semiselective CVP media to isolate the pectolytic bacteria Pectobacterium spp. and Dickeya spp. from different environmental compartments (plants, soil and water). The AG366 pectin, selected after screening six different formulations, was incorporated into single-layer (SL-CVPAG366) and double-layer (DL-CVPAG366) CVP media. Both media were compared with those based on Bulmer, Sigma-Aldrich and Slendid-Burger pectins, using 39 Pectobacterium and Dickeya strains. All strains formed deep cavities on AG366-CVPs, whereas nine did not produce cavities on Bulmer or Sigma-Aldrich media. Recovery rates were similar on DL-CVPAG366, Sigma-Aldrich and Bulmer CVPs for a given taxon, and did not differ significantly between SL- and DL-CVPAG366. Pectolytic bacteria were successfully isolated on both media from field samples of diseased potatoes, carrots, tobacco, onions, radishes and ornamentals. AG366 is thus a high-performance pectin source for the elaboration of CVP media suitable to isolate Dickeya and Pectobacterium. It is also efficient for enrichment purposes in liquid medium. The validation of AG366 as an improved source of pectin to recover the polyphagous Pectobacterium and Dickeya in different environmental compartments is essential given the current worldwide emergence and recrudescence of these bacteria.
Dickeya species: an emerging problem for potato production in Europe
Toth, I.K. ; Wolf, J.M. van der; Saddler, G. ; Lojkowska, E. ; Hélias, V. ; Pirhonen, M. ; Tsror, L. ; Elphinstone, J.G. - \ 2011
Plant Pathology 60 (2011)3. - ISSN 0032-0862 - p. 385 - 399.
soft-rot erwinias - carotovora subsp atroseptica - polymerase-chain-reaction - biovar 3 - temperature tolerances - rapid identification - chrysanthemi strains - pectate medium - 1st report - pcr
Dickeya species (formerly Erwinia chrysanthemi) cause diseases on numerous crop and ornamental plants world-wide. Dickeya spp. (probably D. dianthicola) were first reported on potato in the Netherlands in the 1970s and have since been detected in many other European countries. However, since 2004–5 a new pathogen, with the proposed name ‘D. solani’, has been spreading across Europe via trade in seed tubers and is causing increasing economic losses. Although disease symptoms are often indistinguishable from those of the more established blackleg pathogen Pectobacterium spp., Dickeya spp. can initiate disease from lower inoculum levels, have a greater ability to spread through the plant’s vascular tissue, are considerably more aggressive, and have higher optimal temperatures for disease development (the latter potentially leading to increased disease problems as Europe’s climate warms). However, they also appear to be less hardy than Pectobacterium spp. in soil and other environments outside the plant. Scotland is currently the only country in Europe to enforce zero tolerance for Dickeya spp. in its potato crop in an attempt to keep its seed tuber industry free from disease. However, there are a number of other ways to control the disease, including seed tuber certification, on-farm methods and the use of diagnostics. For diagnostics, new genomics-based approaches are now being employed to develop D. dianthicola- and ‘D. solani’-specific PCR-based tests for rapid detection and identification. It is hoped that these diagnostics, together with other aspects of ongoing research, will provide invaluable tools and information for controlling this serious threat to potato production
|A context for sustainability of natural resource use
Prins, H.H.T. - \ 2006
In: Sporen van een Gedreven Pionier: verhalen bij het afscheid van Helias Udo de Haes / Persoon, G.A., Barendse, G., Bezemer, H., Leiden : Centrum voor Milieuwetenschappen (Universiteit Leiden) - ISBN 9789051911497 - p. 119 - 124.
Evaluation of a PCR kit for the detection of Erwinia carotovora subsp. atroseptica on potato tubers
Frechon, D. ; Exbrayat, P. ; Helias, V. ; Hyman, L.J. ; Jouan, B. ; Llop, P. ; Lopez, M.M. ; Payet, N. ; Perombelon, M.C.M. ; Toth, I.K. ; Beckhoven, J.R.C.M. van; Wolf, J.M. van der; Bertheau, Y. - \ 1998
Potato Research 41 (1998)2. - ISSN 0014-3065 - p. 163 - 173.
A PCR-based kit, Probelia(TM), for the detection of Erwinia carotovora subsp. atroseptica (Eca) on potatoes was evaluated at five laboratories in four countries. The kit is based on DNA-specific PCR amplification followed by detection of amplicons by hybridization to a peroxidase-labelled DNA probe in a microplate. Specificity of the PCR primers for Eca, regardless of serogroups, was confirmed by testing against 246 bacterial, fungal and plant species. Detection limits of the assay varied little between six Eca strains in pure cultures (1.3x102 to 1.5x103 cells ml-1). When Eca-free tuber peel extract from four cultivars was inoculated with known numbers of 15 Eca strains, detection limits were more variable (1.0x101 to 6.2x103 cells ml-1 peel extract), attributed probably to inconsistency in the recovery of DNA during extraction. When the PCR assay was compared with three current commercial Eca detection methods, using naturally contaminated tubers, results matched most closely those from viable counts on a selective medium, the most sensitive method (88%), followed by enrichment ELISA (72%) and last ELISA (30%), the least sensitive method.