Publications
NIBIOs employees contribute to several hundred scientific articles and research reports every year. You can browse or search in our collection which contains references and links to these publications as well as other research and dissemination activities. The collection is continously updated with new and historical material.
2006
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2005
Authors
Lisbeth Mehli Trygve D. Kjellsen Frances M. Dewey Ari M. HietalaAbstract
Strawberry Fragariax ananassa (cv. Korona) was inoculated with Botrytis cinerea by dipping berries in a conidial suspension. The colonization of the pathogen was monitored with real-time PCR, ELISA and ergosterol assays, the first showing the highest sensitivity. The expression of pathogen -tubulin and six polygalacturonases (Bcpg1-6) and three host defense genes (polygalacturonase inhibiting protein (FaPGIP) and two class II chitinases) were monitored with real-time RT-PCR. The maximum transcript levels of the host defense genes occurred at 16 hours post inoculation (hpi), at the presumed initial penetration stage.The unique transcript profile of Bcpg2 over the 96-hour-long incubation time and its high transcript levels relative to those of the other Bcpgs at 8 to 24 hpi suggest that the gene has a specific role in the penetration stage.Bcpg1 was constitutively expressed at a relatively high level in actively growing mycelia throughout the experimental period. Comparison of the transcript profiles indicated that Bcpg1 and Bcpg3-6 were co-ordinately regulated.
Authors
Magnus Karlsson Ari M. Hietala Harald Kvaalen Åke Olson Halvor Solheim Jan Stenlid Carl Gunnar FossdalAbstract
No abstract has been registered
Abstract
This paper describes the use of quantitative real-time PCR for monitoring colonization of birch wood (Betula pubescens) by the white-rot fungus Trametes versicolor in an EN113 decay experiment. The wood samples were harvested after 4, 8, 12, 16 and 20 weeks of incubation.The mass loss was in the range of 440%. Chitin and ergosterol assays were conducted for comparison. Second-order polynomial fits of the mass loss of decayed wood versus chitin, ergosterol and DNA gave correlations (r2) of 0.87, 0.61 and 0.84, respectively. Compared to the other two assays employed, real-time PCR data correlated best with the relative mass loss of decayed samples 48 weeks after inoculation, while the saturation and decline of DNA-based estimates for fungal colonization 1620 weeks after inoculation indicated that the DNA assay is not suited for quantification purposes in the late stages of decay.The impact of conversion factors, extraction efficiency, inhibitory compounds and background levels in relation to the three detection assays used is discussed.
Authors
Nina Jøhnk Ari M. Hietala Carl Gunnar Fossdal David B. Collinge Mari-Anne NewmanAbstract
To study the mechanisms of inducible disease resistance in conifers, changes in transcript accumulation in roots of Norway spruce (Picea abies (L.) Karst.) seedlings exposed to the root rot pathogen Ceratobasidium bicorne Erikss. and Ryv. (anamorph: Rhizoctonia sp.) were monitored by differential display (DD). Because C. bicorne attacks root tips, a desiccation treatment was added to exclude genes induced by pathogen-related desiccation stress. The DD analysis was defined by the use of 11 sets of primers, covering about 5% of the transcriptome. A comparison of gene expression in control, desiccation- and pathogen-stressed roots revealed 36 pathogen-induced gene transcripts. Based on database searches, these transcripts were assigned to four groups originating from spruce mRNA (25 transcripts), rRNA (five transcripts), fungal mRNA (two transcripts) and currently unknown cDNAs (four transcripts). Real-time PCR was applied to verify and quantify pathogen-induced changes in transcript accumulation. Of the 18 transcripts tested, nine were verified to be Norway spruce gene transcripts up-regulated from 1.3- to 66-fold in the infected roots. Four germin-like protein isoforms, a peroxidase and a glutathione S-transferase, all implicated in oxidative processes, including the oxidative burst, were predicted from sequence similarity searches. Seven class IV chitinase isoforms implicated in fungal cell wall degradation and a nucleotide binding site-leucine rich repeat (NBS-LRR) disease resistance protein homologue related to pathogen recognition were identified. Several transcript species, such as the NBS-LRR homologue and the germin-like protein homologues, have not previously been identified as pathogen-inducible genes in gymnosperms.
Abstract
In spring 2002, extensive damages were recorded in southeast Norway on nursery-grown Norway spruce seedlings that had either wintered in nursery cold storage or had been planted out in autumn 2001. The damages were characterised by leader shoot dieback and necroses on the upper or lower part of the 2001-year-shoot. Gremmeniella abietina and Phomopsis sp. were frequently isolated from the diseased seedlings. RAMS (random amplified microsatellites) profiling indicated that the G.abietina strains associated with diseased nursery seedlings belonged to LTT (large-tree type) ecotype, and inoculation tests confirmed their pathogenicity on Norway spruce. Based on sequence analysis of the internal transcribed spacer (ITS) regions of ribosomal DNA, the Phomopsis strains associated with diseased seedlings do not represent any characterized Phomopsis species associated with conifers. Phomopsis sp. was not pathogenic in inoculation tests, this implying it may be a secondary colonizer. ITS-based real-time PCR assays were developed in order to detect and quantify Gremmeniella and Phomopsis in the nursery stock. We describe here the Gremmeniella - associated shoot dieback symptoms on Norway spruce seedlings and conclude that the unusual disease outburst was related to the Gremmeniella epidemic caused by the LTT type on large pines in 2001.
Authors
Carl Gunnar Fossdal Ari M. HietalaAbstract
The root-rot causing fungus Heterobasidion annosum can attack both spruce and pine trees and is the economically most damaging pathogen in northern European forestry. We have monitored the Heterobasidion annosum S-type (fairly recently named H. parviporum) colonization rate and expression of host chitinases and other host transcripts in Norway spruce material with differing resistances. Transcript levels of three chitinases, representing classes I, II and IV, were monitored with real-time PCR. We have also transferred a Class IV chitinase to Arabidopsis as well as its promotor in GFP and YFP reporter constructs. Ramets of two 33 -year-old clones differing in resistance were employed as host material and inoculation and wounding was performed. Multiplex real-time PCR detection of host and pathogen DNA was also performed to follow the colonization of the host tissues by the pathogen and the collapse in host DNA levels in infected regions. Host defense transcript levels, as an indicator of the host defense response, were monitored with singleplex real-time PCR. Three days after inoculation, comparable colonization levels were observed in both clones in the area immediately adjacent to inoculation. Fourteen days after infection, pathogen colonization was restricted to the area immediately adjacent to the site of inoculation for the strong clone (589), but had progressed further into the host tissue in the weak clone (409). Transcript levels of the class II and IV chitinases increased following wounding or inoculation, while the transcript level of the class Ichitinase declined following these treatments. Transcript levels of the class II and class IV chitinases were higher in areas immediately adjacent to the inoculation site in 589 than in similar sites in 409 three days after inoculation, suggesting that the clones differ in the rate of pathogen perception and host defense signal transduction. This an earlier experiments using mature spruce clones as substrate indicate that it is the speed of the host response and notmaximum amplitude of the host response that is the most crucial component in an efficient defense in Norway spruce toward pathogenic fungi such as H. annosum.
Authors
Nina Jøhnk Ari M. Hietala Carl Gunnar Fossdal Mari-Anne Newman David B. Collinge Harald KvaalenAbstract
Stilbene synthases make the backbone of stilbenes in a single enzymatic step. Many stilbenes are stressinduced antimicrobial phenolics, believed to work in disease resistance. In conifers, stilbenes are found in pine (Pinus), spruce (Picea) and a few other genera.Stilbene synthase isoforms in pine use cinnamyl-CoA to form pinosylvin, these are termed pinosylvin synthases, whereas stilbene synthases in spruce use pcoumaryl- CoA to form resveratrol and are sometimes termed resveratrol synthases.Pinosylvin has been found to be more effective than resveratrol in inhibiting fungal growth and wood decay (Seppnen et al. 2004), and pathogens of non-pinosylvin producing species have been found to be less tolerant of pinosylvin than pine pathogens (Seppnen et al. 2004). In the present study, Norway spruce (Transformation of Norway spruce with the pinosylvin synthase gene, PSS1) was transformed using the biolistic technique with a gene encoding pinosylvin synthase, PSS1, from Scots pine and the E. coli nptII antibiotic resistance gene.Vector constructs carrying PSS1 in sense and antisense, as well as control vectors without PSS1 were transferred into two embryogenic cell lines of Norway spruce, 11703-B63 and 186-3C. Selection condition for transgenic tissue was conferred by nptII in combination with the antibiotic geneticin. Geneticin resistant lines were recovered from all transformation events, a total of 55 lines.NptII was detected by PCR analysis in many of these lines, the majority derived from the cell line 11703 B63. However, nptII protein was detected in just five lines, and several lines of evidence indicate that the transgenic lines obtained in this study might be chimaeras.Fifty-six seedlings were successfully regenerated from antibiotic resistant lines, 50 of these were derived from cell line 11703 B63. All seedlings died during cold storage before further testing could be carried out.