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Endophytes in Changing Environments - Do Group, Dresden University of Technology, Pienner Str

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Endophytes in Changing Environments - Do Group, Dresden University of Technology, Pienner Str Short Communication - doi: 10.3832/ifor0932-006 ©iForest – Biogeosciences and Forestry (1) Molecular Physiology of Woody Plants Endophytes in changing environments - do Group, Dresden University of Technology, Pienner Str. 7, D-01737 Tharandt we need new concepts in forest (Germany); (2) Institute of Silviculture and Forest Protection, Dresden University of management? Technology, Pienner Str. 8, D-01737 Tharandt (Germany) Doris Krabel (1), Kristin Morgenstern (1), Sven Herzog (2) @ Doris Krabel ([email protected] dresden.de) Received: Dec 17, 2012 - Accepted: Feb 05, The occurrence of endophytic fungi, hosted by living tissues of forest trees 2013 seems to be a common phenomenon. Numerous studies show that these colo- nists are mostly symptomless or even live in a symbiotic relationship to the Citation: Krabel D, Morgenstern K, Herzog host plant. Our investigations on Douglas-fir and Rhabdocline needlecast show S, 2013. Endophytes in changing environ- that Rhabdocline pseudotsugae (Sydow), which has been described exclusively ments - do we need new concepts in forest as an obligatory needle pathogen up to now, is able to persist symptomless in management? iForest 6: 109-112 [online different types of plant tissues and therefore an endophytic lifestyle has to be 2013-03-05] URL: http://www.sisef.it/ assumed. Whether this lifestyle is part of the infection strategy of the fungus is iforest/contents/?id=ifor0932-006 still unclear. However, examples of other wood associated fungi lead us to the Communicated by: Marco Borghetti hypothesis that environmental such as climate conditions are able to trigger the phenomenon of changing from a mutualist to a virulent parasite. problematic under certain host-related stress Keywords: Douglas-fir, Endophytes, Rhabdocline Needlecast, Climate Change conditions. Material and methods Introduction needles. Finally the needles become brown Material and methods are following Mor- It has been well described by Stone (1987), and die (Stephan 1981, Butin 1996). A re- genstern & Krabel (2013). In short: buds, Carroll & Carroll (1978), McCutcheon et al. peated infection causes a significant decrease needles, cambial meristem as well as em- (1993) and Sieber (2007) that for Douglas- in growth or even the death of the infected bryos were sampled from a 20-year-old con- fir Rhabdocline parkeri (order Heliotiales, trees (Stephan 1981). Up to now it seems to trolled cross-breeding population. Around Kirk et al. 2001) is the dominant needle en- be clear that ascospores infect the sprouting 94% of the trees showed clear symptoms of dophyte with strict host specificity. This needles and afterward they directly penetrate infection by some type of pathogenic needle fungus infects single epidermal cells, which through the needle epidermis into the host cast. Additionally apparently healthy looking die due to the infection. The microorganism cells (Van Vloten 1932, Lyr 1958, Stephan trees from the Forest Botanical Garden Tha- persists in the needle as a multicellular thal- 1980, 1981, Butin 1996). The current state randt were used as a negative control (non- lus without any further growth. In contrast to of knowledge on the morphology and life- infected). Needles with fruiting bodies of Rhabdocline parkeri, the closely related spe- cycle of the fungus is mainly based on ma- Rhabdocline pseudotsugae were collected in cies Rhabdocline pseudotsugae, Rhabdocli- croscopic and microscopic investigations of May 2011 serving as positive control. Em- ne oblonga, Rhabdocline obovata, Rhabdo- characteristic fungal fruiting bodies, spores bryos were dissected from seeds which ori- cline epiphylla and Rhabdocline weirii are and needles showing symptoms of infection. ginated from a controlled cross-breeding of fungi which induce Rhabdocline needlecast, The present study is the first investigation interior and coastal varieties of Douglas-fir. one of the most economically important dis- which verifies Rhabdocline pseudotsugae in Genomic DNA was isolated from all types eases of Douglas-fir (Catal et al. 2010), be- different type of plant material other than of plant tissues described above. For isola- cause it can cause substantial losses in Chri- needles. There are no previous investigations tion of genomic plant DNA an extraction stmas tree plantations (Chastagner 2001). In which show that the fungus shows an endo- protocol, modified after Doyle & Doyle 1930 the pathogen of Rhabdocline needle- phytic lifestyle. Our report is focused on the (1987) was used (Morgenstern & Krabel cast was first described for Germany by v. transmittance of the pathogen beside the re- 2013). For extraction of fungal DNA the Geyr (1930). A detailed description of dis- gular distribution via ascospores, placed in DNeasy Plant Mini Kit (Qiagen GmbH, ease symptoms and life cycle can be found at the context of the idea that Rhabdocline Hilden, Germany) was used. Based on stu- Van Vloten (1932), Stephan (1981) and pseudotsugae behaves in part as a latent pa- dies by Catal (2002), a nested PCR protocol Butin (1996). Rhabdocline pseudotsugae thogen and that this way of life can become was established for the detection of Rhabdo- which is classified into the class of Ascomy- cota is a highly specialized parasite whose one-year life cycle is closely linked to the Tab. 1 - Screening of different type of plant material (varieties and tissue). seasonal growth cycle of Douglas-fir needles Material (Van Vloten 1932). Small yellow-green Number of samples infected (number of genotypes Variety spots are the first indication of the disease. with Rhabdocline pseudotsugae They occur in autumn on this year’s needles. x sampling location) During winter, the spots become necrotic 3 x needles, cambium, viridis x glauca 1 x needles, cambium positive; buds negative and brown. The characteristic orange-yellow buds 1 x needles and cambium positive, buds negative to rust-colored fruiting bodies mature in 1 x needles, cambium, buds positive May/June on the needle surface. High air hu- 11 x needles viridis 11 x negative midity promotes the process of ascospore re- 4 x needles glauca 4 x negative lease and infection of the young sprouting 8 x embryo glauca x viridis 3 x positive, 5 x negative 15 x embryo viridis x glauca 5 x positive, 10 x negative © SISEF http://www.sisef.it/iforest/ 109 iForest (2013) 6: 109-112 Krabel D et al. - iForest 6: 109-112 cline pseudotsugae. Primers ITS1F, ITS4, Schulz & Boyle (2005), Stone & Petrini are known to adopt a non-pathogenic beha- RPP1 and RPP4 were used according to (1997) and Stone et al. (2004). vior under limited oxygen ratio and/or nutri- Catal (2002). PCR conditions are described Although the endophyte-plant interactions ent, while they may switch to parasitism in detail by Morgenstern & Krabel (2013). are products of evolutionary development when the above conditions change (e.g., by For the examination of embryos the Phire (Sieber 2007), the lifestyle (mutualism, com- wounding - Oses et al. 2008). In this case the Plant Direct PCR Kit (Finnzymes, Part of mensalism, parasitism) of a microorganism, host is latent infected by “non-active patho- Thermo Fisher Scientific, Espoo, Finland) which is living inside a plant is not always gens” until internal cell conditions change was used. PCR was carried out with primer obvious and the question may rise for what and the endophyte becomes virulent. The la- pair ITS1F/ITS4 according to the instruc- is such a relationship good for? A more or tent infection hypothesis is proposed by tions of the manufacturer in a total volume less mutualistic relationship occurs in the Boddy & Rayner (1983). This hypothesis is of 20 µl. case the endophyte provides the host plant closely connected to the assumption that a Fragment analysis of PCR products was with some ecological advantages. On the certain density of tissue colonization is re- carried out with the ALFexpress II sequencer other hand, some apathogenic endophytes quired for changing into a pathogenic beha- (Amersham Pharmacia Biotech Europe may become virulent (Brown et al. 1998, vior. Sieber (2007) reports that as soon as GmbH, Freiburg, Germany) in a polyacry- Misaghi & Dondelinger 1990) under certain colonization density of the endophyte rea- lamide gel (ReproGel High Resolution, GE environmental conditions or by mutation ches a certain threshold the plant organ (e.g., Healthcare Europe GmbH, Munich, Ger- (Freeman & Rodrigez 1994). In general, it is needles) may die. This threshold can be rea- many). suspected that the endophyte-plant interac- ched very soon in case the living conditions tion is as prevalent as the mycorrhiza-plant of the host become unfavorable, e.g., by a Results and discussion interaction, with advantages for both part- lack of light in dense stands. ners (Carroll 1988, Sieber 2007). By the pro- As stated by Sieber (2007), depending on General aspects of an endophytic life- duction of specific metabolites and biologi- the selection pressure, it may also be the case style cally active chemicals which strengthen the that a pathogenic endophyte may change into Hawksworth et al. (1995) suggested that plant, it reduces the level of herbivore dama- non-pathogenic and back again. before using the term endophyte one should ge, e.g., gall insects. A successful interaction clearly define it. We will follow the defini- can also be characterized by an increased Hints for an endophytic lifestyle of tion of Saikkonen et al. (1998, 2004) which resistance of the host against pathogens by Rhabdocline pseudotsugae and its con- defines endophythes
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