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Frequency of fungi (percentage colonized roots/percentage total isolates) isolated from root segments of diseased Pinus sylvestris and Picea abies bare-root seedlings from forest nurseries, afforested clear-cuts and farmland in Lithuania
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Fungi colonizing decayed roots of Pinus sylvestris and Picea abies seedlings were assessed by pure-culture isolation and direct sequencing of DNA extracted from roots collected from three environments: bare-root forest nurseries; afforested clear-cuts; and abandoned farmland. Pure-culture isolation from 1500 roots collected from 480 seedlings (240...
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... those, 77 (88·5%) were identified at least to genus level. Fungi most frequently isolated were the ascomycetes and deuteromycetes: Fusarium oxysporum, Nectria radicicola, Nectria macrodidyma, Trichoderma harzianum, Phialocephala fortinii, Penicillium spinulosum, Trichoderma viride and Zalerion varium (Table 2). Considerable fungal diversity was detected in decayed roots, even within a single root system. ...Context 2
... pooled, direct sequencing and isolation detected a total of 131 fungal taxa, 92 (70·2%) of which were identified at least to genus level (Tables 2 and 3). The overlap between the two methods was very low (S S = 0·19; Fig. 2). ...Context 3
... overlap between the two methods was very low (S S = 0·19; Fig. 2). Only 14 (10·7%) of the taxa were both sequenced and isolated; 44 (33·6%) were detected exclusively by sequencing; and 73 (55·7%) exclusively by isolation (Tables 2 and 3). ...Context 4
... in fungal communities between the environments were also confirmed by low values of Sorensen similarity indices, varying between 0·22 and 0·39 for pine, and between 0·09 and 0·26 for spruce (Fig. 3a,b). On the other hand, to some extent such low indices have arisen due to high overall diversity and the presence/absence of occasional taxa, while the dominant fungi were more abundant in different environments (Tables 2 and 3; Fig. 4a,b). Fungal diversity in decayed roots of both tree species was similar, and a total of 87 taxa of fungi was found in pine, and 93 taxa in spruce (Fig. 3c). ...Context 5
... diversity in decayed roots of both tree species was similar, and a total of 87 taxa of fungi was found in pine, and 93 taxa in spruce (Fig. 3c). Among these, 39 were unique for pine, 44 for spruce, and only 48 (36·6%) were found on both tree species, indicating that decayed roots of both tree species in most cases were colonized by different fungi (Tables 2 and 3). As stated above, the species accumulation curves did not reach asymptotic values (Figs 2 and 3), indicating that further fungal taxa would have been discovered with greater sampling effort. ...Context 6
... use of either of these methods on its own would have resulted in very different descriptions of the composition and richness of fungal taxa occupying decayed roots (Fig. 2). Fungi detected by direct sequencing were seldom or never isolated into pure culture (Tables 2 and 3), indicating that some might be unculturable. For example, Chalara sp. ...Context 7
... most common fungi isolated from decayed roots in nurseries in the present study (Table 2; Fig. 4a) were also commonly isolated from diseased conifer roots in other forest nurseries in Europe ( Unestam et al., 1989;Ericson et al., 1991;Lilja et al., 1992;Kacprzak, 1997). This may suggest that similar means for control of root diseases could be applicable. ...Context 8
... all sites, except for pine from the clear-cuts, the taxon most frequently detected by direct sequencing of decayed tissue was P. fortinii, but it was seldom isolated (Tables 2 and 3; Fig. 4b). In contrast, in a previous study P. fortinii was the most commonly isolated taxon from healthy mycorrhizal conifer seedling roots (from 29% of over 8000 root tips), and was only seldom recorded by direct sequencing (from 0·8% of 150 root tips) (Menkis et al., 2005). ...Context 9
... dominant taxa detected in decayed roots of both pine and spruce seedlings were very similar, and included F. oxysporum, N. radicicola, N. macrodidyma, P. fortinii, T. harzianum and P. spinulosum (Tables 2 and 3), together comprising 68·6% of the total isolated/sequenced fungal community. Despite this, the overall fungal communities, as indicated by Sorensen similarity indices (S S = 0·54; Fig. 3c), differed between tree species, suggesting that the many taxa (63·4%) detected exclusively in either pine or spruce might specifically inhabit one or the other host. ...Context 10
... general, highly diverse fungal communities were found, and the species accumulation curves indicate that more taxa are likely to be found with increased sampling effort (Figs 2 and 3). In the present study c. 30% of the taxa remained unidentified (Tables 2 and 3). This is not surprising in that only c. 5% of the fungi have been described so far (Hawksworth, 1991(Hawksworth, , 2001). ...Similar publications
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... Among the other most common fungi, several OTUs of Fusarium were detected ( Table 2). Fungi from the genus Fusarium are ubiquitous in different environments [37,38] and include economically important plant pathogens, which may cause tree diseases in both managed and natural forest ecosystems [39,40]. For example, F. sporotrichioides is a soil-borne pathogen, which is often observed in cold climate zones such as in Northern Japan, Northern USA, or Northern Europe. ...
We investigated the diversity and occurrence of wood wasps in Lithuania and determined communities of associated fungi. Trapping of wood wasps resulted in three different species, including Sirex juvencus, Urocerus gigas, and Tremex fuscicornis. Fungal culturing from adult females of T. fuscicornis mainly resulted in fungi from the genera Penicillium and Trichoderma. High-throughput sequencing of ITS2 rDNA resulted in 59,797 high-quality fungal sequences, representing 127 fungal OTUs. There were 93 fungal OTUs detected in U. gigas, 66 in S. juvencus, and 10 in T. fuscicornis. The most common fungi were Fusarium sporotrichioides (63.1% of all fungal sequences), Amylostereum chailletii (14.9%), Penicillium crustosum (7.8%), Microascus sp. 2261_4 (5.0%), and Pithoascus ater (2.1%). Among these, only A. chailletii was found in all three insect species with the highest relative abundance in U. gigas (15.2%), followed by S. juvencus (7.7%), and the lowest in T. fuscicornis (0.3%) (p < 0.0003). Correspondence analysis of fungal communities showed a distant placement of different species of wood wasps, indicating that fungal communities in each of these were largely different. In conclusion, the study showed that the economically important tree pathogen A. chailletii was among the most common fungal OTUs vectored by siricid wood wasps.
... Forest nurseries, owing to the intensive seedling production, stand out as highly favourable environments for fungal pathogens. The dense cultivation systems create conditions favourable for the establishment of fungal pathogens, and the large monocultures increase the risk for rapid spread of fungal infections among the seedlings (Menkis et al. 2006). ...
... Damping-off and root rot diseases are often seed-transmitted and reduce germination or damage young emerging seedlings (Cram & Fraedrich 2010;Lilja & Poteri 2013). The most important pathogens causing damping-off diseases in conifer seedling production belong to the genera of Fusarium, Phytophthora, Pythium, Rhizoctonia, Cylindrocladium, and Neonectria (Sutherland et al. 2002;Menkis et al. 2006;Cram & Fraedrich 2010;Lilja & Poteri 2013). Shoot blight, caused by S. conigenus is another seedtransmitted disease that can also damage current-year shoots (Figure 2) (Sutherland et al. 2002). ...
Fungal infections in forest nurseries can significantly affect the economic sustainability of tree seedling production. Increased knowledge of local fungal communities, especially prevalent pathogens, could improve preventative and targeted control strategies, ultimately reducing the usage of fungicides. This thesis aimed to acquire new knowledge and advance a general understanding of seedborne, airborne, and foliar fungal communities in Swedish forest nurseries. The studies were conducted in five forest nurseries. Fungal communities were assessed from Picea abies, Pinus sylvestris, and Larix sp. seeds differentiated between the surface and the tissue (paper I), non-symptomatic P. sylvestris needles (paper II), and deposited spores (paper IV) using high-throughput sequencing of the ITS2 rDNA region. Results showed that seedborne fungal communities were distinguished between the seed surface and the seed tissue, where fungi from the seed surface showed a strong host-affinity and regional dependence. This result suggests that the movement of seeds between different geographic regions, including frequently traded seeds, could be a potential source of the spread and introduction of fungal species. The foliar and airborne fungal communities comprised high species richness and showed clear temporal shifts over the seedling growing seasons. The experiments included microbial treatments to control fungal pathogens and stimulate seedling growth on P. sylvestris seedlings. However, the use of microbial treatments was not found to impact the seedling growth or the disease incidence, which was generally low, and had no impact on the foliar fungal communities. All the fungal communities showed a high prevalence of nursery fungal pathogens, e.g., Cladosporium sp., Botrytis cinerea, Phoma herbarum, and Sydowia polyspora. In this thesis, the disease incidence of Diplodia sapinea in P. sylvestris seedlings was confirmed (paper III). Additionally, low abundances of D. sapinea were detected from the seeds and the deposited airborne spores. The findings in this thesis may contribute to developing disease management strategies in forest nurseries owing to a better understanding of fungal communities, thereby highlighting the importance of managing fungal pathogens to maintain healthy tree seedling production.
... 해되고 있다 [5]. 특히 Fusarium spp.는 전 세계적으로 양묘장에 흔히 발생하는 곰팡이병 병원균으 로, 독일가문비와 같은 침엽수에 모잘록병 및 뿌리썩음병을 발생시켜 피해를 유발하는 것으로 알려져 있다 [6,7] 일반적으로 양묘장에서 곰팡이병 방제를 위해 합성농약인 살균제가 사용되고 있다 [11]. ...
... For species identification, representative cultures from each morphological morphotype were subjected to sequencing of ITS rDNA. Isolation of DNA, amplification, and sequencing followed the methods described by Menkis et al. [32]. Amplification by PCR was performed using primers ITS1F [33] and ITS4 [28]. ...
In 2019, the bark beetle Ips amitinus (native to central Europe) was identified in southwestern Siberia at a distance exceeding 2500 km east of its previously known easternmost location in the European part of Russia. In Siberia, its invasive populations are characterised by high abundance and harmfulness. Here, I. amitinus accomplishes primary attacks on standing vital trees of Pinus sibirica with a lethal outcome. This invasion has already resulted in massive dieback in stands of pine over a large geographic territory. By, 2021, the invaded area was estimated to cover at least 31,200 km 2. The objectives of this study were to investigate fungi associated with/vectored by I. amitinus in its invasive area in southwestern Siberia and wood decay fungi that cause root and butt rots to P. sibirica. This led to the following conclusions: (i) DNA analysis of sixty adult beetles of Ips amitinus collected from P. sibirica in southwest Siberia revealed the presence of 143 fungal taxa; (ii) species richness was significantly higher in beetles collected from dead branches than from (more recently infested) dying branches; (iii) fungal communities were >90% dominated by yeasts, among which the most common were Nakazawaea holstii, Kuraishia molischiana, and N. ambrosiae; (iv) entomopathogenic Beauveria bassiana s.l. was the most common fungus isolated from dead/mycosed beetles of I. amitinus, followed by Lophium arboricola and four Ophiostoma spp.; and (v) Heterobasidion parviporum was the most common decay fungus detected, which was causing heart rot in stems of P. sibirica.
... Root rot fungi, which represent a serious threat to nurseries, restrict the generation of dry matter and specifically injure plants, according to Aigbe and Remison [22]. Favorable circumstances include abundant rainfall, high spring and summer temperatures, and weak soil structure promote the growth of pathogen fungal colonies [23,24]. ...
... mm in diameter. Brown and barrel shaped Monilioid cells arising from the sclerotia with measured 15-38 × 10-15μmin size [23,24,41]. ...
Root rot disease is one of the most serious diseases that significantly reduces the plant's productivity, commonly known as collar rot, foot rot, or damping off. The main symptom of this condition is damping off. The newly emerging seedlings collapse and die in areas with severe infection, and the majority of seedlings may die before or after emergence. Due to Root rot disease in C. Deodara which is our state tree, the damage caused by it to the seedlings there was urgent need to identify and study the disease so as to develop proper management strategies. In Himachal Pradesh, a field survey was carried out to know the disease incidence of root rot pathogen in the Cedrus growing regions in district Solan. The survey included the Kanaghat, Solan, and Chail mountains. The Kandaghat range covered Kiari, Karol, and Bisha nurseries, the Solan range Shilli, Nauni nurseries, and the Chail range Chiunth, Chail, and Gaura nurseries were all covered. During the survey of root rot pathogen of Cedrus deodara, 10.90 to 62.15%incidence was recorded in district Solan nurseries. Maximum incidence of the root rot was recorded in Kandaghat range (37.35%) followed by Solan range (33.85%) and Chail range (30.93%), respectively. Amongst nurseries found that,Bisha nursery had maximum incidence (62.15%) followed by Nauni (48.46%) and Chiunth nurseries (44.65), respectively. Cultural and morphological characteristics of root rot pathogen were studied and identified as Rhizoctonia solani. ITS1/ITS4 markers were employed for the molecular identification of the pathogen that caused the root rot in Cedrus deodara, which was confirmed to be binucleate Rhizoctonia AG-E. For the root rot pathogen, isolation frequency was also observed in three solan ranges. Isolation frequency of Chail range comprising of two ranges showed the maximum of 37.71%followed by Kandaghat (34.42%) and Solan (19.37%), respectively. Numerous pathogens were isolated, however, three pathogen, binucleate Rhizoctonia AG-E, Fusarium oxysporum, and Phoma exigua were found to occur most frequently. Binucleate Rhizoctonia AG-E was the most often isolated of the three primary pathogens, occurring at a maximum frequency of 50.23%, followed by P. exigua (18.98%) and F. oxysporum (7.49%). Three different methods were used to test the pathogenicity of binucleate Rhizoctonia AG -E for their ability to produce root rot symptoms. The soil infestation inoculation method recorded the highest disease incidence (83.48%) followed by the root dip (34.53%) and stem application (23.76%), respectively.
... Currently, F. oxysporum is considered a species complex that can cause serious problems not only in agriculture and horticulture, but also in forestry, as it is involved in the mortality of seedlings of Scots pine (Pinus sylvestris) in nurseries, but also kills the buds of oak shoots [4] or other forest tree species such as beech or ash [19,20]. In nurseries, F. oxysporum sensu lato causes pre-emergence and post-emergence damping off seedlings with root death and stem cankers [19,[21][22][23][24][25][26][27][28][29][30][31][32]. ...
Phytopathogenic fungi can cause plant diseases that are difficult to control, including mass mortality of some tree species. The Fusarium oxysporum complex (sensu lato) is one of the most dangerous groups of phytopathogenic fungi, causing the death of conifer species, including Pinus sylvestris seedlings in forest and ornamental nurseries. Recently, non-chemical methods of plant protection have become the basis of integrated pest management (IPM) in the European Union (EC Directive). The possibility of protection of pine seedlings against the pathogen F. oxysporum using active substances from wood-destroying fungi commonly found in forests was examined. Methanolic extracts of Fomitopsis pinicola, Ganoderma applanatum, and Trametes versicolor were found to contain substances effective in both prevention and treatment of infected seedlings. G. applanatum and T. versicolor showed particular biological activity in increasing plant resistance. Efficacy, especially of the extract of F. pinicola, increased with concentration. Further field trials are needed to confirm the results obtained in laboratory tests on plant protection.
... and Alternaria alternata, along with Botrytis cinerea and Cladosporium cladosporioides. Each of these species are common environmental saprotrophs and have been reported previously in studies of fungi present in stumps of conifers [18][19][20][21][22][23][24][25][26][27], although A. alternata and B. cinerea are also well-known pathogens [27]. ...
The presence and frequency of decay fungi in living Abies cilicica and Abies bornmülleriana was examined from 200 healthy-looking trees in 40 different stands in western regions of Turkey. Totally, 965 and 1059 fungal isolates were obtained from A. cilicica and A. bornmülleriana, respectively using increment bore cores. Isolates were identified by PCR amplification of the ITS region and sequencing. In addition, decay fungi were detected directly from wood samples using species-specific primers. The most abundant decay fungi isolated were Heterobasidion abietinum, Hyphodermella rosae, Cerrena unicolor, Stereum hirsutum and Bjerkandera adusta. Heterobasidion abietinum was mostly found in cores taken from the stem base of the trees. Multiplex PCR results yielded Stereum sp., Phellinus sp. and Ganoderma lucidum directly in DNA extracted from the bore cores. Species that are members of Ascomycota such as Cladosporum herbarum, Trichoderma sp., Phoma sp., Penicillium sp. and Alternaria alternata were also isolated from both tree species.
... Pathogenic fungi were least common in the roots and included representatives from genera such as Neonectria, Hymenoscyphus, Alternaria or Ganoderma (Table 4). Although many of these are generalists and commonly found in tree roots [106], H. fraxineus is a pathogen of Fraxinus spp. in Europe and is not associated with P. abies. The detection of H. fraxineus in different samples of the study was probably due to the presence of its propagules on the surface of different tissues (needles, shoots, or roots) and in the soil as the surface of our samples was not sterilised, and the disease caused by this fungus is active in the area [107]. ...
Climate change, which leads to higher temperatures, droughts, and storms, is expected to have a strong effect on both health of forest trees and associated biodiversity. The aim of this study was to investigate the diversity and composition of fungal communities associated with the functional tissues and rhizosphere soil of healthy-looking Picea abies to better understand these fungal communities and their potential effect on tree health in the process of climate change. The study sites included 30 P. abies stands, where needles, shoots, roots, and the rhizosphere soil was sampled. DNA was isolated from individual samples, amplified using ITS2 rRNA as a marker and subjected to high-throughput sequencing. The sequence analysis showed the presence of 232,547 high-quality reads, which following clustering were found to represent 2701 non-singleton fungal OTUs. The highest absolute richness of fungal OTUs was in the soil (1895), then in the needles (1049) and shoots (1002), and the lowest was in the roots (641). The overall fungal community was composed of Ascomycota (58.3%), Basidiomycota (37.2%), Zygomycota (2.5%), Chytridiomycota (1.6%), and Glomeromycota (0.4%). The most common fungi based on sequence read abundance were Aspergillus pseudoglaucus (7.9%), Archaeorhizomyces sp. (3.6%), and Rhinocladiella sp. (2.0%). Pathogens were relatively rare, among which the most common were Phacidium lacerum (1.7%), Cyphellophora sessilis (1.4%), and Rhizosphaera kalkhoffii (1.4%). The results showed that the detected diversity of fungal OTUs was generally high, but their relative abundance varied greatly among different study sites, thereby highlighting the complexity of interactions among the host trees, fungi, and local environmental conditions.
... Among these, Venturia is a destructive plant pathogen that can result in scab diseases (Bock et al., 2018), and Chalara and Mycena cause ash dieback (Tsouvalis, 2019) and coffee leaf spots (Avelino et al., 2007), respectively. Nectria has been detected in the decayed roots of Pinus sylvestris and Picea abies (Menkis et al., 2006) and causes woody plant dieback and canker disease (Yang et al., 2019). The enrichment of specific functional guilds in plant roots during secondary succession may have important influences on plant health and the process of secondary succession, and more specific studies should be conducted in the future. ...
In this study, we provide fungal community structure analysis of three contrasting sites in secondary succession, and try to identify variables that shape fungal community composition. Soil and roots were sampled from three sites representing a secondary succession from grassland, to shrubland to forest. We analyzed the fungal community and potential ecological functions based on high-throughput sequencing and the FUNGuild database. Fungal α-diversity in the nonrhizosphere and bulk soils of the secondary forest was significantly higher than that in the grassland, while the opposite result was observed in roots, and no significant difference was found in fungal α-diversity in the rhizosphere soil. Fungal β-diversity in the soils and roots varied significantly with succession. FUNGuild data show that the relative abundance of arbuscular mycorrhizal (AM) fungi and dung saprotroph decreased with succession; ectomycorrhizal (ECM) fungi, ericoid mycorrhizal and fungal parasite increased with succession; undefined saprotrophs and plant pathogens showed no significant difference during succession. Total phosphorus, available phosphorus and soil nitrogen: phosphorus ratio were the main factors determining fungal β-diversity and fungal functional guilds. Interestingly, the roots of the dominant plants at three succession stages enriched plant pathogens Gibberella, Exserohilum and Venturia, respectively. In conclusion, our result revealed differential change trends in fungal α-diversity and fungal β-diversity in the soils and roots among three succession stages. Moreover, the fungal functional guilds (i.e., ECM, AM, plant pathogen) also showed significant changes during succession. These results could enhance understanding of the mechanism of secondary succession in the subalpine region.
... The material included: (1) pure cultures of isolated fungi; (2) collected adult beetles. First, one isolate per each group of fungal cultures was used for DNA extractions, amplification, and Sanger sequencing following methods described by Menkis et al. [24]. Amplification by PCR was done using ITS1F [25] and ITS4 [26] primers. ...
... Second, isolation of DNA from not surface-sterilized adults of bark beetles was done from 288 individuals. Isolation of DNA was done from each bark beetle separately using the CTAB protocol [24]. Amplification and sequencing of fungal ITS rDNA were carried out as previously described, performing amplification by PCR in two steps: (1) using fungal specific primers NLC2 and NSA3; (2) nested PCR using primers ITS1F and ITS4 [27]. ...
Drought-induced stress and attacks by bark beetle Ips sexdentatus currently result in a massive dieback of Pinus sylvestris in eastern Ukraine. Limited and fragmented knowledge is available on fungi vectored by the beetle and their roles in tree dieback. The aim was to investigate the fungal community vectored by I. sexdentatus and to test the pathogenicity of potentially aggressive species to P. sylvestris. Analysis of the fungal community was accomplished by combining different methods using insect, plant, and fungal material. The material consisted of 576 beetles and 96 infested wood samples collected from six sample plots within a 300 km radius in eastern Ukraine and subjected to fungal isolations and (beetles only) direct sequencing of ITS rDNA. Pathogenicity tests were undertaken by artificially inoculating three-to-four-year-old pine saplings with fungi. For the vector test, pine logs were exposed to pre-inoculated beetles. In all, 56 fungal taxa were detected, 8 exclusively by isolation, and 13 exclusively by direct sequencing. Those included nine ophiostomatoids, five of which are newly reported as I. sexdentatus associates. Two ophiostomatoid fungi, which exhibited the highest pathogenicity, causing 100% dieback and mortality, represented genera Graphium and Leptographium. Exposure of logs to beetles resulted in ophiostomatoid infections. In conclusion, the study revealed numerous I. sexdentatus-vectored fungi, several of which include aggressive tree pathogens.
