The spreading of the S type of Heterobasidion annosum from Norway spruce stumps to the subsequent tree stand

The occurrence of Heterobasidion annosum in stumps and growing trees was investigated on 15 forest sites in southern Finland where the previous tree stand had been Norway spruce (Picea abies) infected by H. annosum, and the present stand was either Scots pine (Pinus sylvestris), lodgepole pine (Pinus contorta), Siberian larch (Larix siberica), silver birch (Betula pendula) or Norway spruce 8–53 years old. Out of 712 spruce stumps investigated of the previous tree stand, 26.3% were infected by the S group and 0.3% by the P group of H. annosum. The fungus was alive and the fruit bodies were active even in stumps cut 46 years ago. In the subsequent stand, the proportion of trees with root rot increased in spruce stands and decreased in stands of other tree species. On average, one S type genet spreading from an old spruce stump had infected 3.0 trees in the following spruce stand, 0.5 trees in lodgepole pine, 0.3 trees in Siberian larch, 0.05 trees in Scots pine and 0.03 trees in silver birch stand. Although silver birch generally was highly resistant to the S type of H. annosum, infected trees were found on one site that was planted with birch of a very northern provenance.     

Controlling and predicting the spread of heterobasidion annosum from infected stumps and trees of picea abies

Two Norway spruce stands with heavy infections of Heterobasidion annosum were clear‐cut in 1957 and 1959 in Sweden. The stumps were extracted, the soil sifted to remove most of the roots, and young Norway spruce were planted. After 25 and 28 years, H. annosum had infected 1 % and 2% of trees on plots where stumps had been removed and 17 % and 12% of the trees on control plots, respectively. Several of the H. annosum clones fruiting on old‐growth stumps were also detected in decayed, standing trees. The same fungal clone was found to be infecting adjacent trees from several old‐growth stumps. In addition to old stumps, stumps from recent thinnings and diseased living trees were traced as infection sources. Their relative importance in spreading disease was estimated. Disease risk predictions based on the distance of a tree from various infection sources correlated well with observed frequencies of rot.     

Intersterilitätsgruppen und Klone von Heterobasidion annosum-Isolaten aus Koniferen- Wurzel- und -Stammfäulen

Intersterility groups and clones of Heterobasidion annosum isolates from root and butt rots of conifers 69 H. annousum heterokaryons from scots pine, Douglas fir, Norway spruce and larch belonged to the P group, three heterokaryons from Norway spruce to the S group. The results of a study with clones of H. annosum isolates from roots of Scotch pine trees in close neighbourhood suggest colonisation by H. annosum (partly) via root contacts.     

Persistence of a biocontrol strain of Phlebiopsis gigantea in conifer stumps and its effects on within‐species genetic diversity

Fungal isolations and genetic fingerprinting were used to determine whether Phlebiopsis gigantea stump treatment against Heterobasidion annosum sl. using a single genotype (Rotstop) would affect the genetic diversity of P. gigantea populations. The survival time of P. gigantea was longer in Norway spruce (Picea abies) stumps compared to Scots pine (Pinus sylvestris) as no isolates were obtained from pine stumps 6 years after treatment, whereas in about half of the spruce stumps the fungus was still present. The usage of Rotstop did not seem to increase the occurrence of the fungus 5 years after the treatment in fresh (1‐year‐old) untreated stumps within the same forest stands. All the isolates from the 6‐year‐old treated spruce stumps were identical in genotype with the Rotstop‐strain, whereas all isolates from the fresh untreated spruce and pine stumps differed from it. Within the treated pine stand, the biocontrol usage seemed to have caused a slight reduction in genetic markers not related to Rotstop, but there were no statistically significant differences between the marker frequencies and the local natural population. Thus, Rotstop is not likely to cause any immediate threat to the genetic diversity of P. gigantea.     

Growth of Phlebiopsis gigantea in wood of seven conifer species

Heterobasidion parviporum and Heterobasidion annosum are widely distributed root‐rot fungi that infect conifers throughout Europe. Infection of conifer stumps by spores of these pathogens can be controlled by treating fresh stumps with a competing non‐pathogenic fungus, Phlebiopsis gigantea. In this study, growth of three Latvian strains of P. gigantea and the biological control agent ‘Rotstop’ strain was evaluated in stem pieces of Norway spruce, Scots pine, lodgepole pine, Douglas‐fir, Weymouth pine, Siberian larch and Sitka spruce. The growth rates of one H. parviporum and one H. annosum isolate were also measured in the same stem pieces. The growth rate of P. gigantea varied greatly in wood of different conifer species. It was higher in the three pine species, lower in Norway spruce and lowest in Sitka spruce and Siberian larch, and in Douglas‐fir, this fungus did not grow. The largest area of wood occupied by P. gigantea was in lodgepole pine. Growth of Latvian isolates of P. gigantea in the wood of Pinus and Picea species was comparable to that of the Rotstop isolate. Consequently, stump treatment with local P. gigantea isolates should be recommended. However, our results suggest that Douglas‐fir stump treatment against Heterobasidion by P. gigantea may be ineffective and other stump treatment methods should be considered.     

Swiss stone pine trees and spruce stumps represent an important habitat for Heterobasidion spp. in subalpine forests

In the Western Italian Alps (WIA), the three European species of the forest pathogen Heterobasidion spp. can coexist in the same area. Heterobasidion parviporum Niemelä & Korhonen and Heterobasidion abietinum Niemelä & Korhonen are normally found in areas with a significant presence of their respective primary hosts, spruce (Picea spp.) and fir (Abies spp.). The host/niche occupied by Heterobasidion annosum (Fr.) Bref. in the region still remains unclear. Although Scots pine (Pinus sylvestris), a major host for this fungal species in other parts of Europe, is abundant in the region, little or no evidence of disease caused by H. annosum is visible in this tree species. Two different, but not mutually exclusive, hypotheses can explain the presence of H. annosum: (1) Scots pines are infected but largely asymptomatic and (2) H. annosum has adapted to different hosts. An analysis of Heterobasidion species was performed in two natural, mixed‐conifer forests using traditional isolation techniques and novel direct molecular diagnosis from wood. In a subalpine stand of mixed spruce (Picea abies), larch (Larix spp.), and Swiss stone pine (Pinus cembra), 18 naturally infected spruces and larches only yielded H. parviporum. A Swiss stone pine in the same stand was extensively colonized by both H. parviporum and H. annosum. In a second subalpine stand, an analysis of 18 spruce stumps and nine Swiss stone pine stumps yielded both H. parviporum and H. annosum isolates. Pine stumps had been mostly colonized by H. parviporum prior to tree felling, suggesting that this species may be secondarily infected by the locally predominant Heterobasidion species (i.e. H. parviporum). Results of our analysis also indicated that primary colonization of spruce stumps (e.g. through basidiospores) was caused by both H. parviporum and H. annosum, while secondary infection of such stumps was mostly because of H. parviporum.     

Effectiveness of treatment of Norway spruce stumps with Phlebiopsis gigantea at different rates of coverage for the control of Heterobasidion

The natural establishment of the root and butt rot causing fungus Heterobasidion annosum s.l. on Norway spruce (Picea abies) thinning stumps treated with Phlebiopsis gigantea was investigated on seven sites in southern Sweden. The trees were cut during summertime and the stumps were treated with different patterns simulating the effect of mechanical stump treatment with a single‐grip harvester. Sampling was conducted 3 and 12 months after treatment. At both samplings, the best control was obtained when 100% of the stump surface was covered by P. gigantea: in contrast, untreated control stumps showed the highest incidences of H. annosum s.l. infection at both sampling times. However, 30 and 26% of the fully covered stumps at the first and second samplings, respectively, were diseased, and question the efficacy of treating Norway spruce stumps with this biological control agent in Sweden.     

Variation in growth of Heterobasidion parviporum in a full-sib family of Picea abies

Abstract

Heterobasidion parviporum (Fr.) Niemelä & Korhonen and Heterobasidion annosum (Fr.) Bref. sensu lato are some of the major forest pathogens in the northern hemisphere causing root and butt rot to conifers. The relative susceptibility to H. parviporum was investigated in a full-sib family of Norway spruce [Picea abies (L.) Karst.] by inoculating a set of 252 cloned progenies from a controlled cross. Four ramets of each progeny were used and the 2-year-old rooted cuttings were incubated for 6 weeks under greenhouse conditions. The condition of the cuttings was assessed visually and all the plants were in excellent vigour with no mortality recorded during the experiment. To score the relative susceptibility, lesion length in the inner bark and fungal growth in the sapwood were measured. Among the progenies, significant differences were found for fungal growth in the sapwood (p<0.0005). There was no significant difference for lesion length; however, there was a significant positive correlation between fungal growth and lesion length. The broad-sense heritability was 0.11 for fungal growth. This shows that the genetic component for susceptibility to H. parviporum can be detected even within a full-sib family of Norway spruce and that there is a potential for mapping quantitative trait loci for this trait in Norway spruce.     

Two studies to assess the risk to Pinus sylvestris from Heterobasidion spp. in southern Sweden

Abstract

Pine stumps are not being treated against Heterobasidion spp. in Sweden. To determine whether they should be, the frequency of stump infections and the species of Heterobasidion involved were investigated in nine newly thinned pure Scots pine (Pinus sylvestris L.) stands in southern Sweden. The incidence of Heterobasidion was measured in roots of standing Scots pine in another 15 stands. Infections by both H. annosum (Fr.) Bref. s.s. and H. parviporum Niemelä & Korhonen were numerous in stumps six months after a summer thinning. The pathogen, mostly H. annosum, was found in 44 of 60 sampled root systems, from 14 of the 15 stands. Twenty of the infected pines were assessed as healthy on the basis of crown symptoms while 24 trees had defoliated crowns. Infected root systems were most frequent among trees with thin crowns in stands on former agricultural land, where previous thinnings had been carried out during the growing season when airborne spores are plentiful. The study suggests that stump treatment may be a profitable way to reduce disease development in Scots pine monocultures on sandy soils as well as in mixtures with Norway spruce [Picea abies (L.) Karst.] on any soil.     

Modelling the incidence of Heterobasidion annosum butt rots and related economic losses in alpine mixed naturally regenerated forests of northern Italy

A total of 2257 recently felled stumps were sampled from 22 mixed naturally regenerated forest stands in the Aosta Valley, western Italian Alps, and examined for Heterobasidion annosum butt rots. Disease incidence ranged from 6% to 71% depending on sites, and H. annosum accounted for 92% of the total number of diseased stumps. H. annosum incidence was significantly higher (Chi‐squared multiple comparison tests, p < 0.05) on Norway spruce (44%) than on the other tree species (silver fir = 18%, larch = 12% and Scots pine = 6%). Based on the information on the airborne inoculum composition of the fungus, all the three European species of H. annosum were present, with a variable frequency, depending on site. A partial least squares regression analysis showed that the relative abundance of Norway spruce and four variables describing the size of trees were the best predictors for the incidence of H. annosum butt rots. A model having these variables as predictors was developed (r² = 0.75; p < 0.001) and successfully validated on five additional forest stands. An estimate of the losses in yield and value of timber associated with the disease, i.e. direct losses, was also carried out in the forests included in the survey. The percentage of direct financial losses was either comparable or lower than the average level of disease incidence.     

Initial reactions in sapwood of Norway spruce and Scots pine after wounding and infection by Heterobasidion parviporum and H. annosum

The xylem surface of seedlings, stem material and roots of Norway spruce (Picea abies) and Scots pine (Pinus sylvestris) were inoculated with strains of Heterobasidion annosum s. str. and H. parviporum s. str. The depth of necrosis in wounded spruce increased at a linear rate for at least seven weeks of incubation, but the rate of necrotic spread was significantly faster in infected wounds. In wounded pine the necrosis was maintained at a more superficial level for several weeks. Both spruce and pine sapwood were initially infected by hyphae of both species. In spruce, the hyphae advanced at a constant rate behind the necrotic front. On the contrary, after 1–2 weeks living H. parviporum hyphae were rare in pine rays. Heterobasidion annosum hyphae survived in pine rays, phloem and tracheids, despite a heavy accumulation of phenolics and resins and were able to penetrate into the sapwood at a linear rate although slower than infections in spruce. Histochemistry and quantitative estimates demonstrated that peroxidase activity was initially higher in spruce sapwood than in pine. Within three days of incubation, the activity in spruce sapwood disappeared concurrently with deepening necrosis. However, in pine, in both control and infected samples, there was a significant increase in peroxidase activity in the area surrounding the superficial necrosis, up to the wound surface and in the cambium and phloem around the wound. After wounding and infection, the content of soluble protein increased significantly in wood of older trees but not in seedlings. Infection resulted in an increased formation of lipophilic extractives in both spruce and pine but to a significantly greater degree in the latter, whereas the amount of hydrophilic compounds decreased in both. High‐performance liquid chromatography (HPLC) analyses of lipophilic extracts showed that inoculation of pine with the two species of Heterobasidion increased the amounts of pinosylvin, its monomethylether and several other phenolics as also resinous compounds. The results obtained may be relevant in explaining the known higher resistance of Scots pine to H. parviporum.     

Resinicium bicolor; a potential biological control agent for Heterobasidion annosum

After introductory competition tests in the laboratory, Resinicium bicolor was used as a potential control agent for the conifer root rot fungus Heterobasidion annosum. Greenhouse pathogenicity tests with R. bicolor on 4-year-old seedlings of Norway spruce (Picea abies) and Scots pine (Pinus sylvestris) showed moderate incidence of infection. In three places in Sweden, four test areas were chosen for field experiments in first-rotation plantations and also in old forest sites of Norway spruce. Wood blocks, pre-inoculated with one strain of R. bicolor, were buried in the soil beside stumps at 0, 1, 2, 3 and 4 months after thinning using various spatial designs. In two of the test areas, half of the stumps were treated with a suspension of H. annosum conidia from one strain by surface spraying. After 2–3 years, stump roots were investigated and the length of growth of both species were noted. The identity of mycelia reisolated from and wood debris in the test areas were confirmed by somatic compatibility tests with the original strains. The strain of R. bicolor released was recovered from all over the test area; the released H. annosum strain was only reisolated from the conidia sprayed stump roots. R. bicolor had little effect on the growth and occurrence of H. annosum. Potential control of disease spread may arise, however, from occlusion of the pathogen from outer parts of roots.     

Outdoor exposure of untreated Scots pine (Pinus sylvestris L.) and Norway spruce [Picea abies (L.) Karst.] wood samples

Abstract

Untreated Scots pine (Pinus sylvestris) and Norway spruce (Picea abies) samples were exposed above ground in a durability test for 6 years. The samples consisted of three pieces of wood, 22×95×500 mm, screwed together; two pieces lengthwise with a third piece overlapping. Weight was measured, to calculate moisture content (MC), and samples checked regularly for cracks and fungal growth. Parameters investigated were heartwood/sapwood (pine), annual ring orientation (spruce), stand site, annual ring width and density. Stand site, annual ring width and density had no influence on MC or fungal growth for either pine or spruce. Spruce samples with vertical annual rings had fewer cracks than samples with horizontal annual rings. Pine sapwood samples had a high MC and a large amount of rot fungi, while heartwood had a lower MC and no rot. Most spruce samples were similar to pine heartwood, except from a few samples that had high MC and fungal growth. Those were all sawn from the outer part of the log. Therefore, it can be stated that spruce sawn from the inner part has almost the same properties as pine heartwood, while spruce from the outer part of the log has similar properties to pine sapwood.     

In vitro interactions between bacteria isolated from Sitka spruce stumps and Heterobasidion annosum

Culture medium composition affected antagonism by bacterial isolates from Sitka spruce (Picea sitchensis) stumps against Heterobasidion annosum. Fifty percent of bacterial isolates inhibited H. annosum growth on sporulation agar or yeast–dextrose–peptone agar; only 10% of isolates caused inhibition on both media. Proportions of isolates inhibiting H. annosum varied with stump age; fewer isolates from 4‐ or 6‐year‐old stumps exhibited antagonism than isolates from older or younger stumps. Fifteen isolates showing antagonism on sporulation agar were tested against H. annosum in spruce wood cubes. None of the bacterial isolates alone caused a significant weight reduction in inoculated cubes. Relative inoculation times of bacterial isolates and H. annosum had an effect on weight loss in interactions; simultaneous inoculation with isolates and H. annosum inhibited weight loss caused by H. annosum compared with bacteria‐free controls. Inoculation with bacterial isolates 10 days before H. annosum had no effect on the decay rate. In contrast, inoculation with H. annosum 10 days before bacteria increased weight loss of cubes by 200% relative to cultures lacking bacteria. The effect of a mixed bacterial inoculum on weight change in 0.2‐mm spruce wood slips co‐inoculated with H. annosum, Resinicium bicolor, Hypholoma fasciculare, Stereum sanguinolentum or Melanotus proteus differed between different fungi.     

A trial of disodium octaborate tetrahydrate for the control of Heterobasidion annosum

The extent of heartwood infection of Sitka spruce Picea sitchensis stumps following inoculation with basidiospores of Heterobasidion annosum was significantly reduced by application of 2% and 4% solutions of disodium octaborate tetrahydrate (DOT) immediately after felling. Infection of sapwood was so rare in all treatments, including controls, that the effectiveness of DOT in this tissue could not be judged. The results of this trial suggest that DOT may be toxic to H. annosum at lower levels of borate loading than previously thought possible. Over the 10 months of the experiment the borate loading in the stumps appeared to reach an equilibrium that was independent of the initial concentration applied. Stump-surface fruiting of Melanotus proteus, a potential fungal competitor of H. annosum, was not inhibited by these treatments.     

Stump infection by Armillaria in first-rotation conifers

Information about the entry of Armillaria into first-rotation pine and spruce stands was obtained by searching for infected stumps, rhizomorph systems or trees that had been killed. In pines Armillaria foci were very rare. In pure Norway spruce Armillaria lutea and A. mellea were detected in stumps but rhizomorphs did not extend into the soil; in Norway spruce mixed with oak, by contrast, A. lutea sometimes produced extensive rhizomorph systems. In Sitka spruce small groups of trees had been killed by A. ostoyae. All foci investigated in conifers contained different genotypes of Armillaria and probably originated from spore infection of stumps created by thinning. Some implications of these findings are discussed.     

Biological control of Heterobasidion annosum (Fr.) Bref. (Fomes annosus) in Finland

Fresh stumps of Norway spruce (Picea abies) were inoculated with Peniophora gigantea (Fr.) Massee, Botrytis cinerea Pers. ex Fr., Gliocladium deliquescens Sopp, Trichoderma viride Pers. ex Fr. ex S. F. Gray aggr. and Verticicladiella procera Kendrick in order to study their effect on infection by Heterobasidion annosum (Fr.) Bref. The organisms colonizing spruce stumps under natural conditions were identified by collecting samples in the field. In addition, the effect of season of the year, air temperature, precipitation and relative air temperature on fungal colonization of the stumps was investigated.     

Induced resistance of Norway spruce,variation of phenolic compounds and their effects on fungal pathogens

Three clones of Norway spruce (Picea abies) were studied for their response to mass‐inoculation with the blue‐stain fungus Ceratocystis polonica. The effect of different pretreatments (fungal inoculation and wounding) before mass‐inoculation was investigated for their possible role in an acquired resistance reaction. Pretreated trees showed enhanced resistance to the subsequent mass‐inoculation relative to control trees that received no pretreatment. Furthermore, the fungal colonization of inoculated trees was less than that of wounded trees. The phenolic content of the bark, analysed by RP‐HPLC, was compared in trees receiving different treatments. Trees inoculated with C. polonica had higher average concentration of (+)‐catechin, taxifolin and trans‐resveratrol than wounded trees. Both inoculated and wounded trees had higher average concentrations of these compounds than control trees. The effect of the phenolic extract of Norway spruce bark on the growth of the root rot fungus Heterobasidion annosum and the blue‐stain fungi C. polonica and Ophiostoma penicillatum were investigated in vitro. Heterobasidion annosum was not negatively affected, and the extracts had fungistatic effects on the blue‐stain fungi. The growth of O. penicillatum was more inhibited than the growth of the more aggressive C. polonica.     

Early infection by Heterobasidion annosum in Larix x eurolepis seedlings planted on infested sites

The incidence of Heterobasidion annosum was examined in three young hybrid larch (Larix × eurolepis) plantations in southern Sweden that were established after felling Norway spruce (Picea abies) stands heavily infected by the fungus. The incidence of H. annosum was 7 %, 33 %, and 70 %, respectively, in the 2-, 3-, and 5-year-old plantations. In all plantations, H. annosum was the most frequent rot-causing fungus (81 %) and all isolates tested belonged to the P-intersterility group. No visible external differences between infected and healthy seedlings were found on any site. Both tap and lateral roots were infected by H. annosum. The results suggest that when hybrid larch is planted on sandy soils after clear felling of Norway spruce heavily infected by H. annosum, it is very susceptible to infection by H. annosum. Furthermore, transfer of H. annosum from infected stumps to hybrid larch occurs during the first few years after planting on scarified sites. In this study, scarification may have exacerbated the root disease situation and actually led to more early infections by distributing pieces inoculum across the sites.     

Distribution of Heterobasidion annosum intersterility groups in Poland

Specimens of Heterobasidion annosum were collected in 104 different stands in 43 regions of Poland. Pure cultures originating from 439 collections were identified in mating tests. Three intersterility groups, P, S and F, of H. annosum were found. Their occurrence in Poland was connected with the natural distribution of the main hosts: Pinus sylvestris, Picea abies and Abies alba, respectively. P was the most common intersterility group of H. annosum in Poland, causing mortality in Scots pine plantations and root rot in older stands. It was also isolated from Betula pendula, P. abies, Larix decidua, Fagus sylvatica and Carpinus betulus. The S group was present in the southern and north‐eastern parts of the country, causing root and butt rot mostly in spruce stands. The F group occurred in the south of Poland, in the mountains, highlands and lowland up to the northern border of the distribution of fir. It was found only on stumps, old dead trees and logs. There was no evidence of damage caused by the F group on A. alba trees.