Rhizomorph production and stump colonization by co‐occurring Armillaria cepistipes and Armillaria ostoyae: an experimental study

In managed spruce forests, Armillaria cepistipes and A. ostoyae are efficient stump colonizers and may compete for these resources when they co‐occur at the same site. The aim of this experiment was to quantify the mutual competitive ability of the two Armillaria species in producing rhizomorphs and in colonizing Norway spruce (Picea abies) stumps. Five isolates of A. cepistipes and two isolates of A. ostoyae were simultaneously inoculated pair‐wise into pots containing a 4‐year‐old spruce seedling. For comparison, each isolate was also inoculated alone. One year after inoculation, stumps were created by cutting down the seedlings. Six months after creation of the stumps, rhizomorph production and stump colonization were assessed. Armillaria spp. were identified from 347 rhizomorphs and 48 colonized stumps. Armillaria cepistipes dominated both as rhizomorphs in the soil and on the stumps. Nevertheless, A. ostoyae was relatively more frequent on the stumps than in the soil and A. cepistipes was relatively more frequent in the soil than on the stumps. In both species, the ability to colonize the stumps in simultaneous inoculations was significantly reduced compared with single inoculations. In respect to rhizomorph production, simultaneous co‐inoculations had a slightly stimulatory effect on A. cepistipes and no significant effect on A. ostoyae. Our study suggests a rather neutralistic co‐existence of A. cepistipes and A. ostoyae as rhizomorphs in the soil. Concerning the ability to colonize stumps, the two species experience a mutual negative effect from the interaction, probably because of interspecific competition.     

Comparison of the virulence of Armillaria cepistipes and Armillaria ostoyae on four Norway spruce provenances

The basidiomycetes Armillaria cepistipes and Armillaria ostoyae frequently occur in the same forest stand. In this study, we determined the virulence of 20 isolates of A. cepistipes and 16 isolates of A. ostoyae on four different provenances of 2‐year‐old Norway spruce (Picea abies). Within 30 months after inoculation, 1.1 and 19.1% of the seedlings inoculated with A. cepistipes and A. ostoyae, respectively, had died or were dying. The incidence of dead and dying seedlings varied between 3 and 49% among the A. ostoyae isolates. The virulence of an isolate was positively correlated to its ability to produce rhizomorphs. One Norway spruce provenance showed significantly lower susceptibility to A. ostoyae than the other three. Rhizomorphs of both Armillaria species were attached to the root surface. The attached rhizomorphs of A. ostoyae, however, were associated with significantly more lesions. The virulence of the isolates was not correlated with their wood‐degrading capability for either of the Armillaria species.     

Variation in virulence among isolates of Armillaria ostoyae

The virulence of Armillaria ostoyae isolates from coastal (16) and interior (33) British Columbia, elsewhere in North America (eight) and Europe (six) was assessed on 2‐year‐old Douglas‐fir seedlings in pots during a 3‐year trial. Isolates from most geographical locations infected similar proportions of seedlings, had similar average damage scores and killed a similar percentage of diseased seedlings. Isolates from the coastal region had a significantly higher probability than interior isolates that a diseased seedling received a damage score > 3 on a 1–5 scale, and coastal isolates killed a higher proportion of diseased seedlings than interior isolates. The mean damage score for isolates that had been in culture for 20–25 years was about 25% lower than that for recently collected isolates. The results indicate that the higher incidence and longer duration of mortality in the southern interior of British Columbia compared to the coast can not be attributed to greater virulence of interior isolates of A. ostoyae.     

Distribution,ecology and host range of Armillaria species in Albania

Species of Armillaria were identified from 645 isolates obtained in a nation‐wide survey in Albania. The material was collected from ca. 250 permanent plots, established for monitoring forest health, and from forests and orchards attacked by Armillaria. Armillaria mellea s.s. occurred on several coniferous and broadleaved trees in most areas examined, although it was absent above 1100–1200 m in northern Albania. This species damaged Abies and Quercus spp. and, to a lesser extent, other forest trees. Armillaria mellea was also commonly recorded causing damage in orchards and vineyards. Armillaria gallica was a common saprophyte or weak pathogen in coniferous and deciduous forests at altitudes from 600 to 1600 m, and less commonly on oaks at lower altitudes. Armillaria ostoyae was rare in central and southern Albania, but common in northern Albania, causing significant damage to pine and other conifers, mostly at altitudes from 600 to 1800 m. Armillaria cepistipes was recorded at altitudes from 800 to 1800 m as a saprophyte or weak pathogen on conifers and deciduous trees, mostly in beech and silver fir forests. Armillaria tabescens was found in oak forests at altitudes from sea level to 900 m. In orchards, A. tabescens occasionally attacked almond and pear trees. Armillaria borealis was found in a few locations in northern Albania, at altitudes from 800 to 1800 m.     

Ecology and distribution of Armillaria species in Norway

The occurence of Armillaria species was assessed in Norway, enabling the northern‐most distribution of this genus to be determined in Europe. Four Armillaria species were found in Norway. Armillaria borealis was the most common species occurring on woody vegetation to the permafrost zone (ca. 69°N). Armillaria cepistipes was present in southern and central Norway, but was not found further than 66°N. Armillaria solidipes and Armillaria gallica were rare, found at only one locality each; 59°40′ and 59°32′, respectively. Armillaria species were found on 14 hosts, but there was no significant difference between occurrence of A. borealis and A. cepistipes on declining and dead trees. Phylogenetic analyses separated each species into separate clades. All isolates of A. borealis, except one, and most isolates of A. solidipes were in separate clades. However, a subclade within the A. borealis clade was formed of two A. ostoyae and one A. borealis isolates. Two small A. cepistipes genets were found in a declining oak stand.     

Influence of Armillaria root rot on the foliar nutrients and growth of some coniferous species

Armillaria root rot induces a reduction in the concentrations of N, P, K, Mg and Na; an increase in the levels of Ca, Mn, Fe and Zn; and a decrease in the annual height growth of the diseased trees of four coniferous species. The data indicates that these changes are caused by interferences in the metabolism of the infected host.     

Distribution and ecology of Armillaria species in Greece

Five Armillaria species were identified in a nationwide survey in Greece. Armillaria mellea was present in coniferous and broad-leaved forests in most of the areas examined, except the high altitudes (above 1100 m) of the mountains of north Greece. It was found to cause significant damage in fir forests as well as in fruit orchards and vineyards. Armillaria gallica was common in coniferous and broad-leaved forests in the high altitudes of central and northern Greece, predominating in the beech forests. The fungus was a weak parasite or a saprophyte of forest trees and was occasionally found on cultivated plants. Armillaria ostoyae was not found in southern and central parts of the country, but it has a wide distribution in the mountain forests of northern Greece and causes significant damage on fir, black pine, Scots pine and spruce. Armillaria cepistipes was recorded at high altitudes (1400–1800 m) on two mountains of northern Greece, mostly as a saprophyte in coniferous and broad-leaved forests. Armillaria tabescens was rare in the forests of Greece; it was found to cause disease in almond tree orchards.     

Ecology of Armillaria species on conifers in Japan

Distribution, host preference and pathogenicity of Japanese Armillaria species on conifers were investigated on the basis of field collections of 65 isolates. We identified seven Armillaria species from 19 conifer species including six major Japanese plantation conifers using mating tests and sequences of the translation elongation‐1 α gene. Armillaria mellea, Armillaria ostoyae, Armillaria cepistipes and Armillaria sinapina were frequently collected, whereas Armillaria nabsnona, Armillaria tabescens and a biological species Nagasawa’s E were rare. On the basis of host condition when the isolates were collected, A. mellea, A. ostoyae, A. cepistipes and A. tabescens are considered as moderate to aggressive pathogens of conifers in Japan.     

Identification of Armillaria species in Japan using PCR-RFLP analysis of rDNA intergenic spacer region and comparisons of Armillaria species in the world

Armillaria species from Japan were characterized using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) of the intergenic spacer region-1 (IGS-1) of ribosomal DNA (rDNA). Eleven different digestion patterns by restriction endonuclease Alu I were found among 70 isolates of seven Armillaria species in Japan. Isolates within Armillaria nabsnona, A. ostoyae, A. cepistipes, and Japanese biological species E showed the same Alu I digestion patterns. Five Alu I patterns were detected for A. gallica, three patterns for A. mellea, and two patterns for A. tabescens. Seven Armillaria species in Japan were clearly distinguished by using the profiles obtained when PCR products were digested with Alu I, Msp I, and Hae III restriction enzymes. There was considerable variability of Alu I restriction sites within the IGS-1 between the isolates of five Armillaria species, A. gallica, A. nabsnona, A. cepistipes, A. mellea, and A. tabescens, in Japan and those of their European and North American counterparts.     

Ecology of Armillaria species in managed forests and plantations in Serbia

The distribution of Armillaria species was investigated in Serbian forest ecosystems, in relation to the main host species attacked, forest‐types, geography and altitude. In total, 388 isolates were identified from 36 host species in 47 sites. Armillaria gallica was the most commonly observed species with the widest distribution and with an altitudinal range of 70–1450 m, it was the dominating Armillaria species in lowland alluvial forests and in Quercus and Fagus forests at higher elevations. Armillaria mellea occurred in Quercus spp. – dominated forests in the north and central regions at 70–1050 m. Sixty‐eight per cent of the A. mellea isolates were collected from living hosts, most commonly in declining conifer plantations. Armillaria ostoyae was distributed in the cooler coniferous forest types and plantations in the Dinaric Alps in the south of Serbia, at 850–1820 m. Armillaria cepistipes was found in the eastern and southern hilly and mountainous regions of the country, at 600–1900 m. Most isolates were obtained from conifers and rhizomorphs in the soil around decaying stumps. Armillaria tabescens was found only on dead oak material in the northern and eastern regions of the country at altitudes lower than 600 m.     

Identification of European Armillaria species by analysis of isozyme profiles

Studies were carried out to test the possibility of identifying European Armillaria species by using isozyme patterns. Twenty-two different enzymes were used to analyse the haploid and diploid mycelium extract of Armillaria borealis, Armillaria cepistipes, Armillaria gallica, Armillaria mellea, Armillaria ostoyae and Armillaria tahescens. Tests for fumarase (E.C. 4.2.1.1.), aconitase (E.C. 4.2.1.3.), leucine-amino peptidase (E.C. 3.4.11.1.), isocitrate dehydrogenase (E.C. 1.1.1.42.), shikimic dehydrogenase (E.C. 1.1.1.25), glucose-6-P-dehydrogenase (E.C. 1.1.1.49.), malic enzyme (E.C. 1.1.1.40.), 6-P-gluconic dehydrogenase (E.C. 1.1.4.4.), pectin esterase (E.C. 3.1.1.11.), and pectic lyase (E.C. 4.2.99.3.) did not reveal enzyme activity. Isozyme profiles of acid phosphatase (E.C. 3.1.3.2.), phospho-gluco-isomerase (E.C. 5.3.1.9.), peroxidase (E.C. 1.11.1.7.), polyphenoloxidase (E.C. 1.14.18.), malic dehydrogenase (E.C. 1.1.1.37.), glutamic dehydrogenase (E.C. 1.4.1.3.) and superoxide dismutase (E.C. 1.15.1.1.) were ineffective for species identification. In contrast, esterase (E.C. 3.1.1.1.), glutamic-oxalacetic transaminase (2.6.1.1.), phospho-gluco-mutase (E.C. 2.7.5.1.), alcohol dehydrogenase (E.C. 1.1.1.1.), and polygalacturonase (E.C. 3.2.1.15.) isoenzyme patterns showed enough polymorphism to allow the identification of the different Armillaria species. However, it is necessary to compare several enzyme profiles for a conclusive identification. Intraspecific crosses of A tabescens were confirmed by the presence of a heteromeric isozyme pattern of alcohol dehydrogenase and phospho-gluco-mutase.     

Distribution of Armillaria species in upland Ozark Mountain forests with respect to site,overstory species composition and oak decline

Armillaria root disease is a contributing factor to oak decline in the Ozark Mountains of central USA. We have identified Armillaria gallica, Armillaria mellea, and Armillaria tabescens in Quercus‐Carya‐Pinus forests of the region. Presence/absence patterns of each Armillaria species as well as all possible Armillaria species combinations were analysed by contingency tables and/or stepwise logistic multiple regressions with principal characteristics of the studied sites and forest stands, both quantitative and qualitative: geographic land‐type association, bedrock type, landform position, slope direction (aspect), soil type and soil surface stone cover, down woody debris, abundance and basal area of woody vegetation and decline mortality by species. Most decline mortality consisted of two red oak species (section Erythrobalanus, Quercus coccinea and Quercus velutina), which also were most sensitive to Armillaria infection. Site characteristics related to the distributions of Armillaria species and decline mortality were also related to the preponderance of Q. coccinea and Q. velutina, regional vegetation history (i.e. conversion of Pinus echinata stands to hardwoods), and the different strategies of territory acquisition and spread of the Armillaria species involved. The presence of A. gallica may reduce the activity of more virulent Armillaria species.     

Post-fire resprouting ability of 15 non-dominant shrub and tree species in Mediterranean areas of NE Spain

Post-fire resprouting ability of the non-dominant tree and shrub species of the Mediterranean Basin has not yet been experimentally tested, although this group contributes to maintain the richness of Mediterranean plant communities. In this study, we have analyzed the post-fire recovery ability of 15 woody species that occur in relatively low abundance in dry and sub humid Mediterranean areas in NE of Spain. The main goals have been: (i) to determine experimentally the post-fire resprouting ability of these species and (ii) to compare the abundance of these species in areas affected by wildland fires and in unburned areas. We have observed a high resprouting ability after prescribed burning of most species except for Juniperus communis and J. phoenicea which showed a null resprouting. As the species with high resprouting ability showed similar presence in burned and unburned areas, we can conclude that wildfires are not a factor that constrains the presence of these species in Mediterranean woodlands. However, we found a reduction in the abundance of J. communis and J. phoenicea at the regional level after wildland fires.     

Penetration of Picea sitchensis root bark by Armillaria mellea,Armillaria ostoyae and Heterobasidion annosum

Penetration of root bark tissues of Picea sitchensis by Armillaria ostoyae, Armillaria mellea and Heterobasidion annosum was examined in the absence of wounds, in superficial wounds (rhytidome tissues removed to expose the secondary phloem) and in wounds to the depth of the vascular cambium (deep wounding). Both species of Armillaria penetrated bark without prior wounding, but neither species formed rhizomorphs in this treatment. Armillaria ostoyae penetrated to 39 cell layers in depth by 48 days after inoculation of unwounded bark, whereas A. mellea penetrated 25 cell layers in the same time. Armillaria mellea penetrated superficial wounds significantly more rapidly than did A. ostoyae. Both species produced rhizomorphs within wounded host tissues. Inoculation of deep wounds with Armillaria resulted in a greater depth of bark necrosis with A. mellea than with A. ostoyae. In the absence of wounding, H. annosum failed to penetrate root bark tissues, but in both superficial and deep wounds hyphae penetrated beyond the ligno–suberized boundary zone (LSZ) by 12 days after inoculation. Where no inoculations were made, superficial or deep wounding led within 25 days to the restoration of a structurally continuous LSZ, and by day 48 the wound periderm (WP) was fully differentiated. In inoculated wounds, however, formation of the LSZ and WP was delayed or inhibited in most trees, particularly following inoculation with A. ostoyae or A. mellea. Suberization in the LSZ and WP remained diffuse and discontinuous 48 days after inoculation. Moreover, the presence of WP did not prevent further penetration of the tissues by the pathogens. Variations between trees in the depth of pathogen penetration were noted, possibly indicating differing susceptibilities of individual host genotypes. The possible host factors involved in resistance to penetration of root bark tissues by Armillaria and Heterobasidion are discussed.     

Identification of European Armillaria species by restriction-fragment-length polymorphisms of ribosomal DNA

Identification of European Armillaria species using ribosomal DNA (rDNA) restriction-fragment-length polymorphisms (RFLPs) was studied. A total of 44 Armillaria isolates representing five European biological species were examined. Whole-cell DNAs were digested with one or two (double digest) restriction endonucleases and probed with a cloned plasmid carrying one complete rDNA repeat copy of Saccharomyces carlsbergensis. Applying the restriction endonuclease Bgl II in combination with either Eco RI, Bam Hl or Hin dIII, it was possible to detect rDNA RFLPs allowing differentiation between all five European Armillaria species investigated in the study. The most conclusive results were obtained in the rDNA/Ava II RFLP pattern. All biological species showed unique Ava II banding patterns. Generally speaking, the interspecific similarities were around 42% and lower, indicating a distinct species separation. The analysis of rDNA RFLP patterns by a restriction-enzyme-rDNA-probe combination (for instance, Ava II or Bgl II/Hin dIII; probe pMY 60) is a practical means of identifying European Armillaria species for further taxonomic, phylogenetic and host-pathogen interaction studies.     

Molecular‐based identification and phylogeny of Armillaria species from Serbia and Montenegro

Armillaria causes problems of root rot, kill trees and decay wood in the forests of Serbia and Montenegro, but the species involved have not hitherto been identified. The aim of this study was to identify field isolates collected on 25 localities. Identification was based on restriction fragment length polymorphism (RFLP) analysis of intergenic spacer 1 (IGS1) region and comparisons of IGS1 sequence with those available on NCBI database. Phylogenetic analysis was performed on sequence information from selected isolates to determine possible interrelationships between isolates with different banding patterns and previously identified tester isolates of five European Armillaria species. Five Armillaria species were identified in 90 isolates obtained from forests in Serbia and Montenegro. Armillaria gallica was most frequently isolated, followed by A. cepistipes, A. mellea, A. ostoyae and A. tabescens; two isolates remained unidentified. Restriction digestion of IGS1 amplification products with AluI produced 10 RFLP patterns. Patterns G4 (400, 250, 180) for A. gallica and pattern X (400, 180, 140) for isolates 74 and 79 are reported for the first time in European isolates. Eight RFLP patterns were observed after restriction with TaqI. Two patterns each were observed for A. ostoyae and A. gallica, and one each for A. cepistipes, A. mellea, A. tabescens and isolates 74 and 79. Parsimony analyses based on the IGS1 region placed the isolates into four clades: one including A. mellea, the second containing A. gallica–A. cepistipes isolates, while isolates of A. ostoyae and A. borealis were in the third clade. Armillaria tabescens differed from all annulate species. Phylogenetic analysis supported the conclusion that European Armillaria species are closely related and separated from a common ancestor in the near past. According to this survey five European Armillaria species are present in the forests of Serbia and Montenegro, while A. borealis is not present in the studied ecosystems.     

The influence of carbon nutrition on Armillaria ostoyae growth and phenolic degradation

Concentrations of 1, 5 and 25 g C L^?1 as glucose, fructose and sucrose were added to basal media containing 0 or 1.0 mM catechol orwa-hydroxybenzoic acid. Media were poured in petri dishes and inoculated with one of three isolates of Armillaria ostoyae(Romagn.) Herink. Armillaria ostoyae isolates usually had greater colony size and biomass yield when glucose was the carbon source as compared to fructose or sucrose. When compared to A. ostoyae growth on basal media alone, 1.0 mM catechol and /wvi-hydroxybenzoic acia added to basal media inhibited A. ostoyae growth. In a second experiment, catechol andpara-hydroxybenzoic acid degradation by three A. ostoyae isolates growing on 1, 5 and 25 g C L^?1 glucose, fructose or sucrose were measured radiometrically. Carbon source and concentration had no effect on the degradation of pzra-hydroxybenzoic acid or catechol by Armillaria ostoyae. Results of these experiments suggests that A. ostoyae can grow faster when at high carbon concentration but cannot more effectively degrade catechol or para-hydroxybenzoic acid.     

黑龙江省牡丹江林区蜜环菌的调查与鉴定

在黑龙江省牡丹江地区10个林业局内进行了蜜环茵生物种的调查和标本采集，共采集分离出41个菌株，用东北5个蜜环茵生物种与之交配测定的结果表明这些菌株大部分为高卢蜜环菌，少数为CBS F。     

Laccase isoenzyme patterns of European Armillaria species from culture filtrates and infected woody plant tissues

Polyacrylamide isoelectric focusing with specific staining for laccase activity was used to characterize laccase from European Armillaria species (Armillaria ostoyae, Armillaria mellea, Armillaria gallica, Armillaria cepistipes). The enzyme was extracted from culture media either supplemented, or not, with pine sawdust, and also from Pinus pinaster naturally infected by A. ostoyae, or artificially inoculated with A. mellea and A. ostoyae. Some differences in banding patterns were found for Armillana isolates according to the species and the culture media, but a common band at pI = 3.4 was found in all the extracts tested, independently of their origin (culture filtrate or wood).     

Foraging ability and growth performance of four subtropical tree species in response to heterogeneous nutrient environments

Considering the ubiquitous heterogeneity in spatial distribution of soil nutrients, we conducted a pot experiment to investigate the foraging traits and growth performance of four important subtropical tree species in a heterogeneous nutrient environment. The tested species exhibited large differences in foraging traits as well as growth benefits obtained from root foraging. Pinus massoniana, Schima superba and Liriodendron chinese all showed a higher degree of root morphological plasticity (expressed as relative fine root mass difference; RFRMD) than Cunninghami lanceolata (P < 0.05). P. massoniana exhibited the largest degree of morphological plasticity, followed by L. chinese and S. superba, whereas there were no significant differences in RFRMD among the three species. S. superba was the only species that exhibiting both morphological plasticity and physiological plasticity. Both P. massoniana and S. superba exhibited greater whole-seedling biomass and high sensitivity in response to nutrient heterogeneity, resulting from both types of plasticity. In contrast, both types of root plasticity were poor for C. lanceolata, resulting in poor growth benefit in heterogeneous environments. As for L. chinese, the root proliferation in nutrient-rich patches occurred at the expense of depressed root growth in other parts of the root system, leading to the lack of increment in total root biomass and nutrient absorption in heterogeneous environments. Our results provide insight and practical advice for silviculture and forest management.