Response of Compensatory-Fertilized Pinus sylvestris to Infection by Heterobasidion annosum

The effects of compensatory fertilization on the infection and growth rate of Heterobasidion annosum were studied in a 40-yr-old, naturally regenerated Scots pine (Pinus sylvestris L.) stand in south-eastern Finland. The treatments were: (1) unfertilized control, (2) a compound fertilizer containing P, K, Ca, Mg, S, Cu, Zn and B, (3) as treatment 2 but with nitrogen, (4) as treatment 3 but with limestone, (5) a mixture of nitrogen and limestone composed on the basis of needle analysis, and (6) wood ash. Pine roots were inoculated on two occasions four and five growing seasons after fertilization using four different strains of the P type of H. annosum. Two years after inoculation, the fungus was alive in 28.9% of the 135 inoculated roots. Fungus survival was highest in the control trees (50%) and poorest in the treatments 2 and 3 (10.5% and 16.7%, respectively). These two treatments differed significantly from the control. Total mycelial extension during the 2-yr study period was 16.4, 2.1, 4.5, 18.1, 15.2 and 5.3 cm in treatments 1, 2, 3, 4, 5 and 6, respectively. Only treatment 2 differed significantly from the control (p<0.05, df=23). The result suggests that the application of slow-release compound fertilizers without supplementary limestone may increase the resistance of Scots pine to infection by H. annosum. The nitrogen-free compound fertilizer may also retard the spread of annosum root rot in infected pine stands.     

Compartmentalization of decay associated with Heterobasidion annosum in roots of Pinus resinosa

Decay associated with Heterobasidion annosum was compartmentalized in roots of Pinus resinosa.     

Growth rates of Heterobasidion annosum s.s. and H. parviporum in functional sapwood of Pinus sylvestris and Picea abies

Growth rates of H. annosum s.s. and H. parviporum were investigated in the functional sapwood of young Pinus sylvestris and Picea abies plants as an indicator of the relative susceptibilities of the hosts to these pathogens. The stems of 520 five‐year‐old P. abies and 321 four‐year‐old P. sylvestris plants were inoculated and the extent of infection determined 16 weeks later. H. annosum s.s. grew further than H. parviporum in P. sylvestris sapwood, while in P. abies, no differences between the two Heterobasidion spp. were found. Both H. annosum s.s. and H. parviporum spread faster in the sapwood of P. abies than in P. sylvestris. There was high within‐host species variation in growth rates for both P. sylvestris and P. abies suggesting it may be possible to identify tree genotypes with lower susceptibility.     

Biological control of Heterobasidion annosum on Pinus pinea by Phlebiopsis gigantea

The efficacy of a native isolate of Phlebiopsis gigantea in protecting the stumps of Pinus pinea against spore infection by Heterobasidion annosum s.str. was investigated. In preliminary tests carried out in stem pieces of P. pinea in confined environment, the efficacy of the isolate was compared with Rotstop^®, the commercial formulation prepared from a north European P. gigantea strain. Both showed a fully protective effect against artificial inoculation of H. annosum. Moreover, in stump treatment experiments carried out in a coastal P. pinea forest close to Rome, the native P. gigantea was effective against heavy airborne inoculum of H. annosum. Some natural infection by P. gigantea occurred in the stumps, but it was unable to control the pathogen. Random amplified microsatellite analysis allowed to recognize the presence of different P. gigantea strains in the forest.     

Pathogenicity of Heterobasidion annosum (Fr.) Bref. sensu stricto on coniferous tree species in Turkey

Two‐year‐old seedlings of Pinus brutia, P. brutia var. eldarica, Pinus pinea and 3‐year‐old seedlings of Pinus radiata, Pinus sylvestris, Pinus nigra and Cedrus libani were inoculated on the lower stem with isolates of Heterobasidion annosum s.s. collected from the Black Sea and Mediterranean regions of Turkey. In total, 315 seedlings were inoculated in April 2014 and incubated in a growth chamber for 7 weeks at 18–20°C. All isolates were pathogenic on the seven different hosts and had the ability to grow in living sapwood. The isolates had a greater growth on C. libani, P. sylvestris and P. radiata seedlings compared to plants of the other species tested. The least affected species were P. brutia and P. nigra. The isolates originating from the Black Sea region caused longer lesions on the hosts. Overall mortality during 7 weeks of incubation was 4%.     

Infection of Picea sitchensis and Pinus contorta stumps by basidiospores of Heterobasidion annosum

Infection of Pinus contorta and Picea sitchensis stumps by basidiospores of Heterobasidion annosum is extremely variable, both within and between sites, but in general P. sitchensis stumps are less sus-ceptible than those of P. contorta. Measurement of the cross-sectional area occupied by H. annosum on each stump provides a more sensitive test of species susceptibility than assessment of the proportion of stumps infected. P. sitchensis stumps become infected on a variety of soils but there is evidence to suggest that infection may be reduced by high rainfall. In some infected stumps, H. annosum is confined to the lower stump tissues. Its absence from the upper portion of the stump may be due to replacement by other micro-organisms or, alternatively, physical conditions in the upper stump tissues may prevent its continued survival after infection has taken place. In both species, but more commonly in P. sitchensis, some stumps remain alive for at least two years after felling, particularly on peat soils, due to the presence of root grafts with neighbouring trees. Results for P. sitchensis suggest that infection occurs more readily in living stumps than in those which die rapidly after felling. The viability of H. annosum basidiospore suspensions can be determined more accurately and more rapidly on a selective agar medium than on conifer stem sections.     

North American P type of Heterobasidion annosum shows pathogenicity towards Pinus halepensis in Italy

During a survey on the distribution of Heterobasidion root rot, the North American P group of H. annosum was recorded causing root rot on Aleppo pine, Pinus halepensis, in the gardens of a historical villa in Rome. This is the first report of Heterobasidion root rot on P. halepensis in its natural distribution area, the Mediterranean Basin.     

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.     

Intraspecific variation in Heterobasidion annosum for growth in sapwood of Picea abies and Pinus sylvestris

Two greenhouse experiments were conducted to study intraspecific variation in growth of the root rot fungus Heterobasidion annosum in living host sapwood. In experiment 1, Scots pine (Pinus sylvestris) and Norway spruce (Picea abies) seedlings were inoculated with H. annosum isolates, 14 each of the S-and P-intersterility groups, collected from various parts of Sweden. In pine, the P-group isolates were more virulent than the S-group isolates both in terms of infection frequency, induced mortality rate (p < 0.05), and fungal growth in sapwood (p < 0.05). In spruce, the P-group isolates were also more virulent on average, but the difference was not statistically significant. Both S and P isolates had a higher infection frequency and a significantly longer sapwood growth on spruce than on pine. The P-group caused higher mortality on pine than on spruce. The length of the lesion in the inner bark was strongly correlated with fungal growth in spruce, but not in pine where the lesions were short or absent. In experiment 2, ten Norway spruce clones were inoculated with 18 S-isolates, originating from nine live-decayed trees and from nine spore-infected stumps in a single Norway spruce stand. The objective was to test whether any selection for growth rate in sapwood was detectable among individuals of H. annosum originating either from stumps or trees. The results gave no support for such selection since no difference in sapwood growth between the two groups of isolates was found.     

Resistance of Picea abies seedlings to infection by Heterobasidion annosum in relation to drought stress

Norway spruce (Picea abies) seedlings, 2–4 years old, were subjected to different degrees of stress. Drought stress affected the predisposition to infection by Heterobasidion annosum in the same way in root wood, stem wood and stem bark. At high water potentials (?3 to ?5 bar) resistance was high, at intermediate levels (- 5 to - 15 bar) it was low but increased again at water potentials below - 8 to - 15 bar. Neither light nor oxygen deficiencies appeared to have adverse effects on resistance. Total length of unsuberized roots was a sensitive indication of strain on the seedlings.     

Systemic effects of Heterobasidion annosum s.s. infection on severity of Diplodia pinea tip blight and terpenoid metabolism in Italian stone pine (Pinus pinea)

Three-year-old seedlings of Pinus pinea L. were inoculated near the stem base with one of two Heterobasidion annosum (Fr.) Bref. sensu stricto (s.s.) strains belonging to two populations: the North American P-group (NAm-P) and the European P-group (Eur-P). The NAm-P strain caused smaller H. annosum stem lesions than the Eur-P strain. Three weeks after the stem inoculations with H. annosum, apical shoots were inoculated with Diplodia pinea (Desmaz.) J. Kick. Basal stem infection with H. annosum resulted in D. pinea causing longer necrotic lesions in the shoots, indicating systemic induced susceptibility (SIS) to this shoot blight pathogen. Furthermore, stem induction with the NAm-P strain resulted in higher susceptibility to D. pinea than stem induction with the Eur-P strain. Total terpene accumulation was suppressed by about 50% in the shoots under attack by D. pinea when seedlings were induced with H. annosum. Total terpene concentration in shoots inoculated with D. pinea was negatively correlated with lesion size, both overall and by stem treatment. Stem base inoculation with H. annosum induced whole-plant changes in ter-pe-noid profiles, but these were not associated with the SIS phenotype. We discuss our findings on modulation of systemic response of P. pinea to fungal attack in the context of tripartite ecological interactions.     

The North American P group of Heterobasidion annosum s.l. is widely distributed in Pinus pinea forests of the western coast of central Italy

The distribution of the North American P group of Heterobasidion annosum s.l., recently reported from a Pinus pinea forest in the surroundings of Rome, was studied using mating tests and DNA fingerprinting (mitochondrial DNA, random amplified microsatellite technique and two group‐specific markers). This fungus is present in several forests and small plantations along the Tyrrhenian coast of the Italian peninsula, within an area approximately 100 km long, extending from Fregene in the north to the National Park of Circeo in the south, and 27 km wide including the city of Rome. In pine forests of Castelporziano, Castel Fusano and Anzio, where US troops resided during the Second World War, the North American P group is more frequent than the European P group. The low number of mating alleles in the Italian population of the North American P group supports the hypothesis of its origin from a small number of introductions. The near 100% sexual compatibility between the North American and European P groups, together with inconsistencies in results obtained with different identification methods of these groups, suggests that hybridization between the North American and European P populations occurs occasionally.     

Responses of in vitro grown pine seedlings to infection by four strains of Heterobasidion annosum1

Scots pine seedlings growing in pure cultures were inoculated with 4 isolates of Heterobasidion annosum. Significant variation in resistance to this pathogen was observed within and among progenies of pine trees selected at two different locations. Mycorrhizal synthesis occurred more frequently in cases where seedlings were inoculated with less pathogenic isolates of the pathogen.     

Growth response of Pinus pinaster Ait. to climatic variables in central Spanish forests

? The effect of climatic variables (temperature and precipitation) on radial growth of the Mediterranean Maritime pine (Pinus pinaster Ait.) was studied using dendrochronological techniques in the Iberian Peninsula.

? Ten tree-ring width chronologies, along the central distribution area of the species, were built. Chronology variability was analysed using Principal Component Analysis (PCA) for the period 1952–2005.

? The first principal component (PC1) explained 56% of tree-growth variability. Tree-growth association with climate was analysed at regional and local scales using correlation coefficient and bootstrapped response functions.

? Radial growth at both scales was positively correlated with rainfall during and prior to the growing season at all sites, and with summer rainfall before the growing season at five sites. Mean temperature effect changed according to the sampling site, from non-significant at the highest sites to significant (positive relationship in winter) at the lowest sites. Growth season temperature also had a negative effect.

? The Kalman filter was applied to estimate changing association between growth and climate over-time. Results suggested a change in association, initiated in the 80s, from non-significant to significant (*p < 0.05) at six of the sampling sites.

? Pinus pinaster is an accurate species for analysing tree-growth association with climate and for studying plant behaviour under global change conditions.

     

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.     

Liberation of Heterobasidion annosum conidia by airflow

The effect of speed and humidity of the airflow and the míst sprayed with airflow on the liberation of conidia of Heterobasidion annosum from pure cultures was studied in a small windtunnel. The conidia were liberated more easily in humid than in dry airflow. Liberation was most abundant when mist was added to the airflow. Threshold speed for the liberation of conidia was calculated to vary between 1.8 m/s and 3.9 m/s. According to the results, conidia of H. annosum may be liberated into the air by wind gusts associated with high humidity or mist and thus the conidia may have a role in aerial dispersal, although this is probably small compared with basidiospores.     

F group of Heterobasidion annosum found in Poland

The F intersterility group of Heterobasidion annosum, specialized to Abies species and earlier reported from southern parts of Europe, was identified from Abies alba and Picea abies in the Carpathian Mountains in southern Poland, close to northern limit of the natural distribution of A. alba. However, the fungus is not common in this area and seems to be primarily a saprophyte, growing on stumps and dead trees.     

Heterobasidion annosum group S identified in north-eastern China

Samples of Heterobasidion annosum were collected in two stands of mixed forest in Changbaishan Nature Reserve in north-eastern China. Pure cultures were isolated from 15 specimens growing on Abies, Picea, Pinus and Populus. Mating tests showed that all these collections belong to the S inter-sterility group. The compatibility of Chinese homokaryons with S homokaryons from Europe was 100%. Clamp connections appeared almost equally in both strains of each pairing, and no distinct demarcation line formed in the pairings. The mating frequency of Chinese isolates with the European F group was also high, approximately 94%. These matings, however, were usually unilateral and a more or less distinct demarcation line was present in most of the pairings. The Chinese isolates were completely intersterile with the European P group. The S group of H. annosum in north-eastern China appears to have similar morphology and ecology as in Europe. It is common on Abies and Picea, but is not a serious pathogen in the pristine forests of north-eastern China.     

Influence of copper on root growth and morphology of Pinus pinea L. and Pinus pinaster Ait. seedlings

We assessed the effects of Cu on root growth and morphology of stone pine (Pinus pinea L.) and maritime pine (Pinus pinaster Ait.) seedlings grown in culture solutions supplied with 0.012 (control), 0.1, 1 or 5 micro M CuSO(4). The presence of 5 micro M Cu in the nutrient solution completely inhibited root growth of both species within 3 days. In both species, taproot elongation was reduced in the presence of 1 micro M Cu, although partial growth recovery occurred after 7 days of treatment. The presence of 0.1 micro M Cu in the culture solution slightly enhanced root elongation in P. pinaster, but did not significantly influence root elongation in P. pinea. In both species, root weight per unit length increased in response to Cu exposure, and in P. pinaster, root diameter was significantly increased. The Cu treatments also affected lateral root number and length. In the presence of 1 micro M Cu, both species formed only short lateral primordia. The 1 micro M Cu treatment increased the lateral root index (number of roots per cm of root length) of P. pinaster, but decreased that of P. pinea, compared with control values. Neither the 0.1 nor 1 micro M Cu treatment had a significant effect on the mitotic index of either species. We conclude that cell elongation is more sensitive to Cu than cell division. Cell membrane damage, as indicated by Trypan blue staining, occurred after 10 days of exposure to 1 micro M Cu.     

Isozyme and RAPD polymorphisms in Heterobasidion annosum in Italy

Isozyme and random amplified polymorphic DNA (RAPD) polymorphisms were used to study variability in a group of 41 isolates from the Italian population of Heterobasidion annosum. The isolates belonged to the intersterility groups P and S, and particularly to the group that is most widely distributed in Italy, group F. Isozyme analysis was effective in identifying the three intersterility groups and revealed a high degree of genetic divergence within the P group isolates; the mannose phosphate isomerase (MPI-2) locus was diagnostic in the attribution of isolates to the more correlated F and S groups. RAPDs were detected following amplification by the polymerase chain reaction (PCR). 74 RAPD fragments, obtained through amplifications with eight primers, were scored. Isolates from the 3 intersterility groups were clearly divergent based on analysis of RAPD markers. However, a similarity index calculated for the isolates within the F population indicated a high uniformity of the isolates collected throughout the Italian peninsula.