Infection of Scots pine seedlings by Gremmeniella abietina during summer under different inoculum potential

The connection between natural conidia dispersal of Gremmeniella abietina (A‐type) and the disease incidence and severity in first and second year conifer seedlings of Pinus sylvestris was studied in central Finland. The seedlings where exposed to natural infection for a 3‐week period throughout the growth season, followed by a 3‐week incubation period in a growth chamber to promote symptom expression. In second year seedlings the infection periods in May–June had a similar effect as regards disease outbreak and roughly half of the successful infections occurred during these periods. However, the first year seedlings were more severely diseased after the infection period in late July/early August, although the inoculum density was lower during this time compared with the period in late June/early July. This result is consistent with earlier studies. The effect of age and growth phase of the seedlings, temperature sum and the number of conidia on the disease occurrence is discussed and compared with the results of earlier studies in which artificial inoculation was employed.     

Infection experiments with Gremmeniella abietina on seedlings of Norway spruce and Scots pine

Conidia of Gremmeniella abietina infected and caused disease symptoms in annual shoots of both Scots pine (Pinus sylvestris) and Norway spruce (Picea abies) seedlings. In Norway spruce shoots the infection remained largely latent, with only a few seedlings showing symptoms. Mycelial growth inside the shoots was faster in Scots pine than in Norway spruce and was favoured by low temperature in both hosts. The shoots of Norway spruce seedlings had higher endophyte populations than those of Scots pine, and the populations were decreased by low temperatures. Reductions in the normal epiphytic or endophytic flora by acid mist treatments seemed to favour the development of G. abietina.     

Studies of the life cycle of Gremmeniella abietina on Scots pine in southern Sweden

Aspects of the life cycle of Gremmeniella abietina (Lagerb.) Morelet were studied from 1988 to 1990 in stands of Pinus sylvestris L., 16–32 years old, in southern Sweden, initiated in 1988 with a widespread outbreak of the disease. Pycnidia started to release conidia in late spring and apothecia began to release ascospores in summer. Latent infections could still be detected one year after their establishment by cultivation of healthy looking shoots on agar petri dishes. G. abietina appeared to have a mainly biennal life cycle, as most spores were released two years after infection of the shoot.     

Effects of nickel and copper on growth and mycorrhiza of Scots pine seedlings inoculated with Gremmeniella abietina

Scots pine (Pinus sylvestris) seedlings were planted in soil originating from two localities with different background levels of nickel and copper. In addition, some of the seedlings were exposed to additional nickel (20.5 mg Ni/l of soil) or copper (63.5 mg Cu/l of soil), or a combination of both Ni and Cu, via soil without direct shoot exposure during their second growing period. The seedlings were either irrigated with spring water (pH 6) or got only natural rain during the whole field experiment. All seedlings were inoculated with conidia of a shoot‐pathogen Gremmeniella abietina during their third growing season, and harvested the following spring. Lengths of shoots of different year‐classes were used as growth estimates. In roots, the proportion of fungal (assumedly mycorrhizal) biomass was estimated by measuring ergosterol concentration. Guajacol peroxidase activity was measured. Short roots were classified into two groups according to their condition and the composition of the mycorrhizal community was expressed as a proportion of morphotypes in the roots. The seedlings exposed to additional Ni had higher shoot growth than the seedlings in the other treatments. The mean Ni concentration in the roots of seedlings exposed to additional Ni was 79 p.p.m. and in other seedlings 16 p.p.m. Additional Ni also decreased the frequency of clearly senescent short roots and the proportion of the mycorrhizal morphotype with the thinnest mantle. These results indicate that the Ni exposure levels used in this experiment had some positive effects on the seedlings. The relative fungal biomass was about 6% lower (p = 0.0981) in the fine roots of seedlings treated with additional Cu. The mean Cu concentration in the roots of seedlings exposed to additional Cu was 256 p.p.m. and in other seedlings 29 p.p.m. Peroxidase activity, which was used as a general stress indicator in this study, was not affected by any of the treatments. The shoot growth and the relative biomass of fungi in the fine roots were positively correlated in all seedlings, and this correlation was stronger in seedlings exposed to additional Ni that were not irrigated compared with seedlings not exposed to additional Ni that were irrigated. The frequency of asymptomatic infections of G. abietina was positively correlated with the proportion of senescent short roots in the irrigated seedlings but not in not‐irrigated seedlings. The general condition of seedlings may be an important factor for infection by G. abietina when moisture is high enough for the fungi to infect seedlings by conidia.     

Susceptibility of Six Scots Pine Clones to the Pathogenic Fungus Gremmeniella abietina

Genotypic variation in susceptibility to Gremmeniella abietina, an economically important fungal pathogen of conifers, was studied by artificially inoculating 23 - yr - old grafted plants of six Scots pine (Pinus sylvestris L.) clones in a seed orchard. Two fungal isolates were used. The mycelium was inserted into the current year shoots in late winter and the length of the necrotic lesion caused by the fungus was measured the next spring. The growth and male flower production of the experimental shoots were also measured. The development of symptoms did not vary among the clones, but the location of the grafts within the orchard statistically significantly affected the length of necrosis. The clones differed significantly in regard to height and the production of male and female flowers. Differences in flowering intensity among the clones were not connected with the development of G. abietina. At the level of the single shoot, the production of male flowers correlated positively with the length of necrosis. The length of necrosis correlated negatively with the length of current and previous year shoots. The ability of the two fungal isolates to cause necrosis differed significantly.     

Development of fruiting bodies of large tree type of Gremmeniella abietina var. abietina and timing of infection on Scots pine inn northern Finland

The formation and maturing of the large tree type Gremmeniella abietina var. abietina fruiting bodies and their sporulation were investigated for 3 years on Scots pine (Pinus sylvestris) in northern Finland. This was done by monthly assessment of shoots in the field and in the laboratory. Infection caused by G. abietina var. abietina was dated on Scots pine by monthly covering with pollination bags and exposing branches during the growing season. Pycnidia appeared between August and September, 1 year after infection, and they started to release conidia between late June and early July, 2 years after infection. Fresh pycnidia and microconidia were formed during the following August and September in the infected shoots. The causal large tree type of G. abietina var. abietina did not produce apothecia on branches within 3 years of infection. Monthly covering and exposing branches showed that infection took place mainly between June and July.     

Occurrence of Gremmeniella abietina damage on Scots pine along a pollution gradient from Monchegorsk nickel smelter to western Lapland

Gremmeniella abietina damage was studied at stand and tree level on the Kola Peninsula and in northern Finland. At stand level, damage was recorded in four areas in Finland, while only individual damaged trees were found in Russia. According to the results of branch analysis, there was no sign of endemic epidemics in any of the areas. There was a significant negative correlation between the average G. abietina damage and modelled SO₂ deposition and temperature sum along the gradient. The results suggest that G. abietina is distributed all over northern Finland and the Kola Peninsula in natural stands of Scots pine, and that the direct effect of SO₂ deposition from the Monchegorsk nickel smelter on the damage may be minimal.     

Susceptibility of six pine species to Gremmeniella abietina isolates from Spain

Inoculation experiments were performed in order to evaluate the virulence of Gremmeniella abietina isolates from Spain on the main pine species planted in the Iberian Peninsula, as well as the influence of seedling age on this virulence. Two different experiments were carried out with four isolates of G. abietina from Spain. The greenhouse experiments consisted of seedling inoculations. One‐ and 2‐year‐old seedlings of the following five pine species were used: Pinus halepensis, Pinus pinea, Pinus pinaster, Pinus sylvestris and Pinus uncinata; also, 1‐year‐old seedlings of P. nigra were inoculated. The relative necrosis length (RNL) caused by the pathogen after 130 days was used as a response variable. The laboratory experiments were performed on 2‐ to 6‐year‐old internodes of the above pine species excluding P. uncinata. The necrosis length after 6 weeks of incubation was measured. The results have shown that all G. abietina isolates were pathogens on seedlings of these six pine species and seedlings of P. halepensis were consistently the most susceptible ones, although it is important to take into account that all the isolates used in the present work were isolated from P. halepensis, the only pine species in Spain where G. abietina has been recovered up to now. The susceptibility of the other pine species depended on the age of the seedlings.     

Formation and growth of stem cankers caused by Gremmeniella abietina on young Pinus contorta

From 1990 to 1995, the formation and growth of stem cankers caused by Gremmeniella abietina on Pinus contorta var. latifolia was studied in three stands in northern Sweden. The stands were planted in 1976–80. The total number of cankers on 756 trees that were individually followed increased from 233 to 477 during the 5‐year period. With 42.0% of the cankers, the pathogen entered through or from the base of diseased branches, and 33.6% through visually undamaged bark. Most of the cankers were within 100 cm of the ground. In one of the three areas, the cankers were evenly distributed within 180 cm of the ground. The frequency of cankers facing north exceeded those facing south. The average vertical length of cankers had increased, 55.6% of cankers had increased their percentage coverage of the stem girth; 13.8% had fully girdled the stem. At two of the sites, there was a negative correlation between canker coverage of the stem circumference and tree height increment.     

Conidial germination and formation of necrosis in pine seedlings by Gremmeniella abietina at low temperatures

The germination of conidia of G. abietina on water agar reached the same germination percentage at 0°C as at 5 °C to 12.5 °C, but required more time. The fungus caused necrosis even at - 4 °C in pine seedlings.     

Growth reactions of a Gremmeniella abietina isolate and Scots pine embryogenic tissue cultures differ in a host–parasite in vitro system

Summary The aim of this study was to screen Scots pine embryogenic tissue culture material to Gremmeniella abietina. According to the results Scots pine (Pinus sylvestris) embryogenic tissue lines (27 lines) could be ranked based on significant differences in G. abietina effects estimated by changes in glucosamine concentrations, growth, and dry matter contents of inoculated tissue pieces. The ranking order of the extreme tissue lines stayed basically similar, regardless of the variable measured. These findings imply a possibility to use the ranked embryogenic tissue lines in more detailed investigations of interactions between G. abietina and its host. This is the first report of using embryogenic tissue cultures in studying host–parasite interaction between Scots pine and G. abietina.     

Infection of Norway spruce container seedlings by Gremmeniella abietina

First‐ and second‐year containerized Norway spruce seedlings were inoculated with conidia of type A (large tree type) and type B (small tree type) of Gremmeniella abietina var. abietina at different times during the summer. The appearance of symptoms after artificial inoculation and natural infection on spruce seedlings were recorded the following spring and compared with the disease symptoms on Scots pine seedlings. The proportion of diseased seedlings after inoculation reached as high as 80%. The susceptible period during the summer began later on the first‐year seedlings than on the second‐year seedlings, and was similar for the pine seedlings. Susceptibility of first‐year seedlings was highest in August and on second‐year seedlings in July. The accumulated temperature sum, relative humidity and height growth for first‐ and second‐year seedlings was assessed. Natural infection in 2002 caused more disease on pine than on spruce seedlings. Experimental thinning of seedlings had no effect on disease incidence. In a preliminary comparison between the ability of A and B types to cause disease in Norway spruce seedlings, type B caused more damage than type A after inoculation. However, type A caused a high disease frequency in other experiments in this study. Symptoms on Norway spruce seedlings often first occurred in the mid‐section of the shoot, and were similar to those observed on pine seedlings: needles turned brown, starting at the needle base, in the spring following inoculation. On first‐year spruce, diseased needles were shed rapidly, in contrast to a slower rate of shedding on first‐year pine seedlings. Pycnidia developed about 2 years after inoculation (on pine 1 year after inoculation). On Norway spruce seedlings the lower part of the shoot, including the lateral shoots, often remained alive. The experiments show that G. abietina can cause disease on containerized Norway spruce seedlings under nursery conditions in Finland. The coincidence of spore dispersal, seedling susceptibility and predisposing factors are important in disease development.     

Effects of foliar potassium concentration on morphology, ultrastructure and polyamine concentrations of Scots pine needles

We examined the effects of three foliar potassium concentrations (high, intermediate and low) on the morphology, ultrastructure and polyamine concentrations of current-year and 1- and 2-year-old needles of 30-year-old Scots pine (Pinus sylvestris L.) trees. Foliar K concentration had only a slight effect on needle morphology. The sclerenchyma cell walls were thinner, the xylem area was larger, and the resin ducts were smaller in needles with a low K concentration than in needles with a high or intermediate K concentration. In addition, the bundle sheath cells were collapsed in needles having a low K concentration. The secondary growth of phloem tissue and the mesophyll area were greater in needles with a high or intermediate K concentration than in needles with a low K concentration, possibly indicating greater production of photoassimilates in these trees. At the ultrastructural level, mesophyll cells with enlarged central vacuoles and small vacuoles containing electron-dense material were common in needles having a low K concentration. Enlargement of the central vacuole coincided with an exponential increase in putrescine concentration in needles with a low K concentration, suggesting that the central vacuole may function as a storage site for putrescine.     

Weevil feeding on Scots pine affects germination of Endocronartium pini

The aim of this study was to assess the effect and importance of the feeding of the pine top weevil (Pissodes piniphilus) on the germination of Endocronartium pini (syn. Peridermium pini) in Scots pine (Pinus sylvestris), and thus to establish the possibility of the E. pini infection via insect woundings. Germination tests were therefore carried out on current and previous year needle and phloem extracts. Elucidation of the importance of pathogen infection for the insect's feeding preference was also required; for this purpose feeding preference tests were carried out with healthy and infected pine branches using the pine top weevil as a test insect. Weevil feeding increased the germination of E. pini aeciospores on pine extracts. Germination on previous year annual-shoot extracts was lower than that on current year annual-shoot extracts. The advance of the growing season increased this trend, but weevil feeding increased germination on extracts from older annual shoots to the levels found on extracts from current annual shoots. Spores germinated equally well on needle extracts and on phloem extracts. The weevils ate more often on infected branches than on healthy branches. E. pini infections may occur via woundings on branches and weevil feeding may facilitate this.     

Gremmeniella abietina,Phacidium infestans,and other causes of damage in alpine,young pine plantations in Norway

Field studies of damage and mortality were carried out through 16 years in six young plantations, mostly of Pinus contorta, but also P. sylvestris and P. sibirica. Differences between pine species, P. contorta provenances, sites, and soil treatments are reported and discussed, as are influences of environment, climatic conditions, snow cover, and nutrient deficiency. Gremmeniella abietina was by far the most important cause of mortality; there were great differences between the sites as to development and seriousness. Other important causes of mortality and damage were Phacidium infestans, elk (Alces alces L.), voles, and snow pressure.     

Tissue and cell injuries caused by chlorine water to Scots pine needles

The effects of chlorine water on mesophyll cells of Scots pine (Pinus sylvestris) were studied with light and electron microscopy. The treatment caused necrotic flecks formed by collapsed cells. Less injured cells showed increase in number of plastoglobuli, swelling of the thylakoids and vacuolization of the cytoplasm.     

How timing of stem girdling affects needle xylem structure in Scots pine

While needles represent a proportionally large fraction of whole-plant hydraulic resistance, no studies to date have investigated how source–sink disturbances affect needle xylem structure. In this study, we evaluated structural changes in xylem in current-year needles of Scots pine 227 and 411 days after stem girdling (hereafter referred to as DAG). Maximum and minimum tracheid lumen diameters and therefore also the size of tracheid lumen areas increased in needles 227 DAG compared to control needles. In contrast, tracheid dimensions were similar in needles 411 DAG as in the control needles, but smaller xylem area and lower number of tracheids resulted in the lower theoretical needle hydraulic conductivity of those needles. Several needle xylem parameters were intercorrelated in both control and girdled trees. These observed changes provide a new understanding of the processes that occur following a source–sink disturbance. Considering anatomical parameters such as the number of tracheids, tracheid dimension, or needle xylem area, which are rarely described in physiological studies, could be helpful, for example, in understanding to tree hydraulic systems or for modeling gas exchange. Finally, empirical equations were developed to calculate needle theoretical hydraulic conductivity and the number of tracheids in needles using an easily measurable parameter of needle xylem area.     

Tree mortality caused by Gremmeniella abietina in a subalpine afforestation in the central Alps and its relationship with duration of snow cover

Tree survival and causes of mortality were studied in an experimental afforestation in the upper subalpine forest zone in the Swiss Alps. A total of 59.8% of Pinus cembra and 45.6% of Pinus mugo were killed by Gremmeniella abietina during the first 20 years after planting, compared to 1.5% of Larix deddua trees. The mortality rates caused by G. abietina were highly correlated with the duration of snow cover in spring. Tree losses were lowest at sites where the snow melted early and highest at sites where the snow ablation was delayed in spring. Tree mortality varied greatly between years. In the year after the coldest summer of the observation period mortality due to G. abietina infections was highest, suggesting high susceptibility of trees in poor condition. Phaddium infestans, which was the second most important factor for mortality in P. cembra, killed trees irrespective of their condition. Other biotic and abiotic causes of tree mortality had negligible influence compared with the impact of G abietina and P. infestans. Excluding anthropogenic impacts, the diverse spatial pattern of forested and treeless sites close to the subalpine timberline may predominantly result from the action of parasitic fungi, depending on the ablation pattern of the snow cover in spring.     

Defence responses induced in embryogenic cultures of Norway spruce by two fractions of Gremmeniella abietina mycelia

We have examined defence responses in embryogenic cell suspension cultures of Norway spruce (Picea abies L.) elicited by intracellular protein and wall fractions (PF and WF, respectively) prepared from mycelia of the fungus Gremmeniella abietina (Lagerberg), focusing on changes in levels of phenolics (soluble and cell wall‐bound), polyamines (free and soluble conjugates) and activities of enzymes that catalyse important steps in their metabolism: phenylalanine ammonia‐lyase (PAL, EC 4.3.1.5) and diamine oxidase (DAO, EC 1.4.3.6), respectively. Treatment with both preparations induced increases in the spruce cells total contents of phenolic acids and polyamines but they responded more rapidly to WF than to PF. The contents of soluble (especially benzoic acid derivatives) and cell wall‐bound (especially ferulic acid) phenolic acids and soluble conjugates of putrescine (mainly amides of hydroxycinnamic acids), started to increase relative to controls (C) within 4 h of the addition of the WF preparation and, more weakly, within 8 h of exposure to the PF preparation. These changes correlated with the induction of PAL activity in spruce cells at 4 and 8 h following the addition of WF and PF preparations, respectively. Increases in the spruce cell’s DAO activity were also detected after just 4 h exposure to WF, and after 24 h exposure to PF. These results demonstrate that both intracellular protein and cell wall fractions prepared from A. abietina mycelium elicit changes in the metabolism of phenylpropanoids and polyamines, substances that are known to be heavily involved in the defence responses of plants to pathogens.