Susceptibility of four conifer species to Gremmeniella abietina

G. abietina causes severe dieback in pole-stage stands of Scots pine in Britain. The susceptibility of ll provenances of Scots pine and of three other conifer species was tested by inoculations. Significant differences in susceptibility were found amongst Scots pine provenances, with those originating from a native Scottish pinewood at Loch Maree showing the highest levels of disease. Corsican pine was shown to be the most susceptible species tested although sporulation was greatest on Scots pine. Norway spruce was also shown to be susceptible when inoculated late in the growing season. Infection in lodgepole pine was negligible. High levels of β-phellandrene were tentatively linked with resistance among trees in a single stand of Scots pine.     

Physiological and morphological variation of Gremmeniella abietina from Spain

Gremmeniella abietina isolated from Pinus halepensis in Spain was characterized based on disease symptoms and conidia morphology. The disease symptoms, which became more evident in the spring, included drying up of needles and branches with some distortion of terminal twigs, resulting in dieback and sometimes to death of trees of all sizes. Variation in conidia size and growth rate was investigated between 15 isolates from Spain, and the conidia morphology was compared with four isolates from Finland, four from Switzerland and three from the US. Great variation occurred between Spanish isolates both in growth rates and in conidia size. The growth rates of the isolates were greatest on malt agar amended with pine needle extract and at 15°C. The length and the width of the conidia of the isolates from Spain ranged between 10.7–44.8 and 1.5–4.4 μm, and most of them had three septa. The results suggest that the isolates from Spain do not belong to the Alpine biotype, and the disease symptoms caused by these isolates resembled those of the European biotype.     

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.     

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.     

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.     

Susceptibility of different provenances of Pinus sylvestris,Pinus Contorta and Picea abies to Gremmeniella abietina

In a randomized block factorial experiment, 1200 seedlings from four provenances, each of Pinus sylvestris, Pinus contorta and Picea abies were inoculated with conidia (2 × 10⁴ and 1 × 10⁶ conidia/seedling) of Gremmeniella abietina (Brunchorstia pinea), isolated from P. contorta plantations in northern Sweden. A further 600 seedlings were left as controls. The occurrence of symptoms and the extension of dead tissues on the annual shoots were recorded 13 months after inoculation. Only the higher spore dose resulted in significant infection. P. sylvestris and P. contorta seedlings were equally susceptible (53% infected), and significantly more infected than P. abies seedlings (39%). The annual shoots of P. sylvestris and P. abies were affected to 43% and 37% of their length, which was significantly more than 15% of the length of P. Contorta shoots. The frequency of affected seedlings differed between the most southern and northern provenances of P. sylvestris and P. abies, and between the northwestern and the south-eastern provenance of P. contorta Twenty-six months after inoculation, a higher proportion of P. contorta seedlings than P. sylvestris and P. abies seedlings had recovered, and a lower proportion of P. contorta than P. sylvestris and P. abies seedlings had died. This paper discusses why P. sylvestris was found to be more susceptible to G. abietina than P. contorta in this experiment, while the reverse is found in plantations in northern Sweden.     

The ability of Gremmeniella abietina to hydrolyze polygalacturonic acid

Gremmeniella abietina grew well in media with pectin as the sole carbon source. The fungus secreted at least one enzyme with polygalacturonase activity, and this was induced by addition of pectin to the culture medium.     

In vitro growth of Gremmeniella abietina isolates (European race) at different temperatures

The mycelial growth of 24 isolates of types A and B of Gremmeniella abietina (Lagerb.) Morelet, collected in northern Finland and the Kola peninsula, Russia, was studied on malt agar plus pine needle extract at 18°C and 5°C. Great variation occurred within both types, and the results suggest that several isolates per stand are needed to represent one local isolate, if responses of G. abietina mycelia are studied on artificial media.     

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.     

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.     

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.     

Rapid living crown reduction caused by Gremmeniella abietina affects foliar nutrient concentrations of Scots pine

The foliar chemistry of diseased and healthy trees was studied one growing season after severe reduction in living crown caused by Gremmeniella abietina in four young Scots pine (Pinus sylvestris) stands. Sample trees were chosen pairwise on the basis of the living crown length: a diseased tree with about 50% live crown reduction and a healthy tree in each pair. Fifteen elements were determined in the youngest healthy needles on the lateral top shoots of each sample tree. Diseased trees had higher foliar boron, manganese and sodium concentrations and lower magnesium, iron, zinc, copper, potassium, nitrogen and sulphur concentrations compared to the healthy trees. Foliar calcium, aluminium, phosphorus, carbon and hydrogen concentrations did not differ between the diseased and healthy trees, except for P and A1 in two of the stands when the stands were analysed separately. Significant correlations between the needle element concentrations and crown ratio (length of the living crown/tree height) were found especially for B (increasing B with decreasing crown ratio) and for Mg, Fe and Zn (decreasing concentrations with decreasing crown ratio). The effect of G. abietina-induced living crown reduction on tree nutrient status and the role of these mineral nutrients in the susceptibility are discussed.     

An outbreak of Gremmeniella abietina in central Finland

G. abietina has damaged P. sylvestris forests in central Finland. The denser the stand, the greater the number of recoverable trees. Artificially regenrated stands were more infected than naturally regenerated ones. Forest fertilization decresed the proportion of recoverable trees.     

Differential reaction in Pinus spp. to inoculation by various isolates of Ascocalyx (Gremmeniella) abietina

Stem inoculations of 3 species of pole-size pines with 4 diverse isolates of Ascocalyx (Gremmeniella) abietina revealed Pinus pinea to be more susceptible than either P. pinaster or P. nigra in terms of canker length. Isolate type and geographical aspects considered more briefly, also have some effect.     

Effects of associated fungi Sclerophoma pythiophila and Cenangium ferruginosum on Gremmeniella abietina dieback in Spain

The interactions between Gremmeniella abietina and either Sclerophoma pythiophila or Cenangium ferruginosum, fungi frequently isolated from diseased twigs along with G. abietina, were studied under laboratory (dual cultures) and greenhouse conditions (double‐inoculations). Virulence of each species was also evaluated in greenhouse experiments by means of single‐inoculations. In vitro interactions were assessed on Petri dishes containing malt agar with pine needle extract, and greenhouse experiments were performed on 1‐year‐old Pinus halepensis seedlings. In vitro growth of G. abietina was inhibited by both fungi when grown in dual culture. In single‐inoculations, G. abietina caused the greatest necrosis length on P. halepensis seedlings, followed by S. pythiophila, whereas C. ferruginosum did not cause significant necrosis. In double‐inoculations, C. ferruginosum was able to reduce the length of necrosis caused by G. abietina on the P. halepensis seedlings. In contrast, necrosis length was greater in seedlings inoculated with both S. pythiophila and G. abietina than in those inoculated with G. abietina alone. Therefore, S. pythiophila seems to play a role in disease expression caused by G. abietina on P. halepensis in Spain.     

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.     

Comparison of Gremmeniella abietina isolates from Pinus sylvestris and Pinus contorta in terms of conidial morphology and host colonization

Gremmeniella abietina isolates from Pinus contorta in northern Sweden produced, in vitro, shorter conidia with fewer septa compared with isolates from Pinus sylvestris in the southern part of the country. After mycelial inoculation of shoots with G. abietina isolates from both host species, the resulting necroses were longer in P. sylvestris than in P. contorta. Keeping seedlings in artificial mild winter climate or detaching shoots from the seedling before inoculation caused longer necroses. No host specificity in colonization was found. Isolates from P. sylvestris caused longer necroses than did isolates from P. contorta, and both types of isolates caused longer necroses in P. sylvestris than in P. contorta. The differences found between the two G. abietina populations probably reflect regional variation in the fungus.     

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.     

Hybridization between two biotypes of Gremmeniella abietina var. abietina in artificial pairings

Summary The ability of the European biotypes A and B of Gremmeniella abietina var. abietina to hybridize in artificial pairings was tested. Using three types of molecular markers progenies with parental markers mixed indicating that the two biotypes can hybridize were observed. However, the resulting progeny had a low rate of successful germination, grew poorly on artificial medium and one was most probably a partial diploid or heterokaryon. Thus, both biotypes are genetically distinct populations and should be described as separate species as soon as sufficient discriminating information is available.     

Resistance of Pinus contorta to the European race of Gremmeniella abietina

Pinus contorta seedlings, together with Pinus resinosa and Pinus banksiana seedlings, were planted adjacent to 25‐year‐old red pine trees infected by the European (EU) race of Gremmeniella abietina. Resistance to this race was assessed over 5 years. All P. resinosa seedlings were dead after that period while 65% of P. contorta and 86% of P. banksiana seedlings appeared resistant to the disease. The tip blight that occurred on P. contorta was slightly longer than that observed on P. banksiana. In microscopy, one, two, or even more suberized boundaries were seen to be initiated near the surface of the shoot at the base of healthy needles where they extended downward in the direction of the vascular cambium. Suberized boundaries occasionally crossed the xylem and joined together in the pith region to form continuous barriers around necrotic tissues. However, in most cases, these suberized barriers were not continuous across the shoot and compartmentalization was then completed by other barriers mainly constituted of parenchyma cells and xylem tracheids that accumulated phenolic compounds. Meristematic‐like cells were observed adjacent to the necrophylactic periderm. Tissue regeneration, restoration of cambial activities and formation of traumatic resin canals also seemed to be associated with the defence system of P. contorta against the EU race of G. abietina.