Effect of plantation design on stem‐infecting form of rust in willow biomass coppice

Planting of genotype mixtures is a major strategy for the non‐chemical control of willow rust in short‐rotation coppice willows grown for biomass. In 1997 and 1998, the relative severity and spatiotemporal distribution of stem canker damage by Melampsora spp. on Salix viminalis `Bowles Hybrid' in a field trial containing monocultures and two design mixtures (random or regular) of three or five genotypes were assessed. Sphaerellopsis filum, a hyperparasite of Melampsora spp. was detected in all plots during the summer (1998) between the time of the two canker assessments. The percentage of rust pustules with this hyperparasite was initially greater in monoculture plantings than in mixtures in June, but by July, more than 92% of pustules were parasitized, irrespective of plantation design. The stem rust assessments showed that, in general, there were fewer cankers per stool in plots containing mixtures of willow genotypes compared with monoculture plantings. The distribution of rust cankers was different between monocultures and willow mixture designs, with differences between years for monocultures.     

Genetic diversity of the hyperparasite Sphaerellopsis filum on Melampsora willow rusts

The non‐specific rust hyperparasite Sphaerellopsis filum occurs naturally on Melampsora rusts of many species of the genus Salix as well as on a large range of other rust genera worldwide. To study the genetic diversity of the hyperparasitic fungus 77 S. filum isolates collected from rusts on willow clones from plantations, clone collections and natural habitats of different sites were investigated using polymerase chain reaction ‐ restriction fragment length polymorphism (PCR‐RFLP) analysis of the rDNA internal transcribed spacer regions including 5.8S rDNA and sequence analysis. Additionally, strains from Melampsora poplar rusts (4) and strains of Puccinia abrupta from Parthenium hysterophorus (5) and of P. obscura from Bellis perennis (1) were used for comparisons. Results of genetic analysis demonstrated distinct variation within the S. filum isolates. Two main groups with more than 32% difference between their nucleotide sequences were distinguished, indicating two taxa within S. filum. Within the first main group three profiles (I, II, III) were detected. The differences between these profiles were about 12%. The variation within each profile was very low (less than 2%). The second main group comprised two profiles (IV, V), which differed in 12 to 16% of their nucleotide positions. The isolates of group IV possessed a higher variation (up to 5%) within the group than those of the first main group (I, II, III). Group V was only represented by a single isolate. Neither interrelations between the S. filum profiles and the Melampsora genotypes nor a spatial distribution could be detected. It is remarkable that the six strains of S. filum from Puccinia rusts belong to one subgroup.     

Rust resistance in Salix induced by inoculations with avirulent and virulent isolates of Melampsora larici‐epitea

Six Salix clones and four Melampsora larici‐epitea isolates were used in two leaf‐disc inoculation experiments to determine whether disease severity was affected by the presence of both virulent and avirulent rust genotypes. In the first experiment, an equal amount of urediniospores of a virulent isolate and an avirulent isolate was applied simultaneously using four levels of spore suspension. In the second, the willows were inoculated with one isolate first and then, after 3 days, with another using two spore concentrations. In the first experiment, overall rust spore production was reduced by 48.6% in inoculations with mixed inocula compared with that in the inoculations with single virulent isolates. In the second experiment, 20 of the 36 combinations involving pre‐inoculations with avirulent isolates showed significant reduction in spore production. The suppressive effects on rust sporulation were more obvious at the higher inoculum densities and on the clones S. × calodendron and S. × mollissima‘Q83’.     

Comparing Chilean and Swedish rust populations: possible impact of long‐distance transport of Melampsora rust

As willow (Salix) cultivation grows globally, worldwide monitoring of pathotypes for Melampsora rust, the most serious disease in willow, becomes increasingly important. We compared the pathotype composition of the Melampsora rust population in Chile with that in Sweden and assessed inocula exchange for the rust between the continents. For pathotyping rust isolates, a willow differential was used (a standardized set of willow test clones) to obtain clone‐virulence patterns. These patterns were used to identify virulence components involved in determining which willow clones a given individual pathogen isolate (genotype) can successfully attack. Each virulence component detected earlier in Sweden's rust population was present in Chile's rust population. Using isoenzyme analysis to classify willow species in Chile, a low genetic variation of Salix viminalis (an introduced willow species) was found, compared with the endemic species Salix humboldtiana. Comparison of rust populations for the two countries supports the present hypothesis that intercontinental inocula exchange can be a significant determinant of local pathotype structure, and consequently can be important for willow‐resistance breeding.     

Compatible and Incompatible Reactions of Melampsora Rust on Willow Leaves

Spore germination, growth and appresssoria formation for Melampsora willow rust were studied for compatible and incompatible hosts to elucidate penetration specificity and prehaustorial events in the infection process. Rust originating from Salix dasyclados was inoculated on a compatible (S. dasyclados) and an incompatible (Salix daphnoides) clone and, for comparison, on the non-host plants tomato, tobacco and poplar. The same experimental procedure was also carried out for rust originating from poplar. Rust development was followed for 5 days at 20 h intervals using a microscope with fluorescent equipment. The study showed that the Melampsora rust can develop on, and penetrate, the leaf regardless of whether the rust was inoculated on compatible or incompatible willow plants. However, the fungus was able to infect and reproduce itself only in compatible interactions. For willow rust in the prehaustorial stage, the study indicated that specific recognition mechanisms were unnecessary to start the infection process.     

Life cycle of Melampsora larici‐epitea on short‐rotation cultures of Salix burjatica

The host alternation of Melampsora larici‐epitea occurring on Salix burjatica in experimental short‐rotation cultures in Finland was studied by inoculation tests. Larix sibirica was tested as the most probable alternate host. It was inoculated with rust basidiospores, and the life cycle of the pathogen was succesfully completed by inoculating the willow with aeciospores developed on larch needles.     

Spread of Cryphonectria hypovirus1 into 45 vegetative compatibility types of Cryphonectria parasitica on grafted American chestnut trees

A mixture of hypovirulent strains of Cryphonectria parasitica, including four white (European) strains infected with Cryphonectria hypovirus1 (CHV1), was used in 1982 and 1983 to inoculate natural blight cankers located within a zone ranging from the ground to 183 cm on grafted American chestnut trees. These four white strains belonged to three vegetative compatibility (vc) types. Using pigmented, single‐spore colonies from white isolates, 48 vc types were identified among 110 white isolates recovered in 1996, 1998, and 1999 from cankers located outside the inoculated zone. Twenty‐five of the 48 white vc types consisted of two or more isolates. The 25 major white vc types were vegetatively incompatible with all four of the original white hypovirulent strains, providing evidence for spread of CHV1 but not for spread of the original inoculated strains. Forty‐five vc types represent the minimum number of `new' vc types into which CHV1 had spread. The ratio of white vc types to white isolates tested (S/N) and Shannon diversity index were 0.436 and 3.64, respectively. The spatial pattern of white vc types on the grafts was found to be non‐random (p=0.019). White single‐spore colonies of white isolates were placed into four cultural morphology (CM) groups. The two largest groups contained 37 (CM group 3) and 33 (CM group 1) isolates. Single‐spore colonies from the original, white inoculated strain, Ep 49, were classified into CM groups 3 and 1, and colonies of Ep 51 W were classified into CM group 1.     

Symptoms in slash pine seedlings following inoculation with the cone rust fungus Cronartium strobilinum

Six‐week‐old seedlings of slash pine (Pinus elliottii) were inoculated with sporidia of Cronartium strobilinum, which is the cause of southern cone rust disease, by wounding the apical tips of the seedlings. Pycnial sori developed abundantly within 1 month of inoculation on the epicotyl and primary needles. The progress of the pathogen was confirmed histologically within the cells of the stem and haustoria were observed near the nucleus of the cells. In the needles, pycnial pustules developed and the fungus was abundant in the mesophyll.     

Rust scorings in a plantation of Salix viminalis clones during ten consecutive years

Among pests and diseases occurring in coppiced willow plantations established to produce biofuel, Melampsora rust is the most widespread and severe. In the present study, the variation in rust attacks in a set of Salix viminalis clones was investigated. The aim was to look for trends possibly arising from changes in rust genotype composition, to get some idea about the risks of rust overcoming resistance bred into new willow varieties. Rust levels on 92 S. viminalis clones belonging to 18 different families were scored once each year during 1987–96 in a plantation in southern Sweden. Rust scorings in 1993 deviated most from the average scoring pattern of the clones. Furthermore there was a successive change, such that the longer the time interval between scorings the lower was the correlation between rust scores. Willow families were identified that deviated from the others over time. Various explanations for inconsistency in rust scores over time are discussed.     

Pathogenicity of Thekopsora areolata from seed orchards in Finland on Prunus spp. and Picea abies

Pathogenicity of cherry‐spruce rust, Thekopsora areolata, was investigated by inoculations with aeciospores from seven Norway spruce, Picea abies, seed orchards that had suffered from successive severe rust epidemics in the 2000s in Finland. Detached leaves of Prunus spp. were inoculated in the laboratory using aeciospores from cones of various ages. In the greenhouse, live Prunus padus plants were inoculated, and possible autoecism of the rust was tested by inoculations of Picea abies seedlings. Thirty‐five spore sources from the seed orchards formed uredinia on Prunus spp. in the laboratory 2 weeks after incubation, but no telia developed. In the greenhouse, uredinia developed on live P. padus, but no rust symptoms, cankers or sporulation were detected on Picea abies. Thus, no evidence of autoecism was observed among the T. areolata populations and therefore all populations, suggesting all those tested from the Finnish seed orchards were heteroecious.     

Variation in leaf phenolics of field‐cultivated willow (Salix myrsinifolia) clones in relation to occurrence of Melampsora rust

Concentrations of potential antifungal phenolics in field‐cultivated willow (Salix myrsinifolia) clones were analysed during three growing seasons, and correlated to the occurrence of Melampsora rust. Consistent relationships between phenolics and rust were not found across the experimental years. There was significant clonal and temporal variation in phenolic content and rust frequency. Levels of some phenolics varied considerably within a sequence of four full‐grown leaves, but the variation in rust occurrence within the same leaf sequence was nonsignificant. The results suggest that the possible association between willow phenolics and rusts is not straightforward, and emphasize the importance of long‐term studies to investigate the chemical basis of willow rust interactions.     

Rust severity in bioenergy willow plantations treated with additional nutrients

A 3‐year field study was carried out to determine the effect of wastewater irrigation and previous differences in mineral fertilization on the occurrence of willow leaf rust (Melampsora epitea). The experiment was conducted in two energy forest plantations: one designed for wastewater purification and the other as a mineral fertilization experiment. The severity of leaf rust on different clones and sites with different treatments was assessed by counting the number of uredinia per leaf unit area. Generally, plants irrigated with wastewater consistently had more leaf rust, irrespective of the study years or willow clones. Previous mineral fertilization had mixed effects on different clones 2 years after the last application. Three years after the last fertilizer application, however, no impact of the treatment on rust disease development was detected. In general, the rust levels differed from year to year probably due to climate. In this study, no correlation was detected between shoot age and rust severity, whereas climate and treatments strongly influenced leaf rust levels on some willow clones.     

Screening of clones of Betula pendula and B. pubescens against two forms of Melampsoridium betulinum leaf rust fungus

Leaf discs from seedlings of six clones of B. pendula Roth and two clones of B. pubescens Ehrh. were inoculated with two urediniospore isolates of M. betulinum Kleb. The field collection iso-lates were obtained from B. pendula and B. pubescens growing in the field. The B. pendula rust was more specialized than the B. pubescens rust. The clones of B. pubescens showed partial resis-tance against the B. pendula rust, while the B. pubescens isolate was compatible to both birch species. Some interclonal variation was also found in both birch species. The results support an earlier suggestion by Klebahn that M. betulinum has two formae speciales.     

澳大利亚ECM菌在尾叶桉上的筛选及其接种效应

报道４３种澳大利亚及２种国产外生菌根菌株在广东尾叶桉人工林上的筛选试验的初期研究结果。分别对试验林６、１２和２４个月的生长情况进行了调查,数据分析结果表明,与未接种对照林木相比,接种试验林在树高和地径（或胸径）生长量上均表现出显著的接种优势（Ｐ〈０．０１）;接种林３次调查平均树高生长量最大增幅分别为２６．１％、３３．７％和２７．４％,地径（或胸径）最大增幅分别为１９．２％、２１．５９％和１９．３２％;２４个月时每公顷材积最大幅为５６．８％。Ｅ０７１０（Ｌａｃｃａｒｉａ）、Ｈ４６７０（Ｌａｂｙｒｉｎｔｈｏｍｙｃｅｓ）、Ｅ４５０１（Ａｍａｎｉｔａ）、Ｅ２０１３（Ｌａｃｃａｒｉａ）、Ｃ９３０１（Ｐｉｓｏｌｉｔｈｕｓ）、Ｈ１２７２（Ｓｃｌｅｒｏｄｅｒｍａ）、Ｈ１１９４（Ｔｈａｘｔｅｒｏｇａｓｔｅｒ）、Ｈ０６０３（Ｓｃｌ     

Short rotation coppice willow in Northern Ireland since 1973: development of the use of mixtures in the control of foliar rust (Melampsora spp.)

A review is given of the use of clonal mixtures in the development of disease control strategies in short rotation coppice willow in Northern Ireland. Salix burjatica‘Korso’ had been grown successfully for over 10 years when, in 1985, Melampsora epitea var. epitea caused serious problems. Although fungicides were effective, their use was not practical for environmental and economic reasons. Therefore, in 1987 large scale field experiments were initiated to investigate the use of clonal mixtures as a disease control strategy. Increased yields were consistently recorded from mixed stands when compared to either the mean yield of component clones or the individual yields of any of the component clones grown in monoculture. Part of this increased yield was due to a reduction in the impact of rust disease. Investigations are currently being conducted on the effect of number of clones within a mixture, the best clones to use and the optimum planting density. It is essential that there is a range of susceptibilities to M. epitea var. epitea pathotypes within the components of the mixture.     

Genetic relationships among genetically distinct forms of Melampsora larici‐epitea and related species based on AFLP data

Amplified Fragment Length Polymorphism (AFLP) was used to study genetic relationship among leaf rust fungi (Melampsora spp.) occurring on Salix species in short‐rotation coppice. Special interest was paid to a newly identified rust fungus found on S. viminalis in southern Sweden, morphologically similar to M. larici‐epitea but with a distinct DNA profile. Genetic distances among 40 Melampsora isolates collected from S. viminalis, S. dasyclados, S. viminalis x dasyclados, S. daphnoides, S. acutifolia and M. larici‐populina were calculated based on 101 AFLP markers. Neighbour‐joining analysis revealed the presence of six clusters, which corresponded exactly to predefined groups, namely three formae speciales of M. larici‐epitea, a stem‐infecting form of Melampsora on S. viminalis, the newly identified Melampsora on S. viminalis and a group consisting of two isolates from the poplar rust M. larici‐populina. All six clusters were well supported by bootstrap analysis (84 to 100% support). The newly identified Melampsora on S. viminalis was indicated to be genetically separated from M. larici‐epitea as well as from the stem‐infecting form of Melampsora.     

Geographical distribution of Erwinia salicis strains,the cause of watermark disease of willows

Eighty-six isolates of Erwinia salicis, the causal agent of watermark disease of willow, were obtained from culture collections and collected from various willow plantations in south-east England. These isolates were characterized by their reaction in a competitive enzyme-linked immunosorbent assay, by enzyme electrophoresis, and for lysis by a panel of five bacteriophage. Eighty-one isolates had similar characteristics and clustered at a level of similarity of at least 60%. Five isolates were found to have less than 20% similarity to the other E. salicis. Antiserum raised against E. salicis (NCPPB 2535) was found to be specific to E. salicis and did not cross-react with other bacteria tested. No geographical clustering of electrotypes was found: however, Dutch and English isolates could be distinguished by bacteriophage typing. The local distribution of electrotypes collected from diseased trees suggests that the bacterium is spread via the propagating material and that tree to tree spread is rare.     

Susceptibility and lesion development in Scots pine saplings infected with Peridermium pini in northern Finland

Susceptibility of Pinus sylvestris provenances from northern Finland to Peridermium pini and lesion development caused by the rust were investigated by inoculating 768 saplings. Resin‐top disease symptoms on the stem including stem swelling, occurrence of spermatial fluid and rust aecia, were recorded over a period of 9 years. The disease incidence was low (3.6%), varying within the range of 0–9.4% among pine provenances and 3.4–3.9% among spore sources in 1994–2002. Aecia were produced for the first time 2–4 years after inoculation, appearing mainly in early June, 2–9 years after inoculation. The aecia ruptured in late July to early August, and the overall sporulation lasted for 1–8 years among the pine provenances. Spermatial fluid was observed regularly in the year prior to aecia development 2–7 years after inoculation. Annual fresh swelling was observed for 1–6 years after inoculation. No statistically significant differences occurred neither between the pine provenances, the spore sources nor the years with regard to all the major disease variables. The results suggest that Scots pine provenances from northern Finland are highly resistant to P. pini regardless of the pine provenance or spore source.     

A new species of Melampsora rust on Salix elbursensis from Iran

A rust fungus was found causing stem cankers on 1‐ to 5‐year‐old stems of Salix elbursensis in the north west of Iran. The rust also forms uredinia on leaves and flowers of the host willow. Light and scanning electron microscopy revealed that the new rust is morphologically distinct from several Melampsora species occurring on the willows taxonomically close to S. elbursensis, but indistinguishable from Melampsora larici‐epitea. Examination of the internal transcribed spacer (ITS) region of the ribosomal DNA suggested that the rust fungus is phylogenetically close to Melampsora allii‐populina and Melampsora pruinosae on Populus spp. Based on both the morphological characteristics and the ITS sequence data, the rust is described as a new species –Melampsora iranica sp. nov.     

Clonal response in Salix to Melampsora rusts in short rotation coppice plantations

The appearance of Melampsora spp. rust on a range of Salix spp. clones used for short rotation coppice was recorded. The progress of the disease and its subsequent effect on leaf fall was also recorded. Rust pustules from all the clones described were examined using the Stereoscan Elec-tron Microscope. This revealed no differences in uredospore architecture which could be consis-tently related to host.