木麻黄与根系微生物关系研究进展

木麻黄是我国沿海防护林基干林带中的主栽树种,与土壤中的微生物可以形成共生关系,提高自身耐干旱、抗贫瘠、抗盐碱等适应恶劣环境的能力。土壤中的细菌和真菌能够与木麻黄根系形成稳定的相互作用的微生态系统,其中微生物主导的物质代谢循环是植物获取营养的主要来源之一,反之木麻黄的根系分泌物可以为微生物的生长提供养分。文中综述了木麻黄与根系微生物之间相互作用的相关研究,尤其是接种弗兰克氏菌（Frankia）及菌根菌提高木麻黄抗逆性方面的研究进展,对木麻黄与根系微生物体系的促生、抗逆机制进行了探讨和展望。     

Litter of an alien tree, <Emphasis Type="Italic">Casuarina equisetifolia</Emphasis>, inhibits seed germination and initial growth of a native tree on the Ogasawara Islands (subtropical oceanic islands)

We investigated the effects of the accumulation of litter of an alien tree, Casuarina equisetifolia, on the initial establishment of Schima mertensiana, a tree native to the Ogasawara (Bonin) Islands, in the northwestern Pacific Ocean, in field and greenhouse experiments. The field experiment compared emergence of seedlings in forests dominated by C. equisetifolia with those in native forests, with and without litter removal. The greenhouse experiment compared seedling germination and biomass among 12 treatments that included two soil types (collected from the C. equisetifolia and native forests), two litter types (C. equisetifolia and native litter), and three amounts of litter (5, 15, or 45 g/pot). Significantly fewer S. mertensiana seedlings emerged at sites dominated by C. equisetifolia than at sites of native species in the field experiment. Litter removal tended to increase seedling emergence. Fewer and smaller S. mertensiana seedlings germinated with greater litter cover regardless of soil type or litter type in the greenhouse experiment. Our results demonstrate that C. equisetifolia litter suppresses germination and initial growth of S. mertensiana, and that the main cause of the suppression is the greater amount of litter accumulated under C. equisetifolia forest floor rather than factors such as the nature of the soil or litter between native and alien species.     

Survival,growth, wood basic density and wood biomass of seven-year-old Casuarina species/provenances grown at Kongowe,Kibaha, Tanzania

A performance comparison of seven-year-old individuals of 13 Casuarina species/provenances in terms of survival, growth (diameter, height and volume), wood basic density and wood biomass was undertaken at Kongowe, Kibaha, Tanzania. The trial was laid out using a randomised complete block design with four replications. The results showed significant differences (P < 0.0001) in all parameters. Casuarina equisetifolia from Montazah National Park, Egypt, had the lowest untransformed survival (48.8%), whereas C. equisetifolia from Wagait Tower (North Timor), Indonesia, had the highest survival (87.5%). Casuarina equisetifolia from Montazah National Park, Egypt, had the lowest mean diameter (8.6 cm) while the C. junghuhniana provenance from Timor, Indonesia (seedlot no. 19489) had the highest diameter (14.8 cm). Casuarina equisetifolia from Montazah National Park, Egypt, had the lowest mean height (16.2 m), volume (22.1 m³ ha^?1) and biomass production (23.5 t ha^?1), whereas C. equisetifolia from Hadsamira Songkhla, Thailand, had the highest mean height (24.3 m), volume (66.4 m³ ha^?1) and biomass production (72.9 t ha^?1). In terms of wood basic density, the C. junghuhniana provenance from Timor, Indonesia (seedlot no. 19489) had the lowest (617 kg m^?3), whereas C. equisetifolia from Wagait Tower (North Timor), Indonesia, had the highest (731 kg m^?3). Ordinal ranking for the best-performing species/provenances revealed that the best two provenances were C. equisetifolia from Hadsamira Sonkhla, Thailand, and C. junghuhniana from Timor, Indonesia (seedlot no. 19491). The two poorest-performing provenances were C. equisetifolia from Montazah National Park, Egypt, and C. equisetifolia from Cotonou, Benin. The outstanding species/provenances are recommended for pilot planting at Kibaha and similar sites.     

Characterization of Corynelia uberata Fr., a putative fungal pathogen of Podocarpus falcatus in Ethiopian forests

Corynelia spp. are ascomycetes belonging to the order Coryneliales and are thought to be obligate parasites of trees in the Podocarpaceae. The aims of this study were to determine the disease intensity of Corynelia infection on Podocarpus falcatus in Ethiopian forests and verify the identity of Corynelia spp. from Ethiopia and other countries using morphological and molecular methods. Disease surveys were conducted in P. falcatus forest areas at Adaba‐Dodola, Bushoftu, Menagesha, Shashamane and Wondo Genet in Ethiopia between 2009 and 2011, and samples were collected for morphological and molecular studies. Additional dried specimens morphologically collected as C. uberata, C. portoricensis and C. tropica from Podocarpus species in Kenya, South Africa, Puerto Rico and New Zealand were also characterized. Morphologically, the South African specimen (F‐006479) of C. uberata had significantly larger ascospores when compared with the other specimens. There was a high sequence similarity (99–100%) in the internal transcribed spacer and 5.8S (ITS‐5.8S) region among the studied C. uberata sequences. Cloning and amplification of the insert spanning partial small ribosomal unit (SSU) and ITS‐5.8S regions of ribosomal DNA validated the unidentified ITS‐5.8S region as the sequence of C. uberata by inferring the reference sequence of SSU rDNA of C. uberata in GenBank. Both neighbour‐joining and/or maximum parsimony methods placed ITS‐5.8S and SSU rDNA sequences of Corynelia spp. at the basal position of the clade Eurotiomycetidae. C. uberata was found to be a potential pathogen on leaves, fruits and young stems of P. falcatus in Ethiopia.     

Improving drought tolerance of <Emphasis Type="Italic">Casuarina equisetifolia</Emphasis> seedlings by arbuscular mycorrhizas under glasshouse conditions

Effects of mycorrhizal association on improving tolerance of host plant under stress environments have received attentions in recent years. In this paper, six isolates of AMF (arbuscular mycorrhizal fungus) were inoculated to Casuarina equisetifolia seedlings under glasshouse conditions to investigate the effects of AMF on growth and drought tolerance of host plants. All the six isolates which belong to Glomus showed high mycorrhizal colonization (88.5–96.0%) with C. equisetifolia seedlings. Seedlings were subjected to drought stress without watering for 7 days and survival of the seedlings inoculated with Glomus caledonium Gc90068, G. versiforme Gv9004 and G. caledonium Gc90036 increased by 36.6, 23.3 and 16.6%, respectively compared with uninoculated seedlings. Limited influence of AMF on seedling height growth was found, but the effects of AMF on total biomass increment were very significant; the increment ranged from 25.7 to 118.9% compared with uninoculated treatment, and it was noted that AMF exerted more influences on root biomass than shoot biomass. Based on the changes in physiological and biochemical parameters among different treatments caused by drought stress, it was concluded that AM (arbuscular mycorrhizal) associations improve the drought tolerance of C. equisetifolia seedlings by means of some physiological and biochemical responses, such as lowering permeability of plasma membrane and MDA (malondialdehyde) contents, enhancing concentrations of P nutrition, soluble sugar, soluble protein and activities of POX (Peroxidase) of C. equisetifolia seedlings.     

New and unexpected host associations for Diplodia sapinea in the Western Balkans

Diplodia sapinea is an important pathogen of pine trees in plantations and urban areas in many parts of the world. This pathogen has recently also been isolated from diseased Cedrus atlantica, C. deodara and Picea omorika planted as ornamentals across the Western Balkans. The aim of this study was to consider the host range of D. sapinea in Serbia and Montenegro. Diplodia sapinea was identified from a broader collection of Botryosphaeriaceae from the Western Balkans region, based on the DNA sequence data for the internal transcribed spacer (ITS) rDNA and the translation elongation factor 1α (TEF 1‐ α). The D. sapinea isolates were obtained from sixteen tree species in the genera Abies, Cedrus, Chamaecyparis, Juniperus, Picea, Pinus, Pseudotsuga and Fagus. Four species represented new hosts in the Balkans, and this is the first report of D. sapinea from F. sylvatica anywhere in the world. Pathogenicity tests were conducted on the tree hosts from which D. sapinea was isolated, as well as on P. abies, Thuja occidentalis, Prunus laurocerasus, Eucalyptus grandis and P. patula. Inoculations were made on seedlings in the field, in the greenhouse or on freshly detached branches. Inoculations on P. pungens, P. omorika, P. abies, P. menziesii, A. concolor, P. nigra and P. sylvestris resulted in death of the seedlings 5–16 weeks after inoculation. Diplodia sapinea produced lesions on J. horizontalis and P. patula seedlings and F. sylvatica cut branches. Reciprocal inoculations showed that D. sapinea is not a pine‐specific pathogen, causing disease on tree species, including those from which it had not been isolated. Not surprisingly, the pathogen was most aggressive on some species of Pinaceae.     

Extent of colonization by Raffaelea quercivora of artificially inoculated living and gamma‐ray‐sterilized seedlings of two Japanese and three American oak species

Mass mortality of Fagacean tree species caused by Raffaelea quercivora has occurred widely in Japan. Because conidia or other propagules of the pathogen have not been found in infected trees, pathogen spread is assumed to occur primarily by hyphae. To clarify the relationship between hyphal growth of the pathogen within trees and their vessel arrangements, we examined two native Japanese oaks, Quercus crispula and Quercus glauca, and three exotic American oaks, Quercus coccinea, Quercus palustris and Quercus rubra. Quercus glauca is a radial‐porous species, whereas the other four species have a ring‐porous wood structure. Hyphal growth within inoculated potted living seedlings and in cut, sterilized stem segments of these species was examined microscopically after fungal inoculation. Water conductance in the seedlings was examined using transverse stem sections. The proportion of non‐conductive sapwood in Q. crispula, Q. coccinea and Q. palustris differed between inoculation and control treatment, being much higher in inoculated seedlings. The proportions were positively correlated with the extent of the hyphal growth. In sterilized stem segments, the extent of fungal colonization varied among the foreign ring‐porous species Q. coccinea, Q. palustris and Q. rubra. It is hypothesized that the extent of colonization by R. quercivora reflects the extent of non‐conductive sapwood irrespective of tree species, but is little affected by vessel arrangements.     

Fine root dynamics of oak saplings in response to Phytophthora cinnamomi infection under different temperatures and durations

The belowground effects of Phytophthora cinnamomi on 1‐year‐old saplings of two common oak species in mid‐Atlantic US forests, white (Quercus alba) and red oak (Q. rubra), were examined after incubation in pathogen‐infested soilless potting mix. Fine root lengths (0–1.5 mm in diameter) of both oak species were quantified after incubation at successive 30‐day intervals up to 300 days, for a total of 10 incubation periods. In addition, colony‐forming units (CFU) of P. cinnamomi were quantified after white oak saplings were incubated in infested soilless potting mix at different temperature/duration combinations that reflect soil conditions present in the mid‐Atlantic United States. Impact of P. cinnamomi on fine root lengths of red and white oak saplings varied considerably over time. Significant periods of fine root loss occurred primarily during spring, when bud break and leaf flush began for both oak species. Red oaks had 17% fine root loss on average, while white oaks appeared more resistant to P. cinnamomi infection with a 2% decrease in fine roots over the course of the experiment. Phytophthora cinnamomi CFU declined significantly with exposure to all incubation temperatures except 8°C. This was in contrast to in vitro experiments, where the optimum temperature for mycelial growth was determined to be 21°C and above. Significant fine root loss caused by P. cinnamomi depended on plant phenology and the oak species tested. Extreme soil temperatures have a significant adverse impact on temporal changes of P. cinnamomi population.     

Volatile organic compounds (VOC) as biomarkers for detection of Ceratocystis platani

Ceratocystis platani causes canker stain of plane trees, and it represents a serious disease of Platanus spp. both in the United States and Europe. Current chemical or biological controls do not effectively manage C. platani, so new preventive methods need to be developed in order to limit this pathogen spreading. In this work, we have characterized the main volatile organic compounds (VOC) emitted in vitro from pure cultures of C. platani and other common pathogenic fungal species of hosts plants growing in the same ecosystems as plane trees. We found that C. platani emitted a similar blend of VOC compared with phylogenetically similar species C. populicola. In particular, C. platani was characterized by emission of isoamyl acetate and isobutyl acetate while C. populicola by ethyl acetate and isobutyl acetate, which were not released by any of the other out‐group fungal species grown on the same medium. Moreover, following a targeted approach based on the main VOC found in vitro, we have successfully validated in vivo that VOC uniquely emitted by C. platani (i.e. isobutyl acetate along with isoamyl alcohol) were released from the bark of plane trees following C. platani inoculation. Our results highlight the possibility to exploit VOC emitted specifically by C. platani as biomarkers to recognize Platanus x acerifolia plants infected by this pathogen.     

Field survey,isolation, identification and pathogenicity of Phytophthora species associated with a Mediterranean‐type tree species

Corymbia calophylla (marri), a keystone tree species in the global biodiversity hot spot of southwestern Australia, is suffering decline and mortality associated with a canker disease caused by the endemic fungus Quambalaria coyrecup. Phytophthora species are frequently isolated from the rhizosphere of C. calophylla, and a hypothesis is that Phytophthora root infection is predisposing C. calophylla to this endemic canker pathogen. Field surveys were conducted in both anthropogenically disturbed and undisturbed C. calophylla stands, from where a total of 100 rhizosphere soil samples, from both healthy and cankered trees, were collected. Phytophthora species were isolated from 26% of the samples collected, with Phytophthora incidence significantly higher on disturbed stands than in natural forests (73% and 27%, respectively). Five Phytophthora species were recovered, including P. cinnamomi, P. elongata, P. multivora, P. pseudocryptogea and P. versiformis. Under‐bark inoculations with the Phytophthora isolates caused significant lesion lengths in excised C. calophylla stems. Corymbia calophylla response to pot infestation trials in the glasshouse varied between Phytophthora species and isolates, with isolates of P. cinnamomi and P. multivora causing a significant reduction in seedling root volume and often leading to seedling death. This study demonstrates that root disease caused by Phytophthora species, especially P. cinnamomi and P. multivora, has the ability to adversely affect C. calophylla health. This study leads the way to do a dual inoculation trial with the canker pathogen Q. coyrecup, and different Phytophthora species to investigate if Phytophthora root infection predisposes C. calophylla to this canker disease.     

Control of Hymenoscyphus fraxineus,the causal agent of ash dieback,using composting

Viability of Hymenoscyphus fraxineus inocula following temperature treatments for different exposure times was examined in vitro and in aerated flask‐ and large‐scale composting tests using green waste. After an exposure for up to 10 days at 20°C, 97.3% of H. fraxineus mycelium and pseudosclerotia plate cultures remained viable. No viability was detected following a 3‐day exposure to 40°C or a 1‐day exposure to 45°C although pseudosclerotia were more tolerant than mycelium to an exposure to 35°C. Primordial apothecia of H. fraxineus emerged from 62%–100% of infected ash rachises collected from two infected sites and stored at 4°C for 0–5 months; exposure to compost for up to 10 days at 20°C did not affect this emergence. No emergence of H. fraxineus apothecia was observed from ash rachises that were exposed to compost at 45°C for 1 day or at 35°C or 40°C for 3 days in flasks or at 40°C for 1 day or at 30°C for 5 days in a large‐scale composting system. Based on a fitted model, estimates of the survival of H. fraxineus inoculum in infected ash rachises exposed to compost at 50°C for 1 day were 0.081% of that in the untreated H. fraxineus ash rachis inoculum. Increasing loss in viability of H. fraxineus inoculum in infected ash rachises during longer and warmer exposures to compost at 35°C–45°C corresponded with a reduced concentration of pathogen DNA detected in the rachises using real‐time PCR. However, exposure of rachises to compost at >53°C resulted in a smaller reduction in pathogen DNA detected than exposure to compost at lower temperatures, possibly due to the inhibition of enzymatic degradation of DNA at elevated temperatures.     

Direct quantitative real‐time PCR assay for detection of the emerging pathogen Neonectria neomacrospora

The epidemic outbreak in northern Europe of Neonectria neomacrospora, the causal agent of dieback in Abies spp., led the European and Mediterranean Plant Protection Organization (EPPO) to include the pathogen on its alert list in 2017. Effective monitoring of this pathogen calls for a rapid and sensitive method of identification and quantification. A probe‐based real‐time PCR (qPCR) assay based on the β‐tubulin gene was developed for the detection and quantification of N. neomacrospora in infected wood samples, and directly for ascospores. This study presents the first published species–specific molecular detection assay for N. neomacrospora. The analytical specificity was validated on taxonomically closely related fungal species as well as on 18 fungal species associated with the host (Abies sp.). The analytical sensitivity was tested on naturally infected wood, on purified pathogen DNA in a matrix of host DNA and on N. neomacrospora ascospores for detection of airborne inoculum. The latter was tested both with a DNA extraction step prior to qPCR and without DNA extraction by direct qPCR on collected ascospores. The assay was specific to N. neomacrospora, with a sensitivity of 130 fg purified DNA, or 10 ascospores by direct qPCR. Omitting DNA extraction and amplifying directly on unpurified ascospores improved assay sensitivity significantly.     

Rapid detection of Fusarium circinatum propagules on trapped pine beetles

Pitch canker is a destructive disease of pine caused by the fungus Fusarium circinatum. This taxon is listed as a quarantine fungus for several regional plant protection organizations throughout the world. Whereas long‐distance spread of the disease is made possible through the trade of infected pine seeds, local spread is caused by aerial dispersion or insect transportation of the fungal conidia. Developing a reliable and efficient tool to detect of F. circinatum in insects would be very useful to monitor the local spread of the pathogen. This tool would also provide the means to assess the range of insect species that could serve as potential vector of the fungus. A DNA extraction protocol was optimized and combined with a real‐time PCR test to detect F. circinatum on pine beetles. Using artificially contaminated Ips sexdentatus, it was shown that the test was able to detect down to 10 F. circinatum conidia per individual, and 20 conidia per batch of 10 insects, which is below the lowest inoculum load occurring in nature. With this technique, several batches of up to 10 insects may be analysed simultaneously, with a timescale for analysis reduced to <5 h and without the need for expertise in Fusarium taxonomy. This tool may be useful to monitor potential spread of the pathogen across regions. Using this method, to date, despite F. circinatum foci occurred in Northern Spanish regions across the border in France, the pathogen was not found on I. sexdentatus.     

Infection approach of Diplodia sapinea,the causal agent of tip blight of Pinus tabulaeformis in China

To clarify the infection approach of Diplodia sapinea, a pathogen that causes tip blight of Pinus tabulaeformis, the infection process of the pathogen in needles was observed using scanning electron microscopy (SEM). In addition, the disease incidence on branches damaged by Aphrophora flavipes (Hemiptera: Cercopidae) and Dioryctria splendidella (Lepidoptera: Pyralidae) in the forest was also investigated. Then, branches and needles of P. tabulaeformis were inoculated using the D. sapinea spore suspension under indoor and field conditions. The results showed that the damage caused by A. flavipes could aggravate the occurrence of tip blight of P. tabulaeformis to some extent. Moreover, the pathogen could also penetrate 1‐, 2‐ and 3‐year‐old pine needles through stomata in the field. The pathogen infected the 1‐year‐old branches first and then gradually spread to 2‐ and 3‐year‐old branches.     

First report of Calonectria montana causing damping‐off disease on pine and spruce seedlings in Europe

In 2015–2016, damping‐off symptoms were observed on Pinus sylvestris and Picea abies seedlings in four forest nurseries in Poland. Some Calonectria isolates were obtained from symptomatic seedlings by culturing pieces of necrotic tissue on malt extract agar.Morphological and molecular analyses revealed that isolates represent Calonectria montana, a recently described species from natural forests in China. The isolates represent the first detection of this species outside of Asia. Pathogenicity tests demonstrated that C. montana is a severe pathogen of P. sylvestris seedlings.     

Ceratocystis smalleyi colonization of bitternut hickory and host responses in the xylem

Colonization of Carya cordiformis sapwood by Ceratocystis smalleyi and subsequent host defence responses following artificial inoculation were investigated using anatomical and histological techniques. Hyphae of C. smalleyi were observed in all sapwood xylem features confirming the ability of the pathogen to invade and colonize the xylem tissues of the host species. The fungus was isolated from within and at the margins of discoloured sapwood areas at 2 and 12 months after inoculation. General host defence responses that included vessel occlusion with gels or tyloses, lipid accumulation, and production of phenolic compounds were observed in xylem tissues of inoculated C. cordiformis stems. Pectic substances, lipids, and to a rare extent, phenolic compounds were detected in vascular gels. The lipid‐rich barriers observed likely prevent lateral expansion of the fungus in the sapwood. Furthermore, lack of fungus sporulation within vessels may restrict axial spread of the fungus. C. smalleyi appears to be a limited vascular wilt pathogen of bitternut hickory based on these observations and previously reported sap flow reduction correlated with multiple infections in artificially inoculated trees.     

Pseudodidymella fagi and Petrakia deviata: Two closely related tree pathogens new to central Europe

Pseudodidymella fagi is a leaf blotch pathogen of Fagus crenata in Japan. This pathogen is now reported for the first time on F. sylvatica in Switzerland and Germany. Species identity was verified by morphological assessment of the asexual and sexual morphs and by comparing the internal transcribed spacer (ITS) sequence of the type material from Japan and of freshly collected samples from Europe. ITS sequences proofed to be completely identical. The asexual morph Pycnopleiospora of Ps. fagi is formed on necrotic leaf spots during summer and autumn. In early spring, its sexual morph is formed in the litter of F. sylvatica. The connection between sexual and asexual morphs was verified by sequencing the ITS region of single conidium and ascospore isolates. The pathogenicity of Ps. fagi on F. sylvatica was tested by inoculations on detached leaves in vitro and Koch's postulates were fulfilled. The second pathogen we report in central Europe for the first time is Petrakia deviata, which causes a leaf blotch disease of field maple (Acer campestre). This species was collected only once in the central Caucasus region in 1929 and was never found again after its first discovery. Now it was rediscovered in two different locations in Switzerland on field maple (A. campestre) and on the new host Norway maple (A. platanoides). Species identity was verified by morphology and by comparison with the ITS sequence of the holotype specimen and freshly collected samples. Phylogenetic analysis based on ITS sequences suggested that Ps. fagi and P. deviata are closely related to each other. Whether these species were simply overlooked so far, profit from climate change, or represent newly introduced invasive species remains to be studied. Moreover, deeper phylogenetic analysis using multiple sequence markers should be conducted to verify species identities.