Blue-stain Fungi Research Articles

In Situ Antimicrobial Properties of Sabinene Hydrate, a Secondary Plant Metabolite.

The objective of this research was to investigate natural products for their potential against pathogenic microorganisms. Sabinene hydrate (SH), a monoterpenoid, is synthesised by numerous different plants as a secondary metabolite. At present, there is a lack of definite investigations regarding the antimicrobial activity of SH itself and its different isomers. The antimicrobial effects of commercially available SH (composed mainly of trans-isomer) were evaluated within a range of concentrations in three types of contact tests: solid and vapor diffusion and the macro-broth dilution method. Moreover, the effects of SH on the rate of linear growth and spore germination were also examined. Ethanolic SH solutions were tested against an array of microorganisms, including blue-stain fungi (Ceratocystis polonica, Ophiostoma bicolor, O. penicillatum), frequently originating from bark beetle galleries; three fungal strains (Musicillium theobromae, Plectosphaerella cucumerina, and Trichoderma sp.) isolated from a sapwood underneath bark beetle galleries (Ips typographus) on spruce (Picea abies) stems; Verticillium fungicola, isolated from diseased I. typographus larvae; two Gram-positive bacteria (Bacillus subtilis and Staphylococcus aureus), two Gram-negative bacteria (Escherichia coli and Pseudomonas aeruginosa); five yeasts (Candida albicans, C. krusei, C. parapsilosis, Saccharomyces cerevisiae, and Rhodotorula muscilaginosa), and two saprophytic fungi (Aspergillus niger and Penicillium notatum). In solid agar disc diffusion tests, Gram-positive bacteria exhibited greater susceptibility to SH than Gram-negative bacteria, followed by yeasts and fungi. The most resistant to SH in both the disc diffusion and broth macro-dilution methods were P. aeruginosa, A. niger, and Trichoderma sp. strains. Blue-stain fungi and fungi isolated from the Picea sapwood were the most resistant among the fungal strains tested. The minimum inhibition concentrations (MICs) generated by SH and determined using a disc volatilization method were dependent on the fungal species and played an important role in the development of microorganism inhibition. The two Gram-positive bacteria, B. subtilis and S. aureus (whose MICs were 0.0312 and 0.0625 mg/mL, respectively), were the organisms most susceptible to SH, followed by the Gram-negative bacterium, E. coli (MIC = 0.125 mg/mL) and two yeasts, C. albicans and C. kruei (MIC was 0.125 mg/mL and 0.25 mg/mL, respectively). C. parapsilosis (MIC = 0.75 mg/mL) was the yeast most resistant to SH. The investigation of antimicrobial properties of plant secondary metabolites is important for the development of a new generation of fungicides.

Local carbon reserves are insufficient for phloem terpene induction during drought in Pinus edulis in response to bark beetle-associated fungi.

Stomatal closure during drought inhibits carbon uptake and may reduce a tree's defensive capacity. Limited carbon availability during drought may increase a tree's mortality risk, particularly if drought constrains trees' capacity to rapidly produce defenses during biotic attack. We parameterized a new model of conifer defense using physiological data on carbon reserves and chemical defenses before and after a simulated bark beetle attack in mature Pinus edulis under experimental drought. Attack was simulated using inoculations with a consistent bluestain fungus (Ophiostoma sp.) of Ips confusus, the main bark beetle colonizing this tree, to induce a defensive response. Trees with more carbon reserves produced more defenses but measured phloem carbon reserves only accounted for c. 23% of the induced defensive response. Our model predicted universal mortality if local reserves alone supported defense production, suggesting substantial remobilization and transport of stored resin or carbon reserves to the inoculation site. Our results show that de novo terpene synthesis represents only a fraction of the total measured phloem terpenes in P. edulis following fungal inoculation. Without direct attribution of phloem terpene concentrations to available carbon, many studies may be overestimating the scale and importance of de novo terpene synthesis in a tree's induced defense response.

Ophiostoma haidaense, sp. nov., a new member of the O. piceae species complex associated with yellow-cedar, Callitropsis nootkatensis

ABSTRACT Ophiostoma haidanensis is described as a new species of the Ophiostoma piceae complex isolated from yellow-cedar (Callitropsis nootkatensis (D. Don) Oerst. ex D.P. Little) sapwood in the Haida Gwaii island archipelago and the North Coast of British Columbia, Canada. The fungus is characterized by the production of a typical sporothrix-like asexual morph but is distinguished morphologically from other members of the O. piceae species complex by its large, multiseptate primary conidia. Phylogenetic analysis of DNA sequences from the nuc rDNA internal transcribed spacer region ITS1-5.8S-ITS2 (ITS) and the β-tubulin (BTUB) and translation elongation factor 1-α (TEF1) genes supports the inclusion of O. haidensis as a distinct member within the O. piceae complex. To our knowledge, this is the first report of a blue stain fungus infecting yellow-cedar, an ecologically, culturally, and economically important conifer naturally distributed along the coastal forests of the Pacific Northwest in North America.

Promjena boje površina drva izloženih vremenskim utjecajima prije i nakon tretmana željezovim (II) sulfatom

Outdoor wood is exposed to various factors that can be described as weathering and cause the wood to grey. The ageing processes can vary greatly depending on the exposure. Parts of wood that are not exposed to external factors, e.g. under the overhanging, are less exposed, and the weathering process is therefore slower. This can be accelerated by solutions based on iron ions. In this way, the wood greys quickly and evenly. However, after iron treatment, the colour also depends on the previous exposure of wood to weathering. In our study, the colour change was observed as a function of weathering time. Before treatment, the samples were exposed to outdoor weathering for different periods of time and then treated with a 5 % solution of pure iron (II) sulphate and commercial iron (II) sulphate. It was determined that the pre-weathering time affected the final colour change, as the samples exposed for five weeks before treatment have comparable colour to naturally weathered wood. At the beginning of exposure, iron (II) sulphate limits mould growth, but after two months, staining fungi develop on the treated samples as well. The growth of blue stain fungi on the treated samples did not significantly affect the colour and visual appearance of the wood treated with iron-based solutions.

Palmitoleic acid as a coordinating molecule between the invasive pinewood nematode and its newly associated fungi.

Symbiotic microorganisms are ubiquitous on the body surface or internal tissues of invertebrates, providing them with benefits. Developing symbiotic relationships requires synchronization of developmental stages and physical proximity of partners. Therefore, the identification of metabolites that coordinate the reproduction of symbiotic partners is essential. This study demonstrates that palmitoleic acid (C16: 1) coordinates bilateral propagation by regulating the synchronization of reproduction between the invasive pinewood nematode (PWN) and its newly associated blue-stain fungus, Sporothrix sp.1. When the PWN fed on Sporothrix sp.1, there was a significant increase in lipid metabolism gene expression and metabolite abundance. Through further investigations, it highlighted a significant enhancement in the reproduction of the PWN through direct acquisition of C16: 1, which was abundantly present in Sporothrix sp.1. Furthermore, the PWN biosynthesized C16: 1 through the involvement of the stearoyl-CoA 9-desaturase gene fat-5 and its hormone nuclear receptor nhr-80, which was clarified to promote the egg-laying capacity of females. Moreover, it is worth noting that the production of C16: 1 was significantly higher by the associated fungus Sporothrix sp.1 to enhance sporulation during the spore formation phase compared to the hypha growth phase. Thus, by coordinating the fecundity and spore production, the key lipid metabolite C16: 1 facilitates the rapid and successful colonization of a mutually beneficial symbiotic relationship between the invasive PWN and the native Sporothrix sp.1 within the host. This finding emphasizes the significant role of metabolite sharing and its function in promoting partner synchronization within symbiotic relationships.

Defensive response of evolutionarily naïve Pinus sylvestris to the mountain pine beetle fungal associate Grosmannia clavigera in comparison to Pinus ponderosa

Mountain pine beetle (MPB, Dendroctonus ponderosae) is a destructive pest of pine forests in western North America. This insect is currently expanding its range across the Canadian boreal forest towards eastern North America, where a suite of novel pine species will be encountered. One species of pine without prior association with MPB is Pinus sylvestris (Scots pine), which is native to Europe and naturalized in parts of central and eastern North America. Here, we take advantage of a unique opportunity in the Black Hills of South Dakota where an isolated, planted, and mature stand of P. sylvestris and native Pinus ponderosa (ponderosa pine) co-exist within the range of MPB. We conducted a punch-inoculation experiment to determine the chemical response of P. sylvestris from a blue-stain fungus associated with MPB, Grosmannia clavigera, and compared the response to that of P. ponderosa. We found that P. sylvestris had a higher localized monoterpene response than P. ponderosa in response to inoculation, but a lower sesquiterpene response. Among the significant monoterpenes associated with MPB behavior, limonene, 3-carene, and myrcene had a larger localized response in P. sylvestris than P. ponderosa; lower levels of 4-allylanisole were found in P. sylvestris. Fungal inoculation did not induce a stronger terpenoid response than mechanical wounding without inoculation, indicating that P. sylvestris responds to mechanical damage similarly as to fungal inoculation. Pinus sylvestris may provide one alternative plantation species for timber production in the Great Lakes Region following mountain pine beetle incursion, however, more evaluation is needed to determine the role of this species in future plantings.

Susceptibility of surface-modified superhydrophobic wood and acetylated wood to mold and blue stain fungi

The susceptibility of surface-modified wood, surface-modified acetylated wood and acetylated wood to mold and blue stain fungi was investigated. The surface modifications were based on fluorinated and non-fluorinated silicone nanofilaments for increased hydrophobicity. Results showed an increased mold resistance of the surface-modified superhydrophobic wood with mold appearing later or with less intensity on the modified surfaces than on the untreated wood in accelerated mold chamber tests due to the increased water resistance of the samples. All acetylated wood samples exhibited good mold resistance as the available water in acetylated wood was reduced. The surface modifications on acetylated wood had a slightly negative effect on mold resistance due to side effects from the modification. The surface-modified wood showed high blue stain fungi coverage, whereas almost no blue stain fungi were observed on the acetylated wood and surface-modified acetylated wood. The surface-modified superhydrophobic wood showed high mold coverage after conditioning in a high-humidity environment or after exposure to UV irradiation. Meanwhile, the acetylated wood and surface-modified superhydrophobic acetylated wood showed a small amount of mold coverage in these conditions.

Semiochemicals produced by fungal bark beetle symbiont Endoconidiophora rufipennis and the discovery of an anti-attractant for Ips typographus.

Bark beetles vector symbiotic fungal species into their host trees during mass attacks. The symbiotic relationship with blue stain fungi of the Ascomycetes, including genera of Endoconidiophora (syn. = Ceratocystis), promotes successful establishment whereby the microbes help to overcome the host trees' defence and degrade toxic resins. This is the first study to evaluate both the volatile emissions from an insect-associated blue stain fungus over time and the insect response in a field trapping experiment. Volatile emissions from isolates of Endoconidiophora rufipennis (ER) were collected by solid-phase microextraction (SPME) and analysed by gas chromatography-mass spectroscopy (GC-MS) over a period of 30 days. This virulent North American fungus is closely related to E. polonica, a symbiotic fungus known from Eurasian spruce bark beetle Ips typographus.Nine volatiles were emitted by ER in substantial amounts: isoamyl acetate, sulcatone, 2-phenethyl acetate, geranyl acetone, geranyl acetate, citronellyl acetate, (R)- and (S)-sulcatol, and (R)-sulcatol acetate. A late peaking compound was geranyl acetone. In the field trapping experiment, three of the fungal volatiles (geranyl acetone, 2-phenethyl acetate and sulcatone) were tested in combination with a synthetic aggregation pheromone for I. typographus. Traps with geranyl acetone attracted lower numbers of I. typographus compared to traps with 2-phenethyl acetate, sulcatone or the pheromone alone as a control. The results showed that geranyl acetone acts as an anti-attractant and may act naturally on I. typographus as a cue from an associated fungus to signal an overexploited host.

Application and characterization of a novel PVDF-HFP/PVP polymer composite with MoO3 nanowires as a protective coating for wood

The coatings on wood must sometimes give aesthetic and basic protection to wooden elements and prevent the development and transmission of microorganisms. Several polymers containing different nanoparticles have already been offered to day for this purpose. The research presents a novel poly(vinylidene fluoride-co-hexafluoropropylene) (PVDF-HFP)/polyvinylpyrrolidone (PVP) polymer composite with MoO3 nanowires with the ability to form coating films on wood. The films of the developed coating exhibit elastic behaviour, which depends on the coating film thickness [tested wet film thicknesses (90, 180 and 360) µm]. The coating showed the ability to interact well with the surface of common beech (Fagus sylvatica L.) wood, in terms of wetting (contact angles of 15.6°), fast spilling on the surface, good penetration of the coating in wood structure and formation of up to 40 µm-thick films with excellent pull-off adhesion strength (6 MPa). An increased roughness of wood coated with C + MoO3 was a consequence of wood etching by the dimethylformamide solvent present in the coating. Moreover, the presence of C + MoO3 on wood made it considerably more hydrophobic, with contact angle of water raising to 123° from initially 46° measured on uncoated wood. The irradiation of wood surfaces with ultra-violet light resulted in visible colour changes on both uncoated and coated wood. The wood coated with C + MoO3 has a good resistance to water, alcohol and dry heat (grade 3 to 4). The antimicrobial testing showed that the presence of MoO3 in the coating plays an important role in the resistance of the coated wood to blue-stain fungi and mould development. The developed PVDF-HFP/PVP/MoO3 coating has an excellent ability to interact with the wood surface and has the potential to be used as a protection for wood in sensitive environments.

Transcriptomic analysis of the fungus Graphilbum sp. in response to the pine wood nematode.

Graphilbum species are important blue stain fungi associated with pine trees and are widely distributed throughout Asia, Australia, and North Africa. Pine wood nematode (PWN) primarily feed on ophiostomatoid fungi such as Graphilbum sp. in wood, the population of PWNs was increased, and incomplete organelle structures were observed in Graphilbum sp. hyphal cells following exposure to PWNs. In this study, we showed that Rho and Ras were involved in the MAPK pathway, SNARE binding and small GTPase-mediated signal transduction, and their expression was upregulated in the treatment group. However, the expression of the Rab7 involved in MAPK and small GTPase-mediated signal pathway was downregulated in the treatment group. Thus, further research is needed to study the MAPK pathway and related Ras and Rho genes in Graphilbum sp. associated with the PWN population. Overall, transcriptomic analysis clarified the basic mechanisms of mycelial growth in Graphilbum sp. fungus used as a food source by PWNs.

Blue-stain development on Norway spruce logs under alpine conditions

Discoloration of the sapwood caused by blue-stain fungi on conifer logs during interim storage causes significant loss to the forest industry. The fungal infection is often associated with bark beetle attacks because the spores are transmitted by the beetles. They can also be disseminated by rain-splash and moist air. While there are methods to protect logs from sap-stain in wood yards, this is often not possible in the forest for practical and regulatory reasons. Timing of harvesting and timely transportation are often the only ways to prevent blue-stain. To estimate the urgency of transportation, knowledge of the growth of blue-stain fungi and its dependence on weather conditions is of great interest. The proportion of discolored sapwood on Norway spruce logs was recorded along a time series, together with weather data in two field experiments conducted in spring and summer at two alpine sites in Austria. A predictive model was developed to estimate the proportion of blue-stained sapwood based on the temperature sum to which the logs were exposed. After harvest in March, there was a time lag of 82 and 97 days at the two respective sites, caused by initially low temperatures, before discoloration started. In contrast, sap-stain occurred 14 days after the harvest in June, when warm conditions prevailed from the start. The nonlinear least square regression model can help to estimate a window of opportunity to transport wood before it loses its value and serves as a sub model for lead time estimation within logistic decision support systems.

Fungal Communities of the Pine Wilt Disease Complex: Studying the Interaction of Ophiostomatales With Bursaphelenchus xylophilus.

Considered one of the most devastating plant–parasitic nematodes worldwide, Bursaphelenchus xylophilus (commonly known as pinewood nematode, PWN) is the causal agent of the pine wilt disease in the Eurasian coniferous forests. This migratory parasitic nematode is carried by an insect vector (Monochamus spp.) into the host tree (Pinus species), where it can feed on parenchymal cells and reproduce massively, resulting in the tree wilting. In declining trees, PWN populations are strongly dependent on fungal communities colonizing the host (predominantly ophiostomatoid fungi known to cause sapwood blue-staining, the blue-stain fungi), which not only influence their development and life cycle but also the number of individuals carried by the insect vector into a new host. Our main aim is to understand if PWN-associated mycobiota plays a key role in the development of PWD, in interaction with the PWN and the insect vector, and to what extent it can be targeted to disrupt the disease cycle. For this purpose, we characterized the fungal communities of Pinus pinaster trees infected and non-infected with PWN in three collection sites in Continental Portugal with different PWD temporal incidences. Our results showed that non-infected P. pinaster mycoflora is more diverse (in terms of abundance and fungal richness) than PWN-infected pine trees in the most recent PWD foci, as opposed to the fungal communities of long-term PWD history sites. Then, due to their ecological importance for PWN survival, representatives of the main ophiostomatoid fungi isolated (Ophiostoma, Leptographium, and Graphilbum) were characterized for their adaptative response to temperature, competition in-between taxa, and as food source for PWN. Under the conditions studied, Leptographium isolates showed promising results for PWN control. They could outcompete the other species, especially O. ips, and significantly reduce the development of PWN populations when compared to Botrytis cinerea (routinely used for PWN lab culturing), suggesting this to be a natural antagonist not only for the other blue-stain species but also for the PWN.

Insights into the Role of Fungi in Pine Wilt Disease.

Pine wilt disease (PWD) is a complex disease that severely affects the biodiversity and economy of Eurasian coniferous forests. Three factors are described as the main elements of the disease: the pinewood nematode (PWN) Bursaphelenchus xylophilus, the insect-vector Monochamus spp., and the host tree, mainly Pinus spp. Nonetheless, other microbial interactors have also been considered. The study of mycoflora in PWD dates back the late seventies. Culturomic studies have revealed diverse fungal communities associated with all PWD key players, composed frequently of saprophytic fungi (i.e., Aspergillus, Fusarium, Trichoderma) but also of necrotrophic pathogens associated with bark beetles, such as ophiostomatoid or blue-stain fungi. In particular, the ophiostomatoid fungi often recovered from wilted pine trees or insect pupal chambers/tunnels, are considered crucial for nematode multiplication and distribution in the host tree. Naturally occurring mycoflora, reported as possible biocontrol agents of the nematode, are also discussed in this review. This review discloses the contrasting effects of fungal communities in PWD and highlights promising fungal species as sources of PWD biocontrol in the framework of sustainable pest management actions.

Endemic Jeffrey Pine Beetle Associates: Beetle/Mite Fungal Dissemination Strategies and Interactions That May Influence Beetle Population Levels.

Fungal and mite associates may drive changes in bark beetle populations, and mechanisms constraining beetle irruptions may be hidden in endemic populations. We characterized common fungi of endemic-level Jeffrey pine beetle (JPB) in western USA and analyzed their dissemination by JPB (maxillae and fecal pellet) and fungivorous mites to identify if endogenous regulation drove the population. We hypothesized that: (1) as in near-endemic mountain pine beetle populations, JPB’s mutualistic fungus would either be less abundant in endemic than in non-endemic populations or that another fungus may be more prevalent; (2) JPB primarily transports its mutualistic fungus, while its fungivorous mites primarily transport another fungus, and (3) based on the prevalence of yeasts in bark beetle symbioses, that a mutualistic interaction with blue-stain fungi present in that system may exist. Grosmannia clavigera was the most frequent JPB symbiont; however, the new here reported antagonist, Ophiostoma minus, was second in frequency. As hypothesized, JPB mostly carried its mutualist fungus while another fungus (i.e., antagonistic) was mainly carried by mites, but no fungal transport was obligate. Furthermore, we found a novel mutualistic interaction between the yeast Kuraishia molischiana and G. clavigera which fostered a growth advantage at temperatures associated with beetle colonization.

Termite Presence and Feeding on Loblolly Pine Wood Differs Among Four Root-Infecting Bluestain (ophiostomatoid) Fungal Species.

Bark beetles and root weevils can impact forests through tree death on landscape scales. Recently, subterranean termites have been linked to these beetles via the presence of bluestain fungi (Ascomycota: Ophiostomataceae), which are vectored to trees by beetles. However, only a small subset of bluestain species have been examined. Here, we tested whether termite-bluestain association patterns in the field reflect termite feeding preference in laboratory choice trials. We documented the presence of four bluestain fungi (Leptographium procerum (W.B. Kendr.), L. terebrantis (Barras & Perry), Grosmannia huntii (Rob.-Jeffr.), and G. alacris (T.A. Duong, Z.W. de Beer & M.J. Wingf.) in the roots of 2,350 loblolly pine trees in the southeastern United States and whether termites were present or absent on these roots and paired this with laboratory choice feeding trials. Termites were found 2.5-fold on tree roots with at least one bluestain fungus present than tree roots without bluestain fungi. Although termites in this study and others were associated with L. procerum, L. terebrantis, and marginally G. huntii, termites only showed preferential feeding on wood inoculated with G. huntii in laboratory trials. This suggests that increased termite presence on wood with bluestain fungi may be driven by factors other than increased wood palatability. Termites could thus disproportionately affect wood turnover rates for specific pools (e.g., bark beetle and root weevil attacked trees) and in some cases (e.g., G. huntii) accelerate wood decomposition. This study supports the growing evidence that the association between subterranean termites and bluestain fungi is spatially and taxonomically widespread.

Seasonal variation in Norway spruce response to inoculation with bark beetle-associated bluestain fungi one year after a severe drought

In 2018, up to 4 million m3 Norway spruce was killed by the spruce bark beetle Ips typographus in Sweden. The event was unique for Sweden, in terms of both affected volume and the fact that it was triggered by severe drought stress, not by ample availability of relatively defenseless storm-felled trees. The outbreak continued in 2019 and 2020, each year with twice as many trees killed as in 2018. The aim of this study was to quantify seasonal variation and potential lag-effects in tree defense capacity the year after a severe drought stress. Inoculation with a bark beetle-associated bluestain fungus, repeated four times with one-month-intervals between May and August 2019, were carried out at three field sites with spruce provenances of Swedish and East European origin representing early and late bud burst, respectively.All sites had experienced moderate to severe drought stress in 2018, and site-specific defense capacity correlated positively with the cumulative precipitation two months before inoculation. Sites with two-month precipitation levels <100 mm had larger necrotic lesions in the phloem following inoculation, an indication of lower tree defense capacity. Lesion size did not differ between provenances, and all trees were able to confine fungal infection successfully.There were some seasonal differences in necrotic lesion size, with the sites Skärsnäs and Norberg having significantly larger lesions in June than in May, and site Lugnet having large lesions also in May. Lesions were generally smaller in July and August than in June.The cross-sectional area and number of traumatic resin ducts was measured in sapwood samples from one site, Lugnet, to quantify an additional aspect of tree defenses. The area of resin ducts produced in May and June were larger than that in July and August. This is in line with a positive correlation between lesion area and resin duct area, indicating that a stronger fungal infection following inoculation in spring triggered a stronger induced defense response. The East European provenances had more resin ducts than Swedish provenances, but the area of resin ducts did not differ significantly between provenances.

Variation in Methyl Jasmonate-Induced Defense Among Norway Spruce Clones and Trade-Offs in Resistance Against a Fungal and an Insect Pest.

An essential component of plant defense is the change that occurs from a constitutive to an induced state following damage or infection. Exogenous application of the plant hormone methyl jasmonate (MeJA) has shown great potential to be used as a defense inducer prior to pest exposure, and could be used as a plant protection measure. Here, we examined (1) the importance of MeJA-mediated induction for Norway spruce (Picea abies) resistance against damage by the pine weevil Hylobius abietis, which poses a threat to seedling survival, and infection by the spruce bark beetle-associated blue-stain fungus Endoconidiophora polonica, (2) genotypic variation in MeJA-induced defense (terpene chemistry), and (3) correlations among resistance to each pest. In a semi-field experiment, we exposed rooted-cuttings from nine different Norway spruce clones to insect damage and fungal infection separately. Plants were treated with 0, 25, or 50 mM MeJA, and planted in blocks where only pine weevils were released, or in a separate block in which plants were fungus-inoculated or not (control group). As measures of resistance, stem area debarked and fungal lesion lengths were assessed, and as a measure of defensive capacity, terpene chemistry was examined. We found that MeJA treatment increased resistance to H. abietis and E. polonica, but effects varied with clone. Norway spruce clones that exhibited high constitutive resistance did not show large changes in area debarked or lesion length when MeJA-treated, and vice versa. Moreover, insect damage negatively correlated with fungal infection. Clones receiving little pine weevil damage experienced larger lesion lengths, and vice versa, both in the constitutive and induced states. Changes in absolute terpene concentrations occurred with MeJA treatment (but not on proportional terpene concentrations), however, variation in chemistry was mostly explained by differences between clones. We conclude that MeJA can enhance protection against H. abietis and E. polonica, but the extent of protection will depend on the importance of constitutive and induced resistance for the Norway spruce clone in question. Trade-offs among resistances do not necessarily hinder the use of MeJA, as clones that are constitutively more resistant to either pest, should show greater MeJA-induced resistance against the other.

Legacy of bark beetles (Dendroctonus spp.) on soil carbon and nitrogen cycling seven years after forest infestation

Between years 2007 and 2013, an outbreak of pine- and spruce-bark beetles (Dendroctonus spp.) has decimated coniferous forests of the northwestern United States. Beetles introduced blue stain fungus to the tree xylem that caused blockage and discontinuation of plant water conductance. One of the immediate disease symptoms was premature and immediate drop-off of all needles rich in nitrogen (N). Soil surface-accumulated needle litter in combination with elevated soil water levels, have led to accelerated N and carbon (C) mineralization. Understanding of the soil C and N cycling during forest transition from “standing dead trees” to “snagfall and forest regrowth” is still unclear. Specifically, the potential lasting legacy of beetle-caused surface-deposited tree litter is of importance in order to assess long-term forest health. The main objective of this study was to assess whether soil C and N mineralization continues to be affected by litter chemistry in this transitional period of post beetle disturbance ecosystem recovery. Mineral soil (0–10 cm) and corresponding surface tree litter were obtained from beneath dead and live tree clusters in three forest types in Medicine Bow-Routt National Forest in Wyoming, USA. All forests were located in the same watershed, grew on similar soils, experienced up to 35% of forest mortality and consisted of: Engelmann spruce (Picea engelmannii), subalpine fir (Abies lasiocarpa) mixed with lodgepole pine (Pinus contorta), and lodgepole pine alone. Results suggest that tree litter uniformly affected mineral soils across all forest types regardless of beetle infestation. In laboratory incubations, the presence of litter doubled mineral soil heterotrophic respiration. While litter had the most uniform effect on soils, beetle infestation and forest type resulted in a number of differences. Soils beneath dead tree clusters continued to have elevated inorganic N concentrations when normalized by total N, which was twice as high compared to soils beneath live tree clusters across all forest types. The lowest NH4+ concentrations were observed in soils beneath lodgepole pine trees, which was accompanied by the highest C to N ratio of microbially available substrates. In conclusion, the legacy of forest die-offs caused by bark beetle infestation was still observable seven years later during the early transitional period. This confirms ecosystem modeling simulations that, while entering decadal stage of forest recovery, forest structure may differ depending on the forest type, and that the forest types that contain lodgepole pines may encounter an earlier onset of potential N limitation.

The Mitogenomes of Ophiostoma minus and Ophiostoma piliferum and Comparisons With Other Members of the Ophiostomatales.

Fungi assigned to the Ophiostomatales are of economic concern as many are blue-stain fungi and some are plant pathogens. The mitogenomes of two blue-stain fungi, Ophiostoma minus and Ophiostoma piliferum, were sequenced and compared with currently available mitogenomes for other members of the Ophiostomatales. Species representing various genera within the Ophiostomatales have been examined for gene content, gene order, phylogenetic relationships, and the distribution of mobile elements. Gene synteny is conserved among the Ophiostomatales but some members were missing the atp9 gene. A genome wide intron landscape has been prepared to demonstrate the distribution of the mobile genetic elements (group I and II introns and homing endonucleases) and to provide insight into the evolutionary dynamics of introns among members of this group of fungi. Examples of complex introns or nested introns composed of two or three intron modules have been observed in some species. The size variation among the mitogenomes (from 23.7 kb to about 150 kb) is mostly due to the presence and absence of introns. Members of the genus Sporothrix sensu stricto appear to have the smallest mitogenomes due to loss of introns. The taxonomy of the Ophiostomatales has recently undergone considerable revisions; however, some lineages remain unresolved. The data showed that genera such as Raffaelea appear to be polyphyletic and the separation of Sporothrix sensu stricto from Ophiostoma is justified.

Evaluation of Stem-Injected TREE-äge® (4% Emamectin Benzoate) for Protecting Western White Pines (Pinus monticola) from Mountain Pine Beetle (Dendroctonus ponderosae Hopkins)(Coleoptera: Curculionidae: Scolytinae)

The protection of high-value trees against bark beetles and the development of alternatives to bole sprays is a priority for the tree manager. The objective of this study was to evaluate stem-injected TREE-äge® (emamectin benzoate [EB]) as a protective treatment for western white pines (Pinus monticola Dougl. ex D. Don) against mountain pine beetle (MPB, Dendroctonus ponderosae Hopkins). Treatment efficacy was based solely on tree mortality as per Shea protocols (i.e., ≥ 60% check vs. ≤ 20% treated tree mortality). Our first experiment was installed in 2007 and included trees stem-injected with TREE-äge and untreated controls. Bole application of S-(-)-verbenone and green leaf volatile (GLV) blend was included for observational comparison. Pressure from MPB was heavy, as indicated by the number and timing of control tree mortality (90%). Strip attacks by MPB in TREE-äge trees indicated that the impacts of EB, and by inference its distribution, were inconsistent. In 2009, the injection protocol was revised to improve EB distribution in the phloem via closer injection points. In the 2009 TREE-äge-treated trees, adult beetle mining stopped when they contacted phloem and was insufficient to cause tree death by girdling. Blue-stain fungi colonized the sapwood of trees in both studies. Isolates from autopsied trees treated with TREE-äge alone were subsequently identified as Grosmannia clavigera and Leptographium longiclavatum (Ophiostomatales: Ascomycota), species that can incite tree mortality. In 2013, we revised our protocol to include GLV plus verbenone or propiconazole with TREE-äge, wherein these treatments proved effective in protecting trees against MPB and their associated pathogenic fungi.