华山松大小蠹和共生真菌分泌酶组成分析

对华山松大小蠹消化道和共生真菌胞外蛋白和酶类组成分析的结果表明 ,华山松大小蠹消化道和共生真菌胞外均可分泌多种酶类 ,其中华山松大小蠹消化道具自身分泌纤维素酶、酯酶、蛋白酶和淀粉酶的能力 ,但不具分泌漆酶等木质素分解酶的能力 ,因此 ,其虽然可以直接利用寄主华山松韧皮部和木质部边材组织内的纤维素、淀粉、多糖、酯类和蛋白等营养物质 ,但不能分解利用木质素类营养。而共生真菌除具有胞外分泌纤维素酶、酯酶、蛋白酶和淀粉酶的能力外 ,还能够在寄主华山松组织内木质素的诱导下分泌漆酶 ,以充分利用华山松韧皮部和木质部内纤维素、木质素、淀粉和脂类物质。此外 ,华山松大小蠹消化道和共生真菌胞外分泌的纤维素酶、酯酶、淀粉酶和蛋白酶还存在结构和营养利用方面的差异。     

华山松大小蠹共生真菌对寄主树木的影响

华山松大小蠹共生真菌（ Ｏｐｈｉｏｓｔｏｍａ ｓｐ ．和Ｌｅｐｔｏｇｒａｐｈｉｕ ｍ ｓｐ ．） 是克服寄主华山松树木抗性系统的先导和致死寄主树木的主要原因。华山松大小蠹共生真菌通过对寄主树木木质部树脂道泌脂细胞的分解和菌丝在寄主树木树脂道内的大量繁殖,堵塞寄主树木树脂道,使寄主华山松泌脂系统失去活性和抗性功能。同时,随共生真菌在寄主树木木质部内的发育,菌丝分解寄主木质部薄壁细胞,使寄主树木木质部物质代谢紊乱,导致寄主华山松迅速死亡     

无菌松材线虫对10年生黑松致病性的研究

2001年9月,用无菌的松材线虫对10年生黑松进行接种.2002年8月,检查黑松发病情况,并对接种的松树进行线虫和细菌的再分离.试验发现:接种的8株黑松11个月后仍有5株表现正常,没有出现任何症状,而从其木质部却能分离到数量较多的无菌松材线虫.结果说明:无菌的松材线虫虽然能侵入松树木质部,但在不携带细菌的情况下对松树是没有致病力的.     

华山松大小蠹对华山松韧皮部营养和矿物元素的利用

对健康和蓝变真菌侵染的华山松韧皮部以及华山松大小蠹排泄物中粗蛋白、粗脂肪、粗纤维、淀粉、还原性糖、可溶性总糖、游离态氨基酸和Na,K,Ca,Mg,Fe,Cu,Mn,Zn等8种元素进行分析,结果表明:华山松大小蠹对寄主华山松营养物质的利用率依次为:还原性糖＞粗脂肪＞可溶性总糖＞粗纤维＞淀粉＞粗蛋白.华山松韧皮部的17种游离态氨基酸中含有华山松大小蠹必需的9种氨基酸;华山松大小蠹对寄主华山松韧皮部组织内的Na,K,Ca,Mg,Fe,Cu,Mn,Zn等8种矿物元素需求量存在明显差异,其中对K元素的需求量最大,而对Cu,Mn,Zn 3种元素的需求量较小.同时,华山松大小蠹共生真菌能够加速华山松木质部和韧皮部中营养的分解转化,为华山松大小蠹在被害华山松韧皮部和木质部间的生存和发育创造条件,并加速被害华山松树势的衰弱和死亡.     

纵坑切梢小蠹蓝污真菌侵害云南松树组织解剖观察

通过对云南松树进行纵坑切梢小蠹（Tomicus piniperda）蓝污真菌（Leptogramphium yunnanense）接种试验，对云南松受害组织进行了形态解剖观察，验证了蓝污真菌对云南松的致病作用。蓝污真菌发现于受害木质部和韧皮组织中的各类细胞中，在木质部，蓝污区域呈现出以心材为中心的扇形分布。在韧皮组织内，蓝污真菌主要沿着细胞轴向在垂直方向上生长，并可通过孔纹进入到水平方向上的相邻细胞内。本项研究揭示了该蓝污真菌在云南松树干韧皮和木质部组织，细胞内的生长和蔓延规律，进一步表明纵坑切梢小蠹蓝污真菌L.yunnanense是云南松的一种病原真菌。     

沙地云杉菌根真菌的鉴定及其促生作用

筛选和鉴定沙地云杉菌根真菌,对沙地云杉育苗以及种群恢复具有重要意义.本研究从沙地云杉根段筛选获得8株菌根真菌,经ITS基因测序比对,分别属于镰刀菌属、皮伞属等5属,其中皮伞属为优势菌属.其中4株菌能够在磷酸三钙固体P KO平板上生长,1株能够在硅酸盐固体平板上生长.盆栽实验表明,接种菌株Marasmius scorod...     

内生炭疽菌在茶树体内的分布及其内生特性

以茶树的一种优势种内生炭疽菌为研究对象,通过系统分离,研究其在不同茶树组织和不同龄期茶叶中的分布;并通过人工接种,研究炭疽菌的侵入过程、侵染菌丝的分布和真菌的再分离,以明确该真菌在茶树组织中的内生性.结果表明:该菌主要分布于茶树的叶片和枝条组织中,而在茶树根、主茎、花和果实等组织中没有分离获得该种炭疽菌.从茶树新芽到叶片展开、成熟的过程中,内生炭疽菌的分离率逐渐上升,但之后随着叶片的老化,分离率又逐步下降.人工接种试验表明,炭疽菌一般自茶树表皮细胞之间直接侵入寄主细胞,但侵染菌丝受到限制,一般只有几个细胞,分布于寄主细胞之间或细胞内.在炭疽菌的侵染点,有87%的寄主细胞表现原生质颗粒聚集和原生质沉积现象.在荧光显微镜下,有67.5%的真菌侵染点寄主细胞表现自发荧光现象,表明在这些细胞中有多酚类物质的积累.接种120 d后,茶树苗没有表现明显的病害症状,并能在人工接种的所有茶苗组织中再分离获得炭疽菌;采用涂抹接种的茶树苗茎部的分离率可高达41.17%.内生炭疽菌在茶树的不同组织中的分布具有明显的组织专化性;在炭疽菌的感染过程中,茶叶细胞表现一定的抗性反应,炭疽菌的内生性可能是真菌-茶树相互作用达到平衡的一种表现.     

松材线虫对数种针叶树的致病性

过去人们一直认为,日本松材线虫的寄主仅限于松属的树木,但在 1978年高温干旱天气,发现也危害绿化树种雪松、欧洲云杉,这就有必要重新明确线虫的寄主范围.笔者就此问题进行了研究,认为该线虫对松属以外的数种针叶树亦有致病性.本文概要报道了这一研究成果.实验方法(一)供试树种 5—8年生,树高约1米的日本冷杉、库页冷杉、日本落叶松、鱼鳞松、欧洲云杉、日本五针松以及8年生、树高2米的雪松等7种.各种树供试株数:接种20株,对照2—7株.(二)供试材线虫 采用从接种羽化的     

蓝变木材基底可可球二孢菌丝复合材料的性能

为了提高易蓝变木材的利用率,利用蓝变菌可可球二孢（Lasiodiplodia theobromae）和蓝变的欧洲赤松（Pinus sylvestris）、橡胶木（Hevea brasiliensis）碎料以及两者混合物制成的基底制备菌丝复合材料,并探讨含水率变化对菌丝复合材料性能的影响。结果表明：培养14 d后,三种基底表面均被菌丝覆盖,颜色显著变黑。增加板坯含水率促进了菌丝复合材料内发生木质素再聚合和美拉德反应,提高了菌丝复合材料的弯曲强度。蓝变欧洲赤松基底、蓝变橡胶木基底以及混合基底菌丝复合材料的弯曲强度分别提高了64%、221%和74%。与蓝变欧洲赤松基底相比,蓝变橡胶木基底更适合可可球二孢菌丝的生长,因此橡胶木基菌丝复合材料的弯曲强度更高,可以达到ANSI A208.1-2016“Particleboard”对轻质刨花板的弯曲强度要求。菌丝表面的疏水蛋白使材料表面接触角增大,疏水性增加,而板坯含水率的提高对材料的疏水性和耐水性有削弱作用。     

树木抗性与小蠹虫生存策略的进化

小蠹虫对寄主树木的入侵危害不仅受到树木原生性抗性的直接影响 ,而且小蠹虫在寄主树木组织内的定居繁衍也受到寄主树木诱导性抗性的制约。寄主树木抗性的强弱决定小蠹虫入侵寄主树木的数量 ,也促使小蠹虫与特定的真菌共生 ,真菌通过对寄主树木生理生化代谢的影响 ,以削弱寄主树木抗虫性 ,从而为小蠹虫在寄主树木组织内的定居创造必备的生存条件。同时小蠹虫为适应真菌的共生和寄主树木的抗虫性 ,促使小蠹虫生存繁衍策略高度进化 ,从而形成小蠹虫—共生真菌—寄主树木三者协同进化共生的关系     

Mechanical injury and fungal infection induce acquired resistance in Norway spruce

Norway spruce trees (Picea abies (L.) Karst.) pretreated by wounding and fungal infection showed highly enhanced resistance to a subsequent challenge inoculation with the pathogenic bluestain fungus Ceratocystis polonica (Siem.) C. Moreau. This is the first time the effectiveness of the constitutive and inducible defenses has been shown to depend on prior wounding and infection in conifers, although such acquired resistance has previously been found in several angiosperms. Trees that were pretreated with a combination of 12 bark wounds (1.6 x 10 cm), four fungal inoculations and four sterile inoculations 1-15 days before mass inoculation with C. polonica at 400 inoculations per square meter over a 0.8 m stem section had significantly shorter necroses in the phloem, less bluestained sapwood, and less dead cambium than untreated control trees. Pretreatment with four fungal or sterile inoculations alone did not lead to enhanced resistance. Pretreatment by bark wounding alone seemed to provide an intermediate degree of resistance compared to bark wounding, fungal inoculations and sterile inoculations combined. All trees had a marked increase in the number of resin ducts in the year of inoculation compared with previous years, suggesting that formation of traumatic resin ducts play an important role in the development and maintainance of enhanced resistance.     

Inducible anatomical defense responses in Norway spruce stems and their possible role in induced resistance

Norway spruce (Picea abies (L.) Karst.) trees were preinoculated with a sublethal dose of the blue-stain fungus Ceratocystis polonica Siem. (C. Moreau) 1 to 52 weeks before they were mass inoculated with the same fungus. Trees pretreated 1 week before mass inoculation had similar, severe symptoms of fungal infection as the control trees. Pretreatment 3, 6 or 9 weeks before mass inoculation resulted in effective protection of the trees, reducing pathogenic symptoms by 63-90% relative to the control trees, whereas pretreatment 52 weeks before mass inoculation gave intermediate protection (44-71% reduction in symptoms). Thus, pretreatment induced resistance to the blue-stain fungus in Norway spruce by a process that requires more than 1 week to become activated and protects trees for at least one year after pretreatment. Pretreatment induced formation of traumatic resin ducts (TDs) in the sapwood and swelling and proliferation of polyphenolic parenchyma cells (PP cells) in the phloem. Trees pretreated 3-9 weeks before mass inoculation had more TDs and showed greater swelling of existing PP cells than control trees or trees pretreated 1 week before mass inoculation. We conclude that induced disease resistance in Norway spruce is probably associated with PP cell activation and TD induction, because resistance was enhanced within the same time frame as the induction of these defense responses.     

Variation in Tree Size and Resistance to Ceratocystis polonica in a Monoclonal Stand of Picea abies

Tree resistance to the pathogenic blue-stain fungus Ceratocystis polonica was studied in a monoclonal stand of Norway spruce [ Picea abies (L.) Karst.] in relation to tree social status and diameter at breast height (DBH). The DBH distribution of the 33-yr-old stand ranged from 5 to 35 cm. There were clear differences in tree height between the suppressed (DBH 7.4-10.3 cm), codominant (DBH 11.8-17.4 cm) and dominant (DBH 18.6-23.9 cm) tree classes. The resistance was tested by mass inoculating trees with a low (400 inoculations m -2 , 60 cm inoculation belt) or high (400 inoculations m -2 , 120 cm inoculation belt) dosage. The small, suppressed trees were more susceptible to inoculation than the codominant and dominant trees, based on the amount of blue-stained and occluded sapwood, lesion length and dead cambium/phloem. A threshold in tree social status or tree size may be important in the overall resistance to fungal infection.     

Methyl jasmonate and oxalic acid treatment of Norway spruce: anatomically based defense responses and increased resistance against fungal infection

To study the effect of chemical pretreatment on conifer resistance, 13-year-old Norway spruce (Picea abies (L.) Karst.) trees were treated with methyl jasmonate (MJ) or oxalic acid (OxA) on the outer bark and inoculated with the pathogenic blue-stain fungus Ceratocystis polonica (Siem.) C. Moreau 4 weeks later. Both chemicals significantly reduced symptoms of fungal infection, but MJ was more effective than OxA (51 versus 18% reduction in length of necrotic lesions in the phloem relative to untreated control trees). Anatomical examination of treated stem tissues showed that MJ induced extensive formation of traumatic resin ducts in the xylem and extra polyphenolic parenchyma (PP) cells in the secondary phloem between the cambium and the regular annual PP cell layer. No traumatic resin ducts were formed after treatment with OxA, and the coverage of extra PP cells in OxA-treated tissues was not significantly higher than in the controls. The anatomically based defense reactions induced by MJ were similar to the reactions observed after pathogen infection, mechanical wounding and bark beetle attack. Neither MJ nor OxA had apparent phytotoxic effects on Norway spruce at the concentrations used, with needle and stem tissues of all trees appearing normal without visible symptoms of toxicity. However, trees treated with MJ had 30% less radial sapwood growth than control trees. In conclusion, MJ treatment of Norway spruce appears to have practical potential as a tool for increasing plant resistance to fungal infection, but with a modest reduction in sapwood growth.     

Intra- and inter-provenance variability in phloem phenols of Picea abies and relationship to a bark beetle-associated fungus

One hundred Norway spruce (Picea abies (L.) Karst.) clones (three ramets per clone) were analyzed for phloem phenol composition and concentration before and 10 days after wound inoculation with sterile malt agar. Fifty clones (Experiment 1) belonged to the same provenance, whereas the remaining clones (Experiment 2) belonged to five provenances from three geographic areas. In Experiment 2, two additional ramets from the same clones were mass inoculated with Ceratocystis polonica (Siem.) C. Moreau to quantify the resistance of each clone. Tree response to wound inoculations was characterized by increased catechin concentration in both experiments, accompanied by increases in astringin and decreases in piceid in Experiment 1. In both experiments, we observed a diverse group of phenolic compounds whose concentrations increased (catechin, astringin) or did not vary (taxifolin glucoside) in response to wound inoculations, whereas concentrations of a homogeneous group of stilbene compounds decreased (piceid) or did not vary (isorhapontin, unidentified stilbene). In Experiment 2, provenances from the alpine and Hercynian-Carpatic areas differed from provenances from the Baltic area with respect to the relative importance of these two groups of compounds, further indicating that the two groupings of phenolic compounds structure the Norway spruce populations. Eighty days after mass inoculation, the percentage of healthy sapwood, which was taken as a measure of tree resistance, indicated that clones from the Baltic area were less resistant to mass inoculations than clones from the alpine and Hercynian-Carpatic areas. We conclude that the degree of resistance of Norway spruce trees to mass inoculations with a bark beetle-associated fungus can be predicted based on the diversity of constitutive phloem phenols and the ability to induce phenol synthesis in response to wounding.     

Pathogenicity of Ceratocystis resinifera to Norway spruce

The blue‐stain fungus Ceratocystis resinifera colonizes wounds on living Picea spp. and other conifers in Europe and North America. Little is known regarding the pathogenicity of this fungus and consequently, four Norwegian C. resinifera isolates were inoculated on to Norway spruce (Picea abies) using two different techniques. These included single‐point inoculations on young trees (two inoculations per tree on 14‐year‐old trees) and mass‐inoculations on older trees (～200 inoculations per tree on 34‐year‐old trees). In both experiments, C. resinifera induced minor symptoms that in most cases did not differ significantly from inoculation with sterile agar. The virulent blue‐stain fungus C. polonica, which was inoculated for comparative purposes, induced extensive symptoms, causing 83% dead cambium circumference and 82% blue‐stained sapwood, and long necrotic lesions in the phloem. The results suggest that C. resinifera is non‐pathogenic or only mildly pathogenic to Norway spruce and does not present a threat to these trees.     

Methyl jasmonate treatment of mature Norway spruce (Picea abies) trees increases the accumulation of terpenoid resin components and protects against infection by Ceratocystis polonica, a bark beetle-associated fungus

When conifers such as Picea abies Karst. (Norway spruce) are attacked by insects or pathogens, they often respond by producing increased quantities of terpenoid oleoresin. This response can be mimicked in young P. abies seedlings by treatment with methyl jasmonate (MJ). In this study, we determined the effects of MJ on terpenoids and other chemical defenses of mature P. abies, and investigated whether this treatment protected trees against attack by the blue-stain fungus Ceratocystis polonica (Siem.) C. Moreau, the most important fungal associate of the spruce bark beetle Ips typographus L. Methyl jasmonate treatment induced the formation of traumatic resin ducts in the developing xylem, enhanced resin flow and stimulated increased accumulation of monoterpenes, sesquiterpenes and diterpene resin acids. However, only minor changes were detected in terpene composition in response to MJ treatment and no changes in soluble phenolic concentration were measured. There was much variability in the timing and degree of response to MJ among clones. The observed chemical and anatomical changes in response to MJ treatment were correlated with increased resistance to C. polonica, suggesting that terpenoid oleoresin may function in defense against this pathogen.     

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.     

Induced resistance of Norway spruce,variation of phenolic compounds and their effects on fungal pathogens

Three clones of Norway spruce (Picea abies) were studied for their response to mass‐inoculation with the blue‐stain fungus Ceratocystis polonica. The effect of different pretreatments (fungal inoculation and wounding) before mass‐inoculation was investigated for their possible role in an acquired resistance reaction. Pretreated trees showed enhanced resistance to the subsequent mass‐inoculation relative to control trees that received no pretreatment. Furthermore, the fungal colonization of inoculated trees was less than that of wounded trees. The phenolic content of the bark, analysed by RP‐HPLC, was compared in trees receiving different treatments. Trees inoculated with C. polonica had higher average concentration of (+)‐catechin, taxifolin and trans‐resveratrol than wounded trees. Both inoculated and wounded trees had higher average concentrations of these compounds than control trees. The effect of the phenolic extract of Norway spruce bark on the growth of the root rot fungus Heterobasidion annosum and the blue‐stain fungi C. polonica and Ophiostoma penicillatum were investigated in vitro. Heterobasidion annosum was not negatively affected, and the extracts had fungistatic effects on the blue‐stain fungi. The growth of O. penicillatum was more inhibited than the growth of the more aggressive C. polonica.     

Low-density Ceratocystis polonica inoculation of Norway spruce (Picea abies) triggers accumulation of monoterpenes with antifungal properties

Among the most devastating pests of Norway spruce (Picea abies) are the European spruce bark beetle (Ips typographus) and the associated pathogenic blue-stain fungus Ceratocystis polonica. Following attack and colonization, the beetle and the fungus must cope with induced host chemical defenses, such as monoterpenes that are generally thought to be toxic to both symbionts. The goal of this study was to better understand the response of Norway spruce following C. polonica inoculation at low density that does not overwhelm the tree and to identify monoterpenes mobilized toward the fungus. We inoculated healthy mature trees and monitored monoterpene profiles 2, 3, and 5 months post-inoculation. We also exposed three different C. polonica strains to the most abundant or significantly up-regulated monoterpenes to determine differences in monoterpene toxicity and resistance among strains. Total monoterpene levels, including limonene, were increased at 2 and 3 months after inoculation and had dropped after 5 months. In in vitro assays, all monoterpenes were inhibitory to C. polonica. Limonene and β-pinene were the most potent inhibitors of fungal growth. The extent of inhibition varied between the three strains tested. These results showed a defense response of Norway spruce to C. polonica, in which limonene may play a critical role in inhibiting the spread of the fungus. We also showed that differences between strains of C. polonica must be taken into account when assessing the role of the fungus in this bark beetle–symbiont system.