高温胁迫对刺芹侧耳菌丝生长及其抗棘孢木霉能力的影响

耐高温性是食用菌抗逆性评价的重要指标。选取5个刺芹侧耳菌株为材料,通过观测常温培养和高温胁迫后恢复培养菌落的生长速率、生长势、恢复生长时间、菌丝显微形态特征以及受棘孢木霉侵染情况,研究高温胁迫对刺芹侧耳菌丝生长及其抗棘孢木霉侵染能力的影响。结果表明,刺芹侧耳不同菌株对高温胁迫的响应存在差异。供试PDA培养的菌丝经高温胁迫(37℃,2d)后,菌丝恢复生长时间2–3d,生长势变壮或无变化,顶端菌丝细胞长度、直径、菌丝体分支频率和菌丝生长速率等均呈现不同程度的降低。菌丝生长速率变化与其抗棘孢木霉能力的下降呈正相关。在此基础上,模拟生产试验发现,长满菌丝的菌棒经高温胁迫(35℃,7d)后,接种棘孢木霉菌丝的侵染率变为100%,接种棘孢木霉孢子的侵染率变为30%;未经高温胁迫的菌棒,接种棘孢木霉孢子/菌丝,未发生侵染。本研究表明,高温胁迫导致刺芹侧耳菌丝抗棘孢木霉侵染能力的下降。本研究将为科学指导食用菌菌种繁育、菌棒生产提供数据支撑,为食用菌育种提供基础数据。     

非生物胁迫因子对棘孢木霉嗜铁素及钙调磷酸酶基因表达的影响

木霉菌(Trichoderma spp.)嗜铁素具有吸收、贮存和胞内运输铁的功能,对菌体自身的生长发育有重要作用,也是重要的生防促生因子,但嗜铁素的产量会受到环境中多种胁迫因子的影响。真菌的钙调磷酸酶基因能够调控外源离子的胁迫反应,但钙调磷酸酶信号通路与嗜铁素合成之间的关系并不明确。以棘孢木霉菌(Trichoderma asperellum)T6为材料,研究了NaCl、CaCl_2及pH对棘孢木霉菌T6嗜铁素产量以及嗜铁素合成关键基因(SidA)与钙调磷酸酶催化亚基基因(CnA)表达量的影响。结果表明,高浓度NaCl和CaCl_2以及高pH均能抑制嗜铁素的产生。利用半定量PCR分析得知,CnA基因与SidA基因之间存在一定的负调控关系,该结果与钙调磷酸酶专性抑制剂环孢菌素A能够提高嗜铁素产量的结果相一致。     

高耐砷真菌的分离及其耐砷能力

从湖南石门县及郴州市重金属矿周边地区采集的6个砷污染土壤样品中,初步分离得到13株具有较强耐砷能力的真菌菌株,其中3株真菌的耐砷能力最强,经形态与分子鉴定分别为微紫青霉(Penicillin janthinellum)、尖孢镰刀菌(Fusarium oxysporum)和棘孢木霉(Trichoderma asperellum).培养试验表明：微紫青霉、尖孢镰刀菌和棘孢木霉分别在30000 mg·L^-1、30000 mg·L^-1和20000 mg·L^-1的砷胁迫平板中表现出很好的菌落生长状况;在砷浓度分别为0～50、0～50和0～80 mg·L^-1的液态培养基中培养2 d后,尖孢镰刀菌、棘孢木霉和微紫青霉的生物量均随砷浓度增加而增加,且在相应最高浓度(50、50、80 mg·L^-1)下的生物量显著高于不加砷的对照.此外,高浓度砷对该3株真菌的产孢能力没有影响.     

农杆菌介导生防棘孢木霉菌转化体系的优化

目的:实现棘孢木霉菌T4的遗传转化并优化其转化体系.方法:以潮霉素抗性为选择标记,利用农杆菌转化法介导转化棘孢木霉菌.结果:潮霉素基因成功整合到受体菌基因组中,转化子抗性基因可稳定遗传.结论:最优的转化体系和条件为:IM和CM培养基中AS浓度为200 μg/mL,棘孢木霉T4孢子浓度为106/mL,农杆菌浓度为200 μL( OD600约0.8),共培养时间为48 h,转化效率约为50个转化子/106个孢子.     

两种野生灌木幼苗对干旱和盐交叉胁迫的生理响应

长枝木蓼(Ateaphaxis viegata)和刺叶锦鸡儿(Caragana acanthophulla)是乌鲁木齐周边植被组成的重要种,在植被恢复中具有潜在价值。该文通过盆栽控水控盐法研究两种野生灌木幼苗在不同程度的干旱和盐交叉胁迫下的生理反应。结果表明,干旱和盐交叉胁迫下,长枝木蓼可溶性糖增幅较刺叶锦鸡儿大;轻度交叉胁迫时,两种灌木的可溶性糖含量高于仅干旱胁迫或盐胁迫下的。中度和重度交叉胁迫下,长枝木蓼叶片丙二醛含量增幅高于刺叶锦鸡儿。交叉胁迫下,两种灌木叶片叶绿素含量下降,且长枝木蓼叶片的叶绿素含量降幅大于刺叶锦鸡儿。因膜系统的过氧化作用,使MDA含量升高。轻度干旱胁迫提高了两种野生灌木对盐胁迫的耐受能力。两种灌木能很好地适应中度交叉胁迫,重度交叉胁迫对两种灌木的伤害很大,但也能生存。综合比较各参数发现,刺叶锦鸡儿更耐早,而长枝木蓼更耐盐;综合比较各生理参数发现,刺叶锦鸡儿对干旱和盐胁迫的耐受能力较长枝木蓼强。     

棘孢木霉挥发性次级代谢产物检测及抑菌活性分析

木霉是一类具有重要生防价值的丝状真菌。文中首先对分离自浙江省绍兴市和广东省佛山市共12株棘孢木霉Trichoderma asperellum进行平板拮抗评价,然后采用顶空固相微萃取气质联用法(HS-SPME-GC-MS)检测拮抗性较好的两株菌的挥发性次级代谢产物。结果表明,棘孢木霉ZJSX5003和GDFS1009菌丝生长迅速,对尖孢镰孢菌Fusariumoxysporum抑制率分别达73%和74%。挥发性次级代谢产物主要是醇类和酮类,其中包含异丁醇、异戊醇、3-甲基-3-丁烯-1-醇、3-羟基-2-丁酮、2,3-丁二醇和6-正戊基-2H-吡喃-2-酮(6-PAP)。进一步通过体外抑菌试验,证实6-PAP具有较好的抑制尖孢镰孢菌的效果,为开发以木霉菌代谢产物如6-PAP为主要成分的生防制剂提供指导。     

玉米耐盐分子机制研究进展

玉米是我国总产量最大的农作物,它对盐胁迫耐受性较差.玉米主产区日益加剧的土壤盐渍化已成为导致玉米产量和品质下降的主要环境胁迫之一.因此,研究玉米的耐盐机制,改良玉米的耐盐性对保障我国乃至世界玉米生产的可持续发展有重要意义.土壤中盐浓度过高会导致玉米根系外围土壤溶液的水势降低,引起渗透胁迫,而盐离子(Na+、Cl-等)的...     

马铃薯晚疫病生防木霉菌的筛选及鉴定

采用马铃薯活体筛选法从268株木霉菌中筛选获得两株对致病疫霉有较强抑菌活性的木霉菌株R-5和T-15。这两株木霉菌代谢液可抑制病原菌生长及孢子囊萌发。温室防病试验发现,接种两株木霉菌可以减轻晚疫病的发生。田间试验进一步证明,两株木霉菌对晚疫病具有良好的田间防治效果,防效分别达到了72.4%和70.0%。经分子生物学方法鉴定,两株木霉菌分别为拟康氏木霉和棘孢木霉。实验构建的以活体筛选为基础的生防木霉菌筛选方法是一种可行高效的生防木霉菌筛选方式。     

盐胁迫下植物细胞离子稳态重建机制

土壤盐渍化是困扰世界粮食产量的一大难题。在盐胁迫环境中,植物获得耐盐能力的一个重要策略是建立新的离子稳态(ionic homeostasis)。盐胁迫下植物细胞离子稳态依赖于膜转运蛋白(泵、载体和离子通道)。利用蛋白质的生化功能分析和突变体功能互补等方法,目前已克隆和鉴定了许多参与离子稳态重建的膜转运蛋白。综述了盐胁迫下植物细胞离子稳态重建的最新研究进展。     

有效霉素A对棘孢木霉的影响及协同防治玉米纹枯病作用

【背景】玉米纹枯病逐渐发展为制约我国玉米持续增产的主要病害,有效霉素A与生防菌木霉均对纹枯病菌具有抗性,但二者各有优缺点。【目的】研究有效霉素A与木霉菌协同作用的可行性,提高对玉米纹枯病的防治水平,实现抗生素类化学农药的减量使用,提高生态环境安全性。【方法】测定有效霉素A处理后的棘孢木霉GDSF1009的细胞壁降解酶及防御反应相关酶活性,烟草叶片验证活性氧及过敏性反应。利用转录组及气相色谱-飞行时间质谱联用分析有效霉素A对GDSF1009基因表达差异及生长代谢的影响,并进行叶片及平皿协同抑菌实验。【结果】有效霉素A对棘孢木霉GDSF1009的几丁质酶、纤维素酶及木聚糖酶的活性无影响,有效霉素A未进入到棘孢木霉菌细胞内。在转录组水平,有效霉素A处理木霉24h和48 h后,与对照相比,差异基因所占比例分别为8.932%和6.779%,有效霉素A对GDSF1009相关的糖类及脂类代谢没有显著影响,仅对氨基酸代谢相关的基因有一些影响,这些基因并不会造成氨基酸代谢的大量变化。同时验证了在二者联合使用时,可以显著地提高玉米纹枯病的防治效果。【结论】有效霉素A对棘孢木霉菌的初生代谢系统是比较安全的,二者协同作用的抗病效果显著高于单因子作用。     

Biochemical and metabolic profiles of Trichoderma strains isolated from common bean crops in the Brazilian Cerrado, and potential antagonism against Sclerotinia sclerotiorum

Some species of Trichoderma have successfully been used in the commercial biological control of fungal pathogens, e.g., Sclerotinia sclerotiorum, an economically important pathogen of common beans (Phaseolus vulgaris L.). The objectives of the present study were (1) to provide molecular characterization of Trichoderma strains isolated from the Brazilian Cerrado; (2) to assess the metabolic profile of each strain by means of Biolog FF Microplates; and (3) to evaluate the ability of each strain to antagonize S. sclerotiorum via the production of cell wall-degrading enzymes (CWDEs), volatile antibiotics, and dual-culture tests. Among 21 isolates, we identified 42.86% as Trichoderma asperellum, 33.33% as Trichoderma harzianum, 14.29% as Trichoderma tomentosum, 4.76% as Trichoderma koningiopsis, and 4.76% as Trichoderma erinaceum. Trichoderma asperellum showed the highest CWDE activity. However, no species secreted a specific group of CWDEs. Trichoderma asperellum 364/01, T. asperellum 483/02, and T. asperellum 356/02 exhibited high and medium specific activities for key enzymes in the mycoparasitic process, but a low capacity for antagonism. We observed no significant correlation between CWDE and antagonism, or between metabolic profile and antagonism. The diversity of Trichoderma species, and in particular of T. harzianum, was clearly reflected in their metabolic profiles. Our findings indicate that the selection of Trichoderma candidates for biological control should be based primarily on the environmental fitness of competitive isolates and the target pathogen.     

一株真菌拮抗细菌Z21的筛选与鉴定及其发酵条件优化

【背景】芽孢杆菌属的许多细菌具有抗逆性强、安全等特点,一直以来都是开发新型活性物质的研究热点。【目的】筛选对食品腐败真菌有抑制作用的细菌,将其开发为天然食品防腐剂。【方法】采用平板分离法、平板对峙法、抑制菌丝生长速率法从空气、竹子内生细菌中筛选真菌拮抗菌,通过形态、生理生化特征及16S rRNA基因序列分析等方法对其进行鉴定,利用正交试验确定其最优生长条件。【结果】筛选到一株对6种常见霉菌均有较强抑制作用的细菌Z21。Z21与甲基营养型芽孢杆菌(Bacillusmethylotrophicus strain CBMB205~T)的相似性最高,且形态特征和生理生化特征与CBMB205~T菌株基本相符。Z21最佳发酵培养基配方和培养条件分别为:葡萄糖20.0 g/L、NaNO_3 20.0 g/L、MgSO_4 3.0 g/L,培养温度为32°C,培养时间为48 h。【结论】Z21为甲基营养型芽孢杆菌(Bacillus methylotrophicus),对黑曲霉、康氏木霉、绿色木霉、少根根霉、易脆毛霉、赭绿青霉的生长具有较强的抑制作用且抑菌效果稳定,为广谱真菌拮抗菌。     

洞庭湖湿地木霉多样性及生防活性

【目的】了解湖南省洞庭湖湿地木霉种类及分布,丰富我国的木霉种质资源,为功能菌株筛选应用奠定基础。【方法】利用ITS序列比对分析结合形态学特征对分离到的木霉菌株进行种类鉴定,构建系统发育进化树分析其亲缘关系。通过菌丝生长速率法测定菌株的平板抑菌能力,根据水解带大小检测菌株的水解酶活性,利用灰色关联度分析筛选综合生防效果较好的木霉菌株。【结果】从52个土样和18个水样中分离得到114株木霉菌株,经鉴定分属16个木霉种类:哈茨木霉、绿木霉、刺孢木霉、土星孢木霉、钩状木霉、拟康宁木霉、短密木霉、深绿木霉、猥木霉、毛细木霉(中国新记录种)、长枝木霉、卵孢木霉、侧耳木霉、加纳木霉、厚木霉及一个疑似新种;哈茨木霉为洞庭湖湿地中的优势种,占总菌株数量的19.30%;16种木霉在系统发育树中归于7个进化支:Harzianum Clade、Virens Clade、Longibrachiatum Clade、Lutea Clade、Viride Clade、Hamatum Clade、Unknown Clade。灰色关联度分析表明,菌株TW21990、QT22040和QT22094的灰色关联度较高,分别为0.849 5、0.798 6和0.732 6,综合生防性状较好。【结论】洞庭湖湿地木霉具有种类多样性和分布多样性,发现了一个中国新记录种毛细木霉和一个疑似新种,哈茨木霉TW21990、长枝木霉QT22040和卵孢木霉QT22094是潜在的优良生防菌株。     

Proline betaine accumulation and metabolism in alfalfa plants under sodium chloride stress. Exploring its compartmentalization in nodules

The osmoprotectant Pro betaine is the main betaine identified in alfalfa (Medicago sativa). We have investigated the long-term responses of nodulated alfalfa plants to salt stress, with a particular interest for Pro betaine accumulation, compartmentalization, and metabolism. Exposure of 3-week-old nodulated alfalfa plants to 0.2 m NaCl for 4 weeks was followed by a 10-, 4-, and 8-fold increase in Pro betaine in shoots, roots, and nodules, respectively. Isotope-labeling studies in alfalfa shoots indicate that [14C]Pro betaine was synthesized from l-[14C]Pro. [14C]Pro betaine was efficiently catabolized through sequential demethylations via N-methylPro and Pro. Salt stress had a minor effect on Pro betaine biosynthesis, whereas it strongly reduced Pro betaine turnover. Analysis of Pro betaine and Pro compartmentalization within nodules revealed that 4 weeks of salinization of the host plants induced a strong increase in cytosol and bacteroids. The estimated Pro betaine and Pro concentrations in salt-stressed bacteroids reached 7.4 and 11.8 mm, respectively, compared to only 0.8 mm in control bacteroids. Na+ content in nodule compartments was also enhanced under salinization, leading to a concentration of 14.7 mm in bacteroids. [14C]Pro betaine and [14C]Pro were taken up by purified symbiosomes and free bacteroids. There was no indication of saturable carrier(s), and the rate of uptake was moderately enhanced by salinization. Ultrastructural analysis showed a large peribacteroid space in salt-stressed nodules, suggesting an increased turgor pressure inside the symbiosomes, which might partially be due to an elevated concentration in Pro, Pro betaine, and Na+ in this compartment.     

Antioxidant enzymatic activity is linked to waterlogging stress tolerance in citrus

Soil flooding constitutes a seasonal factor that negatively affects plant performance and crop yields. In this work, the relationship between oxidative damage and flooding sensitivity was addressed in three citrus genotypes with different abilities to tolerate waterlogging. We examined leaf visible damage, oxidative damage in terms of malondialdehyde (MDA) concentration, leaf proline concentration, leaf and root ascorbate and glutathione contents and the antioxidant enzyme activities superoxide dismutase (EC 1.15.1.1), ascorbate peroxidase (EC 1.11.1.11), catalase (EC 1.11.1.6) and glutathione reductase (EC 1.8.1.7). No differences in the extent of oxidative damage relative to controls were found among genotypes. However, a different ability to delay the apparition of oxidative damage was associated to a higher tolerance to waterlogging. This ability was linked to an enhanced activated oxygen species' scavenging capacity in terms of an increased antioxidant enzyme activity and higher content in polar antioxidant compounds. Therefore, the existence of a direct relationship between stress sensitivity and the early accumulation of MDA is proposed. In addition, data indicate that the protective role of proline has to be considered minimal as its accumulation was inversely correlated with tolerance to the stress. The positive antioxidant response in Carrizo citrange ( Poncirus trifoliata L. Raf. ×  Citrus sinensis L. Osb.) and Citrumelo CPB 4475 ( Poncirus trifoliata L. Raf. ×  Citrus paradisi L. Macf.) might be responsible for a higher tolerance to flooding stress, whereas in Cleopatra mandarin ( Citrus reshni Hort. Ex Tan.), the early accumulation of MDA seems to be associated to an impaired ability for H₂O₂ scavenging.     

Adsorption of Trichoderma reesei cellulase on cellulose: experimental data and their analysis by different equations

The adsorption of cellulase from Trichoderma reesei MCG 77 on Avicel was measured at varying cellulase (2-8 g/L) and Avicel (10-200 g/L) concentrations at pH 4.8 and 50 degrees C. Different mathematical equations were derived for the evaluation of the experimental data. The fraction of cellulase protein that can maximally be adsorbed is 0.96, and 1 g Avicel can bind maximally 0.092 g cellulase protein. The Michaelis constant for the adsorption equilibrium [cellulase] + [Avicel] right harpoon over left harpoon [cellulase Avicel] complex is between 2.0 and 2.3 . 10(-5) mol/L. This value is based on the assumption that cellulase has an average molecular weight of 48.000. The apparent molecular weight of Avicel, i.e., that amount in grams that can bind 1 mol cellulase, is 520,000. Under maximum binding the enzyme covers on Avicel a surface of 42 m(2)/g, and the occupied volume is 0.186 cm(3)/g Avicel.     

Hormonal changes during salinity-induced leaf senescence in tomato (Solanum lycopersicum L.)

Leaf senescence is one of the most limiting factors to plant productivity under salinity. Both the accumulation of specific toxic ions (e.g. Na+) and changes in leaf hormone relations are involved in the regulation of this process. Tomato plants (Solanum lycopersicum L. cv Moneymaker) were cultivated for 3 weeks under high salinity (100 mM NaCl) and leaf senescence-related parameters were studied during leaf development in relation to Na+ and K+ contents and changes in abscisic acid (ABA), cytokinins, the ethylene precursor 1-aminocyclopropane-1-carboxylic acid (ACC), and the auxin indole-3-acetic acid (IAA). Na+ accumulated to a similar extent in both leaves 4 and 5 (numbering from the base of the plant) and more quickly during the third week, while concurrently K+ contents sharply decreased. However, photosystem II efficiency, measured as the F(v)/F(m) ratio, decreased from the second week of salinization in leaf 4 but only at the end of the third week in the younger leaf 5. In the prematurely senescent leaf 4, ABA content increased linearly while IAA strongly decreased with salinization time. Although zeatin (Z) levels were scarcely affected by salinity, zeatin-riboside (ZR) and the total cytokinin content (Z+ZR) progressively decreased by 50% from the imposition of the stress. ACC was the only hormonal compound that increased in leaf tissue coincident with the onset of oxidative damage and the decline in chlorophyll fluorescence, and prior to massive Na+ accumulation. Indeed, (Z+ZR) and ACC contents and their ratio (Z+ZR/ACC) were the hormonal parameters best correlated with the onset and progression of leaf senescence. The influence of different hormonal changes on salt-induced leaf senescence is discussed.     

氧化应激在高原重体力劳动过程中急性高原反应发生中的作用

目的：观察氧化应激在高原重体力劳动过程中急性高原反应（AHAR）发生中的作用。方法：由低海拔（1500m）快速进入高原（3700m）并从事重体力劳动的男性官兵96名,年龄18～35岁。根据AHAR症状评分,分为重度AHAR组（A组,n=24）、轻中度AHAR组（B组,n=47）和无AHAR组（C组,n=25）,在该高度逗留50d后下撤前及返回低海拔（1500m）后12h、15d分别测定血清8．异前列腺素F2a（8-iso-PGF2a）、超氧化物歧化酶（SOD）、丙二醛（MDA）,并与低海拔（1500m）50名健康官兵（D组）比较。结果：A组血清8-iso-PGF2a、MDA[分别为（9．53±0．47）μg／L、（8．91±0．39）μmol／L]水平显著高于B组[分别为（8．34±O．42）μg／L、（7．31±0．32）μmol／L]、C组[分别为（7．02±0．48）μg／L、（6．41±0．23）μmol／L和D组[分别为（5．13±0．56）μg／L、（5．48±0．33）μmol／L]（均P〈0．01）,SOD（52．08±3．44）μ／mL水平显著低于B组（62．27±2．54）μ/mL、C组（71．99±3．35）μ/mL和D组（80．78±3．44）μ/mL,（均P〈0．01）,B组与c组之间和C组与D组之间亦有显著性差异（均P〈0．01）。海拔3700mAHAR总计分与血清8-iso-PGF2α、ⅣⅡ）A呈显著正相关（均P〈0．01）,与血清SOD显著负相关（P〈0．01）;8-iso-PGF2α、MDA与SOD显著负相关（均P〈0．01）。海拔3700m50d,血清8-iso-PGF2α、MDA水平显著高于,SOD水平显著低于海拔1500m12h、15d和D组（均P〈0．01）,海拔1500m12h与15d之间有显著性差异（均P〈0．01）,海拔1500m 15d与D组之间无显著性差异。结论：人体在高原低氧并重体力时氧化应激和氧化．抗氧化失衡与AHAR的发病和程度有密切关系,氧化应激和氧化．抗氧化失衡越严重,AHAR越重。返回低海拔后12h有显著改善,15d恢复到正常水平。