Application of methyl parathion hydrolase (MPH) as a labeling enzyme

Methyl parathion hydrolase (MPH) is an enzyme that catalyzes the degradation of methyl parathion, generating a yellow product with specific absorption at 405 nm. The application of MPH as a new labeling enzyme was illustrated in this study. The key advantages of using MPH as a labeling enzyme are as follows: (1) unlike alkaline phosphatase (AP), horseradish peroxidase (HRP), and glucose oxidase (GOD), MPH is rarely found in animal cells, and it therefore produces less background noise; (2) its active form in solution is the monomer, with a molecular weight of 37 kDa; (3) its turnover number is 114.70 +/- 13.19 s(-1), which is sufficiently high to yield a significant signal for sensitive detection; and (4) its 3D structure is known and its C-terminal that is exposed to the surface can be easily subjected to the construction of genetic engineering monocloning antibody-enzyme fusion for enzyme-linked immunosorbent assay (ELISA). To demonstrate its utility, MPH was ligated to an single-chain variable fragment (scFv), known as A1E, against a white spot syndrome virus (WSSV) with the insertion of a [-(Gly-Ser)(5)-] linker peptide. The resulting fusion protein MPH-A1E possessed both the binding specificity of the scFv segment and the catalytic activity of the MPH segment. When MPH-A1E was used as an ELISA reagent, 25 ng purified WSSV was detected; this was similar to the detection sensitivity obtained using A1E scFv and the HRP/Anti-E Tag Conjugate protocol. The fusion protein also recognized the WSSV in 1 mu L hemolymph from an infected shrimp and differentiated it from a healthy shrimp.

Influence of selected Indian immunostimulant herbs against white spot syndrome virus (WSSV) infection in black tiger shrimp, Penaeus monodon with reference to haematological, biochemical and immunological changes

Immunostimulants are the substances, which enhance the non-specific defence mechanism and provide resistance against the invading pathogenic micro-organism. In order to increase the immunity of shrimps against the WSSV, the methanolic extracts of five different herbal medicinal plants like Cyanodon dactylon, Aegle marmelos, Tinospora cordifolia, Picrorhiza kurooa and Eclipta alba were selected and mixed thoroughly in equal proportion. The mixed extract was supplemented with various concentrations viz. 100 (A), 200 (B), 400 (C), and 800 (D) mg kg(-1) through artificial diets individually. The prepared diets (A-D) were fed individually to WSSV free healthy shrimp Penaeus monodon with an average weight of 8.0 +/- 0.5 g for 25 days. Control diet (E), devoid of herbal extract was also fed to shrimps simultaneously. After 25 days of feeding experiment, the shrimps were challenged with WSSV, which were isolated and propagated from the infected crustaceans. The shrimps succumbed to death within 7 days when fed on no herbal immunostimulant diet (E). Among the different concentrations of herbal immunostimulant supplemented diets, the shrimps fed on diet D (800 mg kg(-1)) significantly (P < 0.0001) had more survival (74%) and reduction in the viral load. Also the better performance of haematological, biochemical and immunological parameters was found in the immunostimulant incorporated diets fed shrimps. The present work revealed that the application of herbal immunostimulants will be effective against shrimp viral pathogenesis and they can be recommended for shrimp culture. (c) 2006 Published by Elsevier Ltd.

Phylogeny of the cuttlerishes (Mollusca : Cephalopoda) based on mitochondrial COI and 16S rRNA gene sequence data

To clarify cuttlefish phylogeny, mitochondrial cytochrome c oxidase subunit 1 (COI) gene and partial 16S rRNA gene are sequenced for 13 cephalopod species. Phylogenetic trees are constructed, with the neighbor-joining method. Coleoids are divided into two main lineages, Decabrachia and Octobrachia. The monophyly of the order Sepioidea, which includes the families Sepiidae, Sepiolidae and Idiosepiidae, is not supported. From the two families of Sepioidea examined, the Sepiolidae are polyphyletic and are excluded from the order. On the basis of 16S rRNA and amino acid of COI gene sequences data, the two genera (Sepiella and Sepia) from the Sepiidae can be distinguished, but do not have a visible boundary using COI gene sequences. The reason is explained. This suggests that the 16S rDNA of cephalopods is a precious tool to analyze taxonomic relationships at the genus level, and COI gene is fitter at a higher taxonomic level (i.e., family).

A preliminary phylogenetic analysis of Metapenaeopsis (Decapoda : Penaeidae) based on mitochondrial dna sequences of selected species from the Indo-West Pacific

Phylogenetic relationships within Metapenaeopsis remain largely unknown. The modern revision of the genus suggests that the shape of the petasma, followed by the presence of a stidulating organ, are the most important distinguishing taxonomic features. In the present study, phylogenetic relationships were studied among seven Metapenaeopsis species from the Indo-West Pacific based on partial sequences of mitochondrial 16S rRNA and cytochrome c oxidase I (COI) genes. Mean sequence divergence was 6.4% for 16S and 15.8% for COI. A strikingly large nucleotide distance (10.0% for 16S and 16.9% for COI) was recorded between M. commensalis, the only Indo-West Pacific species with a one-valved petasma, and the other species with a two-valved petasma. Phylogenetic analyses using neighbor-joining, maximum parsimony, and maximum likelihood generated mostly identical tree topologies in which M. commensalis is distantly related to the other species. Two clades were resolved for the remaining species, one with and the other without a stridulating organ, supporting the main groupings of the recent taxonomic revision. Results of the present study also indicate that the deep-water forms represent a relatively recent radiation in Metapenaeopsis.

Low potential detection of glutamate based on the electrocatalytic oxidation of NADH at thionine/single-walled carbon nanotubes composite modified electrode

A glutamate biosensor based on the electrocatalytic oxidation of reduced nicotinamide adenine dinucleotide (NADH), which was generated by the enzymatic reaction, was developed via employing a single-walled carbon nanotubes/thionine (Th-SWNTs) nanocomposite as a mediator and an enzyme immobilization matrix. The biosensor, which was fabricated by immobilizing glutamate dehydrogenase (GIDH) on the surface of Th-SWNTs, exhibited a rapid response (ca. 5 s), a low detection limit (0.1 mu M), a wide and useful linear range (0.5-400 mu M), high sensitivity (137.3 +/- 15.7) mu A mM(-1) cm(-2), higher biological affinity, as well as good stability and repeatability. In addition, the common interfering species, such as ascorbic acid, uric acid, and 4-acetamidophenol, did not cause any interference due to the use of a low operating potential (190 mV vs. NHE). The biosensor can be used to quantify the concentration of glutamate in the physiological level. The Th-SWNTs system represents a simple and effective approach to the integration of dehydrogenase and electrodes, which can provide analytical access to a large group of enzymes for wide range of bioelectrochemical applications including biosensors and biofuel cells.

黄土丘陵区撂荒对土壤酶活性的影响

为了解侵蚀环境下植被恢复对土壤酶活性的影响,以典型侵蚀环境黄土丘陵区纸坊沟流域生态恢复1至50年撂荒地长期定位试验点为研究对象,选取坡耕地为对照,分析了植被恢复过程中土壤脲酶、磷酸酶、蔗糖酶、淀粉酶、纤维素酶、过氧化氢酶、多酚氧化酶及理化性质的演变特征。结果表明,土壤酶活性前期变化波动较大,后期(20-30a)年变化趋于稳定;尿酶、碱性磷酸酶、蔗糖酶、过氧化氢酶和多酚氧化酶与其他因子相关性相对较强,可以作为评价土壤质量的生物学指标;尿酶、淀粉酶、碱性磷酸酶、蔗糖酶、过氧化氢酶和纤维素酶活性随恢复年限而增加,多酚氧化酶则减少;土壤酶指数可以作为评价土壤质量的方法。

固斑蝰蛇毒C-型凝集素样蛋白Dabocetin及L-氨基酸氧化酶DRS-LAO的研究

本论文研究了从圆斑蛙蛇泰国亚种（Daboia russellii siamensis）蛇毒中纯化的C一型凝集素样蛋白Dabocetin和L一氨基酸氧化酶DRS一AO的理化性质、生物学活性和分子克隆。Dabocetin是分子量约为28扔。的异二聚体蛋白，它由分子量约为15.0kDa和14．5kDa的两个同源亚基以和p共价结合形成。N－末端氨基酸序列比较显示，Dabocetin与目前已知的蛇毒c一型凝集素样蛋白有很高的同源性。即使在终浓度达50．0。叫而时，Dabocetin也不能直接诱导血小板聚集。此外，在终浓度为40.00μg/ml时，Dabocetin几乎不能抑制由AdP，TMVA和stejnulxin诱导的血小板聚集。但是，Dabocetin呈剂量依赖地抑制瑞斯托霉素诱导的血小板凝集，其半数抑制率ICS。值为10.80ug/ml。流式细胞仪分析表明，Dabocetin显著抑制单克隆抗体522与GPIba的结合，提示Dabocetin很可能是一个GPIb结合蛋白。从圆斑蛙蛇的毒腺中克隆到了7个编码不同蛇毒C一型凝集素样蛋白亚基的七DNA（命名为DRs一1至DRs一7）。其中，DRsLS编码Dabocetin的a亚基，DRS一6编码Dabocetin的p亚基。DRs一1和DRS一2很可能是圆斑蛙蛇毒腺中表达的X因子激活剂的两条轻链LCZ和LCI的山NA。DRS一3，DRS毛4和DRSL7可能是圆斑蛙蛇毒腺中表达的C一型凝集素样蛋白p亚基的。NA。DRsLAO是一个新的L一氨基酸氧化酶，比活力为1．98U加噶。十二烷基磺酸钠一聚丙烯酞氨凝胶电泳（SDs-PAGE）分析显示，该酶在还原和非还原条件下均呈现一条蛋白带，表观分子量约为58kDa。N-末端氨基酸序列比较显示，DRS一AO与目前已知的蛇毒L一氨基酸氧化酶有很高的同源性。该酶的最适底物为L一亮氨酸，最适pH为8．8。DRs一Ao呈剂量依赖地抑制扔P和仆IvA诱导的血小板聚集，其半数抑制率ICS。值分别为32.8μg/ml和32.3μg/ml。DRS-LAO对金黄色葡萄球菌（灯Cc25923）和耐甲氧西林金黄色葡萄球菌有较强的抗菌作用。DRs一AO对金黄色葡萄球菌必Tcc25923）的最低抑菌浓度卿C）和最低杀菌浓度耐甲氧西林金黄色葡萄球菌的孤CS。和呱Cg。值分别为18．。林留时和36.0μg/ml；DRSLAo对耐甲氧西林金黄色葡萄球菌的MBC50和MBCg。值分别为36.0μg/ml和72.0μg/ml。通过对DRS一AO的分子克隆，得到了编码DRS-AO的部分cDNA序列。

CO2升高对土壤微生物群落影响的模拟研究

大气CO2浓度升高可以通过植物间接影响土壤生态系统。土壤生态系统的结构和功能改变将影响有机质矿化和营养物质循环，进而可能对CO2浓度升高产生正反馈或负反馈。微生物是土壤生态系统的主体，在对CO2浓度升高的反馈中起着至关重要的作用。本研究以开顶箱系统为平台，采用微生物分子生态学技术和现代酶学技术，通过对长期接受500 ppm CO2的红松幼树、长白赤松幼树和蒙古栎幼树非根际土壤连续两个生长季的测定，系统研究了高浓度CO2对温带森林土壤微生物群落的生物量和微生物活性的影响，检测了土壤微生物群落的结构和功能以及土壤化学性质变化，主要结论如下： （1）高浓度CO2处理提高了土壤有机碳含量。与对照组相比较，红松幼树土壤有机碳含量提高9.4%；长白赤松幼树土壤提高0.6%；蒙古栎幼树土壤提高1.3%。 （2）高浓度CO2处理使土壤磷酸酶（phosphatase）、几丁质酶（1,4-β-acetylglucosaminidase, 1,4-β-NAG）和多酚氧化酶（phenol oxidase）活性发生了显著变化，高浓度CO2使红松土壤 1,4-β-NAG活性提高7-25%，长白松土壤1,4-β-NAG平均活性降低14%，蒙古栎土壤1,4-β-NAG平均活性提高31%。 同时研究还发现，过氧化物酶（peroxidase）和多酚氧化酶(phenol oxidase)活性与微生物量碳和微生物量氮呈显著的正相关。相关分析还显示，土壤湿度与1,4-α-葡萄糖苷酶（1,4-α-glucosidase）活性、 微生物生物量碳和微生物生物量氮呈显著的正相关。 高浓度CO2在不同程度上改变了土壤转化酶活性和脱氢酶活性。高浓度CO2显著提高了红松和长白赤松土壤硝化酶活性；而显著降低反硝化酶活性。 （3）研究发现三种树土壤的真菌和细菌群落存在着季节性演替，并且高浓度CO2熏蒸处理使真菌群落结构发生了显著的变化，表现为一些种群优势度下降，另一些升高。虽然，细菌群落没有如真菌群落变化的明显，但研究中也发现高浓度CO2的确使个别细菌种群的优势度发生了显著改变。 亲缘关系与Calocybe carnea，Magmatodrilus obscurus密切的真菌是红松土壤优势种群，与Humicola fuscoatra关系相近的是长白松土壤的优势种群，并且此三种真菌的季节性变化不显著。研究发现高浓度CO2使红松土壤中亲缘关系与Pachyella clypeata，Cochlonema euryblastum，Lepiota cristata，Eimeriidae sp.， Trichoderma sp.相近的种群的丰富度显著提高，使蒙古栎土壤中亲缘关系与Serendipita vermifera，Calocybe carnea种群丰富度显著下降，使蒙古栎土壤中与Candida sp.，Magmatodrilus obscurus和Pachyella clypeata亲缘关系密切种群的丰富度显著提高。 （4）三种幼树叶的原位分解培养429天结果显示，红松和长白松凋落物的β-葡萄糖苷酶（1,4-β-glucosidase）和木糖苷酶（1,4-β-xylosidase）活性随着分解而逐渐增加，而这两种酶在蒙古栎凋落物分解过程中保持相对恒定；高浓度CO2显著影响叶凋落物分解磷酸酶（phosphatase），纤维二糖酶（cellobiohydrolase）, 几丁质酶(1,4-β-NAG)，多酚氧化酶（phenol oxidase）和过氧化物酶（peroxidase）的活性。研究发现，凋落物的生物化学性质变化能引起分解的微生物群落发生变化，进而引起分泌的胞外酶活性变化，科学印证了大气CO2浓度升高“通过影响凋落物质量进而影响分解叶凋落物的微生物群落的结构和功能”的猜测。 不同凋落物之间酶活性差异显著，真菌和细菌群落结构也显著不同。序列与Hyphodiscus hymeniophilus亲缘关系密切的真菌和亲缘关系与Verrucomicrobia bacterium密切的细菌是长白松凋落分解的最优势种群，序列与Lophium mytilinum亲缘关系密切的真菌是红松凋落分解的最优势种群。 另外，研究还发现，高浓度CO2使参与分解红松凋落物Beta proteobacterium OS-15A亲缘关系相近的细菌种群和与Azospirillum amazonense亲缘关系相近的种群丰富度显著降低；使与Luteibactor rhizovicina亲缘关系相近的种群和与Luteibactor rhizovicina亲缘关系相近的种群显著提高。高浓度CO2使定殖于长白松凋落物上Hyphodiscus hymeniophilus亲缘关系相近的种群和与Bionectria pityrodes亲缘关系相近的种群显著提高，而使与Neofabraea malicorticis亲缘关系相近的种群和与Hyphodiscus hymeniophilus亲缘关系相近的种群显著下降。

花椒的化感作用对凋落物分解的影响及机理研究

花椒（Zanthoxylum bungeanum）是川西干旱河谷地区重要的经济作物，化感作用是花椒连作障碍的重要原因之一。花椒凋落物分解是影响花椒林地土壤肥力及花椒产量的重要因素，因此系统研究花椒化感作用是否对花椒凋落物的分解产生影响可以为解决花椒连作障碍导致的产量下降等问题提供科学的理论依据。本文通过室内模拟实验研究了花椒凋落物的四个分解动态以及分解后凋落物浸提液对花椒林地土壤性质的影响；通过野外盆栽实验研究了花椒凋落物浸提液对花椒幼苗的生长、花椒凋落物的质量及土壤性质的影响。最终从生理生化角度揭示了花椒的化感作用对凋落物分解的影响机理，为深入解决花椒连作障碍问题、对花椒凋落物采取有效的人工措施提供了科学的理论依据。主要的研究结果如下： 1. 室内分解实验证明，花椒凋落物在分解的60 d 内分解速率呈现由大到小的变化趋势，并且凋落物分解呈现明显的毒性动态。凋落物在分解的10 d、30 d 时，分解速率较大，30 d 以后分解速率显著降低。凋落物分解的10 d 左右酚酸释放量最大，此时凋落物的毒性最大，凋落物分解到10 d 以后，酚酸释放量逐渐减少，凋落物的毒性也逐渐减小。 2. 四个分解动态的花椒凋落物浸提液对土壤化学性质产生了显著的影响。花椒凋落物在分解的60 d 内，其浸提液使土壤pH值均显著的增加。分解0 d 的凋落物浸提液显著的降低了土壤铵态氮的含量，抑制了纤维素分解菌的生长；分解60 d 的凋落物浸提液显著的降低了土壤酚酸含量，增加了土壤有效磷的含量；分解30 d 和60 d 的凋落物浸提液均显著的促进了好气性纤维素分解菌的生长。这说明花椒凋落物在分解过程中呈现出明显的毒性动态：凋落物分解的初期毒性作用较大，随着分解的继续进行特别是在分解的30 d 以后，其毒性作用慢慢降低。 3. 花椒凋落物浸提液对花椒幼苗表现出明显的化感作用。不同浓度的浸提液对花椒幼苗地上及地下生物量、叶面积均产生了显著的抑制作用，并且随着浸提液浓度的升高抑制作用加强。凋落物浸提液对叶片厚度的影响较小，只有Y1对叶片厚度的生长抑制作用显著。 4. 花椒的化感作用改变了凋落物的质量，并对凋落物分解产生了显著的影响。对花椒幼苗用不同浓度的凋落物浸提液进行处理，Y1使凋落物有机碳含量、木质素含量、C/N、木质素/氮显著降低，纤维素含量显著升高；Y3使凋落物有机碳含量、木质素含量、C/N、木质素/氮显著升高。花椒凋落物质量的改变显著的影响了凋落物的分解，凋落物的分解速率大小依次为：Y1（10.15 a-1）> Y2（8.71 a-1）> CK（6.41 a-1）> Y3（5.08 a-1）。 5. 花椒的化感作用改变了土壤性质，并对凋落物分解产生了显著的影响。对花椒幼苗用不同浓度的凋落物浸提液处理的同时，也改变了土壤性质。不同浓度的凋落物浸提液显著的升高了土壤pH值、有机碳含量。各种浓度的凋落物浸提液对土壤多酚氧化酶的活性均起到了显著的促进作用。凋落物浸提液Y1对土壤纤维素分解酶的活性、细菌和真菌的生长也具有显著的促进作用。土壤性质的改变显著的影响了凋落物的分解，凋落物的分解速率大小依次为：Y1（10.30 a-1）>Y2（9.60 a-1）>CK（6.41 a-1）>Y3（6.29 a-1）。 6. 不论是凋落物质量发生改变还是土壤性质发生改变，在凋落物分解的整个过程中，C元素始终处于单调净释放的状态，并且C释放量与分解速率成显著的正相关，即凋落物分解越快，凋落物C释放量越大。凋落物分解过程中，均出现了酚酸大量释放的情况，并与凋落物分解速率成显著正相关。凋落物分解后的木质素含量、木质素/氮均增加，并且随着浸提液浓度的升高，凋落物木质素含量、木质素/氮升高。 Zanthoxylum bungeanum is an important economic crop in dry valley of the Minjiang river (Sichuan, Southwest China), but allelopathy is one of the important reasons for its continuous cropping. Zanthoxylum bungeanum litter decomposition affects Zanthoxylum bungeanum soil fertility and its output. So systemically investigate if allelopathy affects litter decomposition could provide the scientific methods to solve the problem of output fall caused by the continuous cropping. In this paper, the releasing dynamics of phenolic acid during Zanthoxylum bungeanum litter decomposition (0, 10, 30 and 60 days) and the effects of its aqueous extract on soil chemical properties were investigated via the laboratory study. Effects of Zanthoxylum bungeanum litter aqueous extract on the growth of young Zanthoxylum bungeanum seedlings, litter qualities and the soil qualities were investigated via the field study. Finally, we open out the action manner of Zanthoxylum bungeanum allelopathic effect on the litter decomposition, and provide the theoretical basis to solve the Zanthoxylum bungeanum continuous cropping. The main results showed that: 1. The laboratory litter decomposition experiment showed a trend of decomposition rate from large to small and an occurrence of phytotoxicity with clear dynamic patterns during Zanthoxylum bungeanum litter decomposition. The litter decomposition rate was larger at the tenth and 30th day during 60-day litter decomposition and gradually decreased after 30 days of litter decompostion. The releasing quantity of the litter phenolic acid was the highest at the tenth day, and here, the litter toxicity was the biggest. The releasing quantity of the litter phenolic acid gradually decreased after 10 days of litter decomposition, so the phytotoxicity of litter was gradually decreased with the litter decomposition. 2. The Zanthoxylum bungeanum litter aqueous extract after four decomposition stages had significantly effect on the soil chemical qualities. The pH value in soil was significantly increased in litter aqueous extract of four decomposition stages. The NH+4-N concentration was significantly decreased in soil amended with litter aqueous extract of 10-day decomposition which inhibited the growth of Aerobic cellulose-decomposer. The growth of soil Aerobic cellulose-decomposer was promoted by the litter aqueous extract of 30-day decomposition. Available phosphorus concentration was significantly increased and phenolic acid content was significantly decreased in soil amended with litter aqueous extract of 60-day decomposition which promoted the growth of Aerobic cellulose-decomposer. The study results showed an occurrence of phytotoxicity with clear dynamic patterns during Zanthoxylum bungeanum litter decomposition. The phytotoxicity of litter was the largest at the initial stage, but the phytotoxicity gradually decreased with the litter decomposition, especially after 30 days of decomposition. 3. The field study indicated that the Zanthoxylum bungeanum litter aqueous extract had significant allelopathic effects on the growth of young seedlings.Different concentration aqueous extract had signinficant inhibiting effects on biomass and leaf area of young seedlings. The inhibiting effect on the biomass strengthened with the litter aqueous extract concentration augment. Litter aqueous extracts had less effect on the leaf thickness, and only Y1 had significant inhibiting effect on the leaf thickness. 4. The Zanthoxylum bungeanum allelopathy had significant effect on the litter qualities and the litter decomposition. Treating the young Zanthoxylum bungeanum seedlings with different concentration of litter aqueous extracts, the leaf litter organic C, lignin, C/N and lignin/N all decreased and the cellulose content increased under Y1 treatment. The leaf litter organic C, lignin, C/N and lignin/N all increased under Y3 treatment. So the litter decomposition was significant affectded by the litter qualities, and the litter decomposition rate was Y1(10.15 a-1)> Y2(8.71 a-1) > CK(6.41 a-1) > Y3(5.08 a-1). 5. The Zanthoxylum bungeanum allelopathy had significant effect on the soil qualities and the litter decomposition. Treating the young Zanthoxylum bungeanum seedlings with different concentration of litter aqueous extracts, also changed the soil qualities. Different concentration of litter aqueous extracts had significant effects on the soil pH and organic C content. Every concentration of litter aqueous extracts accelerated the soil Polyphenol Oxidase activity and Y1 accelerated the soil Cellulase activity, the number of soil bacteria and fungi. So the litter decomposition was significant affected by the soil qualities, and the litter decomposition rate was Y1(10.30 a-1) > Y2 (9.60 a-1) >CK(6.41 a-1)>Y3(6.29 a-1)。 6. Whether the litter or soil qualities changed, the litter C element at the state of release at all times during the litter decomposition, and the release quantity increased with the decomposition rate augment. Litter released plentiful total penolics content during decomposition, and the release quantity had the positive correlation with the litter decomposition rate. The litter lignin content and the lignin/N all increased with the litter aqueous extracts concentration augment after litter decomposition.

土壤微生物和酶对大气CO2浓度与温度升高的响应

土壤微生物(Soil microbes)是生态系统的重要组成部分，它参与土壤中复杂有机物质的分解和再合成，也参与C、N、S、P等的循环。土壤酶(Soil enzyme)是土壤中具有生物活性的蛋白质，它与微生物一起推动着土壤的生物化学过程，并在树木营养物质的转化中起着重要的作用。鉴于土壤微生物和土壤酶对环境变化的敏感性，它们在CO2浓度和温度升高时的反应将在很大程度上影响森林生态系统的结构和功能。因此，要全面评价大气CO2浓度和温度升高对整个生态系统的影响，有必要对CO2浓度和温度升高条件下的土壤微生物的反应进行深入的研究与探讨。本文应用自控、封闭、独立的生长室系统，研究了川西亚高山岷江冷杉（Abies faxoniana）根际、非根际土壤微生物数量，红桦（Betula albosinensis）根际微生物数量以及根际、非根际土壤酶活性对大气CO2浓度(环境CO2浓度+350±25μmol·mol-1，EC)和温度(环境温度+2.0±0.5℃，ET)升高及两者同时升高(ECT)的响应。结果表明： 1) EC和ET显著增加岷江冷杉根际微生物数量，但不同微生物种类对EC和ET的反应有所差异。6、8和10月，岷江冷杉根际微生物数量与对照(CK)相比，EC处理的根际细菌数量分别增加了35%、164%和312%，ET处理增加了30%、115%和209%；EC和ET处理对根际放线菌和根际真菌数量影响不显著。ECT处理的根际放线菌数量分别增加了49%、50%和96%，根际真菌数量增加了151%、57%和48%；而ECT对根际细菌数量影响不显著。EC、ET和ECT处理对岷江冷杉土壤微生物总数的根际效应明显，其R/S值分别为1.93、1.37和1.46(CK的R/S值为0.81)。 2) 红桦根际微生物数量对EC、ET和ECT的响应不同。生长季节(5~10月)，高密度的红桦根际细菌数量与CK 相比，EC的根际细菌数量分别增加28%、33%、423%、65%、43%和79%，而低密度的红桦根际细菌数量增加不显著。ET能显著增加根际细菌数量(7~10月)，其中高密度的根际细菌数量分别增加了377%、107%、35%、22%，而低密度的根际细菌数量分别增加了27%、27%、64%、48%；ECT对两个密度水平下根际细菌数量均未产生有显著的影响。高、低密度的红桦根际放线菌和根际真菌数量与 CK 相比，EC显著增加了低密度的红桦根际放线菌数量，而对高密度的根际放线菌数量无显著影响；ET和ECT对高低密度的红桦根际放线菌数量均未产生显著影响。EC和ET对高低密度的根际真菌数量也无显著影响，而ECT却显著增加了高低密度的根际真菌数量。 3) EC、ET和ECT处理的低密度红桦根际微生物(细菌、放线菌和真菌)数量没有显著高于或低于高密度根际微生物数量，表明短期内密度对红桦根际微生物数量不产生影响。 4) 不同种类的氧化还原酶对EC、ET和ECT的响应不同。5~10月，EC的红桦根际过氧化氢酶活性是CK 的1.44、1.06、1.11、1.10、1.12和1.24倍，差异显著(6月除外)；ET和ECT处理根际过氧化氢酶活性无显著增加。EC的红桦根际多酚氧化酶活性比CK显著增加；ET的根际多酚氧化酶活性显著高于CK(8月除外)。ECT的根际多酚氧化酶活性高于CK，差异不显著。EC的根际脱氢酶活性分别增加了46%、40%、133%、48%、17%和26%，差异显著。5~7月，ET和ECT的根际脱氢酶活性高于CK的脱氢酶活性，而8~9月则相反，差异性均不显著。 5) EC、ET和ECT对不同种类的水解酶的影响不同。EC能显著增加红桦根际脲酶活性，5~10月分别增加了29%、42%，、70%、67%、59%和57%。ET和ECT 对根际脲酶活性未产生显著影响。EC显著提高根际转化酶活性，5、6和9月EC的根际转化酶活性分别比CK高51%、42%和40%。5和10月，ET的根际转化酶活性低于CK，而其余月份却高于CK，但均具有显著性差异。ECT的根际转化酶活性与CK的根际转化酶活性有显著性差异(9月除外)，5、6和7月的根际转化酶活性分别提高了94%、198%和67%。 6) 与CK相比，EC、ET和ECT的非根际土壤微生物数量以及非根际土壤酶活性均无显著提高。EC、ET和ECT的过氧化氢酶、脲酶的根际效应明显，而多酚氧化酶和脱氢酶根际效应不明显。EC和ECT的转化酶根际效应明显，而ET的转化酶根际效应不明显。 It is well known that atmospheric CO2 concentration and temperature are increasing as a consequence of human activities. In past decades, considerable efforts had been put into investigating the effects of climate change on processes of forest ecological system. In general, studies had been mainly focused on the effects of elevated atmospheric CO2 on plant physiology and development, litter quality, and soil microorganisms. Studies showed that there was variation in the responses of root development and below-ground processes to climate between different plant communities. Since the concentration of CO2 in soil was much higher (10~50 times) than in the atmosphere, increasing levels of atmospheric CO2 may not directly in fluence below ground processes. Betula albosinensis and Abies faxoniana, as the dominated tree species of subalpine dark coniferous forest in the western Sichuan province, which play an important role in the structure and function of this kind of forest ecosystem. In our study, effects of elevated atmospheric CO2 concentration (350±25μmol·mol-1), increased temperature (2.0±0.5℃) and both of the two on the number of rhizospheric microbe and rhizospheric enzyme activity were studied by the independent and enclosed-top chamber’ system under high-frigid conditions. Responses of rhizospheric bacteria, actinomycetes and fungi number of Betula albosinensis and Abies faxoniana under different densities(high density with 84 stems·m-2, low density with 28 stems·m-2 ), and rhizospheric enzyme activity of Betula albo-sinensis to elevated CO2 concentration and increased temperature were analyzed and discussed. The results are as the following, 1) In comparion with the control, the numbers of rhizospheric bacteria of Abies faxoniana were increased by 35%, 164% and 312% significantly in June, August and October respectively of EC, and were increased by 30%, 115% and 209% respectively of ET.However the effect of EC and ET on rhizospheric actinomycetes and fungi was not significant. The number of rhizospheric actinomycetes of ECT were increased significantly by 49%, 50% and 96% respectively, and the increment of rhizospheric fungi were 151%, 57% and 48% respectively .The effect of ECT on rhizospheric bacteria was not significant. Rhizospheric effect of soil microbe for all treatments was significant, with the R/S of 1.93, 1.27 and 1.46 for EC, ET and ECT, respectively. 2) Treatment EC improved the number of rhizospheric bacteria of Betula albosinensis under high density significantly in comparison with the control, over the growing season, the greatest increment of rhizospheric bacteria was from July. However, EC had no effect on the number of rhizospheric bacteria under low density. Except May and June, treatment ET improved the number of rhizospheric signifcantly. The effect of treatment ECT on the number of rhizospheric bacteria under different densities was not significant. Of treatment EC, the number of rhizospheric actinomycetes of Betula albosinensis under low density were increased significantly, however, treatment EC did not stimulate the number of rhizospheric actinomycetes under high density. Simultaneously, treatment ET and ECT did not stimulate the number of rhizospheric actinomycetes. Finally, in treatment ECT, the number of rhizospheric fungi under high density were increased significantly, however treatment EC and ET did not stimulate the number of rhizospheric fungi under different densities. 3) Of treatment EC, ET and ECT, the number of rhizospheric microbe of Betula albosinensis under low density were not more or fewer than that of microbe under hign density along the growing season, which showed that plant density had no effect on the nmber of microbe. 4) From May to October, 2004，rhizospheric catalase activity of Betula albosinensis of treatment EC was 1.44, 1.06, 1.11, 1.10, 1.12 and 1.24 times as treatment CK respectively, and the difference was statistically significant(except June). Treatment ET and ECT did not increase rhizospheric catalase activity significantly. In treatment EC, the rhizospheric pohyphenol oxidase activity was higher than treatment CK significantly. The rhizospheric pohyphenol oxidase activity of treatment ET was higher than CK significantly (except August). The rhizospheric pohyphenol oxidase activity of treatment ECT was higher than CK, but the difference was not statistically significant. Over the growing period, the rhizospheric dehydrogenase activity were increased 46%, 40%, 133%, 48%, 17% and 26% respectively by treatment EC, and the difference was statistically significant. From May to July, the rhizospheric dehydrogenase activity in treatment ET and ECT was higher than CK, but from August to October, the rhizospheric dehydrogenase activity was lower than CK, the difference was not significant. 5) Treatment EC increased rhizospheric urease activity significantly, from May to October, rhizospheric urease activity were increased 29%, 42%, 70%, 67%, 59% and 57% respectively by EC. Treatment ET and ECT had no effect on rhizospheric urease activity. Treatment EC improved rhizospheric invertase activity significantly, in May, June and September, the rhizospheric invertase activity of treatment EC were increased 51%, 42% and 40% in comparison with the control. Except May and October, the rhizospheric invertase activity of treatment ET was markly higher than CK. The rhizospheric invertase activity of treatment ECT was significantly different from CK (except September), in May, June and July treatment ECT increased rhizospheric invertase activity by 94%, 198% and 67% respectively. 6) In comparison with the control, treatment EC, ET, and ECT had no effect on the number of non-rhizospheric microbe and non-rhizospheric enzyme activity. Rhizospheric effect of catalase and urease for all treatments was significant, but rhizospheric effect of pohyphenol oxidase and dehydrogenase was not significant. Rhizospheric effect of invertase of EC and ECT was significant, but rhizospheric effect of invertase of ET was not significant.

花椒叶和土壤浸提液对土壤微生物、土壤酶及土壤化学性质的影响

花椒（Zanthoxylum bungeanum Maxim.）是川西地区重要的经济植物，化感作用是花椒连作障碍的原因之一，而花椒凋落物和根系分泌物对土壤质量的影响是花椒化感作用的一个重要方面。系统研究花椒如何影响土壤有助于深入理解和解决花椒连作障碍。本文主要以大红袍（10a生）花椒叶和种植过花椒的土壤的浸提液浇灌花椒幼苗进行试验，分析叶浸提液与土壤浸提液对非花椒生长土壤中土壤微生物、土壤酶及土壤化学性质的影响。主要结果如下： 1.花椒叶浸提液和土壤浸提液减少了土壤中微生物的种类、组成和数量。本试验中未施加浸提液的土样中根际微生物明显高于非根际区，在经过花椒叶浸提液处理后，根际细菌、真菌和放线菌数量以及微生物总数都有所减少，这样将会导致土壤中的有效养分的供给减少，进而可能影响植物的生长。 2．施加花椒叶浸提液和土壤浸提液，以及花椒幼苗的栽种，对不同土样中的土壤酶各有促进和抑制作用。在浸提液处理下，水解酶之间及氧化还原酶之间各存在相互促进作用。 3.施加花椒叶浸提液和土壤浸提液均抑制了根际土中全氮和有机质含量，叶浸提液还抑制了无苗土中全磷含量，土壤浸提液还抑制了无苗土中全氮含量与根际土全磷、有机质含量。但两种浸提液均促进了根际土中有效磷和水解性氮含量、根外土中全磷含量，叶浸提液促进了根际土中全磷含量，土壤浸提液促进了根外土中有效磷含量。全氮和有机质含量的下降可能对植物生长发育不利。 4．土壤化学性质与土壤酶活性在不同土样中有不同的相关性。全氮含量在施加叶浸提液的土样中与蛋白酶活性呈正相关。水解性氮含量在施加叶浸提液的土样中与蛋白酶活性、蔗糖酶活性呈正相关。全磷含量在施加叶浸提液的土样中与多酚氧化酶活性呈正相关；在施加土壤浸提液的土样中与蛋白酶活性、蔗糖酶活性呈正相关，与多酚氧化酶活性呈负相关。有效磷含量在施加叶浸提液的土样中与多酚氧化酶活性呈正相关，与蛋白酶活性呈负相关；在施加土壤浸提液的土样中与蛋白酶活性、过氧化氢酶活性呈正相关。有机质含量在施加叶浸提液的土样中与蛋白酶活性、蔗糖酶活性呈正相关。 Zanthoxylum bungeanum is one of the most important cash crops in Eastern Tibetan Plateau. Allelopathic effects could be one of reasons for Z. bungeanum’s continuous cropping impediment. The effects of secretion of leaf and root of Z. bungeanum on soil quality is a important way of Z. bungeanum’s allelopathic effects. However, allelopathic effect of Z.bungeanum on soil microbes, enzyme activities and chemical property were seldom studied. In this study, leaf and soil extracts of Da Hongpao(DHP), the most common varieties of Z.bungeanum in this area, were used to assess allelopathic effect of Z. bungeanum on soil biology and biochemistry by pot experiments . The main results showed that: 1. The irrigation of two kinds of extracts reduced the species, component and quantity of soil microbes. In rhizosphere soil which irrigated by distilled water, the quantity of soil microbes is significantly different from exoroot soil. In rhizosphere soil which irrigated by leaf extracts, the quantity of bacterial, fungi, actionmycete and gross of microbes were decreased, it may resulted in reduce of Available nutrient in soil, and influenced the growth of plants. 2．The irrigation of two kind of extracts reduced or enhanced the enzyme activities in different soils. Interaction between hydrolytic ferments and redoxases were promoted each other. 3. The irrigation of two kinds of extracts reduced the total N and organic matter in rhizosphere soil. Leaf extracts also reduced the total P in soil without seedling. Soil extracts reduced total N in soil without seedling and total P, organic matter in rhizosphere soil. But both extracts also enhanced available P and hydrolysable N in rhizosphere soil, total P in exoroot soil. Leaf extracts enhanced total P in rhizosphere soil. Soil extracts enhanced available P in exoroot soil. The reduction of total N and organic matter may influence growth of plants. 4．Positive correlations between total N and prolease, hydrolysable N and prolease, hydrolysable N and saccharase, total P and polyphenol oxidase, available P and polyphenoloxidase, organic matter and prolease, organic matter and saccharase, were studied in soil irrigated by leaf extracts. In soil irrigated by soil extracts, there are positive correlations between total P and prolease, total P and saccharase, available P and prolease, available P and catalase, while negative correlation between total P and polyphenoloxidase, available P and prolease, available P and catalase was found.

荞麦(Fagopyrum tataricum Gaertn.和F. esculentum Moench.)对紫外线B辐射的响应研究

近二十多年来，基于对臭氧层衰减、紫外线B（UV-B）增强的担心，研究者希望了解到紫外线辐射对不同作物的影响情况，增强UV-B辐射条件下是否对作物的生长发育、产量质量构成威胁。在本试验中，我们首先探讨了双子叶作物黄瓜（Cucumis sativus）和大豆（Glycine max）对不同紫外波段的生物效应[分别为B-UVA（315-400 nm），N-UVA（315-340 nm），B-UVB（275-400 nm）和N-UVB（290-340 nm），UV-（>400nm）作对照]。我们观察到所有的UV波段处理都使黄瓜和大豆的生长受到抑制，并且细胞受到不同程度的氧化伤害；UV波段处理的作用效果与不同波段的紫外有效生物辐射剂量有关。处理差异在UV-B波段内部和UV-A波段内部同样存在。植物生长UV辐射公式（BSWF）能很好的预测本试验UV-B波段内的平均植物效应，但不能预测UV-A波段的植物效应。短波UV-A的生物作用强于长波UV-A。光合色素的变化与UV波谱差异和种间差异有关。在高的紫外/可见光背景下，UV-A处理同UV-B同样导致光合色素的降低，但黄瓜类胡萝卜素/叶绿素比例升高。与其他研究者的试验结果比较后，我们认为紫外线B辐射的生物效应一致性很高，但紫外线A波段的生物学效应存在较大争议。因此我们在本试验的基础上仅进行荞麦[苦荞(Fagopyrum tataricum Gaertn.)和甜荞(Fagopyrum esculentum Moench.)]对紫外线B波段的响应研究。 我们对苦荞品种-圆籽荞进行了连续两个生长季节的大田半控制试验以观察UV-B辐射对苦荞生长、发育、产量及叶片色素的影响；试验小区进行降低UV-B、近充足UV-B和增强UV-B辐射处理。我们的试验表明在不同强度UV-B辐射下苦荞的生长、地上部生物量积累及最终产量都有所下降，但苦荞的发育加快；当前条件下的日光紫外线B辐射对植物生长和产量也造成负面影响。植物光合色素被日光及增强UV-B辐射降低；UV化合物及卢丁含量在中低剂量的UV-B辐射强度下显著升高，但在高剂量的增强UV-B辐射下短期升高后迅速下降。我们的试验表明苦荞是一个对UV-B高度敏感的作物。苦荞对UV-B的敏感性与UV-B剂量、外界环境因素及生长季节有关。 单个苦荞品种的试验结果使我们认识到外界UV-B辐射已经对苦荞生长发育构成逆境条件，未来全球气候变化条件下增强紫外线B辐射可能使其处于更不利的生长环境中。因此我们有进行了多个种群进行UV-B响应观察并筛选耐性种群。我们对15个苦荞种群进行增强UV-B辐射处理（6.30 kJ m2 UV-BBE，模拟当地25%的臭氧衰减），我们观察苦荞UV-B辐射效应存在显著的种内差异，UV-B辐射对多数种群具有抑制作用，但对一些种群还有刺激作用。我们采用主成分分析方法与作物UV-B响应指数（RI）来评价苦荞作物UV-B辐射耐性。我们发现作物的UV-B耐性不仅与其原产地背景UV-B强度有关，而且与作物相对生长效率、次生代谢产物含量（如卢丁）及其他因素有关。我们观察到苦荞伸展叶总叶绿素变化与UV-B耐性成正相关；室内苦荞幼苗的UV-B辐射致死试验表明：苦荞种群死亡率与其UV-B耐性成负相关。 此外，我们对甜荞的UV-B辐射响应也进行了初步研究。选取美姑甜荞、巧家甜荞和云龙甜荞进行5个梯度的增强UV-B辐射室外模拟试验。我们观察到UV-B辐射显著降低了甜荞的生长、生物量及产量；并严重影响了甜荞的生殖生长，降低了花序数、种子数和结实率；并且UV-B辐射对甜荞的抑制作用存在显著的剂量效应。三种甜荞品种存在显著的种内差异，其中美姑品种UV-B耐性最强，且膜脂受UV-B辐射氧化伤害最小，这与该品种UV-B辐射下较高的GR酶活性、APX酶活性和PPO酶活性、以及含量更高的抗坏血酸有关。甜荞的次生代谢也受到增强UV-B辐射的影响，其香豆酰类化合物在UV-B辐射下升高显著，而槲皮素含量也在高剂量UV-B辐射下有所增加；卢丁含量依赖UV-B辐射剂量而变化，中低剂量UV-B辐射下其卢丁含量逐渐升高，但在高剂量辐射下逐渐下降。 通过对生长在高海拔地区的荞麦作物(苦荞和甜荞)进行的室外研究，我们认识到作物不同品种存在很大的耐性差异，这就为UV-B耐性育种创造了有利条件。进一步加大荞麦种质资源筛选力度并深入认识荞麦抗性机理，在此基础上通过杂交或其他基因融合手段培育抗性品种，对高剂量UV-B辐射地区的荞麦产量的提高将起到重要推动作用，并使荞麦生产能有效应对未来全球气候变化条件下UV-B辐射可能升高的威胁。 During last few decades, due to concern of ozone layer depletion and enhancement of ultraviolet B radiation（UV-B, 280-315 nm), the agronomist want to know the responses of different crop species to UV-B. In the first experiment of our study, the effect of different UV band [B-UVA（315-400 nm), N-UVA（315-340 nm), B-UVB（275-400 nm), N-UVB（290-340 nm）and UV-（>400nm, as control)] on the cucumber（Cucumis sativus）and soybean（Glycine max）were investigated in growth room. Spectra-dependent differences in growth and oxidation indices existed within UV-A bands as well as UV-B bands. The general biological effects of different band were UV- ＜ B-UVA＜ N-UVA＜N-UVB＜B-UVB. The plant growth biologically spectra weighting function（BSWF）matched well with average plant response in UV-B region, but not in UV-A region. Shorter UV-A wavelength imposed more negative impact than longer UV-A wavelength did in both species. The effect on photosynthetic pigment was related to different UV bands and different species. The photosynthetic pigment content was decreased by UV-A spectra as well as UV-B spectra. In comparison with the results of previous studies, we found that the wavelength-dependent biological effect of ultraviolet B radiation has high consistency, but the biological effect of ultraviolet-A radiation was inconsistent. We narrow our following study on the effect of ultraviolet B radiation on the buckwheat（tartary buckwheat and common buckwheat). The tartary buckwheat（Fagopyrum tataricum Gaertn.）cultivars Yuanziqiao was grown in the sheltered field plots for two consecutive seasons under reduced, near-ambient and two supplemental levels of UV-B radiation. The crop growth, photosynthetic pigments, total biomass, final seed yield and thousand-grain weight were decreased by near-ambient and enhanced UV-B radiation, while crop development was promoted by enhanced UV-B radiation. Leaf rutin concentration and UV-B absorbing compound was generally increased by UV-B with the exception of 8.50 kJ m-2 day-1 supplemental levels. Our results showed that tartary buckwheat is a potentially UV-B sensitive species. Study on one cultivars showed that ambient solar radiation had present a stress to tartary buckwheat. This makes it necessary to observe the UV-B response of many cultivars and screen tolerant cultivars. Fifteen populations of tartary buckwheat were experienced enhanced UV-B radiation simulating 25% depletion of the stratospheric ozone layer in Kunming region, and plant responses in growth, morphology and productivity were observed. Principal components analysis（PCA）was used to evaluate overall sensitivity of plant response to UV-B as well as response index. The different populations exhibited significant differences in responses to UV-B. The photosynthetic pigments of young seedlings were also affected significantly under field condition. On the other hand, the healthy seedlings of different populations were exposed to the high level of UV-B radiation in growth chambers to determine the plant lethality rate. The plant tolerance evaluated by multivariate analysis was positively related to total plant chlorophyll change, but negatively related to lethality rate. In other hand, the UV-B responses of the other important cultivated buckwheat species, common buckwheat（Fagopyrum esculentum Moench.), were also studied preliminarily. Three widespread cultivated variety（Meigu, Qiaojia and Yunlong cultivars）were provided with five level of enhanced UV-B radiation outdoors. We observed that the crop growth, development and production were significantly decreased, and reproductive production, like anthotaxy number, seed number and seed setting ratio, was also decreased. Dose-dependent inhibition effect caused by enhanced UV-B radiation also existed in common buckwheat. Significant intraspecific difference existed in those three cultivars. The Meigu cultivars with dwarfed growth and lower production have highest UV-B tolerance as well as lowest damage in cell membrane, this could be associated with profound enhancements of glutathione reductase（GR）activity, ascorbate peroxidase activity and polyphenol oxidase activity as well as higher ascorbic acid concentration. The secondary metabolism was also affected by UV-B radiation, with profound elevation of coumarin compound and moderate increase of quercetin concentration. Rutin concentration was peaked in 5kJ m-2 UV-B. The contrasting effect of UV-B radiation on different populations indicated that there existed abundant genetic resources for selecting tolerant populations of common and tartary buckwheat. Much effort needed be pose on screening of buckwheat germplasm and clarification of mechanism of buckwheat tolerance to UV-B. On this base the tolerant cultivars could be bred by hybridization and other gene transfusion method, this would help increase buckwheat yield in high ambient UV-B region and counteract the effect of possible enhanced UV-B radiation in future.

青杨组(Section Tacamahaca Spach)杨树不同种群对土壤干旱和重金属锰污染的生态生理响应

土壤是人类赖以生存的自然环境和农业生产的重要资源，世界面临的粮食、资源和环境问题与土壤密切相关，目前危害土壤的主要因素是干旱和重金属污染。杨树具有适应性强、生长快和丰产等特性，本论文以青杨组杨树为模式植物，采用植物生态、生理及生物化学等方法，研究杨树对土壤干旱和锰胁迫的生态生理反应以及种群间差异，研究成果可为我国干旱半干旱地区营造人工林、防止沙漠化提供理论依据，也为恢复与重建重金属污染地区退化生态系统提供科学指导。主要研究结果如下： 1. 青海杨不同种群对干旱胁迫的响应差异 干旱胁迫显著降低了两个青海杨种群（干旱种群和湿润种群）生物量积累，包括株高、基径、干物质积累等，通过植物结构的调整，有更多的生物量向根部分配。干旱胁迫还显著降低了叶绿素和类胡萝卜素含量，增加了游离脯氨酸和总氨基酸含量。另一方面，干旱胁迫诱导了活性氧的积累，作为第二信使，激活了抗氧化系统，包括抗坏血酸（ASA）含量和酶系统如超氧化物歧化酶（SOD），愈创木酚过氧化物酶（GPX），抗坏血酸过氧化物酶（APX）和谷胱甘肽还原酶（GR）。这样，杨树既有避旱机制又有耐旱机制，使其在干旱胁迫下有相当程度的可塑性。与湿润种群相比，干旱种群杨树有更多的生物量分配到根部，积累了更多的游离脯氨酸和总氨基酸来进行渗透调节，并且有更有效的抗氧化系统，包括更高含量的ASA 和更高活性的APX 和GR，这些使得干旱种群杨树比湿润种群杨树对干旱有更好的耐性。 2. 喷施硝普钠（SNP）对青海杨阿坝种群干旱胁迫耐性的影响 干旱胁迫显著的降低了青海杨阿坝种群的生长和生物量积累以及叶片相对含水量，还诱导了脯氨酸的合成以进行渗透调节。干旱胁迫下过氧化氢（H2O2）显著累积从而造成对膜脂和蛋白的伤害，使得丙二醛和蛋白羰基含量升高。干旱胁迫下喷施SNP可以减轻干旱胁迫造成的伤害，包括增加叶片的相对含水量，增加脯氨酸和总氨基酸的积累，并激活抗氧化酶系统如SOD，GPX和APX，从而减少丙二醛（MDA）和蛋白羰基（C＝O）的积累，但是在水分良好情况下SNP的效果不显著。 3. 青杨不同种群对锰胁迫的生长与形态响应差异 在同一锰浓度下，干旱种群的耐性指数都要高于湿润种群，这表明青杨对干旱和高锰胁迫具有交叉耐性。两个种群的株高，生物量和叶绿素含量都随锰浓度的升高而逐渐下降。就累积浓度而言，0 和0.1 mM 锰胁迫下，干旱种群积累的锰浓度要高于湿润种群，而在高浓度锰胁迫下(0.5 和1 mM)，湿润种群要高于干旱种群。在0，0.1 和0.5 mM下，锰大多积累在根中，叶片次之，茎中最少。而在1 mM，锰更多的积累在叶片中。就累积总量而言，在各个锰浓度胁迫下，根，茎和叶相比，两个种群青杨都是叶片累积的锰总量要高于根和茎。两个种群间比较，对照中没有显著区别，0.1 mM 锰胁迫下，湿润种群根中累积的锰要高于干旱种群，而在地上部中，干旱种群要高于湿润种群。而0.5 和1 mM 锰胁迫下，根、叶、茎+叶、根+茎+叶中，锰累积总量都是湿润种群高于干旱种群。锰胁迫下，青杨叶片数和叶面积包括总叶面积和平均叶面积都显著降低。叶片横切面的光学显微观察结果表明未经锰胁迫的栅栏组织的细胞饱满，海绵组织发达、清晰；胁迫后杨树叶片栅栏组织细胞出现不同程度的皱缩，海绵组织几乎不可见，此外还发现输导组织在胁迫下密度变小和分生组织严重割裂等现象。 4. 青杨不同种群对锰胁迫的生理与生化响应差异 青杨两个种群脱落酸（ABA）含量在锰胁迫下都显著增加，干旱种群的增幅更大。三种多胺含量在锰胁迫下显示了不同的响应趋势：腐胺在两个种群的各个锰处理下都增加，亚精胺只在干旱种群中显著增加，而精胺除了干旱种群在1 mM 下有所增加外，在锰胁迫下变化很小。谷胱甘肽含量随锰浓度升高而增加，在0.5 mM 锰时达到最高值，1mM 时有所下降。植物络合素（PCs）含量与非蛋白巯基（NP-SH）趋势相似，随锰浓度的升高而增加，且干旱种群中含量要高于湿润种群。锰处理还引起氧化胁迫，表现为过氧化氢和丙二醛含量增加。SOD 活性在湿润种群中，在0 到0.5 mM 锰胁迫下活性升高，但在1 mM 锰胁迫时，其活性有所下降。而在干旱种群中，SOD 活性变化较小，并始终维持在一个较高的水平。APX 活性在两个种群中都随锰浓度的升高而增加，干旱种群活性要高于湿润种群。锰胁迫还显著增加了酚类物质的含量，同时GPX 和多酚氧化酶（PPO）活性也随锰浓度的升高而增加。干旱种群的酚类含量和GPX 与PPO 活性都要高于湿润种群。锰胁迫还改变了氨基酸的含量和构成，根据锰胁迫下浓度变化的不同，可以将游离氨基酸分为三组：第一组包括，谷氨酸，丙氨酸和天门冬氨酸，这一组氨基酸含量在锰胁迫下有所下降。第二组包括缬氨酸，亮氨酸和苏氨酸含量在锰胁迫下基本不变化或变化很小。剩下的氨基酸为第三组，这组氨基酸含量在锰胁迫下显著增加，而根据增加的幅度又可以将它们分为两个亚组，丝氨酸，酪氨酸，苯丙氨酸，组氨酸和脯氨酸，在1 mM 下的含量是对照的4 倍以上。异亮氨酸，赖氨酸，精氨酸和甘氨酸含量在1 mM 下是对照含量的2 倍以下。同时，同一锰浓度下，干旱种群比湿润种群积累的氨基酸含量要高。 Soil is the indispensable environment for human survival and important resource foragriculture development. Food and environmental problems facing the world are all closelyrelated to soil and nowadays it is threatened by many factors, among which drought stress andheavy metal pollution are the most serious ones. Poplars (Populus spp.) are importantcomponents of ecosystem and suitable as a source of fuel, fiber and lumber due to their fastgrowth. In this study, different populations of Section Tacamahaca spach were used as modelplants to investigate the adaptability to drought stress and manganese toxicity and differencesbetween populations from dry and wet climate regions. Our results can provide theoreticalevidence for the afforestation and prevention of desertification in the arid and semi-arid areas,and also can supply scientific direction for the reconstruction and rehalibitation of ecosystemscontaminated by heavy metals. The results are as follows: 1. Differences in ecophysiological responses to drought stress in two contrastingpopulations of Populus przewalskii Drought stress not only significantly affected dry mass accumulation and allocation, butalso significantly decreased chlorophyll pigment contents and accumulated free proline andtotal amino acids. On the other hand, drought also significantly increased the levels ofabscisic acid and reactive oxygen species, as secondary messengers, to induce the entire set ofantioxidative systems including the increase of reduced ascorbic acid content and the activities of superoxide dismutase, guaiacol peroxidase, ascorbate peroxidase and glutathioneredutase. Thus the combination of drought avoidance and tolerance mechanisms conferredpoplar a high degree of plasticity in response to drought stress. Compared with the wetclimate population, the dry climate population showed lower dry matter accumulation andallocated more biomass to root systems, and accumulated more free proline and total aminoacids for osmotic adjustment. The dry climate population also showed more efficientantioxidant systems with higher content of ascorbic acid and higher activities of ascorbateperoxidase and glutathione redutase than the wet climate population. All these made the dryclimate population superior in adaptation to drought stress than the wet climate population. 2. Effect of exogenous applied SNP on drought tolerance in Populus przewalskii Drought stress significantly increased hydrogen peroxide content and caused oxidativestress to lipids and proteins assessed by the increase in malondialdehyde and total carbonylcontents, respectively. The cuttings of P. przewalskii accumulated proline and other aminoacids for osmotic adjustment to lower water potential, and activated the antioxidant enzymes such as superoxide dismutase, guaiacol peroxidase and ascorbate peroxidase to maintain thebalance of generation and quenching of reactive oxygen species. Moreover, exogenous SNPapplication significantly heightened the growth performance of P. przewalskii cuttings underdrought treatment by promotion of proline accumulation and activation of antioxidant enzymeactivities, while under well-watered treatment the effect of SNP application was very little. 3. Morphological responses to manganese toxicity in the two contrasting populations ofPopulus cathayana High concentration of manganese caused significant decrease in shoot height andbiomass accumulation. The tolerance index of the dry climate population was significantlyhigher than that of the wet climate population, suggesting the superior Mn tolerance in theformer and the existence of cross-tolerance of drought stress and high Mn toxicity. Injuries tothe leaf anatomical features were also found as the reduced thickness in palisade and spongyparenchyma, the decreased density in the conducting tissue and the collapse and split in themeristematic tissue in the central vein. As for the Mn concentrations in the plant tissues, under0, 0.1 and 0.5 mM, most of the Mn accumulated in the roots, then leaves, and stem the least, while under 1 mM, most of the Mn accumulated in the leaves. As far as the total amounts ofMn extraction are concerned, the leaf extracted more Mn than the root and stem in the twopopulations under various Mn concentrations. There is no difference between the twopopulations under control. Under 0.1 mM, the wet climate population extracted higher Mn inthe root than the dry climate population, while in the shoot, the dry climate populationextracted much more Mn. Under 0.5 and 1 mM, the wet climate population translocated moreMn both in the root and the shoot than the dry climate population. 4. Physiological and biochemical responses to manganese toxicity in the two contrastingpopulations of Populus cathayana Mn treatment resulted in oxidative stress indicated by the oxidation to lipids, proteinsand DNA. A regulated network of defence strategies was employed for the chelation,detoxification and tolerance of Mn including the enhanced synthesis of ABA and polyamines,the accumulation of free amino acids, especially His and Pro, and the activation of theenzymes superoxide dismutase and guaiacol peroxidase. Contents of non-protein thiol,reduced glutathione, phytochelatins and phenolics compounds and activities of superoxide dismutase, guaiacol peroxidase and polyphenol oxidase also increased significantly forantioxidant or chelation functions. The wet climate population not only accumulated lessabscisic acid, free amino acids, phytochelatins and phenolics compounds, but also exhibitedlower activities of superoxide dismutase, guaiacol peroxidase and polyphenol oxidase thusresulting in more serious oxidative damage and more curtained growth.

模拟增温对川西亚高山人工云杉林土壤碳含量和土壤呼吸的影响

全球气候变化已经成为不争的事实，其中全球变暖是近年来国内外的研究热点之一。土壤碳库作为陆地生态系统最大的碳库，气温升高必然会导致一系列的土壤碳储量和碳通量的变化，这些微小的变化又可能导致大气CO2浓度的变化并强化这种变暖的趋势。目前，土壤碳循环对温度升高的响应仍然是陆地碳循环研究最缺乏的部分，对土壤有机碳动态变化的研究仍存在着很大的不确定性与争议。四川西部的亚高山人工针叶林是青藏高原东部高寒林区的重要组成部分，是研究全球变化对森林生态系统影响的关键地区和重要森林类型。本研究通过采用原位人工模拟增温装置（Open-top chambers，OTCs）对川西米亚罗60年人工云杉林土壤实施增温，研究高海拔地区森林，尤其是人工森林系统下的土壤有机碳 含量、土壤呼吸及土壤酶活性对温度升高的响应。结果表明： 1. 增温处理的660天（2005年11月至2007年9月）期间，增温条件下的平均气温和土壤平均温度分别比对照提高0.43 ℃和0.27 ℃；0~10 cm土壤含水量在增温的不同时期均有不同程度的降低。 2. 土壤蔗糖酶、蛋白酶和脲酶活性在温度升高的不同阶段均有不同程度的提高。在增温处理300天（2006.09）、540天（2007.05）、600天（2007.07）和660天（2007.05）后，0~10 cm层的蔗糖酶活性分别比对照提高了36.36%（P<0.05）、24.31%、14.54%（P<0.05）和7.22%，脲酶活性分别提高了12.90%、24.19%（P<0.01）、34.48%（P<0.05）和14.64%(P<0.05)，蛋白酶活性分别提高了31.37%、1.99%、3.70%和17.80%。10~20 cm层的土壤酶活性也均有不同程度的提高，但均没有显著差异。蔗糖酶、脲酶和蛋白酶活性均呈现出随土层加深而减弱的趋势。 3. 土壤过氧化氢酶和多酚氧化酶活性在增温的第1年内均有不同程度的提高，但在增温的第2年内比对照有所降低。增温300天后（2006.09），过氧化氢酶和多酚氧化酶在0~10 cm层分别比对照增加3.76%和49.25%（P<0.05），10~20 cm层分别增加了5.54%和29.67%。在增温的第2年内，增温540天（2007.05）、600天（2007.07）和660天（2007.09）后，0~10 cm层的过氧化氢酶活性分别比对照降低了27.70%（P<0.05）、4.34%和1.47%，多酚氧化酶活性分别降低了5.86%、11.76%（P<0.05）和7.47%。增温的第2年内，10~20 cm层的过氧化氢酶和多酚氧化酶活性也均有不同程度的降低，但差异均未达到显著水平。不同土层之间相比较，过氧化氢酶活性随土层加深而降低，多酚氧化酶活性随土层加深而增加。 4. 土壤有机碳和有机质在增温的不同阶段，含量比对照均有所降低；且随增温时间的延长，降低的幅度下降。0~10 cm层的土壤有机碳和土壤有机质在增温300天（2006.09）、540天（2007.05）、600天（2007.07）和660天（2007.09）后分别降低了8.69%、4.35%、3.80%和2.44%，差异均未达到显著水平。土壤全氮含量在增温后与对照相比无明显的增加或者降低趋势。增温条件下的土壤C/N比与对照相比有所降低，但在增温各阶段的差异均不显著。10~20 cm层的有机碳、有机质和C/N比也有不同程度的降低趋势，但差异均不显著。不同土层之间相比，0~10 cm层的有机碳、有机质、全氮含量和C/N比均高于10~20 cm层，呈现出随土层加深而降低的趋势。 5. 土壤呼吸速率在增温第1年内，与对照相比明显提高，但在增温处理2年后，与对照相比无显著变化。增温300天（2006.09）和360天（2006.11）后分别提高了13.32%和21.17%，差异显著。增温处理540天（2007.05）到660天（2007.09）期间，与对照相比，不仅没有明显的提升，反而有些月份比对照有所降低，对温度升高的敏感性降低，呈现出对温度升高的适应性。土壤呼吸的日呼吸速率呈现单峰曲线形式，在14:00~20:00期间达到最大值，在4:00~10:00期间具有最低值。土壤呼吸的季节变化，呈现出与外界环境温度相一致的趋势，在7月份（夏季） 最高，11月份（冬季）最低。土壤呼吸与2 cm土壤温度、5 cm土壤温度和空气温度均呈极显著指数相关，与0~10 cm土壤含水量呈线性相关，相关性达到显著水平，但低于土壤呼吸与温度的相关性。 The past century has seen a marked increase in atmospheric carbon dioxide concentrations and a concomitant warming that has drawn scientific attention to the link between global carbon stocks and climate change. In particular, the decomposition and turnover of soil organic matter is recognised as an important determinant of carbon driven climate change. The slightly variation in soil organic carbon will result in the increase of atmospheric carbon dioxide concentrations and reinforce the tendency of warming. The experiment was conducted in Subalpine coniferous forest in western Sichuan province. Subalpine coniferous forest in western Sichuan was a important part of eastern Qinghai-Tibetan Plateau, which play a important role in reseaching the sensitivity of forest ecosystem to climate change. To investigate the effects of elevated temperature on soil organic carbon content, soil respiration rates, and soil enzyme activities in subalpine Picea asperata plantations, a esimulated warming measure was applied with Open-top chambers. The results were as followed: 1) During the period from Nov. 2005 to Sep. 2007, mean air temperature and soil temperature were respectively 0.43℃ and 0.27℃ the ambient higher. Soil water content decreased to different exent in different months in warmed plots than in unwarned plots at depth of 0-10 cm. 2) In general, elevated temperature enhanced the soil enzyme activities of invertase, protease, and urease. In the first year of warming—after 300 days’ treatment (in Sep,2006), the activities of invertase, protease, and urease increased by 36.36%, 12.90% and 31.37% respectively at the depths of 0-10 cm，among which the activity of invertase reached statistic significance. In the second year of warming, invertase activity increased by 24.31% after 540 days’ treament (in May, 2007), 14.54% after 600 days’ treament (in Jul, 2007) and 7.22% after 660 days’ treatment (in Sep, 2007) at the depths of 0-10 cm, and the differences in July and Septemmber were statistically significant. Elveated temperature also increased the activity of urease in the second year of warming and had significant effects in May and July. The activity of protease in warmed plots was also higher than in unwarmed plots at depths of 0-10 cm, but there was no significant difference. Elevated temperature had no significant effects on all soil enzyme acitivities at the depths of 10-20 cm in the first and sencond year. The values of above-mentioned soil enzyme all decreased with soil layers. 3) Eleavted temperature enhanced the activities of catalase and polyphenol oxidase in the first year of warming while they turned out downtrend in the second year. The activity of catalase increased by 3.76% and 5.54% at depths of 0-10 cm and 10-20 cm respectively in the first year—after 300 days’ warming (in Sep, 2006), the differences of which had no statistical significance. The activity of polyphenol oxidase was significantly increased by 49.25% at depths of 0-10 cm and not significantly increased by 29.67% at depths of 10-20 cm after 300 days’ warming. In the second year of warming, the catalase activity was significantly decreased by 27.70% after 540 days’ treament (in May, 2007) and not significantly decreased by 4.34% and 1.47% after 600 days’ (in Jul, 2007) and 660 days’ treament (in Sep, 2007) respectively. The activities of catalase and polyphenol oxidase at depths of 10-20 cm were decreased to different extent, but there was no significant difference. Catalase activity stepped down with soil layers while polyphenol oxidase activity stepped up. 4) Increased temperature in both the first year and the second year resulted tendency of decrease in the contents of soil organic carbon and soil organic matter, and C/N ratios at soil depths of 0-10 cm and 10-20 cm. However, with the prolonged warming, the tendency of decrease gradually tapered off and the extent of decrease in the second year of experiment were lower than that in the first year. The contents of soil organic carbon and soil organic matter were all decreased 8.69% by warming in the first year and dcreased 4.35%, 3.80% and 2.44% in May, July and September of the second year, but no significant difference were found. The C/N ratios increased 8.52% in the first year of warming and had less increment in the second year, all of which were not statistical significant. Eleveated temperature had no obvious effect on the content of tatol N in two year consecutive warming experiment. The contents of soil organic carbon and soil organic matter, total N and C/N ratios all had the tendency of dcreasing with soil layers. 5) Soil respiration rates were significantly enhanced by 13.32% and 21.17% after 300 days’ (in Sep, 2006) and 360 days’ (in Nov, 2006) treament in the first year of warming, but the same showed no obvious difference in the second year of treatment, which was assumed the adaptability of soil respiration with a certain heightened temperature. Diurnal soil resspiration showed a daily variation with a minimum value between 4:00 and 10:00 h and a maximum value between 14:00 and 20:00 h, coinciding with the minimum and maximum values of soil temperature at 2 cm. Soil respiration rates exhibited a pronounced seasonal variation with minimum values in Novmber and a maximum value in July, approximately coinciding with the seasonal variation of air and soil temperature. An exponential function provided the best fit for soil respiration with temperature while a quadric equation was used to estimate the effect of soil moisture on soil respiration, which were all significantly correlated. Soil respiraion rate was more highly correlated with the soil temperature than soil moisture.