淹水时间对水稻土中地杆菌科群落结构及丰度的影响

【目的】通过模拟水稻土淹水过程,探讨地杆菌科(Geobacteraceae)群落结构和相对丰度随淹水时间的动态变化特征,揭示其群落结构和相对丰度变化与微生物Fe(Ⅲ)还原的内在联系。【方法】提取水稻土淹水培养1 h、1 d、5 d、10 d、20 d和30 d后的微生物总DNA,构建地杆菌科16S rDNA克隆文库,采用PCR-RFLP方法分析地杆菌科的群落结构和多样性变化特征,通过Real-time PCR技术测定地杆菌科相对丰度的动态变化。采用厌氧泥浆培养方法,测定水稻土中Fe(Ⅱ)产生量变化。【结果】供试水稻土中,微生物Fe(Ⅲ)还原过程在淹水培养初期变化明显,培养20 d后达到稳定期,最大铁还原潜势为10.16 mg/g,最大反应速率为1.064 mg/(g.d),最大反应速率对应的时间为4.84 d。α多样性指数显示,水稻土中地杆菌科的多样性随淹水时间延长呈现波动性变化,淹水5 d和20 d处理出现2个峰值,而淹水10 d和30 d处理的多样性明显减小。β多样性指数表明淹水过程中群落结构存在明显差异。不同淹水时间共产生了10种地杆菌科优势类型,分别属于Clade 1和Clade 2。Real-time PCR结果表明,地杆菌科与总细菌16S rDNA丰度的比值在淹水培养1 d时最小(1.20%),而20 d时达到最大值(4.54%)。【结论】淹水培养的水稻土中,地杆菌科微生物的多样性和相对丰度的动态变化与微生物Fe(Ⅲ)还原过程密切相关。     

淹水培养过程中水稻土细菌丰度与群落结构变化

水稻土是非常复杂又典型的生态系统, 分析淹水培养过程中水稻土细菌的丰度和群落结构变化规律, 可以客观反映水稻土中细菌群落结构信息, 为深入探讨水稻土细菌微生物对稻田的影响和在生态系统中的作用(营养元素转换、重金属还原与抑制甲烷生成过程等)提供实验基础与理论依据。作者采用淹水非种植水稻土微环境模式系统, 提取水稻土淹水培养1 h和1、5、10、20、30、40、60 d后的微生物总DNA, 利用Real-time PCR和PCR-DGGE (denaturing gradient gel electrophoresis)技术检测了淹水培养过程中细菌丰度与群落结构的变化。结果表明: 淹水水稻土中细菌的丰度在1 d时最大, 并在40 d到达第二个峰值, 说明淹水过程改变了细菌的丰度。基于16S rRNA基因V3区的DGGE图谱分析显示, 淹水过程中细菌的群落结构发生了演替性变化: r-策略生存的细菌仅存在于淹水初期; k-策略生存的细菌存在于淹水后期; r-和k-策略共生存的细菌存在于整个淹水过程中, 淹水后期k-策略的细菌占据优势。淹水培养过程中优势种群多样性指数大体呈现先上升后减小的趋势。主成分分析(PCA)将淹水处理过程分成几类不同的生境, 反映出中、后期细菌群落结构较为稳定; 测序结果表明, 32个优势条带所代表的细菌分别属于厚壁菌门、绿弯菌门、拟杆菌门、变形菌门和酸杆菌门, 且与来自不同地域的水稻土、其他类型土壤、活性污泥以及湖泊沉积物等生态系统的细菌关系密切。     

醉马草免培养内生细菌的多样性

为了解醉马草内生细菌的组成和多样性,采用液氮研磨法提取醉马草总DNA,利用内生细菌16S rRNA基因特异性引物对醉马草总DNA 进行16S rRNA 基因扩增,构建醉马草内生细菌16S rRNA基因文库。对筛选到249个克隆进行Hae Ш酶切分析,得到57个操作分类单元(OTUs)。系统发育分析将其归为4个门:变形杆菌门(Proteobacteria)、厚壁菌门(Firmicutes)、放线菌门(Actinobacteria)、拟杆菌门(Bacteroidetes)以及1个未知类群。其中74%的克隆与可培养细菌中的37个属具有高的16S rRNA基因序列相似性 (97％-100％),其中芽孢杆菌属(Bacillus spp.)、红球菌属(Rhodococcus spp.)、黄杆菌属(Flavobacterium spp.)、黄单胞杆菌属(Xanthomonas spp.)最为丰富。另外26%的克隆序列与GenBank中已存细菌16S rRNA基因序列相似性小于96%。研究结果表明,醉马草中可培养内生细菌丰富,多样并且可能存在一些潜在新物种或新类群。     

碳源和淹水时间对水稻土微生物Fe(Ⅲ)还原能力的影响

以我国6个省的水稻土为供试样品,采用厌氧恒温培养方法,研究了分别以葡萄糖、丙酮酸盐、乳酸盐和乙酸盐为惟一碳源时不同淹水时间土壤微生物群落对Fe(Ⅲ)的还原能力.结果表明：不同淹水时间对Fe(Ⅲ)还原特征值V_max的影响显著,表现为淹水20 d > 30 d > 12 d > 1 d > 5 d,不同淹水时间下水稻土微生物群落结构不同是导致Fe(Ⅲ)还原能力不同的主要原因.不同碳源对微生物铁还原过程有显著影响,葡萄糖和丙酮酸盐在不同淹水时间中始终为优势碳源,其Fe(Ⅲ)还原率分别为88.1%～99.9%和58.0%～97.9%;不同土壤铁还原微生物群落对乳酸盐的利用差距较大,湖南和浙江水稻土在整个淹水周期中Fe(Ⅲ)还原率达到87.1%～100%,而其他土壤则表现为淹水前5 d为5.0%～49.4%,12 d后增加到52.2%～99.9%;乙酸盐处理在不同淹水时间中都表现为随时间推移Fe(Ⅲ)还原率逐渐增大的趋势,其中浙江水稻土的变化最大,在5.3%～75.8%.     

土壤氧气可获得性对双季稻田温室气体排放通量的影响

为探讨土壤氧气可获得性(SOA)对双季稻田温室气体排放的影响,利用静态箱气相色谱法对多种管理措施影响下稻田温室气体排放通量和土壤氧化还原电位(Eh)、pH值及田间淹水深度(H)等3种SOA因子进行了观测。结果表明,甲烷(CH₄)排放最集中的Eh值、pH值和H范围分别为-100-0mV、5 < pH < 6和1-5cm,3个范围内分别观测到48.8%、61.1%和77.0%的CH₄排放,其中H对CH₄排放影响最明显,单独由其就可解释37.8%的CH₄排放通量(P < 0.0001)。对于氧化亚氮(N₂O),观测到较多的负通量,其纯排放最密集的3种SOA因子的范围分别是:0-100mV、5 < pH < 6和1-5cm,而200-300mV是其排放的临界Eh范围,高于此范围N₂O排放极少。厌氧的反硝化过程是双季稻田N₂O产生的主导过程。可为水稻田温室气体排放机理研究提供基础数据。     

硝化抑制剂对蔬菜土硝化和反硝化细菌的影响

土壤N素循环主要是微生物驱动的转化过程,然而对其的驱动与调控机理了解还很不够。选取长沙黄兴镇蔬菜基地两种蔬菜土研究硝化抑制剂(DCD)对N素转化过程及功能微生物的影响。试验通过室内土壤培养,处理为单施尿素(CK)和尿素与硝化抑制剂双氰胺配合施用(DCD),重复3次。在培养过程中系统监测了土壤中NH₄⁺-N、NO₃^--N含量变化,同时采用荧光定量PCR(real-time PCR)方法研究硝化抑制剂对土壤中氮素转化功能基因丰度的影响。结果表明:在培养过程中DCD处理使两个供试土壤的NH₄⁺浓度稳定在较高水平,而NO₃^-浓度则明显低于对照;施用DCD导致土壤中硝化基因amoA丰度显著减少,而对16S rRNA和反硝化基因nirK丰度没有产生明显影响。因此,DCD在菜地土壤中主要通过抑制氨氧化细菌的繁衍来抑制硝化作用。     

丰水期长江感潮河口段网采浮游植物的分布与长期变化

于2009年6、8月对长江口门至江阴的河口段浮游植物进行了拖网采集,共检出浮游植物6门99属239种。其中:硅藻123种,甲藻19种,绿藻和蓝藻各42种,裸藻9种,黄藻4种。河口段网采浮游植物丰度以蓝藻占绝对优势,硅藻次之,两者合计在群落中的比例超过了95%。优势种也主要以蓝藻(水华鱼腥藻Anabaena flos-aquae、柔软腔球藻Coelosphaerium kuetzingiarum、微囊藻Microcystis spp.、颤藻Oscillatoria spp.和席藻Phorimidium spp.)构成,硅藻仅有2种(骨条藻Skeletonema spp.和颗粒直链藻Aulacoseira granulata)。口门内盐度均<0.5,群落基本以淡水类群为主,口门附近则以半咸水类群为主,海水类群主要位于口门外(盐度>13)。随着水温和营养盐水平的升高,8月浮游植物平均丰度(347.75×10⁴ 个/m³)明显高于6月(204.19×10⁴ 个/m³)。根据多维尺度和相似性分析,丰水期长江河口段浮游植物群落组成与分布存在显著(P<0.01)的时空差异。对比20世纪80年代以来的历史资料发现,长江口门内网采浮游植物丰度显著升高,且优势种也从硅藻(骨条藻、直链藻和圆筛藻)转变为蓝藻(颤藻、鱼腥藻和微囊藻)。     

鄱阳湖苔草湿地甲烷释放特征

2009年5月-2010年4月在鄱阳湖南矶湿地国家级自然保护区选择以灰化苔草为建群种的洲滩,设置土壤-植物系统(TC)、剪除植物地上部分 (TJ)2个试验处理,利用密闭箱-气相色谱法测定了鄱阳湖典型苔草湿地的甲烷(CH₄)释放通量。结果表明:1)TC、TJ 2个试验处理CH₄释放速率变化范围分别为-0.094-17.75 mg · m^-2 · h^-1、-0.122-19.16 mg · m^-2 · h^-1,均表现出明显的季节变化规律;2)地表未淹水期间,剪草处理CH₄释放显著高于非剪草处理(t=2.69, P<0.05);地表淹水达到15 cm后,剪草处理CH₄释放明显低于非剪草处理。3)土壤5 cm温度、土壤水分与2处理非淹水期间CH₄释放速率均呈显著正相关,是非淹水期间CH₄通量变化的主要控制因子,2因子能够共同解释非淹水期苔草湿地65%-74%的CH₄通量变异;4)试验期间,苔草湿地CH₄释放量约为12.77 gC/m²,相当于同期土壤有机质分解碳排放量的4%,甲烷释放的碳消耗不足苔草湿地年NPP的1%。     

干旱胁迫下4种常用植物幼苗的光合和荧光特性综合评价

老鸭嘴,山毛豆,假连翘和葛藤是常见的矿山植被恢复植物。采用盆栽控制土壤水分的方法,测定了这 4 种幼苗叶片的光合和荧光指标,以期为矿山植被的恢复植物筛选提供依据。结果表明（1）干旱胁迫处理期间,4 种幼苗叶片的净光合速率（P_n）、气孔导度（G_s）、蒸腾速率（T_r）和胞间CO₂ 浓度（C_i）持续下降,复水8 d后P_n均显著增加,老鸭嘴和山毛豆的G_s和T_r恢复到对照水平,山毛豆和葛藤的C_i恢复到对照水平;假连翘幼苗叶片的气孔限制值（L_s）随干旱胁迫时间的延长而逐渐增加,其他3 种幼苗略降后增加,复水8 d后均恢复到对照水平。（2）干旱胁迫处理期间,4 种幼苗叶片的photosystemⅡ（PSⅡ）光能捕获效率（F_v’/F_m’）、实际光化学量子效率（Yield）和表观电子传递速率（ETR）持续降低,8 d时显著小于对照。复水8 d后,F_v’/F_m’小幅波动,Yield和ETR有所恢复,其中葛藤的Yield和山毛豆的ETR恢复到对照水平;4种幼苗叶片的非化学淬灭（NPQ）持续上升,在第8天显著大于对照,复水8 d后,均持续下降,其中葛藤恢复到对照水平。（3）对光合和荧光指标进行主成分分析表明,干旱胁迫下4 种幼苗的抗旱性能大小顺序为山毛豆 > 老鸭嘴 > 假连翘 > 葛藤。     

胡杨异形叶叶绿素荧光特性对高温的响应

胡杨(Populus euphratica Oliv)是塔里木极端干旱荒漠区优势乔木树种,由于其生长在荒漠环境中,极端高温远高于其它地区,因此研究胡杨对高温胁迫的响应特征对于解释胡杨的抗逆机理与生态适应策略具有极为重要的意义。以胡杨3种典型的异形叶为材料,研究不同温度对其叶绿素荧光特性和能量分配的影响。结果表明,25-45 ℃温度下胡杨异形叶的F₀、F_m、F_v/F_m与F_v/F₀变化不大,尤其F_v/F_m仍能保持在0.78左右,光合反应正常;高温胁迫下(＞45 ℃)F_m、F_v/F₀、F_v/F_m、F'_v/F'_m、qP、ΦPSⅡ、P和ETR均大幅降低;F₀、qN、E显著上升,而D先上升后下降,说明高温抑制了PSⅡ的功能,使PSⅡ反应中心活性下降,QA^-的还原速率加快,光化学电子传递速率降低,某些能量耗散途径受阻,影响了PSⅠ和PSⅡ激发能的平衡分配,最终导致光合机构受损、光合速率降低。胡杨3种异形叶的叶绿素荧光参数随温度升高变幅不同,高温处理下锯齿阔卵形叶各参数均高于卵形叶与条形叶,表明锯齿阔卵形叶比卵形叶和条形叶具有更强的高温耐受能力。用模糊数学的隶属度函数对胡杨3种异形叶的耐热性进行综合评价,锯齿阔卵形叶的耐热性最强。     

Phylogeny of Phaeomollisia piceae gen. sp. nov.: a dark, septate, conifer-needle endophyte and its relationships to Phialocephala and Acephala

Dark, septate endophytes (DSE) were isolated from roots and needles of dwarf Picea abies and from roots of Vaccinium spp. growing on a permafrost site in the Jura Mountains in Switzerland. Two of the isolates sporulated after incubation for more than one year at 4 °C. One of them was a hitherto undescribed helotialean ascomycete Phaeomollisia piceae gen. sp. nov., the other was a new species of Phialocephala, P. glacialis sp. nov. Both species are closely related to DSE of the Phialocephala fortinii s. lat.-Acephala applanata species complex (PAC) as revealed by phylogenetic analyses of the ITS and 18S rDNA regions. Morphologically dissimilar fungi, such as Vibrissea and Loramyces species, are phylogenetically also closely linked to the new species and the PAC. Cadophora lagerbergii and C. (Phialophora) botulispora are moved to Phialocephala because Phialocephala dimorphospora and P. repens are the closest relatives. Several Mollisia species were closely related to the new species and the PAC according to ITS sequence comparisons. One DSE from needles of Abies alba and one from shoots of Castanea sativa formed Cystodendron anamorphs in culture. Their identical 18S sequences and almost identical ITS sequences indicated Mollisia species as closest relatives, suggesting that Mollisia species are highly euryoecious.     

Genetic variation and relationships in Laetiporus sulphureus s. lat., as determined by ITS rDNA sequences and in vitro growth rate

The aim of this study was to characterise the genetic variation and molecular relationships of the brown rot polypore, Laetiporus sulphureus s. lat., from Europe, South America, Africa, and Asia, using ITS sequences of the nu-rDNA and by comparing the growth rate in vitro. In a NJ analysis of the sequences of 130 individuals of L. sulphureus s. lat., eight distinct clusters emerged, supported by BS values of 70-100 %. Within each cluster, the ITS rDNA sequence variation was below 3 %. The sequences were also analysed together with Laetiporus sequences available from GenBank. Results demonstrated the possible presence of L. huroniensis in Europe (invalidly named L. montanus) and L. gilbertsonii in South America, and provided more information on the Pan-American and European distribution of one of the clades, currently known in North America as L. sulphureus. L. conifericola formed a separate distinct clade. Moreover, the analysis revealed two unknown Laetiporus taxa in Korea, one in South Africa, and one in Europe. As L. sulphureus is described from Europe (France), and we show that more than one taxon exist here, it is presently not possible to define L. sulphureus s. str. Certain biological differences between some of the clades (in vitro growth rates, chemical composition, and pigmentation) were demonstrated and discussed.     

Essential oil composition of Hypericum L. species from Southeastern Serbia and their chemotaxonomy

The essential oils of the aerial parts of nine species of Hypericum (Hypericum barbatum, Hypericum hirsutum, Hypericum linarioides, Hypericum maculatum, Hypericum olympicum, Hypericum perforatum, Hypericum richeri, Hypericum rumeliacum and Hypericum tetrapterum), collected from different locations in Southeast Serbia, were obtained by steam distillation and analyzed by GC and GC–MS. The essential oils investigated were characterized by a high content of non-terpene compounds and a low content of monoterpenes. The contents of non-terpenes, monoterpenes and sesquiterpenes in oils of the species H. barbatum, H. richeri and H. rumeliacum (section Drosocaprium) were similar and these oils were characterized by high contents of fatty acids. The oils of H. hirsutum and H. linarioides (section Taeniocarpium) contained a high percentage of n-nonane. There were similarities in contents of non-terpenes and sesquiterpenes in oils of species that belong to the section Hypericum (H. maculatum, H. perforatum and H. tetrapterum). The oil of H. olympicum differed from others by higher terpene content. A comparison was also carried out of the chemical composition of the essential oils from flower, leaf and stem of H. perforatum and it revealed that the highest concentration of non-terpene compounds was found in the flower and stem oil, while a high concentration of sesquiterpenes was characteristic for leaf oil. There were significant differences in the concentrations of the same compounds in the essential oils of H. maculatum, H. olympicum and H. perforatum, collected in different years from the same location which could be explained by seasonal differences. All data were statistically processed with principal component analysis and cluster analysis. The main conclusion from the above data is that genetic and environmental factors both play a role in determining the composition of essential oils of the Hypericum species studied.     

Laboratory evaluation of the chemosterilant lufenuron against the fruit flies Ceratitis capitata, Bactrocera dorsalis, B. cucurbitae, and B. latifrons

Four species of tephritid fruit flies, Ceratitis capitata, Bactrocera dorsalis, B. cucurbitae, and B. latifrons were evaluated for toxic, developmental, and physiological responses to the chemosterilant lufenuron. No significant mortality of laboratory strains of the first three species was observed after their exposure up to 50 μg/mL of lufenuron in agar adult diet, whereas B. latifrons adults fed with 50 μg/mL of lufenuron in the diet caused significant mortality compared to the control. Fertility of C. capitata adults fed on 50 μg/mL lufenuron-fortified diet between 7 and 12 days of age was approximately 46% of the no lufenuron control. Fertility of B. dorsalis and B. latifrons adults fed on 50 μg/mL lufenuron-incorporated diet was about 45% and 62% of the control, respectively. Lufenuron did not significantly affect fertility of B. cucurbitae adults. Lufenuron did not affect fecundity of C. capitata and B. dorsalis. Fecundity of B. cucurbitae and B. latifrons was not evaluated due to difficulty to count the eggs laid deep in the agar diet. Larvae fed on a liquid larval diet with ≤ 0.1 μg/mL of lufenuron were also evaluated. Pupal recovery, adult emergence, adult fliers, mating, egg hatch, and egg production of C. capitata were significantly decreased, while for B. dorsalis, pupal recovery, larval duration and adult emergence were affected. No effect of lufenuron on B. cucurbitae larvae was observed. B. latifrons was not performed because shortage of eggs at the time of this research. Lufenuron is a potential agent for management and control of C. capitata and B. dorsalis.     

Prenylated acetophenones from Melicope obscura and Melicope obtusifolia ssp. obtusifolia var. arborea and their distribution in Rutaceae

Four prenylated acetophenones 2,6-dihydroxy-4-geranyloxyacetophenone (1), 4-geranyloxy-2,6,β-trihydroxyacetophenone (2), 2,6-dihydroxy-4-geranyloxy-3-prenylacetophenone (3), and 4-geranyloxy-3-prenyl-2,6,β-trihydroxyacetophenone (4) have for the first time been isolated from Melicope obscura (1 and 2) and Melicope obtusifolia ssp. obtusifolia var. arborea (3 and 4). The distribution of prenylated acetophenones in Rutaceae is reviewed and the results, including the new records, indicate that prenylated acetophenones are valuable as chemotaxonomic markers for the subfamily Rutoideae, tribe Xanthoxyleae sensu Engler.     

Description of Brenneria roseae sp. nov. and two subspecies, Brenneria roseae subspecies roseae ssp. nov and Brenneria roseae subspecies americana ssp. nov. isolated from symptomatic oak

Gram-negative, facultatively anaerobic bacteria were isolated from symptomatic oak tissue in the UK and USA. Partial gyrB sequencing placed ten strains in the genus Brenneria, with B. goodwinii as the closest phylogenetic relative. The strains were investigated further using a polyphasic approach including MLSA (based on partial gyrB, rpoB, infB and atpD gene sequences), 16S rRNA gene sequencing, DNA–DNA relatedness studies and both phenotypic and chemotaxonomic assays. The MLSA and 16S rRNA gene analyses separated the strains into two groups based on origin, suggesting that they belong to Brenneria as two novel species. However, the DNA–DNA relatedness values revealed a closer relationship between the groups and indicated that they should belong to the same species. As the two groups of strains from the UK and USA can be differentiated from each other phenotypically and by ERIC PCR fingerprints, it is proposed to classify them as novel subspecies of a novel Brenneria species. The name Brenneria roseae sp. nov. (FRB 222^T = LMG 27714^T = NCPPB 4581^T) is proposed, with Brenneria roseae subsp. roseae ssp. nov. (FRB 222^T = LMG 27714^T = NCPPB 4581^T) for the strains from the UK and Brenneria roseae subsp. americana ssp. nov. (FRB 223^T = LMG 27715^T = NCPPB 4582^T) for the strains from the USA.     

The anchor cell initiates dorsal lumen formation during C. elegans vulval tubulogenesis

Tubulogenesis and lumen formation are critical to the development of most organs. We study Caenorhabditis elegans vulval and uterine development to probe the complex mechanisms that mediate these events. Development of the vulva and the ventral uterus is coordinated by the inductive cell-signaling activity of a gonadal cell called the anchor cell (AC). We demonstrate that in addition to its function in specifying fate, the AC directly promotes dorsal vulval tubulogenesis. Two types of mutants with defective anchor cell behavior reveal that anchor cell invasion of the vulva is important for forming the toroidal shape of the dorsal vulval cell, vulF. In fos-1 mutants, where the AC cannot breakdown the basement membranes between the gonad and the vulva, and in mutants in unc-6 netrin or its receptor unc-40, which cause AC migration defects, the AC fails to invade the vulva and no lumen is formed in vulF. By examining GFP markers of dorsal vulval cell fate, we demonstrate that fate specification defects do not account for the aberrant vulF shape. We propose that the presence of the AC in the center of the developing vulF toroid is required for dorsal vulval lumen formation to complete vulval tubulogenesis.     

Chemical markers in Veronica sect. Hebe. II

In a continued chemosystematic investigation of the water-soluble compounds in Veronica sect. Hebe, four additional species were investigated. In comparison to other, Northern Hemisphere (NH) species of Veronica, those belonging to the New Zealand species in sect. Hebe are apparently more variable in chemical content. In addition to the compounds characteristic for NH Veronica, namely mannitol, aucubin, catalpol and 6-O-esters of catalpol as well as some caffeoyl phenylethanoid glucosides (CGPs), Veronica topiaria (syn. Hebe topiaria) also gave an unusual 6-O-ester of aucubin named topiarioside. The former Hebe species Veronica cupressoides and Veronica stenophylla each provided one of the two previously undescribed disaccharide esters named hebitol I and II, respectively, and the former plant also provided a CPG named cuproside, a 6-O-β-glucopyranosyl derivative of the known hebeoside. The last species, namely Veronica hulkeana (syn. Heliohebe hulkeana) only contained compounds common to other species of Veronica. The taxonomic results are discussed and it is concluded that carbohydrate esters are common in sect. Hebe. The data so far obtained indicate that the occurrences of esters of 6-O-rhamnopyranosylcatalpol are confined to the most derived species in the section.