Contributions of ammonia-oxidizing archaea and bacteria to nitrification in Oregon forest soils

Ammonia oxidation, the first step of nitrification, is mediated by both ammonia-oxidizing archaea (AOA) and bacteria (AOB); however, the relative contributions of AOA and AOB to soil nitrification are not well understood. In this study we used 1-octyne to discriminate between AOA- and AOB-supported nitrification determined both in soil-water slurries and in unsaturated whole soil at field moisture. Soils were collected from stands of red alder (Alnus rubra Bong.) and Douglas-fir (Pseudotsuga menziesii Mirb. Franco) at three sites (Cascade Head, the H.J. Andrews, and McDonald Forest) on acidic soils (pH 3.9–5.7) in Oregon, USA. The abundances of AOA and AOB were measured using quantitative PCR by targeting the amoA gene, which encodes subunit A of ammonia monooxygenase. Total and AOA-specific (octyne-resistant) nitrification activities in soil slurries were significantly higher at Cascade Head (the most acidic soils, pH < 5) than at either the H.J. Andrews or McDonald Forest, and greater in red alder compared with Douglas-fir soils. The fraction of octyne-resistant nitrification varied among sites (21–74%) and was highest at Cascade Head than at the other two locations. Net nitrification rates of whole soil without NH₄⁺ amendment ranged from 0.4 to 3.3 mg N kg⁻¹ soil d⁻¹. Overall, net nitrification rates of whole soil were stimulated 2- to 8-fold by addition of 140 mg NH₄⁺-N kg⁻¹ soil; this was significant for red alder at Cascade Head and the H.J. Andrews. Red alder at Cascade Head was unique in that the majority of NH₄⁺-stimulated nitrifying activity was octyne-resistant (73%). At all other sites, NH₄⁺-stimulated nitrification was octyne-sensitive (68–90%). The octyne-sensitive activity—presumably AOB—was affected more by soil pH whereas the octyne-resistant (AOA) activity was more strongly related to N availability.     

Fungal and bacterial N2O production regulated by soil amendments of simple and complex substrates

Fungal N₂O production results from a respiratory denitrification that reduces NO₃⁻/NO₂⁻ in response to the oxidation of an electron donor, often organic C. Despite similar heterotrophic nature, fungal denitrifiers may differ from bacterial ones in exploiting diverse resources. We hypothesized that complex C compounds and substances could favor the growth of fungi over bacteria, and thereby leading to fungal dominance for soil N₂O emissions. Effects of substrate quality on fungal and bacterial N₂O production were, therefore, examined in a 44-d incubation after soils were amended with four different substrates, i.e., glucose, cellulose, winter pea, and switchgrass at 2 mg C g⁻¹ soil. During periodic measurements of soil N₂O fluxes at 80% soil water-filled pore space and with the supply of KNO₃, substrate treatments were further subjected to four antibiotic treatments, i.e., no antibiotics or soil addition of streptomycin, cycloheximide or both so that fungal and bacterial N₂O production could be separated. Up to d 8 when antibiotic inhibition on substrate-induced microbial activity and/or growth was still detectable, bacterial N₂O production was generally greater in glucose- than in cellulose-amended soils and also in winter pea- than in switchgrass-amended soils. In contrast, fungal N₂O production was more enhanced in soils amended with cellulose than with glucose. Therefore, fungal-to-bacterial contribution ratios were greater in complex than in simple C substrates. These ratios were positively correlated with fungal-to-bacterial activity ratios, i.e., CO₂ production ratios, suggesting that substrate-associated fungal or bacterial preferential activity and/or growth might be the cause. Considering substrate depletion over time and thereby becoming limited for microbial N₂O production, measurements of soil N₂O fluxes were also carried out with additional supply of glucose, irrespective of different substrate treatments. This measurement condition might lead to potentially high rates of fungal and bacterial N₂O production. As expected, bacterial N₂O production was greater with added glucose than with added cellulose on d 4 and d 8. However, this pattern was broken on d 28, with bacterial N₂O production lower with added glucose than with added cellulose. In contrast, plant residue impacts on soil N₂O fluxes were consistent over 44-d, with greater bacterial contribution, lower fungal contribution, and thus lower fungal-to-bacterial contribution ratios in winter pea- than in switchgrass-amended soils. Real-time PCR analysis also demonstrated that the ratios of 16S rDNA to ITS and the copy numbers of bacterial denitrifying genes were greater in winter pea- than in switchgrass-amended soils. Despite some inconsistency found on the impacts of cellulose versus glucose on fungal and bacterial leading roles for N₂O production, the results generally supported the working hypothesis that complex substrates promoted fungal dominance for soil N₂O emissions.     

Fractionation of the reference inoculum of epizootic rabbit enteropathy in discontinuous sucrose gradient identifies aetiological agents in high density fractions

Epizootic rabbit enteropathy (ERE) is a major cause of economic loss in intensive rabbit production. Since its first recognition in 1997, much work has been done to determine the pathogenic mechanisms of the disease and to identify the aetiological agent(s). Unfortunately, the quest for aetiology has only met with limited success despite the ability to reproduce the syndrome by inoculation of intestinal contents from field cases. These intestinal inocula contain a huge number of microorganisms which could all be involved in the aetiology of ERE. To decrease the number of putative agents, the French reference inoculum TEC3 was fractionated on a discontinuous sucrose gradient so that seven fractions (supernatant, 10%, 20%, 30%, 40%, 50% and pellet) were obtained. Specific-pathogen-free rabbits were inoculated with three out of these seven fractions (supernatant, 30%, and pellet). The objectives were: (1) to characterise the seven fractions by bacteriological examination; (2) to verify whether the aetiological agent was present in the fractions by inoculation of rabbits; (3) to assign the aetiological agent of ERE to a morphological group of pathogens; (4) to identify a fraction which could replace the reference inoculum TEC3 in applications such as cell cultures or egg inoculation. The results strongly suggest that ERE is a bacterial disease and does not have a viral or parasitic aetiology.     

植烟土壤提取物质对烟株生长及根际土壤细菌多样性的影响

对盆栽烟草外源添加不同浓度植烟土壤提取物质(T1:40μg·mL-1;T2:120μg·mL-1;CK:蒸馏水对照),探讨植烟土壤提取物质对烟草生长及土壤细菌多样性的影响。结果表明,植烟土壤提取物处理使烟株生长受抑制,且随处理浓度的增加受抑制程度显著提高,具体表现为烟株变矮,叶面积变小,光合作用能力降低,且烟草的保护酶系统受到破坏,丙二醛含量随处理浓度加大而增加,T2处理的丙二醛含量是对照的3.44倍。对外源添加物质处理后烟草根际土壤微生物T-RFs分析发现,在对照检测到17个门24个纲,T1处理有14个门21个纲,T2有10个门17个纲;丰富度指数的变化也和门纲的变化一致,随着处理浓度的增加而显著降低。可见外源添加物质处理后,根际土壤细菌群落减少,多样性水平下降。对各处理的根际土壤微生物T-RFs变化与烟株生长变化进行相关性分析表明,在外源添加物质处理的土壤中存在较多的负相关T-RFs片段,且这些片段中较多为病原菌;而正相关的T-RFs片段主要存在于对照土壤中,其中有较多与土壤营养元素循环相关的微生物。本研究结果显示,在外源添加植烟土壤提取物质处理下,烟草的生长受抑制,烟草根际土壤的微生态受到破坏,且随浓度的提升而加重。因此,连作土壤中自毒物质的富集是造成烟草连作障碍的主要原因。关键词烟草连作障碍根际细菌自毒作用T-RFLP     

基于蛋白质反式剪接在大肠杆菌中表达鲎C因子CES多肽

鲎C因子是鲎血细胞中一种对内毒素具有高亲和力的丝氨酸蛋白酶原,可替代鲎试剂用于内毒素检测。鲎C因子N端Cys—EGF．sushil—sushi2（CES）片段中的sushil区域是与内毒素结合的关键部位。本研究在sushil结构域中的特定位点断开CES与内毒素结合的功能片段,然后分别和蛋白质内含肽SspDnaX的N端片段（IN）和C端片段0C）连接,并在大肠杆菌中表达,表达产物经亲和层析纯化、复性以及内毒素去除后,利用蛋白质反式剪接系统,在体外诱导该蛋白质内含肽发生剪接,重新得到C因子的CES蛋白,并检测其活性。实验结果表明此重组CES蛋白具有结合内毒素活性,提示在大肠杆菌系统中开发低成本内毒素检测试剂的可行性。     

Starving the soil of plant inputs for 50 years reduces abundance but not diversity of soil bacterial communities

If soil communities rely on plant-derived carbon, is biodiversity lost when this primary source is removed? Soil microbial and mesofaunal communities at the Rothamsted Highfield site were compared under a mixed grass sward, arable rotation and a section maintained as a bare-fallow for the past 50 years by regular tillage. Organic matter reserves have been degraded and microbial and mesofaunal numbers and mite diversity have declined in this unique bare-fallow site, where fresh carbon inputs have been drastically reduced. However, it supports a species-rich metabolically active bacterial community of similar diversity to that in soil maintained as grass sward. Thus in contrast to soil mesofauna, bacterial diversity (but not abundance) is apparently independent of plant inputs.     

The influence of soil properties on the structure of bacterial and fungal communities across land-use types

Land-use change can have significant impacts on soil conditions and microbial communities are likely to respond to these changes. However, such responses are poorly characterized as few studies have examined how specific changes in edaphic characteristics do, or do not, influence the composition of soil bacterial and fungal communities across land-use types. Soil samples were collected from four replicated (n = 3) land-use types (hardwood and pine forests, cultivated and livestock pasture lands) in the southeastern US to assess the effects of land-use change on microbial community structure and distribution. We used quantitative PCR to estimate bacterial–fungal ratios and clone libraries targeting small-subunit rRNA genes to independently characterize the bacterial and fungal communities. Although some soil properties (soil texture and nutrient status) did significantly differ across land-use types, other edaphic factors (e.g., pH) did not vary consistently with land-use. Bacterial–fungal ratios were not significantly different across the land-uses and distinct land-use types did not necessarily harbor distinct soil fungal or bacterial communities. Rather, the composition of bacterial and fungal communities was most strongly correlated with specific soil properties. Soil pH was the best predictor of bacterial community composition across this landscape while fungal community composition was most closely associated with changes in soil nutrient status. Together these results suggest that specific changes in edaphic properties, not necessarily land-use type itself, may best predict shifts in microbial community composition across a given landscape. In addition, our results demonstrate the utility of using sequence-based approaches to concurrently analyze bacterial and fungal communities as such analyses provide detailed phylogenetic information on individual communities and permit the robust assessment of the biogeographical patterns exhibited by soil microbial communities.     

2,4-二乙酰基藤黄酚产生菌在番茄根部的定殖及对番茄青枯病的防治

2,4-二乙酰基藤黄酚(2,4-DAPG)产生菌是荧光菌对土传病害进行生物防治的主要类群之一。室内筛选结果表明：供试的12个2,4-DAPG产生菌菌株对番茄青枯病菌均有不同程度的抑制作用,其中CPF、10抑菌效果最佳,抑菌带宽为3．5mm;而2P24、5J10次之,抑菌带宽为3．0mm。CPF10和2P24培养原液对番茄胚根的生长均有明显的抑制作用。CPF10和2P24根部定殖结果表明,两菌株均可在番茄幼苗根部大量定殖,根表细菌数量随着时间延长呈下降趋势但仍维持较高数量;而根内细菌数量有明显上升的趋势。CPF10和2P24对番茄青枯病均有一定的防治效果,其中2P24的防治效果最好。而CPF10在所有的处理中变异系数(CV)最小,防治效果最稳定。     

沼泽红假单胞菌的抗氧化酶活性和对紫外线的吸收作用

对不同培养方式下沼泽红假单胞菌的5个菌株的菌体的抗氧化酶活性以及培养液的细胞和上清液对紫外线吸收作用进行了研究。结果表明,5个菌株的活细胞CAT活性普遍以好氧黑暗条件下的培养物高于微好氧光照条件下的,SOD活性则以微好氧光照条件下的培养物高于好氧黑暗条件的。5个菌株在好氧黑暗、微好氧光照培养条件下的细胞液和培养液的上清液在紫外线200～300nm波长范围有明显的吸收峰。     

Effects of substrate sterilization,fungicide treatment,and mycorrhization helper bacteria on ectomycorrhizal formation of pedunculate oak (Quercus robur) inoculated with Laccaria laccata in two peat bare-root nurseries

Summary Pedunculate oak seedlings (Quercus robur) inoculated with the ectomycorrhizal fungus Laccaria lacata were grown for 1 year on fertilized sphagnum peat in two nurseries. Three factors affecting microbial populations in the substrate were studied, fungicide treatment of the seeds, peat disinfection before sowing (methyl bromide or steam pasteurization), and inoculation with mycorrhization helper bacteria. Treatment of acorns with Iprodione had no depressive effect on mycorrhiza formation. Both disinfection techniques were equivalent, stimulating or depressing mycorrhiza formation depending on the initial microflora in the peat. The introduction of two previously selected mycorrhization helper bacteria (one Pseudomonas fluorescens and one unidentified fluorescent pseudomonad), isolated from L. laccata sporocarps associated with Douglas fir—L. laccata ectomycorrhizas in other nurseries, significantly increased the mycorrhizal rate from 30 to 53% of the short roots. The implications of these results for the controlled mycorrhization of planting stocks and the specificity of mycorrhization helper bacteria are discussed.