缢蛏滩涂养殖环境的细菌群落组成及分析

1引言滩涂贝类养殖由于实行生态养殖,目前已成为宁波市海水养殖业的一大热点.近几年来,随着沿海经济的高速发展,滩涂环境污染问题日趋严重,养殖贝类大规模死亡的现象时有发生,给海洋水产经济造成了严重损失.而微生物数量及种群分布与区域环境有着密切联系,它可直接或间接地反映区域环境的营养水平、污染程度及底质状况等.目前有关养殖环境微生物学的研究主要是针对水体及养殖池底泥中的细菌[1,4,7～10],而对养殖滩涂环境细菌的研究很少[15,18,19].为此,本文以宁波市昆亭缢蛏滩涂养殖区为研究对象,从细菌生态学角度对滩涂缢蛏养殖环境进行研…     

池塘氮循环中各种细菌与理化因子的相关性研究

对精养鱼池的水体及淤泥进行多次采样,用统计学分析处理,结果表明：硝化类细菌总体上与各主要理化因子的相关性较强（R＞0．60）,其中氨化菌主要与溶氧相关程度较高;反硝化菌主要与有效磷相关程度较高;亚硝化菌主要与氨氮负相关程度较高;硝化菌主要与亚硝酸盐相关程度较高。硝化类细菌与异养菌之间的相关性则更强,如反硝化菌与厌养菌（R＝0．944,P＝0．001）、拓硝化菌和好氧异养菌（R＝0．832,P＝0．003）皆显正相关,亚硝化菌和厌氧异养菌（R＝－0．76,P＝0．009）显负相关;而在硝化类细菌之间的相关程度却较弱（R＜0．60）,表明池塘硝化类细菌对水质具有一定的调控作用,但相互之间的依赖性不强,各自相对独立地发挥作用。     

The influence of mineral fertilizer combined with a nitrification inhibitor on microbial populations and activities in calcareous Uzbekistanian soil under cotton cultivation

Application of fertilizers combined with nitrification inhibitors affects soil microbial biomass and activity. The objective of this research was to determine the effects of fertilizer application combined with the nitrification inhibitor potassium oxalate (PO) on soil microbial population and activities in nitrogen-poor soil under cotton cultivation in Uzbekistan. Fertilizer treatments were N as urea, P as ammophos, and K as potassium chloride. The nitrification inhibitor PO was added to urea and ammophos at the rate of 2%. Three treatments--N200 P140 K60 (T1), N200 PO P140 K60 (T2), and N200 P140 PO K60 (T3) mg kg(-1) soil--were applied for this study. The control (C) was without fertilizer and PO. The populations of oligotrophic bacteria, ammonifying bacteria, nitrifying bacteria, denitrifying bacteria, mineral assimilating bacteria, oligonitrophilic bacteria, and bacteria group Azotobacter were determined by the most probable number method. The treatments T2 and T3 increased the number of oligonitrophilic bacteria and utilization mineral forms of nitrogen on the background of reducing number of ammonifying bacteria. T2 and T3 also decreased the number of nitrifying bacteria, denitrifying bacteria, and net nitrification. In conclusion, our experiments showed that PO combined with mineral fertilizer is one of the most promising compounds for inhibiting nitrification rate, which was reflected in the increased availability and efficiency of fertilizer nitrogen to the cotton plants. PO combined with mineral fertilizer has no negative effects on nitrogen-fixing bacteria Azotobacter and oligo-nitrophilic bacteria.     

Inactivation by ionizing radiation of Salmonella enteritidis serotype montevideo grown in composed sewage sludge.

S. enteritidis ser. montevideo were grown in composted sewage sludge to levels of approximately 10(9)/g. These bacteria were found to be inactivated by ionizing radiation at approximately the same rate (30 krads/log) as Salmonella species in liquid digested sludge.     

黄土丘陵区不同植被土壤氮素转化微生物生理群特征及差异

采用最大或然计数法(most probable number, MPN)对黄土高原洞子沟流域不同植被恢复阶段土壤氮素微生物生理群(氨化细菌、亚硝化细菌、反硝化细菌)数量分布特征进行了测定,结果表明:1)土壤氨化细菌、亚硝化细菌和反硝化细菌数量随植被恢复而增加,三者最大值分别为最小值的74、4和31倍,其中氨化细菌和反硝化细菌的数量在铁杆蒿群落最低,辽东栎群落最高,亚硝化细菌数量在丁香群落最低,辽东栎群落最高;2)植被恢复对各氮素生理群影响不同,对氨化细菌影响最大,其次分别为反硝化细菌和亚硝化细菌;3)各氮素生理群数量差异较大,氨化细菌>反硝化细菌>亚硝化细菌。研究区氨化细菌占总数的75%-80%,反硝化细菌占20%-25%时,生态系统最为稳定;4)土壤理化性质与各功能菌关系紧密,其中,土壤容重和硝态氮含量与微生物数量相关性最大,全钾、矿化氮和微生物量氮也表现出很大的相关性。     

Cultivation-independent, semiautomatic determination of absolute bacterial cell numbers in environmental samples by fluorescence in situ hybridization.

Fluorescence in situ hybridization (FISH) with rRNA-targeted oligonucleotide probes has found widespread application for analyzing the composition of microbial communities in complex environmental samples. Although bacteria can quickly be detected by FISH, a reliable method to determine absolute numbers of FISH-stained cells in aggregates or biofilms has, to our knowledge, never been published. In this study we developed a semiautomated protocol to measure the concentration of bacteria (in cells per volume) in environmental samples by a combination of FISH, confocal laser scanning microscopy, and digital image analysis. The quantification is based on an internal standard, which is introduced by spiking the samples with known amounts of Escherichia coli cells. This method was initially tested with artificial mixtures of bacterial cultures and subsequently used to determine the concentration of ammonia-oxidizing bacteria in a municipal nitrifying activated sludge. The total number of ammonia oxidizers was found to be 9.8 x 10(7) +/- 1.9 x 10(7) cells ml(-1). Based on this value, the average in situ activity was calculated to be 2.3 fmol of ammonia converted to nitrite per ammonia oxidizer cell per h. This activity is within the previously determined range of activities measured with ammonia oxidizer pure cultures, demonstrating the utility of this quantification method for enumerating bacteria in samples in which cells are not homogeneously distributed.     

Anaerobic mineralization of quaternary carbon atoms: isolation of denitrifying bacteria on dimethylmalonate.

The microbial capacity to degrade simple organic compounds with quaternary carbon atoms was demonstrated by enrichment and isolation of five denitrifying strains on dimethylmalonate as the sole electron donor and carbon source. Quantitative growth experiments showed a complete mineralization of dimethylmalonate. According to phylogenetic analysis of the complete 16S rRNA genes, two strains isolated from activated sewage sludge were related to the genus Paracoccus within the alpha-Proteobacteria (98.0 and 98.2% 16S rRNA gene similarity to Paracoccus denitrificans(T)), and three strains isolated from freshwater ditches were affiliated with the beta-Proteobacteria (97.4 and 98.3% 16S rRNA gene similarity to Herbaspirillum seropedicae(T) and Acidovorax facilis(T), respectively). Most-probable-number determinations for denitrifying populations in sewage sludge yielded 4.6 x 10(4) dimethylmalonate-utilizing cells ml(-1), representing up to 0.4% of the total culturable nitrate-reducing population.     

生物反应器填埋场系统渗滤液的脱氮性能

利用填埋场垂直分布的好氧-缺氧-厌氧的独特生态环境,并采用填埋垃圾上层间歇曝气充氧的方式,研究了生物反应器填埋场系统渗滤液的脱N性能.结果表明,填埋垃圾上层间歇曝气充氧,促进了填埋垃圾层硝化细菌和反硝化细菌的生长,且可使反硝化细菌的数量比普通的填埋垃圾层高4～13个数量级,硝化细菌的最大数量可达到10⁹个·g^-1;营建了填埋场内硝化、反硝化等脱N反应的生物环境,有利于回流渗滤液含N化合物的去除.试验结束时,其渗滤液NH₄⁺-N和TN浓度分别为186和289 mg·L^-1,仅为对照的18%和26％.此外,填埋垃圾上层间歇曝气充氧也有利于填埋垃圾的降解,提高垃圾的稳定化效果.     

Effect of seeding during thermophilic composting of sewage sludge

The effect of seeding on the thermophilic composting of sewage sludge was examined by measuring the changes in CO2 evolution rates and microbial numbers. Although the succession of thermophilic bacteria and thermophilic actinomycetes clearly reflected the effect of seeding, no clear difference was observed in the overall rate of composting or quality of the composted product.     

Molecular and ecological analyses of microbial community structures in biofilms of a full-scale Aerated Up-Flow Biobead process

Molecular and cultivation techniques were used to characterize the bacterial communities of biobead reactor biofilms in a sewage treatment plant to which an Aerated Up-Flow Biobead process was applied. With this biobead process, the monthly average values of various chemical parameters in the effluent were generally kept under the regulation limits of the effluent quality of the sewage treatment plant during the operation period. Most probable number (MPN) analysis revealed that the population of denitrifying bacteria was abundant in the biobead #1 reactor, denitrifying and nitrifying bacteria coexisted in the biobead #2 reactor, and nitrifying bacteria prevailed over denitrifying bacteria in the biobead #3 reactor. The results of the MPN test suggested that the biobead #2 reactor was a transition zone leading to acclimated nitrifying biofilms in the biobead #3 reactor. Phylogenetic analysis of 16S rDNA sequences cloned from biofilms showed that the biobead #1 reactor, which received a high organic loading rate, had much diverse microorganisms, whereas the biobead #2 and #3 reactors were dominated by the members of Proteobacteria. DGGE analysis with the ammonia monooxygenase (amoA) gene supported the observation from the MPN test that the biofilms of September were fully developed and specialized for nitrification in the biobead reactor #3. All of the DNA sequences of the amoA DGGE bands were very similar to the sequence of the amoA gene of Nitrosomonas species, the presence of which is typical in the biological aerated filters. The results of this study showed that organic and inorganic nutrients were efficiently removed by both denitrifying microbial populations in the anaerobic tank and heterotrophic and nitrifying bacterial biofilms well-formed in the three functional biobead reactors in the Aerated Up-Flow Biobead process.     

Inactivation by Ionizing Radiation of Salmonella enteritidis Serotype montevideo Grown in Composed Sewage Sludge

S. enteritidis ser. montevideo were grown in composted sewage sludge to levels of approximately 10⁹/g. These bacteria were found to be inactivated by ionizing radiation at approximately the same rate (30 krads/log) as Salmonella species in liquid digested sludge.     

Activity of microorganisms in activated sludge in relation to the concentration of mercury in sewage

Activated sludge from the refinery of a plant producing synthetic rubber was shown to adsorb mercury ions contained in the sewage. As a result, the content of microorganisms in the sludge as well as the activity of dehydrogenase decreased. The quantity of Pseudomonas aeruginosa cells was 40 x 10(6) per 1 ml in a chemically defined medium without mercury ions, 20 x 10(5) per 1 ml in the water after the first sedimentation tank (0.07 to 0.08 mg Hg per litre), and 10 x 10(6) per 1 ml in the water of sewage common to the whole plant (0.13 to 0.14 mg Hg per litre). The use of the phage-resistant P-aeruginosa T-76 culture adapted to synthetic organic pollutants contained in the sewage of the plant together with the activated sludge of the refinery increased the biological activity of the sludge and improved the quality of purification in terms of dehydrogenase activity and chemical uptake of oxygen.     

Sanitary analysis of crude and composted activated sludge.

This paper presents a sanitary analysis of crude and composted activated sewage sludge from the sewage treatment plants in Slawa near Zielona G6ra in the period from September to December 1999. The composts were made from crude active sludge collected on 16.08.1999 with the addition of sawdust in a proportion of 15% with respect to dry sludge matter, and left in film bags with a capacity of 10 litres. Composting was carried out at a temperature of 20 degrees C for 8 weeks. Sanitary analysis of the activated sludge showed that the total quantity of bacteria, fungi, pathogenic bacteria and parasite ova was higher in the summer period compared to the spring period. The quantity of pathogenic bacteria and parasite eggs exceeded the permitted level in activated sludge designated for agricultural use. Analysis of the suitability of composts made from crude activated sludge showed that the quantity of pathogenic bacteria and parasite ova had been reduced, but was still higher than the permitted value for sewage sludge designated for agricultural use.     

Communities of ammonia-oxidizing bacteria in activated sludge of various sewage treatment plants in Tokyo

We investigated ammonia-oxidizing bacteria in activated sludge collected from 12 sewage treatment systems, whose ammonia removal and treatment processes differed, during three different seasons. We used real-time PCR quantification to reveal total bacterial numbers and total ammonia oxidizer numbers, and used specific PCR followed by denaturing gel gradient electrophoresis, cloning, and sequencing of 16S rRNA genes to analyze ammonia-oxidizing bacterial communities. Total bacterial numbers and total ammonia oxidizer numbers were in the range of 1.6 x 10(12) - 2.4 x 10(13) and 1.0 x 10(9) - 9.2 x 10(10)cellsl(-1), respectively. Seasonal variation was observed in the total ammonia oxidizer numbers, but not in the ammonia-oxidizing bacterial communities. Members of the Nitrosomonas oligotropha cluster were found in all samples, and most sequences within this cluster grouped within two of the four sequence types identified. Members of the clusters of Nitrosomonas europaea-Nitrosococcus mobilis, Nitrosomonas cryotolerans, and unknown Nitrosomonas, occurred solely in one anaerobic/anoxic/aerobic (A2O) system. Members of the Nitrosomonas communis cluster occurred almost exclusively in association with A2O and anaerobic/aerobic systems. Solid residence time mainly influenced the total numbers of ammonia-oxidizing bacteria, whereas dissolved oxygen concentration primarily affected the ammonia-oxidizing activity per ammonia oxidizer cell.     

Watering, Fertilization, and Slurry Inoculation Promote Recovery of Biological Crust Function in Degraded Soils

Biological soil crusts are very sensitive to human-induced disturbances and are in a degraded state in many areas throughout their range. Given their importance in the functioning of arid and semiarid ecosystems, restoring these crusts may contribute to the recovery of ecosystem functionality in degraded areas. We conducted a factorial microcosm experiment to evaluate the effects of inoculation type (discrete fragments vs slurry), fertilization (control vs addition of composted sewage sludge), and watering frequency (two vs five times per week) on the cyanobacterial composition, nitrogen fixation, chlorophyll content, and net CO₂ exchange rate of biological soil crusts inoculated on a semiarid degraded soil from SE Spain. Six months after the inoculation, the highest rates of nitrogen fixation and chlorophyll a content were found when the biological crusts were inoculated as slurry, composted sewage sludge was added, and the microcosms were watered five times per week. Net CO₂ exchange rate increased when biological crusts were inoculated as slurry and the microcosms were watered five times per week. Denaturing gradient gel electrophoresis fingerprints and phylogenetic analyses indicated that most of the cyanobacterial species already present in the inoculated crust had the capability to spread and colonize the surface of the surrounding soil. These analyses showed that cyanobacterial communities were less diverse when the microcosms were watered five times per week, and that watering frequency (followed in importance by the addition of composted sewage sludge and inoculation type) was the treatment that most strongly influenced their composition. Our results suggest that the inoculation of biological soil crusts in the form of slurry combined with the addition of composted sewage sludge could be a suitable technique to accelerate the recovery of the composition and functioning of biological soil crusts in drylands.     

脱氮微生物及脱氮工艺研究进展

脱氮是大部分污水处理系统中不可缺少的一环。由于具有经济高效、工艺简单和无二次污染等显著优势,生物脱氮工艺在最近数十年中备受关注。根据脱氮微生物的生理特性和脱氮机制不同,文中分类综述了近年来生物脱氮工艺的研究进展,重点对比分析了硝化菌、反硝化菌和厌氧氨氧化菌以及以这些菌为基础的不同生物脱氮工艺的优缺点,为复杂污水环境的脱氮工艺选择提供参考。基于微生物脱氮机制,通过合成生物学技术开发高效脱氮菌株,结合不同工艺优点并应用自动化模拟最佳条件,从而建立经济高效的脱氮工艺将是未来发展的重要方向。     

Evaluation of abundance of aerobic bacteria in the rhizosphere of transgenic and non-transgenic alfalfa lines

Fourteen genetically modified lines of alfalfa (Medicago sativa) containing the gene Ov from Japanese quail, coding for a methionine-rich protein ovalbumin, were evaluated for nodulation ability and concentration of aerobic bacteria in the rhizosphere. The transgenic lines were derived from a highly regenerable genotype Rg9/I-14-22, selected from cv. Lucia. On selective media, a higher concentration of ammonifying bacteria, bacterial spores, denitrifying and nitrifying bacteria were observed in the rhizosphere of transgenic clonesand, on the other hand, lower concentration of cellulolytic bacteria and Azotobacter spp. compared with the rhizosphere of non-transgenic clone SE/22-GT2. A statistically significant difference in the concentration of all the bacterial types was found between samples taken from two types of substrates (i.e. sterile vs. nonsterile). Higher bacterial concentration (measured as colony forming units per g soil dry mass) were observed for all tested groups of culturable bacteria in the non-sterile substrate. The presence of Azotobacter spp. was found only in the rhizosphere of plants grown in non-sterile soil in which the highest number of fertile soil particles (97 %) was observed in transgenic clones SE/22-9-1-12 and SE/22-11-1-1S.1. Concentration of bacteria involved in the N cycle in the soil was increased in the rhizosphere of transgenic clones and decreased in the rhizosphere of non-transgenic plants compared with the average value. In spite of some differences in colony numbers in samples isolated from the root rhizosphere of transgenic and nontransgenic alfalfa plants, we could not detect any statistically significant difference between individual lines.     

Heavy metals in sewage sludges contribute to their adverse effects on the arbuscular mycorrhizal fungus<Emphasis Type="Italic">Glomus mosseae</Emphasis>

Applying sewage sludges to agricultural land is a widespread practice because of the sludges’ agronomic value as a source of plant nutrients and organic matter. Nevertheless, sludges often contain micropollutants that can constitute a menace for health and the environment. Arbuscular mycorrhizal fungi are sensitive to sewage sludges that have been spiked, or not, with metallic trace elements (MTE). Here we have investigated if MTE in sewage sludges could be responsible for effects on mycorrhizal development betweenGlomus mosseae andMedicago truncatula. The impact of a dehydrated or composted urban sewage sludge spiked or not with MTE, was tested on spore germination and root colonization byG. mosseae. The sewage sludges depressed both the presymbiotic andin planta stages of development of the mycorrhizal fungus. This negative effect was more related to the metallic pollutant contents of the sludges than to the presence of antagonistic microorganisms or phosphorus.     

Competition for Ammonium between Nitrifying and Heterotrophic Bacteria in Continuously Percolated Soil Columns

Although the absence of nitrate formation in grassland soils rich in organic matter has often been reported, low numbers of nitrifying bacteria are still found in these soils. To obtain more insight into these observations, we studied the competition for limiting amounts of ammonium between the chemolithotrophic ammonium-oxidizing species Nitrosomonas europaea and the heterotrophic species Arthrobacter globiformis in the presence of Nitrobacter winogradskyi with soil columns containing calcareous sandy soil. The soil columns were percolated continuously at a dilution rate of 0.007 h^-1, based on liquid volumes, with medium containing 5 mM ammonium and different amounts of glucose ranging from 0 to 12 mM.A. globiformis was the most competitive organism for limiting amounts of ammonium. The numbers of N. europaea and N. winogradskyi cells were lower at higher glucose concentrations, and the potential ammonium-oxidizing activities in the uppermost 3 cm of the soil columns were nonexistent when at least 10 mM glucose was present in the reservoir, although 10⁷ nitrifying cells per g of dry soil were still present. This result demonstrated that there was no correlation between the numbers of nitrifying bacteria and their activities. The numbers and activities of N. winogradskyi cells decreased less than those of N. europaea cells in all layers of the soil columns, probably because of heterotrophic growth of the nitrite-oxidizing bacteria on organic substrates excreted by the heterotrophic bacteria or because of nitrate reduction at reduced oxygen concentrations by the nitrite-oxidizing bacteria. Our conclusion was that the nitrifying bacteria were less competitive than the heterotrophic bacteria for ammonium in soil columns but that they survived as viable inactive cells. Inactive nitrifying bacteria may also be found in the rhizosphere of grassland plants, which is rich in organic carbon. They are possibly reactivated during periods of net mineralization.     

长江口低氧区异养细菌及氮磷细菌分布

2009年8月15-28日,对长江口低氧高发海域的异养细菌、无机磷细菌、有机磷细菌、反硝化细菌和氨化细菌的空间分布特征进行初步研究.结果表明:氨化细菌数量最高,表层水、底层水和表层沉积物的数量均值分别为307.52×104个·L-1、184.50×104个·L-1和199.97×102个·g-1;其次为异养细菌,数量均值分别为87.35×104 cfu·L-1、86.85×104cfu·L-1和64.26×102cfu·g-1;再次为有机磷细菌,数量均值分别为19.26×104 cfu·L-1、18.82×104cfu·L-1和19.56×102cfu·g-1;无机磷细菌只分布在长江口内和河口南槽至舟山海域,数量均值分别为18.50×104cfu·L-1、31.00×104cfu·L-1和7.17×102cfu·g-1;反硝化细菌分布广,但数量较低,均值分别为3.94×104个·L-1、23.08×104个·L-1和6.22×102个·g-1.相关性分析结果说明:盐度、硝酸盐、磷酸盐、硅酸盐和pH是影响水体和表层沉积物异养细菌、磷细菌和反硝化细菌分布的主要因子;底层水和表层沉积物异养细菌、磷细菌与水温呈显著正相关;底层水异养细菌和有机磷细菌与溶解氧(DO)呈显著正相关;表层沉积物无机磷细菌与DO呈显著正相关,氨化细菌与DO呈显著负相关.聚类分析结果说明:低氧对表层沉积物的细菌群落结构产生影响.