结合代谢组学和转录组学分析恶臭假单胞菌Y-9主动稳定胞内外pH的机制

【目的】揭示恶臭假单胞菌(Pseudomonas putida) Y-9在氨氧化过程中主动调节胞外和胞内pH稳态机制。【方法】在初始pH为7.19和9.40的硝化培养基中培养Y-9生长48 h，利用代谢组学对比分析Y-9氨氧化过程中的显著差异代谢产物并预测解离常数(pKa)；结合转录组学对比分析Y-9氨氧化过程中的显著差异调控基因。【结果】Y-9在初始pH为7.19的相对酸性条件下，产生麦芽糖醇提高胞外pH；通过上调脱氨酶、脱亚胺酶和阳离子转运相关基因在相对酸性环境中的表达来维持细胞内pH稳定性。在初始pH为9.40的碱性条件下，产5-氨基戊酸3和草氨酸等有机酸及酸性物质降低胞外pH；通过调控NADH脱氢酶、细胞色素、ATP合酶和氨基酸转运相关基因的表达来维持细胞内酸度，应对碱性环境。【结论】本研究结果首先发现了Y-9具有稳定胞外pH的能力，探讨了其胞内pH稳态机制，拓展了对微生物与环境相互作用的认知，为进一步认识微生物脱氮过程中系统pH稳定机理提供了理论依据。     

Adaptation of Lactobacillus plantarum IMDO 130201, a wheat sourdough isolate, to growth in wheat sourdough simulation medium at different pH values through differential gene expression

Sourdough is a very competitive and challenging environment for microorganisms. Usually, a stable microbiota composed of lactic acid bacteria (LAB) and yeasts dominates this ecosystem. Although sourdough is rich in carbohydrates, thus providing an ideal environment for microorganisms to grow, its low pH presents a particular challenge. The nature of the adaptation to this low pH was investigated for Lactobacillus plantarum IMDO 130201, an isolate from a laboratory wheat sourdough fermentation. Batch fermentations were carried out in wheat sourdough simulation medium, and total RNA was isolated from mid-exponential-growth-phase cultures, followed by differential gene expression analysis using a LAB functional gene microarray. At low pH values, an increased expression of genes involved in peptide and amino acid metabolism was found as well as that of genes involved in plantaricin production and lipoteichoic acid biosynthesis. The results highlight cellular mechanisms that allow L. plantarum to function at a low environmental pH.     

Escherichia coli acid resistance: tales of an amateur acidophile

Gastrointestinal pathogens are faced with an extremely acidic environment. Within moments, a pathogen such as Escherichia coli O157:H7 can move from the nurturing pH 7 environment of a hamburger to the harsh pH 2 milieu of the stomach. Surprisingly, certain microorganisms that grow at neutral pH have elegantly regulated systems that enable survival during excursions into acidic environments. The best-characterized acid-resistance system is found in E. coli.     

Fungi in acidic fire: A potential source of industrially important enzymes

The microbial life that exists in harsh habitats of low pH possess several unique characteristics, which assign interesting qualities to these microorganisms and enable them to thrive in such a harsh environment. Among microorganisms inhabiting low pH environments, fungi are the second largest reported organisms. These acidophilic fungi are the main source of acid–stable enzymes that could be utilized in many industries including paper, leather making, food and feed industries, where the efficacy of commonly available enzymes is limited by challenges like stability and functional kinetics. The current review discusses the acidophilic fungi with emphasis on their diversity and pH homeostasis mechanisms adopted against low pH environments. In addition, an overview about the acid–stable enzymes obtained from these acidophilic fungi, their main sources and potential applications have also been discussed.     

产嗜铁素砷抗性微生物在砷污染环境中的作用

在砷污染环境中,许多微生物进化出了砷抗性,对地球环境中砷的命运起着决定性的作用。其次,由于自然条件下铁有效浓度低,微生物一般会表达嗜铁素,协助微生物吸收铁。嗜铁素除了与铁结合外,还可与多种金属离子形成稳定的复合物,促进环境中砷酸盐的溶解和亚砷酸盐的氧化。最后,产嗜铁素微生物有促进植物生长和促进或减弱植物吸收砷的可能性。因此,产嗜铁素砷抗性微生物可能具有在砷污染环境的修复中发挥作用的潜力。     

新型杀菌剂在造纸生产流程中的中试研究

造纸白水中的可溶性有机物以及造纸系统的pH值、温度等环境条件都极其有利于微生物生长。当白水中的微生物含量大于10~8CFU/mL,造纸系统就会产生腐浆,进而引起断纸、洞眼等问题。因此造纸系统要使用杀菌剂抑制微生物生长。本研究采用两种新型杀菌剂分别进行了实验室和中试抑菌试验,结果表明在两种杀菌剂协同作用下,造纸系统没有明显的腐浆产生。     

The 'pH optimum anomaly' of intracellular enzymes of Ferroplasma acidiphilum

A wide range of microorganisms, the so-called acidophiles, inhabit acidic environments and grow optimally at pH values between 0 and 3. The intracellular pH of these organisms is, however, close to neutrality or slightly acidic. It is to be expected that enzymatic activities dedicated to extracellular functions would be adapted to the prevailing low pH of the environment (0-3), whereas intracellular enzymes would be optimally active at the near-neutral pH of the cytoplasm (4.6-7.0). The genes of several intracellular or cell-bound enzymes, a carboxylesterase and three alpha-glucosidases, from Ferroplasma acidiphilum, a cell wall-lacking acidophilic archaeon with a growth optimum at pH 1.7, were cloned and expressed in Escherichia coli, and their products purified and characterized. The Ferroplasmaalpha-glucosidases exhibited no sequence similarity to known glycosyl hydrolases. All enzymes functioned and were stable in vitro in the pH range 1.7-4.0, and had pH optima much lower than the mean intracellular pH of 5.6. This 'pH optimum anomaly' suggests the existence of yet-undetected cellular compartmentalization providing cytoplasmic pH patchiness and low pH environments for the enzymes we have analysed.     

高寒草甸黄帚橐吾种群根际/非根际土壤可培养微生物群落特征

黄帚橐吾（Ligularia virgaurea）是高寒草甸常见的毒杂草,被认为是指示一个地区草地植被退化的重要物种,研究其根际/非根际土壤微生物在不同海拔梯度上的群落特征具有重要意义。以甘南州高寒草甸不同海拔梯度黄帚橐吾根际/非根际土壤可培养微生物为研究对象,采用稀释涂布平板法和最大可能数法（MPN）测定了土壤微生物的数量及土壤理化因子的变化。结果表明：细菌在微生物总数中占比最大,根际微生物数量随海拔升高呈先增加后减小的变化,非根际则表现为递增的趋势,微生物功能群在根际和非根际土壤中均逐渐增加;根际土壤的微生物和功能群数量均高于非根际土壤。RDA分析发现,土壤温度、有机碳、电导率、pH、全氮、全磷、速效氮及脲酶对根际/非根际土壤微生物数量及功能群变化影响较大。通径分析可知：根际土壤中,细菌和真菌受速效氮和有机碳影响较大,放线菌主要受土壤温度和电导率的影响;根际土壤固氮菌和氨化细菌决策系数速效氮 > 有机碳 > 全氮;根际和非根际土壤中硝化细菌的影响因子各不相同,根际土壤决策系数最大和最小分别为全磷和全氮,非根际则是pH和脲酶。     

Soil acidification and adaptations of plants and microorganisms in Bornean tropical forests

In tropical forest ecosystems, a paradoxical relationship is commonly observed between massive biomass production and low soil fertility (low pH). The loss and deficiency of soil phosphorus (P) and bases generally constrain biomass production; however, high productivity on nutrient-deficient soils of Bornean tropical forests is hypothesized to be maintained by plant and microorganism adaptation to an acidic soil environment. Proton budgets in the plant–soil system indicated that plants and microorganisms promote acidification to acquire bases, even in highly acidic tropical soils. The nitric and organic acids they produce contribute to the mobilization of basic cations and their uptake by plants. In response to soil P deficiency and the recalcitrance of lignin-rich organic matter, specific trees and fungi can release organic acids and enzymes for nutrient acquisition. Organic acids exuded by roots and rhizosphere microorganisms can promote the solubilization of P bonded to aluminum and iron oxides and its uptake by plants from P-poor soils. Lignin degradation, a rate-limiting step in organic matter decomposition, is specifically enhanced in acidic organic layers by lignin peroxidase, produced by white-rot fungi, which may solubilize recalcitrant lignin and release soluble aromatic substances into the soil solution. This dissolved organic matter functions in the transport of nitrogen, P, and basic cations in acidic soils without increasing leaching loss. In Bornean tropical forests, soil acidification is promoted by plants and microorganisms as a nutrient acquisition strategy, while plant roots and fungi can develop rhizosphere and enzymatic processes that promote tolerance of low pH.     

The effects of bioleaching conditions on the nonferrous metals content in copper-zinc concentrate

The effects of pH and ferrous iron concentration in cultural medium on the bioleaching of copper-zinc concentrate by mesophilic and moderately thermophilic acidophilic microorganisms were studied. It was revealed that the optimum pH for bioleaching in presence of 5 g/L of ferrous iron was 1.4–1.5. It was shown that bioleaching under optimal conditions led to an increase in the copper content in solid phase from 10.1 to 14% and a decrease in the zinc content from 7.4 to 1.4%. The results of the present work demonstrate that acidophilic microorganisms can be used for treatment of complex sulfide concentrates containing copper and zinc.     

Inorganic Species Distribution and Microbial Diversity within High Arctic Cryptoendolithic Habitats

Cryptoendolithic habitats in the Canadian high Arctic are associated with a variety of microbial community assemblages, including cyanobacteria, algae, and fungi. These habitats were analyzed for the presence of metal ions by sequential extraction and evaluated for relationships between these and the various microorganisms found at each site using multivariate statistical methods. Cyanobacteria-dominated communities exist under higher pH conditions with elevated concentrations of calcium and magnesium, whereas communities dominated by fungi and algae are characterized by lower pH conditions and higher concentrations of iron, aluminum, and silicon in the overlying surfaces. These results suggest that the activity of the dominant microorganisms controls the pH of the surrounding environment, which in turn dictates rates of weathering or the possibility for surface crust formation, both ultimately deciding the structure of microbial diversity for each cryptoendolithic habitat.     

Organotin compounds and their interactions with microorganisms.

Organotin compounds are ubiquitous in the environment. The general order of toxicity to microorganisms increases with the number and chain length of organic groups bonded to the tin atom. Tetraorganotins and inorganic tin have little toxicity. Because of their lipophilicity, organotins are regarded as membrane active. There is evidence that the site of action of organotins may be both at the cytoplasmic membrane and intracellular level. Consequently, it is not known whether cell surface adsorption or accumulation within the cell, or both is a prerequisite for toxicity. Biosorption studies on a fungus, cyanobacteria, and microalgae indicates that cell surface binding alone occurred in these organisms, while studies on the effects of TBT (tributyltin) on certain microbial enzymes indicated that in some bacteria TBT can interact with cytosolic enzymes. Microorganism-organotin interactions are influenced by environmental conditions. In aquatic systems, both pH and salinity can determine organotin speciation and therefore reactivity. These environmental factors may also alter selectivity for resistant microorganisms in polluted systems. Tin-resistant microorganisms have been identified, and resistance can be either plasmid or chromosomally mediated. In one TBT-resistant organism, an Altermonas sp., an efflux system was suggested as the resistance mechanism. Biotransformation of organotin compounds by debutylation or methylation has been observed. These reactions may influence the toxicity, mobility, and environmental fate of organotin compounds.     

Environmental mutagens may be implicated in the emergence of drug-resistant microorganisms

The emergence of drug-resistant microorganisms is an important medical and social problem. Drug-resistant microorganisms are thought to grow selectively in the presence of antibiotics. Most clinically isolated drug-resistant microorganisms have mutations in the target genes for the drugs. While any of the many mutagens in the environment may cause such genetic mutations, no reports have yet described whether these mutagens can confer drug resistance to clinically important microorganisms. We investigated how environmental mutagens might be implicated in acquired resistance to antibiotics in clinically important microorganisms, which causes human diseases. We selected mutagens found in the environment, in cigarette smoke, or in drugs, and then exposed Pseudomonas aeruginosa to them. After exposure, the incidence of rifampicin- and ciprofloxacin-resistant P. aeruginosa strains markedly increased, and we found mutations in genes for the antibiotic-target molecule. These mutations were similar to those found in drug-resistant microorganisms isolated from clinical samples. Our findings show that environmental mutagens, and an anticancer drug, are capable of inducing drug-resistant P. aeruginosa similar to strains found in clinical settings.     

土壤pH对玉米与微生物竞争吸收氨基酸的影响

化学合成肥料的大量使用导致土壤pH发生显著变化,但其对植物与根际微生物竞争吸收氨基酸的影响机制尚不明确.本试验通过电解法调节杭州红壤和铁岭棕壤两种土壤pH, 采用外源添加¹⁵N标记甘氨酸短期吸收4 h的方法,研究了pH对玉米及根际微生物竞争吸收氨基酸的影响.结果表明:土壤pH对玉米根和地上部生物量有显著影响,对于红壤,pH为6.48最适宜玉米生长,且玉米地上部¹⁵N丰度和¹⁵N-甘氨酸吸收量也显著高于其他处理;对于棕壤,pH为7.65最适宜玉米生长,其玉米地上部和根系¹⁵N丰度显著低于pH为5.78处理,但¹⁵N-甘氨酸吸收量显著高于其他处理.红壤pH为6.48条件下,其微生物生物量碳相对较高,而棕壤pH为7.65条件下,其微生物生物量碳相对较低.综合根系吸收、转运及微生物竞争吸收的结果,推断红壤在pH为6.48条件下虽然面临着微生物的竞争吸收,但生长于其上的玉米通过提高吸收速率和转移比率提高了氨基酸的吸收量;在pH为7.65的棕壤中,微生物活性较低,降低了与玉米竞争吸收氨基酸的能力,从而增加了玉米对氨基酸的吸收量.     

Release of Inorganic Phosphate from Irradiated Yeast: Radiation Biodosimetry and Evaluation of Radioprotective Compounds

When cells of bakers' yeast, Saccharomyces cerevisiae, were irradiated with ionizing radiation, inorganic phosphate, ninhydrin-reactive material, and substances absorbing at 260 mmu were released into the suspending medium. The amount of inorganic phosphate released depended on the radiation dose and on the temperature and pH during irradiation. The concentration of yeast cells did not affect the phosphate yield per milligram of yeast. It is suggested that the release of phosphate may serve as an index of the total radiation environment (i.e., as a biodosimeter) where radiation inactivation of microrganisms is of primary importance, e.g., in radiation preservation of foods. The somewhat limited range of the yeast biodosimeter (ca. 0.5 to 1.75 Mrad) may be extended by use of other more resistant microorganisms, such as bacterial spores. Compounds which have been reported as protecting microorganisms and mammals against the lethal effect of ionizing radiation also inhibited the radiation-induced release of inorganic phosphate from yeast. This phosphate release system is proposed as the basis for an economical, rapid supplement to screening procedures in the evaluation of radioprotective compounds.     

Characterization of Inorganic Biocarriers That Moderate System Upsets during Fixed-Film Biotreatment Processes

Inorganic matrices were developed for fixed-film bioreactors affording protection to microorganisms and preventing loss of bioreactor productivity during system upsets. These biocarriers, designated Type-Z, contain ion-exchange properties and possess high porosity and a high level of surface area, which provide a suitable medium for microbial colonization. Viable cell populations of 10⁹/g were attainable, and scanning electron micrographs revealed extensive external colonization and moderate internal colonization with aerobic microorganisms. Laboratory-scale bioreactors were established with various biocarriers and colonized with Pseudomonas aeruginosa, and comparative studies were performed. The data indicated that bioreactors containing the Type-Z biocarriers were more proficient at removing phenol (1,000 ppm) than bioreactors established with Flexirings (plastic) and Celite R635 (diatomaceous earth) biocarriers. More significantly, these biocarriers were shown to moderate system upsets that affect operation of full-scale biotreatment processes. For example, subjecting the Type-Z bioreactor to an influent phenol feed at pH 2 for periods of 24 h did not decrease the effluent pH or reactor performance. In contrast, bioreactors containing either Celite or Flexirings demonstrated an effluent pH drop to ~2.5 and a reduction in reactor performance by 75 to 82%. The Celite reactor recovered after 5 days, whereas the bioreactors containing Flexirings did not recover. Similar advantages were noted during either nutrient or oxygen deprivation experiments as well as alkali and organic system shocks. The available data suggest that Type-Z biocarriers represent an immobilization medium that provides an amenable environment for microbial growth and has the potential for improving the reliability of fixed-film biotreatment processes.     

模拟失重对白假丝酵母菌增殖和毒性的影响

【背景】近年来研究发现,失重条件可对一些致病微生物的增殖和毒性产生影响,白假丝酵母菌(Candida albicans)是典型的条件性致病真菌,在太空环境和人体中普遍存在,研究失重条件下白假丝酵母菌的增殖和毒性意义重大。【目的】利用旋转细胞培养系统(Rotary cell culture system,RCCS)模拟失重环境对白假丝酵母菌进行连续传代培养,检测模拟失重环境对白假丝酵母菌增殖情况、毒性以及基因表达的变化。【方法】将白假丝酵母菌接种在旋转生物反应器(High aspect rotating vessel,HARV)中,利用旋转细胞培养系统连续传代培养14 d,然后对菌株进行增殖速率测定、不同pH条件下增殖能力测定、生物膜相对形成能力测定和细胞毒性和动物毒力测定;利用转录组测序技术找出差异表达基因,结合性状分析模拟失重可能对白假丝酵母菌增殖和毒力的影响。【结果】与对照组相比,模拟失重组白假丝酵母菌对数期提前,增殖速率加快,在适宜pH条件下的增殖能力普遍提高,但其生物膜形成能力相对减弱,对LoVo细胞和小鼠的毒性减弱;转录组测序发现,模拟失重组共有280个基因表达差异达1.5倍以上(P0.05),其中248个上调、32个下调。差异基因经基因功能注释(Gene ontology,GO)和京都基因及基因组百科全书(Kyoto encyclopedia of genes and genomes,KEGG)富集分析发现,相关胞膜形成及细胞分裂基因表达上调,生物膜形成、细胞黏附及共生粘连宿主基因表达下调。【结论】模拟失重环境可引起白假丝酵母菌增殖和毒性水平发生变化,相关改变可为研究失重环境对微生物的影响提供参考。     

Bacterial Diversity in the Hyperalkaline Allas Springs (Cyprus), a Natural Analogue for Cementitious Radioactive Waste Repository

The biogeochemical gradients that will develop across the interface between a highly alkaline cementitious geological disposal facility for intermediate level radioactive waste and the geosphere are poorly understood. In addition, there is a paucity of information about the microorganisms that may populate these environments and their role in biomineralization, gas consumption and generation, metal cycling, and on radionuclide speciation and solubility. In this study, we investigated the phylogenetic diversity of indigenous microbial communities and their potential for alkaline metal reduction in samples collected from a natural analogue for cementitious radioactive waste repositories, the hyperalkaline Allas Springs (pH up to 11.9), Troodos Mountains, Cyprus. The site is situated within an ophiolitic complex of ultrabasic rocks that are undergoing active low-temperature serpentinization, which results in hyperalkaline conditions. 16S rRNA cloning and sequencing showed that phylogenetically diverse microbial communities exist in this natural high pH environment, including Hydrogenophaga species. This indicates that alkali-tolerant hydrogen-oxidizing microorganisms could potentially colonize an alkaline geological repository, which is predicted to be rich in molecular H₂, as a result of processes including steel corrosion and cellulose biodegradation within the wastes. Moreover, microbial metal reduction was confirmed at alkaline pH in this study by enrichment microcosms and by pure cultures of bacterial isolates affiliated to the Paenibacillus and Alkaliphilus genera. Overall, these data show that a diverse range of microbiological processes can occur in high pH environments, consistent with those expected during the geodisposal of intermediate level waste. Many of these, including gas metabolism and metal reduction, have clear implications for the long-term geological disposal of radioactive waste.     

Cardiolipin synthase of Arabidopsis thaliana

Functional expression studies in microorganisms showed that the Arabidopsis thaliana gene At4g04870 represents the cardiolipin synthase (CLS) gene encoding a hydrophobic preprotein of 38 kDa with a cleavable signal peptide for the import into mitochondria. CLS of Arabidopsis over-expressed in Escherichia coli has an alkaline pH optimum, a strict requirement for divalent cations and a distinctly lower K(m) for cytidinediphosphate-diacylglycerol than for phosphatidylglycerol. It displayed a preference for both its substrates esterified with unsaturated acyl groups. Solubilization and purification experiments revealed that the protein requires a defined phospholipid environment, particularly the presence of cardiolipin, to acquire its catalytically active conformation.