不同pH值条件下硫酸盐还原菌组成及硫酸盐还原机制分析

【目的】从海洋沉积物中富集获得硫酸盐还原菌群,改变pH值进行培养,分析pH值对硫酸盐还原性质的影响,明确菌群组成和进行硫酸盐还原功能基因预测,探究硫酸盐还原机制。【方法】分析硫酸盐还原菌群在不同pH值条件下的硫酸盐还原率,在此基础上,利用高通量测序技术和PICRUSt软件分析硫酸盐还原菌群优势菌组成及硫酸盐还原相关基因相对丰度。【结果】硫酸盐还原菌群在不同pH值培养条件下的生长和硫酸盐还原率出现显著变化(P<0.01),在pH 5.0时达到峰值,分别为0.34±0.01和96.52%±0.44%。高通量测序数据显示,pH 5.0时菌群丰富度和多样性最高,优势菌属为假单胞菌(Pseudomonas)和芽孢杆菌(Bacillus),相对丰度较高的基因为同化性硫酸盐还原相关基因。【结论】硫酸盐还原菌富集生长的最适pH 5.0,在此条件下的高硫酸盐还原率由同化性硫酸盐还原途径主导,为揭示硫酸盐还原机制提供了实验支持,并拓宽了硫酸盐还原菌实践应用方面的种质资源。     

分离自活性污泥的硫酸盐还原菌用于铅锌冶炼渣重金属污染修复

【背景】从活性污泥中分离出一类具有硫酸盐还原能力的菌株,探讨了其用于铅锌冶炼渣重金属污染修复的可行性。【目的】研究硫酸盐还原菌(Sulfate reducing bacteria)对铅锌冶炼渣中重金属的固化作用。【方法】将从活性污泥中分离出的硫酸盐还原菌接种到铅锌冶炼渣中进行修复,采用X射线衍射、Tessier、电感耦合等离子体发射光谱仪检测、高通量测序等方法进行实验。检测铅锌冶炼渣中矿物组成,以及修复过程中重金属化学形态、各金属离子浓度和微生物群落结构的变化。【结果】修复实验表明,体系中电位降低、pH值提高、各重金属稳定态增加、离子浓度降低且微生物群落结构变化显著,硫酸盐还原菌成为优势菌群。【结论】接种硫酸盐还原菌后铅锌冶炼渣中的重金属原位固化效果显著,从而降低生物可利用性,将恶性循环变为良性循环,所以硫酸盐还原菌可用作重金属污染修复的固化药剂。     

Cr(VI)还原菌的筛选、鉴定及其还原物质分析

【背景】铬污染土壤是我国土壤污染修复的重点治理对象,在众多修复技术中,微生物法因具有简单、经济、无二次污染等特性已成为研究热点,而微生物法中筛选出既能适应污染场地环境又能高效还原Cr(VI)的菌株尤为重要。【目的】筛选适应西北寒旱区高效还原Cr(VI)的菌株,丰富铬还原菌资源库,为铬污染土壤修复奠定基础。【方法】采用富集驯化、分离纯化法进行筛菌;通过形态学和分子生物学相结合的方法对目的菌株进行鉴定;采用傅里叶变换红外光谱法对还原机理进行研究。【结果】菌株G-13有较强的Cr(VI)还原能力,pH 9.0、温度为30°C条件下,60 h对Cr(VI)(100 mg/L)的还原率达到82.8%。经形态学和分子生物学鉴定,菌株G-13为Micrococcus luteus。反应中Cr(VI)的降低伴随着Cr(III)的增加,说明以还原反应为主,并且还原能力与细菌生长呈依赖型关系。对细胞各组分及变性研究表明,胞外酶在还原反应中占主要作用。除Pd~(2+)、Cd~(2+)外,其余金属离子对酶活性无明显抑制作用。通过傅里叶变换红外光谱分析,发现G-13与Cr(VI)结合位点主要为羟基、羰基、羧基、–CH、酰胺基等。【结论】菌株G-13有较强的Cr(VI)还原能力,能为西北寒旱区铬污染土壤修复丰富菌种资源。     

微生物烟气脱硫中硫酸盐还原阶段的限制性因素及其影响

【目的】利用硫酸盐还原菌(SRB)厌氧活性污泥进行烟气脱硫,探索硫酸盐生物还原的最适条件及重金属离子对硫酸盐生物还原的影响,以提高硫酸盐还原阶段的效率。【方法】对取自污水处理厂的SRB厌氧活性污泥进行高浓度硫酸盐胁迫驯化。分析生物脱硫过程中SRB厌氧污泥还原硫酸盐的限制性因素及影响。【结果】在最适生长条件下(pH 6.5,32°C),经驯化获得的SRB厌氧活性污泥有较强的硫酸盐还原能力。Fe2+的适量添加对硫酸盐还原有一定促进作用。SRB厌氧污泥还原硫酸盐的ThCOD/SO42-最适值为3.00,ThCOD=3.33为最适理论化学需氧量,硫酸盐还原率可达72.15%。SRB厌氧污泥还原硫酸盐反应体系中抑制SRB活性的硫化物浓度为300 mg/L。Pb2+和Ni2+在较低的浓度下(1.0 mg/L和2.0 mg/L)对硫酸盐的还原产生较强的抑制作用,而Cu2+在稍高的浓度下(8.0 mg/L)显示出明显的抑制作用。【结论】经驯化,SRB厌氧活性污泥显示出较强的硫酸盐还原能力,具有应用于工业烟气生物脱硫的潜力。去除重金属离子Pb2+、Ni2+和Cu2+可有效解除对硫酸盐生物还原作用的抑制。     

一株兼性氧化亚氮还原菌的还原N2O能力

【目的】从水稻土中分离筛选出一株兼性氧化亚氮还原菌,并探索其在不同条件下还原N_2O的能力,为减少温室气体N_2O的排放提供重要依据。【方法】通过微生物富集培养分离技术从水稻土中分离得到纯菌;利用nosZ基因和16S rRNA的测序分析鉴定菌株;通过测定菌株在不同条件下N_2O的还原量,分析该菌株还原N_2O的能力及调控因子。【结果】经鉴定,该菌株含有nos Z基因,属于假单胞菌属,在温度30°C、厌氧条件下还原N_2O速率高达0.0219μmol/min以上,改变不同温度和氧气浓度后其能力相对减弱,但仍具备较强的还原N_2O作用。【结论】从水稻土中分离筛选得到的兼性氧化亚氮还原菌为假单胞菌,它在不同环境条件下都具备较强的还原N_2O能力,该菌株可能为减少土壤N_2O排放提供新途径,对保障生态环境安全具有重要的应用价值。     

一株木质素降解菌高温驯化及酶学性质的测定

【背景】高温引起的微生物活性降低是限制园林绿化废弃物堆肥过程中木质素降解的主要因素。【目的】驯化一株木质素降解菌芽孢杆菌NO.2,提高其在高温下的微生物活性,探究其生长情况及酶学特性。【方法】采用温度梯度方法驯化菌株,以菌株生长曲线、酶活力、木质素降解率为评价指标,探究驯化前后菌株间差异,以及驯化后菌株所产木质素降解酶的酶促反应温度和pH范围。【结果】与原菌株相比,驯化后菌株在60℃培养时最大生物量间差异不显著;漆酶(laccase,Lac)、锰过氧化物酶(manganeseperoxidase,MnP)和木质素过氧化物酶(ligninperoxidase,LiP)酶活力得到进一步提高,分别提高了30.75%、35.98%和29.62%,木质素降解率提高60.52%。酶学性质研究表明,驯化后菌株所产Lac、MnP和LiP在20-60℃、pH 3.0-9.0范围内酶活力均较高,而且具有较好的稳定性,稳定性依次为Lac>LiP>MnP。【结论】温度梯度驯化方法可有效提高微生物对高温环境的适应性,扩大木质素降解酶的酶促反应温度和pH范围,在进一步自主研制专用降解园林废弃物微生物菌...     

一株泰乐菌素高效降解菌的分离鉴定及其降解特性

【目的】从长期堆放泰乐菌素药渣附近的土壤中分离出泰乐菌素降解菌,并考察其对泰乐菌素的降解特性。【方法】采用梯度驯化、划线分离法筛选出泰乐菌素优势降解菌,通过形态观察、生理生化特征和16S rRNA基因序列分析方法对其进行系统发育分析及菌种鉴定,并考察菌株对泰乐菌素的降解特性。【结果】从长期堆放泰乐菌素药渣的土壤中分离得到1株泰乐菌素高效降解菌,命名为TS1,其为革兰氏阴性杆菌,菌落形态呈圆形,乳白色,表面光滑,不透明,边缘整齐,鉴定为越南伯克霍尔德氏菌(Burkholderia vietnamiensis)。该菌株在温度35°C、pH 7.0的条件下培养72 h,对初始浓度为300 mg/L泰乐菌素的降解率可达99%以上。【结论】说明菌株TS1对泰乐菌素具有良好的降解特性,可用于生物修复被泰乐菌素废渣废水污染的生态环境。     

圈养大熊猫肠道乳酸菌分离及益生特性

【背景】大熊猫数量稀少、繁殖困难,在其生长过程中极易感染消化系统疾病甚至死亡。【目的】分析圈养大熊猫肠道可培养乳酸菌的群落结构与功能,筛选具有益生特性的乳酸菌菌株,为大熊猫消化系统疾病的预防和肠道微生物研究提供理论参考及菌种资源。【方法】采用3种培养基分离大熊猫粪便中的乳酸菌,通过革兰氏染色镜检和过氧化氢试验对分离的菌株进行初步鉴定,基于BOXA1R-PCR图谱遗传多样性选取代表菌株进行16S rRNA基因测序分析并进行主成分分析(principal component analysis, PCA),同时分析乳酸菌菌株安全性和益生特性。【结果】通过初步鉴定共分离获得58株乳酸菌,根据BOXA1R-PCR结果挑选20株菌进行测序,结果显示20株菌分属于明串珠菌属(Leuconostoc)、肠球菌属(Enterococcus)、魏斯氏菌属(Weissella)和链球菌属(Streptococcus)这4个属,主成分分析结果表明不同年龄段的大熊猫肠道乳酸菌群落结构存在差异。20株乳酸菌均不溶血,17株乳酸菌对11种抗生素均敏感;11株菌耐pH值2.0酸性条件,14株菌对0.3%的胆盐具有良好...     

一株耐受低pH、高浓度乳酸及琥珀酸菌株的筛选与鉴定

【目的】通过构建的人工耐酸系统,筛选耐受低pH值、乳酸及琥珀酸的菌株。【方法】构建人工耐酸系统长期驯化菌株,利用不同p H的酸性平板进行筛选,从环境中筛选出一株对低p H值、高浓度乳酸以及琥珀酸有很好耐受性的菌株。通过形态学特征、生理生化特征研究,并结合18S rDNA基因序列分析及分子系统发育树的构建结果,确定菌株的种类。【结果】经过酸性人工系统的长期驯化,筛选分离出一株耐受低pH值、高浓度乳酸以及琥珀酸的菌株WJ-2,经鉴定该菌株为酿酒酵母(Saccharomyces cerevisiae),其最适生长温度为30°C。酸性平板实验显示该菌株能够耐受pH2.5的酸性环境,同时对9%的乳酸及8%的琥珀酸也有很好的耐受性。另外,耐酸菌株WJ-2在pH 2.5、9%乳酸和8%琥珀酸的培养环境中仍能保持相对中性的细胞内pH值。【结论】通过构建人工酸性系统,成功筛选出一株对低pH值、高浓度乳酸以及琥珀酸具有耐受性的菌株——酿酒酵母菌WJ-2,该方法可为筛选具有特定耐受能力菌株提供一个新思路。     

海洋固氮菌和解磷菌的分离鉴定及发酵条件优化

【目的】从西沙喜盐草根际沉积物中分离纯化得到具有高效固氮能力及解磷能力的菌株。优化其发酵培养条件,研究其制备海洋微生物菌剂的可能性。【方法】从形态学特征、生理生化、16S rDNA及功能基因水平进行鉴定,通过乙炔还原法、钼锑抗显色法检测菌株的固氮酶活性和解磷能力,单因素法和响应面法优化其发酵培养条件,溶血试验和急性毒性实验鉴定菌株的安全性。【结果】结果表明,菌株AZ16属于星箭头菌(Sagittula stellate),革兰氏阴性菌,选择性固氮培养基中菌落呈黄圆形黏稠状,固氮酶活性达34.63 nmol C2H2/(mL·h),最适生长条件为:盐度25‰、pH 7.5、温度33°C、接种量5.0%;菌株XT37为海洋芽孢杆菌(Bacillus sp.),革兰氏阳性菌,选择性固氮培养基中菌落呈深黄色圆形褶皱,植酸酶活性达239.49μg/L,最适合生长条件为:盐度25‰、pH 6.7、温度28°C、接种量5.0%。溶血实验和急性毒性实验证明两株菌属实际无毒级别。【结论】两株菌具有高效的固氮解磷功能,以及抗高盐、强碱等环境的能力,安全无毒,因此有潜力应用于多功能混合微生物菌剂的研制。     

Degradation and conversion of somatostatin in normal and diabetic rats in vivo and in vitro

Plasma somatostatin-like immunoreactivity in the portal and jugular veins of streptozotocin diabetic rats was compared with that in normal control rats. In the diabetic group, somatostatin levels in the portal (p less than 0.05) and jugular (p less than 0.01) veins were both elevated compared with those in the control group. Moreover, the degree of elevation was greater in the jugular vein than in the portal vein. To further investigate the role of the liver in the clearance of somatostatin-28 in vivo, 2 micrograms of somatostatin-28 was administered as a bolus into the external jugular vein of intact and functionally hepatectomized rats. The mean half-time of somatostatin-28 was significantly longer in intact diabetic rats than in controls (p less than 0.05). The functional hepatectomy did not cause a significant difference in the half-time in diabetic rats but made it longer in control rats. These results suggest that the longer half-time of somatostatin-28 in diabetic rats in vivo is due to its slower hepatic clearance. The hepatic clearance of somatostatin-28 and somatostatin-14 was further studied in vitro using a recirculating liver perfusion method. The hepatic clearance of 1.2 nM of either somatostatin-28 or somatostatin-14 was significantly lower in diabetic rats than in controls (p less than 0.01). This indicates that elevated plasma somatostatin levels in diabetic rats are caused at least in part by decreased hepatic clearance of somatostatin.(ABSTRACT TRUNCATED AT 250 WORDS)     

Production of arsine and methylarsines in soil and in culture

Arsenate, arsenite, monomethylarsonate, and dimethylarsinate were added to different soils, and evolution of gaseous arsenical products was determined over 3 weeks. Arsine was produced in all three soils from all substrates, whereas methylarsine and dimethylarsine were produced only from methylarsonate and dimethylarsinate, respectively. At least three times more arsine than dimethylarsine was produced in soil incubated with dimethylarsinate. Resting cell suspensions of Pseudomonas and Alcaligenes produced arsine as the sole product when incubated anaerobically in the presence of arsenate or arsenite. In all instances, no trimethylarsine was observed, nor could any evidence be shown for the methylation of any arsenical substrate in soil or in culture. It was concluded that reduction to arsine, not methylation to trimethylarsine, was the primary mechanism for gaseous loss of arsenicals from soil.     

Growth and mitogenic effects of arylphorin in vivo and in vitro

In insects, developmental responses are organ- and tissue-specific. In previous studies of insect midgut cells in primary tissue cultures, growth-promoting and differentiation factors were identified from the growth media, hemolymph, and fat body. Recently, it was determined that the mitogenic effect of a Manduca sexta fat body extract on midgut stem cells of Heliothis virescens was due to the presence of monomeric alpha-arylphorin. Here we report that in primary midgut cell cultures, this same arylphorin stimulates stem cell proliferation in the lepidopterans M. sexta and Spodoptera littoralis, and in the beetle Leptinotarsa decemlineata. Studies using S. littoralis cells confirm that the mitogenic effect is due to free alpha-arylphorin subunits. In addition, feeding artificial diets containing arylphorin increased the growth rates of several insect species. When tested against continuous cell lines, including some with midgut and fat body origins, arylphorin had no effect; however, a cell line derived from Lymantria dispar fat body grew more rapidly in medium containing a chymotryptic digest of arylphorin.     

Antiplatelet and antithrombotic activities of purpurogallin in vitro and in vivo

Enzymatic oxidation of pyrogallol was efficiently transformed to an oxidative product, purpurogallin (PPG). Here, the anticoagulant activities of PPG were examined by monitoring activated partial thromboplastin time (aPTT), prothrombin time (PT), and the activities of thrombin and activated factor X (FXa). And, the effects of PPG on expression of plasminogen activator inhibitor type 1 (PAI-1) and tissue-type plasminogen activator (t-PA) were evaluated in tumor necrosis factor (TNF)-α activated human umbilical vein endothelial cells (HUVECs). Treatment with PPG resulted in prolonged aPTT and PT and inhibition of the activities of thrombin and FXa, as well as inhibited production of thrombin and FXa in HUVECs. In addition, PPG inhibited thrombin-catalyzed fibrin polymerization and platelet aggregation. PPG also elicited anticoagulant effects in mice. In addition, treatment with PPG resulted in significant reduction of the PAI-1 to t-PA ratio. Collectively, PPG possesses antithrombotic activities and offers a basis for development of a novel anticoagulant. [BMB Reports 2014; 47(7): 376-381]     

Growth and reproduction of Arabidopsis thaliana in relation to storage of starch and nitrate in the wild-type and in starch-deficient and nitrate-uptake-deficient mutants

We have investigated the interactions between resource assimilation and storage in rosette leaves, and their impact on the growth and reproduction of the annual species Arabidopsis thaliana. The resource balance was experimentally perturbed by changing (i) the external nutrition, by varying the nitrogen supply; (ii) the assimilation and reallocation of resources from rosette leaves to reproductive organs, by cutting or covering rosette leaves at the time of early flower bud formation, and (iii) the internal carbon and nitrogen balance of the plants, by using isogenic mutants either lacking starch formation (PGM mutant) or with reduced nitrate uptake (NU mutant). When plants were grown on high nitrogen, they had higher concentrations of carbohydrates and nitrate in their leaves during the rosette phase than during flowering. However, these storage pools did not significantly contribute to the bulk flow of resources to seeds. The pool size of stored resources in rosette leaves at the onset of seed filling was very low compared to the total amount of carbon and nitrogen needed for seed formation. Instead, the rosette leaves had an important function in the continued assimilation of resources during seed ripening, as shown by the low seed yield of plants whose leaves were covered or cut off. When a key resource became limiting, such as nitrogen in the NU mutants and in plants grown on a low nitrogen supply, stored resources in the rosette leaves (e.g. nitrogen) were remobilized, and made a larger contribution to seed biomass. A change in nutrition resulted in a complete reversal of the plant response: plants shifted from high to low nutrition exhibited a seed yield similar to that of plants grown continuously on a low nitrogen supply, and vice versa. This demonstrates that resource assimilation during the reproductive phase determines seed production. The PGM mutant had a reduced growth rate and a smaller biomass during the rosette phase as a result of changes in respiration caused by a high turnover of soluble sugars ( Caspar et al. 1986 ; W. Schulze et al. 1991 ). During flowering, however, the vegetative growth rate in the PGM mutant increased, and exceeded that of the wild-type. By the end of the flowering stage, the biomass of the PGM mutant did not differ from that of the wild-type. However, in contrast to the wild-type, the PGM mutant maintained a high vegetative growth rate during seed formation, but had a low rate of seed production. These differences in allocation in the PGM mutant result in a significantly lower seed yield in the starchless mutants. This indicates that starch formation is not only an important factor during growth in the rosette phase, but is also important for whole plant allocation during seed formation. The NU mutant resembled the wild-type grown on a low nitrogen supply, except that it unexpectedly showed symptoms of carbohydrate shortage as well as nitrogen deficiency. In all genotypes and treatments, there was a striking correlation between the concentrations of nitrate and organic nitrogen and shoot growth on the one hand, and sucrose concentration and root growth on the other. In addition, nitrate reductase activity (NRA) was correlated with the total carbohydrate concentration: low carbohydrate levels in starchless mutants led to low NRA even at high nitrate supply. Thus the concentrations of stored carbohydrates and nitrate are directly or indirectly involved in regulating allocation.     

Importance of guanine nitration and hydroxylation in DNA in vitro and in vivo

Guanine (Gua) modification by nitrating and hydroxylating systems was investigated in DNA. In isolated calf thymus DNA, 8-NO(2)-Gua and 8-oxo-Gua were dose-dependently formed with peroxynitrite, and 8-NO(2)-Gua was released in substantial amounts. Myeloperoxidase (MPO) with H(2)O(2) and NO(2)(-) reacted with calf thymus DNA to form 8-NO(2)-Gua dose dependently without release of 8-NO(2)-Gua. The frequency of strand breaks was higher than the sum of 8-NO(2)-Gua and 8-oxo-Gua, particularly in the MPO-treated DNA, indicating the importance of other types of damage. The activation of human neutrophils and lymphocytes with phorbol ester did not induce 8-NO(2)-Gua and 8-oxo-Gua in their nuclear DNA. However, 8-NO(2)-Gua was found in calf thymus DNA co-incubated with activated neutrophils in the presence of NO(2)(-). No significant formation of 8-NO(2)-Gua was found in liver DNA from mice treated with Escherichia coli lipopolysaccharide. The incubation of peroxynitrite or MPO-H(2)O(2)-NO(2)(-)-treated DNA with formamidopyrimidine glycosylase (Fpg) released 8-oxo-Gua, but not 8-NO(2)-Gua, indicating that 8-NO(2)-Gua is not a substrate for Fpg. Although 8-NO(2)-Gua was generated in isolated DNA by different nitrating systems, other types of damage were formed in abundance, and the lesion could not be found reliably in nuclear DNA, suggesting that the biological importance is limited.