Taxonomy of oxalotrophic <Emphasis Type="Italic">Methylobacterium</Emphasis> strains

Most of the oxalotrophic bacteria are facultative methylotrophs and play important ecological roles in soil fertility and cycling of elements. This study gives a detailed picture of the taxonomy and diversity of these bacteria and provides new information about the taxonomical variability within the genus Methylobacterium. Twelve mesophilic, pink-pigmented, and facultatively methylotrophic oxalate-oxidizing strains were included in this work that had been previously isolated from the soil and some plant tissues by the potassium oxalate enrichment method. The isolates were characterized using biochemical tests, cellular lipid profiles, spectral characteristics of carotenoid pigments, G+C content of the DNA, and 16S rDNA sequencing. The taxonomic similarities among the strains were analyzed using the simple matching (S _SM) and Jaccard (S _J) coefficients, and the UPGMA clustering algorithm. The phylogenetic position of the strains was inferred by the neighbor-joining method on the basis of the 16S rDNA sequences. All isolates were Gram-negative, facultatively methylotrophic, oxidase and catalase positive, and required no growth factors. Based on the results of numerical taxonomy, the strains formed four closely related clusters sharing ≥85% similarity. Analysis of the 16S rDNA sequences demonstrated that oxalotrophic, pink-pigmented, and facultatively methylotrophic strains could be identified as members of the genus Methylobacterium. Except for M. variabile and M. aquaticum, all of the Methylobacterium type strains tested had the ability of oxalate utilization. Our results indicate that the capability of oxalate utilization seems to be an uncommon trait and could be used as a valuable taxonomic criterion for differentiation of Methylobacterium species.     

Purification of total DNA extracted from activated sludge

Purification of the total DNA extracted from activated sludge samples was studied. The effects of extraction buffers and lysis treatments (lysozyme, sodium dodecyl sulfate (SDS), sonication, mechanical mill and thermal shock) on yield and purity of the total DNA extracted from activated sludge were investigated. It was found that SDS and mechanical mill were the most effective ways for cell lysis, and both gave the highest DNA yields, while by SDS and thermal shock, the purest DNA extract could be obtained. The combination of SDS with other lysis treatment, such as sonication and thermal shock, could apparently increase the DNA yields but also result in severe shearing. For the purification of the crude DNA extract, polyvinyl polypyrrolidone was used for the removal of humic contaminants. Cetyltrimethyl ammonium bromide, potassium acetate and phenol/chloroform were used to remove proteins and polysaccharides from crude DNA. Crude DNA was further purified by isopropanol precipitation. Thus, a suitable protocol was proposed for DNA extraction, yielding about 49.9 mg (total DNA)/g volatile suspended solids, and the DNA extracts were successfully used in PCR amplifications for 16S rDNA and 16S rDNA V3 region. The PCR products of 16S rDNA V3 region allowed the DGGE analysis (denatured gradient gel electrophoresis) to be possible.     

啤酒废水处理系统高效菌株的分离筛选与分析

从啤酒废水处理系统SBR池中取得的水样,分离纯化得到菌株35株,分别以进站原水和水解酸化池出水作为污水培养基,进行COD降解试验,复筛后分别得到3株COD降解率高且降解效果稳定的细菌12#,15#,23#与15#,25#,31#。将高效菌株与SBR池的活性污泥混合,考察其对啤酒废水的处理效果,结果表明,23#与活性污泥交互作用时在原水污水培养基中摇床培养16h COD降解率达到75.13%;25#与活性污泥交互作用处理水解酸化池出水120h时降解率达到82.40%。对4株高效菌株15#,23#,25#,31#进行16S rDNA序列测定,将所得序列与GenBank中已登录的相近细菌种群的DNA序列对比,得到高度同源性的细菌菌种。     

Pyrrolizidine alkaloid content in <Emphasis Type="Italic">Senecio</Emphasis>: ontogeny and developmental constraints

Summary. The ontogeny of pyrrolizidine alkaloid (PA) synthesis and constraints on defence level during the seedling stage were examined in the annual Senecio vulgaris and the monocarpic perennial Senecio jacobaea. In both species, PAs were actively synthesized from the onset of seedling growth so that juvenile stages did not go through an undefended stage. Roots are known to be the exclusive sites where PAs are produced. Root biomass was the single most important biomass parameter explaining variation in total PAs per seedling. All correlation coefficients between—relative growth rate and PA concentration were negative, but none was significant. However, a significant negative—correlation was found between shoot to root ratio and PA concentration in S. jacobaeaseedlings, suggesting a dilution effect of the PAs. Earlier studies have shown that the shoot to root ratio is positively correlated with relative growth—rate of established S. jacobaea plants. It is therefore suggested that young S. jacobaea plants with a high shoot to root ratio and hence a high growth capacity necessarily have lower PA defence levels than plants with a low shoot to root ratio. Received 10 July 2002; accepted 16 November 2002.     

疏棉状嗜热丝孢菌Thermomyces lanuginosus YNUCC4154耐碱热稳定木聚糖酶的特性及其产生菌的系统发育分析

从云南洱源县热泉中分离到一株产木聚糖酶嗜热真菌Thermomyces lanuginosus YNUCC4154．对其28S rDNA的5‘端约900bp的片段进行了扩增和测序，并与GenBank中最相似的12个分类单位进行了比较，邻近接合法构建的系统发育树显示T.lanuginosus YNUCC4154与几个产木聚糖酶的嗜热真菌关系较近．在30L全自动发酵罐中55℃发酵50h后，木聚糖酶酶活达1032 U mL^-1．该木聚糖酶最适反应温度为73℃，最适pH为6，5，在10-67℃和pH3～12时比较稳定，表明这是一种热稳定耐碱木聚糖酶，在造纸工业中具有良好的应用前景．图7表1参24     

链格孢菌菌株致病性及其遗传差异性

对分离自紫茎泽兰的19个链格孢菌菌株的致病性进行了比较研究，并对这19个菌株和其它来源的3个菌株的5．8S rDNA及其两侧的ITS1区和ITS2区进行了序列分析．结果显示，链格孢菌菌株的致病性差异明显；21株链格孢菌菌株在5．8SrDNA及其两侧的转录问区（ITS区）的序列无差异，而与百日草链格孢菌（Alternaria zinnine）的序列差异显著，表明21株链格孢菌菌株属种内变异．采用AFLP分子标记技术对21株链格孢菌的DNA扩增片段长度多态性进行了分析，结果表明，链格孢菌自然变异群体内存在很高的遗传多样性．链格孢菌菌株的相似性与菌株的地理来源有一定的相关性，但基于AFLP标记划分的AFLP类群与基于寄主病情指数划分的致病力类群问关系不相一致；起始菌株与变异菌株相似性显著，但二者和致病性的关系不显著．图2表1参18     

一株多菌灵降解菌NY97-1的分子鉴定及GFP标记

用PCR方法扩增的多菌灵降解菌NY97-1的16SrDNA片段经TA克隆后,进行序列测定和BLAST同源序列比较分析,确定了其分类地位为短小芽孢杆菌(Bacilluspumilus,B.p).经BamHⅠ酶切的启动子探针载体pUC19-gfp与NY97-1基因组DNA的Sau3AⅠ酶切片段酶连,酶连产物转化E.coliDH5a,建立B.p的启动子基因文库.挑选其中的两个强阳性克隆,亚克隆来自短小芽孢杆菌总基因组的启动子活性片段F4、F5,构建大肠杆菌-短小芽孢杆菌穿梭表达载体pNW33N-F4-gfp、pNW33N-F5-gfp.通过电转化得到gfp在B.p中的两株标记菌株.在荧光显微镜下,观察到了明亮的绿色荧光,证明活性片段F4、F5均具有组成型启动子的功能,实现了gfp基因在B.p中组成型表达,且遗传稳定,为今后研究多菌灵降解菌B.p在自然环境中的定殖、分布及动态变化打下了基础.图3表2参20     

Accounting for Uncertainty in Making Species Protection Decisions

Abstract:  Uncertainty gives rise to two decision errors in implementing the U.S. Endangered Species Act: listing species that are not in danger of extinction and delisting species that are in danger of extinction. I evaluated four methods (minimum standard, precautionary principle, minimax regret criterion, adaptive management) for deciding whether to list or delist a species when there is uncertainty about how those decisions are likely to influence survival of the species. A safe minimum standard criterion preserves some minimum amount or safe standard (population) of a species unless maintaining that amount generates unacceptable social cost. The precautionary principle favors not delisting a species when there is insufficient evidence on the efficacy of state management plans for protecting them. A minimax regret criterion selects the delisting decision that minimizes the maximum loss likely to occur under alternative ecosystem states. When the cost of making a correct decision is less than the cost of making an incorrect decision, the minimax regret criteria indicates that delisting is the optimal decision. Active adaptive management employs statistically valid experiments to test hypotheses about the likely impacts of delisting decisions. Safe minimum standard and minimax regret criterion are not compatible with the U.S. Endangered Species Act. The precautionary principle comes closest to describing how federal agencies make delisting decisions. Active adaptive management is scientifically superior to the other methods but is costly and time consuming and may not be compatible with the U.S. National Environmental Policy Act.     

SO4^2-／TiO2对SO2-C7H16-TiO2复相光化学反应的影响

研究了TiO2与SO2，C7H16复相光化学反应的光催化活性．在SO2-C7H16-TiO2光催化反应过程中，TiO2表面可形成SO4^2-/TiO2结构，它的存在可提高庚烷的光催化氧化速率，利用IR和XPS研究了反应过程中TiO2表面形成的SO4^2-/TiO2结构。     

Alkaloid tolerance in Manduca sexta and phylogenetically related sphingids (Lepidoptera: Sphingidae)

Summary. Nicotine tolerance is well known for Manduca sexta. It also occurs in several other sphingids of the subfamilies Macroglossinae and Sphinginae. Only members of the subfamily Smerinthinae appear to be more susceptible to nicotine intoxication. Phylogenetic trees have been reconstructed from mitochondrial 16S rDNA and nuclear DNA to map nicotine tolerance.?The nicotine binding site of both α-subunits of nicotinic acetylcholine receptors (nAChR) have been amplified and sequenced. No apparent amino acid substitution can be seen in the putative nicotine binding site of the α-subunits of nAChR from nicotine tolerant and nicotine sensitive sphingids. Thus, a simple target-site modification can be ruled out as a cause for nicotine tolerance. This finding agrees with feeding experiments: larvae of M. sexta and other sphingids of the Macroglossinae and Sphinginae not only tolerated nicotine, but also many other alkaloids that affect neuroreceptors other than acetylcholine receptors (nAChR, mAChR).?Only 10 to 20% of nicotine injected into larvae of nicotine-tolerant taxa could be recovered later as free nicotine, nicotine N-oxide or cotinine, i.e., 80 to 90% must have been converted to polar conjugates or degradation products which are not detectable with the methods applied. Usually more than 98% of the recoverable alkaloids were found in the faeces. Excretion reached a maximum 6 h after injection in tolerant taxa. Larvae of Manduca sexta, which were reared on a nicotine-rich diet, showed higher nicotine degradation and faster nicotine elimination than na?ve larvae. Application of the cytochrome P450 inhibitor SKF 525A (proadifen) reduced the formation of nicotine N-oxide and the rate of alkaloid degradation. Thus, an inducible detoxification mechanism, coupled with a rapid and inducible excretion, appear to be a strategy in Sphingidae that helps them to live on host plants rich in otherwise toxic secondary metabolites. Received 23 March 2001; accepted 4 August 2001.