Effect of Competing Electron Acceptors on the Reduction of U(VI) by Desulfotomaculum reducens

The biological reduction of soluble U(VI) to the less soluble U(IV) has been proposed as a strategy to remediate uranium-contaminated sites. However, the majority of the contaminated sites contain, in addition to U(VI), competing electron acceptors (CEAs) that can either enhance or inhibit U(VI) reduction. Desulfotomaculum reducens MI-1 is a sulfate-reducing bacterium able to reduce a variety of electron acceptors including U(VI). We characterized U(VI) reduction by D. reducens in the presence of pyruvate and three CEAs: sulfate, nitrate or soluble ferric iron. In the presence of sulfate or ferric iron and U(VI), cell growth was driven by respiration of the CEA. Nitrate was not used as an electron acceptor for growth and vegetative cells grew instead by fermenting pyruvate. Sulfate remaining after sulfate reduction has ceased or the presence of nitrate did not affect U(VI) reduction. However, in the case of sulfate, the addition of H₂ after the depletion of pyruvate greatly enhanced U(VI) reduction. Contrary to sulfate and nitrate, the presence of Fe(II), the product of Fe(III) reduction, abolished U(VI) reduction. The results from this investigation suggest that this microorganism and others with similar characteristics may play a role in U(VI) bioremediation efforts but only after the soluble Fe(II) produced by Fe(III) reduction has been advected away.     

Sulfate Requirement for the Growth of U(VI)-Reducing Bacteria in an Ethanol-Fed Enrichment

A field experiment at the Oak Ridge Field Research Center has demonstrated the in situ biostimulation of U(VI) reduction with ethanol amendment, but little is known about the stimulated metabolic pathways or composition of the bacterial community mediating the reduction. This work characterized the metabolism and community structure of a sulfate-reducing enrichment developed from sediment from the field site to help address this knowledge gap. Structure was investigated by clone library construction and terminal restriction fragment length polymorphism (T-RFLP) analysis of 16S rDNA. The enrichment used ethanol concomitantly with sulfate, producing acetate. Hydrogen accumulated intermittently. The clone library contained sequences related to Clostridia, Desulfovibrio, Bacteroides, and Synergistes species. The enrichment reduced U(VI), and the reduction rate was 0.055 L/mg volatile suspended solids (VSS)/day. The enrichment's T-RFLP profile was comprised largely of Desulfovibrio-like fragments, and Desulfovibrio species are known to reduce sulfate and U(VI). A second line of enrichments, inoculated from the sulfate-amended enrichment, was maintained without sulfate. After four transfers of the sulfate-free culture, it was found unable to reduce U(VI). This culture's T-RFLP profile was largely comprised of Clostridia-like fragments, and Clostridia ferment ethanol to acetate. The results indicate a sulfate requirement for the growth of U(VI)-reducing organisms in this community.     

The Use of Terminal Restriction Fragment Length Polymorphism (T-RFLP) for the Characterisation of Microbial Communities in Marine Sediments

Terminal Restriction Fragment Length Polymorphism (T-RFLP) of PCR amplified 16S rRNA genes was used to investigate microbial communities in the sediments of Ria Formosa, Portugal. Five replicates of surface sand sediments were collected at an artificial inlet to the sea, between June 2001 and July 2002. Restriction enzymes Msp1 and Hha1 provided 57 different terminal fragments (T-RFs). The sediments were essentially dominated by the same ribotypes throughout the year, with seasonal shifts attributed to minor ribotypes. Principal component analysis of the T-RFs profile revealed no consistent pattern of temporal variation and no consistent grouping of replicate sediment samples. The results suggest that the small-scale spatial variability outweighs the seasonal variability. Phylogenetic affiliations suggested that the dominant bacteria were representatives of the α-Proteobacteria group.     

Temporal variation of magnetotactic bacterial communities in two freshwater sediment microcosms

Magnetotactic bacteria (MTB), which can mineralize nanosized magnetite or greigite crystals within cells, play important roles in biogeochemical processes, for example iron and sulfur cycling, and depositional remanent magnetization acquisitions. Despite decades of research, the knowledge of MTB distribution and ecology is still limited. In the present study, we investigated the temporal variation of MTB communities in freshwater sediment microcosms based on 16S rRNA genes and unifrac analyses. Two microcosms (MY8 and MY11) collected from two separate sites in Lake Miyun (Beijing, China) were analyzed. The majority of retrieved sequences belonged to alphaproteobacterial magnetotactic cocci in both microcosms (representing 64.29% of clones from MY8 and 100% of clones from MY11), whereas so-called ' Magnetobacterium bavaricum '-like MTB affiliated within Nitrospira phylum were exclusively found in microcosm MY8. Over a 3-month period, the temporal variation of MTB communities was evident in both microcosms. In addition, the phylogenetic discrepancy of MTB communities between two microcosms is more prominent than that of the same microcosm at different times, implying adaptation of MTB phylogenetic lineages to specific microenvironments. Among the physical–chemical parameters measured, a strong correlation was shown between nitrate and the main genetic variability of MTB communities, indicating that nitrate may influence the occurrence of MTB phylogenetic lineages in natural environments.     

Microbial reduction of Cr(VI) in the presence of chromate conversion coating constituents

The reduction of Cr(VI) by the metal-reducing bacterium Shewanella oneidensis MR-1 was evaluated, to determine the potential for exploiting Cr(VI) bioreduction as a means of treating chromate conversion coating (CCC) waste streams. Inclusion of Cr(VI) at concentrations ≥1 mM inhibited aerobic growth of S. oneidensis, but that organism was able to reduce Cr(VI) at a concentration of up to 1 mM under anaerobic, nongrowth conditions. S. oneidensis reduced Cr(VI) in the presence of common CCC constituents, with the exception of ferricyanide, when these CCC constituents were included at concentrations typical of CCC waste streams. Ferricyanide inhibited neither aerobic growth nor metabolism under aerobic, nitrate- or iron-reducing conditions, suggesting that the ferricyanide-depended inhibition of Cr(VI) reduction is not due to broad metabolic inhibition, but is specific to Cr(VI) reduction. Results indicate that under some conditions, the activities of metal-reducing bacteria, such as S. oneidensis, could be exploited for the removal of Cr(VI) from CCC waste streams under appropriate conditions.     

Biogeochemical Elimination of Chromium (VI) from Contaminated Water

Ferrous iron [Fe(II)] reductively transforms heavy metals in contaminated groundwater, and the bacterial reduction of indigenous ferric iron [Fe(III)] to Fe(II) has been proposed as a means of establishing redox reactive barriers in the subsurface. The reduction of Fe(III) to Fe(II) can be accomplished by stimulation of indigenous dissimilatory metal-reducing bacteria (DMRB) or injection of DMRB into the subsurface. The microbially produced Fe(II) can chemically react with contaminants such as Cr(VI) to form insoluble Cr(III) precipitates. The DMRB Shewanella algae BrY reduced surface-associated Fe(III) to Fe(II), which in batch and column experiments chemically reduced highly soluble Cr(VI) to insoluble Cr(III). Once the chemical Cr(VI) reduction capacity of the Fe(II)/Fe(III) couple in the experimental systems was exhausted, the addition of S. algae BrY allowed for the repeated reduction of Fe(III) to Fe(II), which again reduced Cr(VI) to Cr(III). The research presented herein indicates that a biological process using DMRB allows the establishment of a biogeochemical cycle that facilitates chromium precipitation. Such a system could provide a means for establishing and maintaining remedial redox reactive zones in Fe(III)-bearing subsurface environments.     

Biogeochemical Elimination of Chromium (VI) from Contaminated Water

Ferrous iron [Fe(II)] reductively transforms heavy metals in contaminated groundwater, and the bacterial reduction of indigenous ferric iron [Fe(III)] to Fe(II) has been proposed as a means of establishing redox reactive barriers in the subsurface. The reduction of Fe(III) to Fe(II) can be accomplished by stimulation of indigenous dissimilatory metal-reducing bacteria (DMRB) or injection of DMRB into the subsurface. The microbially produced Fe(II) can chemically react with contaminants such as Cr(VI) to form insoluble Cr(III) precipitates. The DMRB Shewanella algae BrY reduced surface-associated Fe(III) to Fe(II), which in batch and column experiments chemically reduced highly soluble Cr(VI) to insoluble Cr(III). Once the chemical Cr(VI) reduction capacity of the Fe(II)/Fe(III) couple in the experimental systems was exhausted, the addition of S. algae BrY allowed for the repeated reduction of Fe(III) to Fe(II), which again reduced Cr(VI) to Cr(III). The research presented herein indicates that a biological process using DMRB allows the establishment of a biogeochemical cycle that facilitates chromium precipitation. Such a system could provide a means for establishing and maintaining remedial redox reactive zones in Fe(III)-bearing subsurface environments.     

Influence of calcium on microbial reduction of solid phase uranium(VI)

The effect of calcium on the dissolution and microbial reduction of a representative solid phase uranyl [U(VI)], sodium boltwoodite (NaUO(2)SiO(3)OH . 1.5H(2)O), was investigated to evaluate the rate-limiting step of microbial reduction of the solid phase U(VI). Microbial reduction experiments were performed in a culture of a dissimilatory metal-reducing bacterium (DMRB), Shewanella oneidensis strain MR-1, in a bicarbonate medium with lactate as electron donor at pH 6.8 buffered with PIPES. Calcium increased the rate of Na-boltwoodite dissolution and U(VI) bioavailability by increasing its solubility through the formation of a ternary aqueous calcium-uranyl-carbonate species. The ternary species, however, decreased the rates of microbial reduction of aqueous U(VI). Laser-induced fluorescence spectroscopy (LIFS) and transmission electron microscopy (TEM) collectively revealed that microbial reduction of solid phase U(VI) was a sequentially coupled process of Na-boltwoodite dissolution, U(VI) aqueous speciation, and microbial reduction of dissolved U(VI) to U(IV) that accumulated on bacterial surfaces/periplasm. Under studied experimental conditions, the overall rate of microbial reduction of solid phase U(VI) was limited by U(VI) dissolution reactions in solutions without calcium and limited by microbial reduction in solutions with calcium. Generally, the overall rate of microbial reduction of solid phase U(VI) was determined by the coupling of solid phase U(VI) dissolution, U(VI) aqueous speciation, and microbial reduction of dissolved U(VI) that were all affected by calcium.     

Wolbachia在熊蜂中的双重感染

Wolbachia是一类广泛存在于节肢动物体内细胞质遗传的细菌,它可以通过诱导产雌孤雌生殖、引起细胞质不亲和、遗传雄性的雌性化、雄性致死和增强生殖力等作用方式引起其寄主生殖行为的改变.本文以16S rDNA为标记检测了3种熊蜂不同组织(头,胸,足,卵巢或雄外生殖器)的Wolbachia感染.其中明亮熊蜂Bombus l...     

Cellular and biochemical response to Cr(VI) in Stenotrophomonas sp.

Chromium (Cr)-resistant bacteria isolated from a soil with 6 g kg⁻¹ of Cr were identified based on 16S rRNA gene sequence analysis as a Stenotrophomonas , and designated as JD1. Growth of JD1 was accompanied by transformation of Cr(VI) to Cr(III) in liquid medium initially containing 300 mg L⁻¹ Cr(VI), the maximum concentration allowing growth. JD1 produced the highest levels of a Cr(VI)-binding exopolysaccharide when grown in medium with 100 mg L⁻¹ Cr(VI). The relative exopolysaccharide monosaccharide composition was analysed by HPLC, which showed that rhamnose+galactose was the major component, and that its relative level increased when cells were grown with Cr(VI). JD1 grew as a biofilm on various inert surfaces. Biofilm macromolecular composition analysis indicated that the relative levels of exopolysaccharide and protein were more abundant in biofilms grown in 100 mg L⁻¹ Cr(VI), whereas relative uronic acid levels remained constant. Biofilm cells exposed to Cr(VI) were elongated, grouped in clusters and exopolysaccharide obtained from the biofilm extracellular matrix had an enhanced capacity to bind Cr(VI). Exopolysaccharide production and composition, and biofilm growth are discussed as a mechanism of protection that allows survival during Cr(VI) stress.     

Effect of complexing agents on reduction of Cr(VI) by Desulfovibrio vulgaris ATCC 29579

The reduction of Cr(VI) at the expense of molecular hydrogen was studied using resting cells of Desulfovibrio vulgaris ATCC 29579 in anaerobic resting cell suspensions in MOPS buffer. Bioreduction occurred only in the presence of ligands or chelating agents (CO32-, citrate, NTA, EDTA, DTPA). The stimulatory effect of these ligands on the rate of Cr(VI) reduction was correlated (r = 0.988) with the strength of the ligand/chelate complex of Cr(III). The data are examined with respect to likely solution and redox equilibria in the ionic matrix of the carrier solution, and with respect to the potential for bioremediation of Cr(VI).     

Community structure of Archaea and Bacteria in a profundal lake sediment Lake Kinneret (Israel)

The microbial community structure of an anoxic profundal lake sediment, i.e., subtropical Lake Kinneret, was analysed with respect to its composition by culture-independent molecular methods including terminal restriction fragment length polymorphism (T-RFLP) analysis, comparative sequence analysis, and quantitative real-time PCR. In particular we were interested in the structure, species composition, and relative abundance of the overall microbial community in the methanogenic sediment layer (0-10 cm depth). Pairwise comparison of archaeal and bacterial 16S rRNA gene T-RFLP profiles obtained from three independent samplings indicated stability of the microbial community. The numbers of Archaea and Bacteria, quantified by real-time PCR, amounted to about 10(8) and 10(10) 16S rRNA gene copies cm(-3) sediment, respectively, suggesting that Archaea may account for only a minor fraction (approximately 1%) of the total prokaryotic community. Hydrogenotrophic Methanomicrobiales and acetoclastic Methanosaeta spp. dominated T-RFLP profiles of the archaeal community. T-RFLP profiles of the bacterial community were dominated by Deltaproteobacteria, sulphate reducers and syntrophs in particular. The second most abundant group was assigned to the Bacteroidetes-Chlorobi-group. Only one bacterial group, which was affiliated with halorespiring bacteria of subphylum II of the Chloroflexi, showed variation in abundance within the sediment samples investigated. Our study gives a comprehensive insight into the structure of the bacterial and archaeal community of a profundal lake sediment, indicating that sulphate reducers, syntrophs, bacteroidetes, halorespirers and methanogens are of particular importance in Lake Kinneret sediment.     

Nitrogen-fixing nodules from rose wood legume trees (Dalbergia spp.) endemic to Madagascar host seven different genera belonging to alpha- and beta-Proteobacteria

Although legume biodiversity is concentrated in tropical regions, the majority of studies on legume nodulating bacteria (LNB) are focused on cultivated leguminous plants from temperate regions. However, recent works on tropical regions tend to indicate that the actual diversity of LNB is largely underestimated. In this study, we report the isolation and characterization of 68 nitrogen-fixing root nodule bacteria collected from eight endemic tree species of Dalbergia in Madagascar. The isolates were characterized by (i) restriction fragment length polymorphism (RFLP) analysis of 16S-IGS rDNA, (ii) 16S rDNA gene sequencing and (iii) nodulation tests. Results revealed a wide diversity of bacteria present in the nodules of Dalbergia. Among the 68 isolated bacteria, 65 belonged to Bradyrhizobium, Mesorhizobium, Rhizobium, Azorhizobium and Phyllobacterium from the alpha-class of Proteobacteria, and three isolates belonged to Burkholderia and Ralstonia from the beta-class of Proteobacteria. Our results also show for the first time that a strain belonging to the Burkholderia cepacia complex is able to induce efficient nodules on a legume plant.     

Nonreductive Biomineralization of Uranium(VI) Phosphate Via Microbial Phosphatase Activity in Anaerobic Conditions

The remediation of uranium from soils and groundwater at Department of Energy (DOE) sites across the United States represents a major environmental issue, and bioremediation has exhibited great potential as a strategy to immobilize U in the subsurface. The bioreduction of U(VI) to insoluble U(IV) uraninite has been proposed to be an effective bioremediation process in anaerobic conditions. However, high concentrations of nitrate and low pH found in some contaminated areas have been shown to limit the efficiency of microbial reduction of uranium. In the present study, nonreductive uranium biomineralization promoted by microbial phosphatase activity was investigated in anaerobic conditions in the presence of high nitrate and low pH as an alternative approach to the bioreduction of U(VI). A facultative anaerobe, Rahnella sp. Y9602, isolated from soils at DOE's Oak Ridge Field Research Center (ORFRC), was able to respire anaerobically on nitrate as a terminal electron acceptor in the presence of glycerol-3-phosphate (G3P) as the sole carbon and phosphorus source and hydrolyzed sufficient phosphate to precipitate 95% total uranium after 120 hours in synthetic groundwater at pH 5.5. Synchrotron X-ray diffraction and X-ray absorption spectroscopy identified the mineral formed as chernikovite, a U(VI) autunite-type mineral. The results of this study suggest that in contaminated subsurfaces, such as at the ORFRC, where high concentrations of nitrate and low pH may limit uranium bioreduction, the biomineralization of U(VI) phosphate minerals may be a more attractive approach for in situ remediation providing that a source of organophosphate is supplied for bioremediation.     

稻纵卷叶螟感染Wolbachia的ftsZ基因和16S rDNA基因的序列分析

Wolbachia是一类胞质遗传的内共生菌, 广泛分布于节肢动物和其他动物中, 与宿主的生殖调控密切相关。通过研究迁飞性害虫稻纵卷叶螟Cnaphalocrocis medinalis (Guenée)的Wolbachia感染情况, 为探讨Wolbachia在迁飞性昆虫中的生殖调控和传递方式等提供基础资料。本研究应用Wolbachia的ftsZ基因和16S rDNA基因的特异性引物, 通过PCR扩增的方法对我国20个地区的稻纵卷叶螟样本进行了检测。结果表明: 中国不同地区的稻纵卷叶螟感染Wolbachia的现象较为普遍, 其中浙江温州和江苏扬州样本的感染率最高（90%）; 四川雅安、 湖南长沙和天津宁河样本的感染率最低（40%）。不同地区稻纵卷叶螟的Wolbachia ftsZ基因序列完全一致, 而且不同地区的Wolbachia 16S rDNA基因序列也完全相同。此外, 稻纵卷叶螟感染的Wolbachia ftsZ基因和16S rDNA基因序列与其他物种感染的Wolbachia B群的ftsZ基因序列和16S rDNA基因序列相似性分别在99%~100%和98%~99%之间, 说明我国稻纵卷叶螟感染的Wolbachia隶属B群。研究结果表明, 稻纵卷叶螟感染的Wolbachia类型较为单一, 这也是我国有关稻纵卷叶螟内共生菌Wolbachia的首次研究报道。     

Aerobic Cr(VI) Reduction by Bacteria in Culture and Soil Conditions

We compared the performance of aerobic Cr(VI)-reducing bacteria isolated from Cr(VI)-contaminated soil in pure and mixed cultures of five isolated strains. The mixed culture had increased reduction rates compared to individual cultures. Cr(VI) reduction was observed in sterile soil inoculated with Pseudomonas fluorescens and in non-sterile soil with and without inoculation with P. fluorescens at initial pore water concentrations up to 1,600 mg Cr(VI)/L, whereas in culture the maximum inhibitory concentration was 500 mg Cr(VI)/L. Linear rates of Cr(VI) reduction in non-sterile soil amended with peptone were ～5 to 8 times higher than those observed in the mixed culture. Inoculation of non-sterile soil with P. fluorescens did not further enhance Cr(VI) reduction rates. Our results indicate that evaluation of Cr(VI) reduction capacity in Cr(VI)-contaminated soil for in-situ bioremediation purposes should not be done solely in pure culture. Although the latter may be used initially to assess the effects of process parameters (e.g., pH, temperature), the rate and extent of Cr(VI) reduction should be determined in soil for bioremediation design purposes.     

基于16S rDNA基因序列的泽兰实蝇幼虫肠道细菌多样性分析

【目的】探究泽兰实蝇 Procecidochares utilis 幼虫肠道细菌的多样性。【方法】利用Illumina HiSeq技术对泽兰实蝇幼虫肠道细菌的16S rDNA-V6变异区序列进行测序,应用USEARCH和QIIME等软件整理和统计样品序列数目和操作分类单元（operational taxonomic unit, OTU）数量,分析物种的丰度和Alpha多样性。【结果】共获得1 593 506 对reads,拼接为1 579 372条tags,经过滤后得到的1 572 860条tags聚类为1 341个OTU。总共注释到13个门,4个纲,6个目,7个科,10个属和4个种。其中变形菌门(Proteobacteria)的细菌为优势菌,占99%;在属分类阶元上,沃尔巴克氏体属 Wolbachia 占45%,是优势属。【结论】泽兰实蝇幼虫肠道细菌多样性丰富。相关的种类和丰度信息为后期研究揭示肠道细菌介导泽兰实蝇寄主植物专化性奠定了基础。     

稻纵卷叶螟肠道细菌群落结构与多样性分析

【目的】为明确水稻主要害虫稻纵卷叶螟Cnaphalocrocis medinalis(Guenée)幼虫肠道细菌的群落结构和多样性。【方法】利用Illumina Mi Seq技术对稻纵卷叶螟4龄幼虫肠道细菌的16S r DNA V3-V4变异区序列进行测序,应用USEARCH和QIIME软件整理和统计样品序列数和操作分类单元(operational taxonomic unit,OTU)数量,分析4龄幼虫肠道细菌的物种组成、丰度和多样性。【结果】稻纵卷叶螟4个数量不同的4龄幼虫样本(1,2,3和5头)共得165 386条reads,在97%相似度下可将其聚类为604个OTUs。总共注释到22个门,43个纲,82个目,142个科,204个属,244个种。其中在门水平上,主要优势菌为变形菌门(Proteobacteria)(相对丰度26%~34%)和放线菌门(Actinobacteria)(23%~32%);在纲水平上,主要优势菌为放线菌纲(Actinobacteria)(相对丰度23%~32%)、酸杆菌纲(Acidobacteria)(9%~11%)、α-变形菌纲(Alphaproteobacteria)(10%~13%)、β-变形菌纲(Betaproteobacteria)(6%~8%)和γ-变形菌纲(Gammaproteobacteria)(6%~12%);在科水平上,共有优势菌为类诺卡氏菌科(Nocardioidaceae)、丛毛单胞菌科(Comamonadaceae)、黄单胞菌科(Xanthomonadaceae)、鞘脂单胞菌科(Sphingomonadaceae)和芽单胞菌科(Gemmatimonadaceae)等。在属水平上,4个样本前5位优势属中,类诺卡氏属Nocardioides和鞘脂单胞菌Sphingomonas为共有优势属。稻纵卷叶螟肠道细菌Simpson指数、Shannon指数、Ace指数和Chao指数分别为0.16~0.65,0.94~3.22,212~488和210~490。【结论】稻纵卷叶螟幼虫肠道细菌多样性比较丰富,个体间微生物群落结构和多样性有差异。不同数量样本数据与总体数据有助于综合反映稻纵卷叶螟种群肠道微生物状况。本研究结果为进一步研究稻纵卷叶螟肠道微生物的功能及其在防治中的应用奠定了基础。