scholarly journals Anaerobic reduction of europium by a Clostridium strain as a strategy for rare earth biorecovery

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Maleke Maleke ◽  
Angel Valverde ◽  
Alba Gomez-Arias ◽  
Errol D. Cason ◽  
Jan-G Vermeulen ◽  
...  
Keyword(s):  
Resistance Mechanism ◽  
Growth Conditions ◽  
Protein Characterization ◽  
Intracellular Accumulation ◽  
Carbonatite Complex ◽  
Edx Analysis ◽  
Primary Mineral ◽  
Pyruvate Oxidoreductase ◽  
A Site ◽  
Anaerobic Reduction

Abstract The biorecovery of europium (Eu) from primary (mineral deposits) and secondary (mining wastes) resources is of interest due to its remarkable luminescence properties, important for modern technological applications. In this study, we explored the tolerance levels, reduction and intracellular bioaccumulation of Eu by a site-specific bacterium, Clostridium sp. 2611 isolated from Phalaborwa carbonatite complex. Clostridium sp. 2611 was able to grow in minimal medium containing 0.5 mM Eu3+. SEM-EDX analysis confirmed an association between Eu precipitates and the bacterium, while TEM-EDX analysis indicated intracellular accumulation of Eu. According to the HR-XPS analysis, the bacterium was able to reduce Eu3+ to Eu2+ under growth and non-growth conditions. Preliminary protein characterization seems to indicate that a cytoplasmic pyruvate oxidoreductase is responsible for Eu bioreduction. These findings suggest the bioreduction of Eu3+ by Clostridium sp. as a resistance mechanism, can be exploited for the biorecovery of this metal.

scholarly journals The Escherichia coli cysG promoter belongs to the ‘extended −10’ class of bacterial promoters

1993 ◽  
Vol 296 (3) ◽  
pp. 851-857 ◽  
Author(s):  
T Belyaeva ◽  
L Griffiths ◽  
S Minchin ◽  
J Cole ◽  
S Busby
Keyword(s):  
Escherichia Coli ◽  
Point Mutations ◽  
Growth Conditions ◽  
Upstream Region ◽  
E Coli ◽  
Run Off ◽  
A Site ◽  
Specific Sequences

The Escherichia coli cysG promoter has been subcloned and shown to function constitutively in a range of different growth conditions. Point mutations identify the -10 hexamer and an important 5′-TGN-3′ motif immediately upstream. The effects of different deletions suggest that specific sequences in the -35 region are not essential for the activity of this promoter in vivo. This conclusion was confirmed by in vitro run-off transcription assays. The DNAase I footprint of RNA polymerase at the cysG promoter reveals extended protection upstream of the transcript start, and studies with potassium permanganate as a probe suggest that the upstream region is distorted in open complexes. Taken together, the results show that the cysG promoter belongs to the ‘extended -10’ class of promoters, and the base sequence is similar to that of the P1 promoter of the E. coli galactose operon, another promoter in this class. In vivo, messenger initiated at the cysG promoter appears to be processed by cleavage at a site 41 bases downstream from the transcript start point.

scholarly journals Chromate Reduction by a Pseudomonad Isolated from a Site Contaminated with Chromated Copper Arsenate

2001 ◽  
Vol 67 (3) ◽  
pp. 1076-1084 ◽  
Author(s):  
Jeff McLean ◽  
Terry J. Beveridge
Keyword(s):  
Reductase Activity ◽  
Wood Preservation ◽  
Chromate Reduction ◽  
Anaerobic Conditions ◽  
Toxic Heavy Metal ◽  
Metal Metal ◽  
A Site ◽  
Anaerobic Reduction ◽  
Aerobic And Anaerobic ◽  
Aerobic And Anaerobic Conditions

ABSTRACT A pseudomonad (CRB5) isolated from a decommissioned wood preservation site reduced toxic chromate [Cr(VI)] to an insoluble Cr(III) precipitate under aerobic and anaerobic conditions. CRB5 tolerated up to 520 mg of Cr(VI) liter−1 and reduced chromate in the presence of copper and arsenate. Under anaerobic conditions it also reduced Co(III) and U(VI), partially internalizing each metal. Metal precipitates were also found on the surface of the outer membrane and (sometimes) on a capsule. The results showed that chromate reduction by CRB5 was mediated by a soluble enzyme that was largely contained in the cytoplasm but also found outside of the cells. The crude reductase activity in the soluble fraction showed aKm of 23 mg liter−1 (437 μM) and a V max of 0.98 mg of Cr h−1 mg of protein−1 (317 nmol min−1 mg of protein−1). Minor membrane-associated Cr(VI) reduction under anaerobiosis may account for anaerobic reduction of chromate under nongrowth conditions with an organic electron donor present. Chromate reduction under both aerobic and anaerobic conditions may be a detoxification strategy for the bacterium which could be exploited to bioremediate chromate-contaminated or other toxic heavy metal-contaminated environments.

Mechanisms of Bacterial Survival in Chlorinated Drinking Water

1988 ◽  
Vol 20 (11-12) ◽  
pp. 145-151 ◽  
Author(s):  
Mark W. LeChevallier ◽  
Cheryl D. Cawthon ◽  
Ramon G. Lee
Keyword(s):  
Drinking Water ◽  
Resistance Mechanism ◽  
Fold Increase ◽  
Growth Conditions ◽  
Resistance Mechanisms ◽  
Bacterial Survival ◽  
Nutrient Effects ◽  
Chlorinated Drinking Water ◽  
Bacterial Encapsulation ◽  
Microscope Slides

Experiments showed that attachment of bacteria to surfaces provided the greatest increase in disinfection resistance. Attachment of high nutrient grown, unencapsulated, Klebsiellapneumoniae to glass microscope slides afforded the microorganisms as much as a 150 fold increase in disinfection resistance. Other mechanisms which increased disinfection resistance included: the age of the biofilm, bacterial encapsulation and previous growth conditions (e.g. growth medium, and growth temperature). These factors increased chlorine resistance from two to ten fold. The choice of disinfectant residual was shown to influence the type of resistance mechanism observed. Disinfection by free chlorine was affected by surfaces, age of the biofilm, encapsulation and nutrient effects. Disinfection by monochloramine, however, was only affected by surfaces. Importantly, the research showed that these resistance mechanisms were multiplicative (e.g. the resistance provided by one mechanism could be multiplied by the resistance provided by a second). These results provide important insights to understand the survival of bacteria in chlorinated drinking water supplies.

scholarly journals Copper Stress Induces a Global Stress Response in Staphylococcus aureus and Represses sae and agr Expression and Biofilm Formation

2009 ◽  
Vol 76 (1) ◽  
pp. 150-160 ◽  
Author(s):  
Jonathan Baker ◽  
Sutthirat Sitthisak ◽  
Mrittika Sengupta ◽  
Miranda Johnson ◽  
R. K. Jayaswal ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Staphylococcus Aureus ◽  
Biofilm Formation ◽  
Transcriptional Profiling ◽  
Resistance Mechanism ◽  
Growth Conditions ◽  
Copper Resistance ◽  
Transcriptional Analysis ◽  
Copper Stress ◽  
High Copper

ABSTRACT Copper is an important cofactor for many enzymes; however, high levels of copper are toxic. Therefore, bacteria must ensure there is sufficient copper for use as a cofactor but, more importantly, must limit free intracellular levels to prevent toxicity. In this study, we have used DNA microarray to identify Staphylococcus aureus copper-responsive genes. Transcriptional profiling of S. aureus SH1000 grown in excess copper identified a number of genes which fall into four groups, suggesting that S. aureus has four main mechanisms for adapting to high levels of environmental copper, as follows: (i) induction of direct copper homeostasis mechanisms; (ii) increased oxidative stress resistance; (iii) expression of the misfolded protein response; and (iv) repression of a number of transporters and global regulators such as Agr and Sae. Our experimental data confirm that resistance to oxidative stress and particularly to H2O2 scavenging is an important S. aureus copper resistance mechanism. Our previous studies have demonstrated that Eap and Emp proteins, which are positively regulated by Agr and Sae, are required for biofilm formation under low-iron growth conditions. Our transcriptional analysis has confirmed that sae, agr, and eap are repressed under high-copper conditions and that biofilm formation is indeed repressed under high-copper conditions. Therefore, our results may provide an explanation for how copper films can prevent biofilm formation on catheters.

scholarly journals Pooled genome-wide CRISPR screening for basal and context-specific fitness gene essentiality in Drosophila cells

eLife ◽  
2018 ◽  
Vol 7 ◽  
Author(s):  
Raghuvir Viswanatha ◽  
Zhongchi Li ◽  
Yanhui Hu ◽  
Norbert Perrimon
Keyword(s):  
Cultured Cells ◽  
Growth Conditions ◽  
Vertebrate Lineage ◽  
Site Specific Integration ◽  
Genome Wide ◽  
Dna Libraries ◽  
A Genome ◽  
Context Specific ◽  
A Site ◽  
Drosophila Cells

Genome-wide screens in Drosophila cells have offered numerous insights into gene function, yet a major limitation has been the inability to stably deliver large multiplexed DNA libraries to cultured cells allowing barcoded pooled screens. Here, we developed a site-specific integration strategy for library delivery and performed a genome-wide CRISPR knockout screen in Drosophila S2R+ cells. Under basal growth conditions, 1235 genes were essential for cell fitness at a false-discovery rate of 5%, representing the highest-resolution fitness gene set yet assembled for Drosophila, including 407 genes which likely duplicated along the vertebrate lineage and whose orthologs were underrepresented in human CRISPR screens. We additionally performed context-specific fitness screens for resistance to or synergy with trametinib, a Ras/ERK/ETS inhibitor, or rapamycin, an mTOR inhibitor, and identified key regulators of each pathway. The results present a novel, scalable, and versatile platform for functional genomic screens in invertebrate cells.

scholarly journals Role of TnrA in Nitrogen Source-Dependent Repression of Bacillus subtilis Glutamate Synthase Gene Expression

2000 ◽  
Vol 182 (21) ◽  
pp. 5939-5947 ◽  
Author(s):  
Boris R. Belitsky ◽  
Lewis V. Wray ◽  
Susan H. Fisher ◽  
Dian E. Bohannon ◽  
Abraham L. Sonenshein
Keyword(s):  
Gene Expression ◽  
Bacillus Subtilis ◽  
Nitrogen Limitation ◽  
Glutamate Synthase ◽  
Growth Conditions ◽  
Ammonium Nitrogen ◽  
Bacterial Metabolism ◽  
Major Donor ◽  
A Site

ABSTRACT Synthesis of glutamate, the cell's major donor of nitrogen groups and principal anion, occupies a significant fraction of bacterial metabolism. In Bacillus subtilis, the gltABoperon, encoding glutamate synthase, requires a specific positive regulator, GltC, for its expression. In addition, the gltABoperon was shown to be repressed by TnrA, a regulator of several other genes of nitrogen metabolism and active under conditions of ammonium (nitrogen) limitation. TnrA was found to bind directly to a site immediately downstream of the gltAB promoter. As is true for other genes, the activity of TnrA at the gltAB promoter was antagonized by glutamine synthetase under certain growth conditions.

scholarly journals FORMATION OF SLOW-REACTING SUBSTANCE OF ANAPHYLAXIS IN HUMAN LUNG TISSUE AND CELLS BEFORE RELEASE

1974 ◽  
Vol 140 (5) ◽  
pp. 1133-1146 ◽  
Author(s):  
Robert A. Lewis ◽  
Stephen I. Wasserman ◽  
Edward J. Goetzl ◽  
K. Frank Austen
Keyword(s):  
Lung Tissue ◽  
Cyclic Nucleotides ◽  
Human Lung ◽  
Intracellular Accumulation ◽  
Immediate Hypersensitivity ◽  
Human Lung Tissue ◽  
Additional Dimension ◽  
Cellular Events ◽  
A Site ◽  
Surrounding Fluid

The capacity to extract slow-reacting substance of anaphylaxis (SRS-A) from human lung tissue or cells after immunologic activation, together with the measurement of SRS-A in both the extract and the surrounding fluid, permits study of total SRS-A generation. That the material extracted is SRS-A was established by both differential bioassay and purification. SRS-A accumulation was entirely intracellular after limited IgE-dependent direct or reversed anaphylactic activation. Intracellular accumulation also generally preceded release, with generation of SRS-A continuing well beyond a plateau in the cellular SRS-A level and the release of preformed mediators. The quantity of SRS-A generated after immunologic activation was modulated by the introduction of exogenous cyclic nucleotides, revealing a site of cyclic nucleotide action distinct from that on mediator release. The capacity to determine not only the release of preformed mediators but also the generation of a newly formed mediator, the sum of SRS-A in cells and supernate, adds an additional dimension to the analysis of the cellular events of immediate hypersensitivity.

scholarly journals CcpA-Dependent and -Independent Control of Beta-Galactosidase Expression in Streptococcus pneumoniae Occurs via Regulation of an Upstream Phosphotransferase System-Encoding Operon

2007 ◽  
Vol 189 (14) ◽  
pp. 5183-5192 ◽  
Author(s):  
Greer E. Kaufman ◽  
Janet Yother
Keyword(s):  
Streptococcus Pneumoniae ◽  
Protein A ◽  
Surface Protein ◽  
Growth Conditions ◽  
Spontaneous Mutant ◽  
Galactosidase Activity ◽  
Phosphotransferase System ◽  
Catabolite Control Protein A ◽  
Responsive Element ◽  
A Site

ABSTRACT A spontaneous mutant of Streptococcus pneumoniae strain D39 exhibiting elevated β-galactosidase activity was identified. We determined that the β-galactosidase activity was due to BgaA, a surface protein in S. pneumoniae, and that the expression of bgaA was regulated. Transcription analyses demonstrated expression of bgaA in the constitutive β-galactosidase (BgaAC) mutant, but not in the parent. β-Galactosidase expression was induced in the parent under specific growth conditions; however, the levels did not reach those of the BgaAC mutant. We localized the mutation resulting in the BgaAC phenotype to a region upstream of bgaA and in the promoter of a phosphoenolpyruvate-dependent phosphotransferase system (PTS) operon. The mutation was in a catabolite-responsive element (cre) and affected the binding of CcpA (catabolite control protein A), a key regulator of many carbon metabolism genes. The pts operon and bgaA were cotranscribed, and their transcription was regulated by CcpA. Deletion of ccpA altered β-galactosidase activity, leading to a sevenfold increase in the parent but a fivefold decrease in the BgaAC mutant. The resulting β-galactosidase activities were the same in the two strains, suggesting the presence of a second repressor. The presence of glucose in the growth medium resulted in pts-bgaA repression by both CcpA and the second repressor, with the latter being important in responding to the glucose concentration. Expression of β-galactosidase is important for S. pneumoniae adherence during colonization of the nasopharynx, a site normally devoid of glucose. CcpA and environmental glucose concentrations thus appear to play important roles in the regulation of a niche-specific virulence factor.

scholarly journals Modeling site-specific nucleotide biases affecting Himar1 transposon insertion frequencies in TnSeq datasets

2021 ◽  
Author(s):  
Sanjeevani Choudhery ◽  
A Jacob Brown ◽  
Chidiebere D Akusobi ◽  
Eric J. Rubin ◽  
Christopher M Sassetti ◽  
...  
Keyword(s):  
Markov Models ◽  
A Priori ◽  
Growth Conditions ◽  
Specific Model ◽  
Site Specific ◽  
Transposon Insertion ◽  
Local Sequence ◽  
A Site ◽  
Analytical Approaches ◽  
Insertion Mutants

In bacterial TnSeq experiments, a library of transposons insertion mutants is generated, selected under various growth conditions, and sequenced to determine the profile of insertions at different sites in the genome, from which the fitness of mutant strains can be inferred. The widely used Himar1 transposon is known to be restricted to insertions at TA dinucleotides, but otherwise, few site-specific biases have been identified. As a result, most analytical approaches assume that insertion counts are expected a priori to be randomly distributed among TA sites in non-essential regions. However, recent analyses of independent Himar1 Tn libraries in M. tuberculosis have identified a local sequence pattern that is non-permissive for Himar1 insertion. This suggests there are site-specific biases that affect the frequency of insertions of the Himar1 transposon at different TA sites. In this paper, we use statistical and machine learning models to characterize patterns in the nucleotides surrounding TA sites associated with high and low insertion counts. We not only affirm that the previously discovered non-permissive pattern (CG)GnTAnC(CG) suppresses insertions, but conversely show that an A in the -3 position or T in the +3 position from the TA site encourages them. We demonstrate that these insertion preferences exist in Himar1 TnSeq datasets other than M. tuberculosis, including mycobacterial and non-mycobacterial species. We build predictive models of Himar1 insertion preferences as a function of surrounding nucleotides. The final predictive model explains about half of the variance in insertion counts, presuming the rest comes from stochastic variability between libraries or due to sampling differences during sequencing. Based on this model, we present a new method, called the TTN-Fitness method, to improve the identification of conditionally essential genes or genetic interactions, i.e., to better distinguish true biological fitness effects by comparing the observed counts to expected counts using a site-specific model of insertion preferences. Compared to previous methods like Hidden Markov Models, the TTN-Fitness method is able to classify the essentiality of many small genes (with few TA sites) that were previously characterized as Uncertain.

Hoeflea anabaenae sp. nov., an epiphytic symbiont that attaches to the heterocysts of a strain of Anabaena

2011 ◽  
Vol 61 (10) ◽  
pp. 2439-2444 ◽  
Author(s):  
Bradley S. Stevenson ◽  
Michael T. Suflita ◽  
Blake W. Stamps ◽  
Edward R. B. Moore ◽  
Crystal N. Johnson ◽  
...  
Keyword(s):  
Type Strain ◽  
Optimal Growth ◽  
Growth Conditions ◽  
Rrna Gene ◽  
Polar Lipid Profile ◽  
Growth Requirements ◽  
The Family ◽  
C 30 ◽  
Microaerobic Conditions ◽  
A Site

The heterotrophic, epiphytic, symbiotic bacterial strain WH2KT was previously isolated from a two-member culture in which it was attached to the heterocysts of a strain of Anabaena (SSM-00). Analysis of its 16S rRNA gene sequence demonstrated that the symbiont was most closely related to the type strain of Hoeflea marina (96.9 % similarity), which belongs to the family Phyllobacteriaceae within the order Rhizobiales of the class Alphaproteobacteria. A polyphasic taxonomic study was performed on strain WH2KT, which consisted of irregular rods (2–5 µm long, 0.2 µm wide) that appeared to be narrower at one pole. Optimal growth was obtained in complex media with 15 g sea salts l−1, at 18–34 °C (30 °C optimum) and at pH 6.0–8.0 (optimum pH 6.5). Unknown growth requirements were provided by small amounts of yeast extract but not by standard vitamin and trace metal solutions. Of the substrates tested, WH2KT was able to utilize only acetate, pyruvate, malate and fumarate. Growth was observed only under aerobic and microaerobic conditions, and nitrate was not reduced. No photosynthetic pigments were detected under any of the growth conditions tested. The predominant fatty acids were a summed feature that comprises C18 : 1ω7c, C18 : 1ω9t, C18 : 1ω12t or any combination of these (64.0 %) and an unidentified fatty acid of equivalent chain length 17.603 (13.5 %). The polar lipid profile consisted of diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine, phosphatidylmonomethylethanolamine, phosphatidylcholine, phosphoglycolipid, unknown lipids and an unidentified aminolipid. The only respiratory ubiquinone detected was Q-10. The DNA G+C content of the strain was 58.1 mol%. The organism can form a site-specific attached symbiotic relationship with a species of Anabaena. Based on phylogenetic and phenotypic evidence, it is proposed that strain WH2KT be classified within a novel species of the genus Hoeflea, for which the name Hoeflea anabaenae sp. nov. is proposed. The type strain is WH2KT ( = CCUG 56626T  = NRRL B-59520T).

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