scholarly journals Effect of Nitrate and Acetylene on nirS, cnorB, and nosZ Expression and Denitrification Activity in Pseudomonas mandelii

2009 ◽  
Vol 75 (15) ◽  
pp. 5082-5087 ◽  
Author(s):  
Saleema Saleh-Lakha ◽  
Kelly E. Shannon ◽  
Sherri L. Henderson ◽  
Bernie J. Zebarth ◽  
David L. Burton ◽  
...  
Keyword(s):  
Gene Expression ◽  
Gas Chromatography ◽  
Electron Acceptor ◽  
Pseudomonas Mandelii ◽  
Denitrification Gene ◽  
Reverse Transcription Quantitative Pcr ◽  
Denitrification Activity ◽  
Acetylene Blockage ◽  
Denitrification Genes ◽  
Denitrification Process

ABSTRACT Nitrate acts as an electron acceptor in the denitrification process. The effect of nitrate in the range of 0 to 1,000 mg/liter on Pseudomonas mandelii nirS, cnorB, and nosZ gene expression was studied, using quantitative reverse transcription-quantitative PCR. Denitrification activity was measured by using the acetylene blockage method and gas chromatography. The effect of acetylene on gene expression was assessed by comparing denitrification gene expression in P. mandelii culture grown in the presence or absence of acetylene. The higher the amount of NO3 − present, the greater the induction and the longer the denitrification genes remained expressed. nirS gene expression reached a maximum at 2, 4, 4, and 6 h in cultures grown in the presence of 0, 10, 100, and 1,000 mg of KNO3/liter, respectively, while induction of nirS gene ranged from 12- to 225-fold compared to time zero. cnorB gene expression also followed a similar trend. nosZ gene expression did not respond to NO3 − treatment under the conditions tested. Acetylene decreased nosZ gene expression but did not affect nirS or cnorB gene expression. These results showed that nirS and cnorB responded to nitrate concentrations; however, significant denitrification activity was only observed in culture with 1,000 mg of KNO3/liter, indicating that there was no relationship between gene expression and denitrification activity under the conditions tested.

scholarly journals Effect of pH and Temperature on Denitrification Gene Expression and Activity in Pseudomonas mandelii

2009 ◽  
Vol 75 (12) ◽  
pp. 3903-3911 ◽  
Author(s):  
Saleema Saleh-Lakha ◽  
Kelly E. Shannon ◽  
Sherri L. Henderson ◽  
Claudia Goyer ◽  
Jack T. Trevors ◽  
...  
Keyword(s):  
Gene Expression ◽  
Fold Increase ◽  
Low Ph ◽  
Effect Of Ph ◽  
Liquid Cultures ◽  
Ph Values ◽  
Reverse Transcription Pcr ◽  
Pseudomonas Mandelii ◽  
Denitrification Gene ◽  
Denitrification Activity

ABSTRACT Pseudomonas mandelii liquid cultures were studied to determine the effect of pH and temperature on denitrification gene expression, which was quantified by quantitative reverse transcription-PCR. Denitrification was measured by the accumulation of nitrous oxide (N2O) in the headspace in the presence of acetylene. Levels of gene expression of nirS and cnorB at pH 5 were 539-fold and 6,190-fold lower, respectively, than the levels of gene expression for cells grown at pH 6, 7, and 8 between 4 h and 8 h. Cumulative denitrification levels were 28 μmol, 63 μmol, and 22 μmol at pH 6, 7, and 8, respectively, at 8 h, whereas negligible denitrification was measured at pH 5. P. mandelii cells grown at 20°C and 30°C exhibited 9-fold and 94-fold increases in levels of cnorB expression between 0 h and 2 h, respectively, and an average 17-fold increase in levels of nirS gene expression. In contrast, induction of cnorB and nirS gene expression for P. mandelii cells grown at 10°C did not occur in the first 4 h. Levels of cumulative denitrification at 10 h were 6.6 μmol for P. mandelii cells grown at 10°C and 20°C and 30 μmol for cells grown at 30°C. Overall, levels of cnorB and nirS expression were relatively insensitive to pH values over the range of pH 6 to 8 but were substantially reduced at pH 5, whereas gene expression was sensitive to temperature, with induction and time to achieve maximum gene expression delayed as the temperature decreased from 30°C. Low pH and temperature negatively affected denitrification activity.

Influence of temperature on microbial colonisation of clayey schist as a support media of a submerged filter for groundwater denitrification

2007 ◽  
Vol 55 (8-9) ◽  
pp. 165-172 ◽  
Author(s):  
A. De la Rúa ◽  
B. Rodelas ◽  
J. González-López ◽  
M.A. Gómez
Keyword(s):  
Biofilm Formation ◽  
Effect Of Temperature ◽  
Cold Environments ◽  
Pseudomonas Mandelii ◽  
Different Temperatures ◽  
Influence Of Temperature On ◽  
Denitrification Activity ◽  
Natural Microbiota ◽  
Denitrification Process ◽  
Made In

The effect of temperature on biofilm formation and denitrification activity was evaluated. Assays were made in a lab-scale submerged filter for the denitrification of polluted groundwater, with and without a previous inoculation. The inoculation was carried out with a selected strain of Pseudomonas mandelii. Different temperatures were tested: 5, 10, 20 and 30 °C. Biofilm observations were made, and monitoring of the denitrification capacity of the system was maintained during the experiment. Our results showed that both colonisation of the support material of the filter and biofilm maturity have a dependency with temperature, with an optimum temperature of 20 °C if the system was previously inoculated with the Pseudomonas mandelii strain. For a correct achievement of the denitrification process, a previous inoculation of the system is essential. Although the development of a biofilm from the natural microbiota present in the groundwater is possible, it is not capable to adequately denitrify polluted groundwater. In terms of the correct achievement of the denitrification process, temperature affects the operation of the system at cold environments, although the use of Pseudomonas mandelii strain A103 allows denitrification at 10–30 °C with very good results (above 90% removal), affecting only to the time needed for the stabilisation of the system.

scholarly journals Edaphic factors and plants influence denitrification in soils from a long-term arable experiment

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Ian M. Clark ◽  
Qingling Fu ◽  
Maïder Abadie ◽  
Elizabeth R. Dixon ◽  
Aimeric Blaud ◽  
...  
Keyword(s):  
Gas Production ◽  
Soil Treatment ◽  
Soil Microbiome ◽  
Arable Soils ◽  
Experimental Conditions ◽  
Mineral N ◽  
Nitrate Availability ◽  
Denitrification Gene ◽  
Denitrification Genes

Abstract Factors influencing production of greenhouse gases nitrous oxide (N2O) and nitrogen (N2) in arable soils include high nitrate, moisture and plants; we investigate how differences in the soil microbiome due to antecedent soil treatment additionally influence denitrification. Microbial communities, denitrification gene abundance and gas production in soils from tilled arable plots with contrasting fertilizer inputs (no N, mineral N, FYM) and regenerated woodland in the long-term Broadbalk field experiment were investigated. Soil was transferred to pots, kept bare or planted with wheat and after 6 weeks, transferred to sealed chambers with or without K15NO3 fertilizer for 4 days; N2O and N2 were measured daily. Concentrations of N2O were higher when fertilizer was added, lower in the presence of plants, whilst N2 increased over time and with plants. Prior soil treatment but not exposure to N-fertiliser or plants during the experiment influenced denitrification gene (nirK, nirS, nosZI, nosZII) relative abundance. Under our experimental conditions, denitrification generated mostly N2; N2O was around 2% of total gaseous N2 + N2O. Prior long-term soil management influenced the soil microbiome and abundance of denitrification genes. The production of N2O was driven by nitrate availability and N2 generation increased in the presence of plants.

scholarly journals Selection of reference genes for measuring the expression of aiiO in Ochrobactrum quorumnocens A44 using RT-qPCR

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Dorota M. Krzyżanowska ◽  
Anna Supernat ◽  
Tomasz Maciąg ◽  
Marta Matuszewska ◽  
Sylwia Jafra
Keyword(s):  
Gene Expression ◽  
Quorum Sensing ◽  
Reference Genes ◽  
Significance Threshold ◽  
Expression Studies ◽  
Reverse Transcription Quantitative Pcr ◽  
Gene Expression Studies ◽  
The Stability ◽  
Selection Of

Abstract Reverse transcription quantitative PCR (RT-qPCR), a method of choice for quantification of gene expression changes, requires stably expressed reference genes for normalization of data. So far, no reference genes were established for the Alphaproteobacteria of the genus Ochrobactrum. Here, we determined reference genes for gene expression studies in O. quorumnocens A44. Strain A44 was cultured under 10 different conditions and the stability of expression of 11 candidate genes was evaluated using geNorm, NormFinder and BestKeeper. Most stably expressed genes were found to be rho, gyrB and rpoD. Our results can facilitate the choice of reference genes in the related Ochrobactrum strains. O. quorumnocens A44 is able to inactivate a broad spectrum of N-acyl homoserine lactones (AHLs) – the quorum sensing molecules of many Gram-negative bacteria. This activity is attributed to AiiO hydrolase, yet it remains unclear whether AHLs are the primary substrate of this enzyme. Using the established RT-qPCR setup, we found that the expression of the aiiO gene upon exposure to two AHLs, C6-HLS and 3OC12-HSL, does not change above the 1-fold significance threshold. The implications of this finding are discussed in the light of the role of quorum sensing-interfering enzymes in the host strains.

scholarly journals Transcriptomic and Proteomic Responses of the Organohalide-Respiring Bacterium Desulfoluna spongiiphila to Growth with 2,6-Dibromophenol as the Electron Acceptor

2019 ◽  
Vol 86 (5) ◽  
Author(s):  
Jie Liu ◽  
Lorenz Adrian ◽  
Max M. Häggblom
Keyword(s):  
Gene Expression ◽  
Secondary Metabolites ◽  
Electron Acceptor ◽  
Marine Sponge ◽  
Reductive Dehalogenation ◽  
Important Process ◽  
Organohalide Respiration ◽  
Content Type ◽  
Reductive Dehalogenase ◽  
Significant Difference

ABSTRACT Organohalide respiration is an important process in the global halogen cycle and for bioremediation. In this study, we compared the global transcriptomic and proteomic analyses of Desulfoluna spongiiphila strain AA1, an organohalide-respiring member of the Desulfobacterota isolated from a marine sponge, with 2,6-dibromophenol or with sulfate as an electron acceptor. The most significant difference of the transcriptomic analysis was the expression of one reductive dehalogenase gene cluster (rdh16), which was significantly upregulated with the addition of 2,6-dibromophenol. The corresponding protein, reductive dehalogenase RdhA16032, was detected in the proteome under treatment with 2,6-dibromophenol but not with sulfate only. There was no significant difference in corrinoid biosynthesis gene expression levels between the two treatments, indicating that the production of corrinoid in D. spongiiphila is constitutive or not specific for organohalide versus sulfate respiration. Electron-transporting proteins or mediators unique for reductive dehalogenation were not revealed in our analysis, and we hypothesize that reductive dehalogenation may share an electron-transporting system with sulfate reduction. The metabolism of D. spongiiphila, predicted from transcriptomic and proteomic results, demonstrates high metabolic versatility and provides insights into the survival strategies of a marine sponge symbiont in an environment rich in organohalide compounds and other secondary metabolites. IMPORTANCE Respiratory reductive dehalogenation is an important process in the overall cycling of both anthropogenic and natural organohalide compounds. Marine sponges produce a vast array of bioactive compounds as secondary metabolites, including diverse halogenated compounds that may enrich for dehalogenating bacteria. Desulfoluna spongiiphila strain AA1 was originally enriched and isolated from the marine sponge Aplysina aerophoba and can grow with both brominated compounds and sulfate as electron acceptors for respiration. An understanding of the overall gene expression and the protein production profile in response to organohalides is needed to identify the full complement of genes or enzymes involved in organohalide respiration. Elucidating the metabolic capacity of this sponge-associated bacterium lays the foundation for understanding how dehalogenating bacteria may control the fate of organohalide compounds in sponges and their role in a symbiotic organobromine cycle.

scholarly journals Tool for Quantification of Staphylococcal Enterotoxin Gene Expression in Cheese

2010 ◽  
Vol 76 (5) ◽  
pp. 1367-1374 ◽  
Author(s):  
Manon Duquenne ◽  
Isabelle Fleurot ◽  
Marina Aigle ◽  
Claire Darrigo ◽  
Elise Borez�e-Durant ◽  
...  
Keyword(s):  
Gene Expression ◽  
Reference Genes ◽  
Staphylococcal Enterotoxin ◽  
Enterotoxin Gene ◽  
Food Borne ◽  
Enterotoxin Genes ◽  
Reverse Transcription Quantitative Pcr ◽  
Study Gene Expression ◽  
Yeasts And Molds

ABSTRACT Cheese is a complex and dynamic microbial ecosystem characterized by the presence of a large variety of bacteria, yeasts, and molds. Some microorganisms, including species of lactobacilli or lactococci, are known to contribute to the organoleptic quality of cheeses, whereas the presence of other microorganisms may lead to spoilage or constitute a health risk. Staphylococcus aureus is recognized worldwide as an important food-borne pathogen, owing to the production of enterotoxins in food matrices. In order to study enterotoxin gene expression during cheese manufacture, we developed an efficient procedure to recover total RNA from cheese and applied a robust strategy to study gene expression by reverse transcription-quantitative PCR (RT-qPCR). This method yielded pure preparations of undegraded RNA suitable for RT-qPCR. To normalize RT-qPCR data, expression of 10 potential reference genes was investigated during S. aureus growth in milk and in cheese. The three most stably expressed reference genes during cheese manufacture were ftsZ, pta, and gyrB, and these were used as internal controls for RT-qPCR of the genes sea and sed, encoding staphylococcal enterotoxins A and D, respectively. Expression of these staphylococcal enterotoxin genes was monitored during the first 72 h of the cheese-making process, and mRNA data were correlated with enterotoxin production.

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