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Competition between Ammonia-Oxidizing Bacteria and Benthic Microalgae. Nils Risgaard-Petersen, 2004.The abundance, activity, and diversity of ammonia-oxidizing bacteria (AOB) were studied in prepared microcosms with and without microphytobenthic activity . In the microcosm without alga activity, both AOB abundance, estimated by real-time PCR, and potential nitrification increased during the course of the experiment . AOB present in the oxic zone of these sediments were able to fully exploit their nitrification potential because NH4+ did not limit growth . In contrast, AOB in the alga-colonized sediments reached less than 20% of their potential activity, suggesting starvation of cells . Starvation resulted in a decrease with time in the abundance of AOB as well as in nitrification potential . This decrease was correlated with an increase in alga biomass, suggesting competitive exclusion of AOB by microalgae . Induction of N limitation in the oxic zone of the alga-colonized sediments and O2 limitation of the majority of AOB in darkness were major mechanisms by which microalgae suppressed the growth and survival of AOB . The competition pressure from the algae seemed to act on the entire population of AOB, as no differences were observed by denaturing gradient gel electrophoresis of amoA fragments during the course of the experiment . Enumeration of bacteria based on 16S rRNA gene copies and D-amino acids suggested that the algae also affected other bacterial groups negatively . Our data indicate that direct competitive interaction takes place between algae and AOB and that benthic algae are superior competitors because they have higher N uptake rates and grow faster than AOB . Epoxyalkane:Coenzyme M Transferase in the Ethene and Vinyl Chloride Biodegradation Pathways of Mycobacterium Strain JS60. Nicholas V. Coleman, 2003.Mycobacterium strains that grow on ethene and vinyl chloride (VC) are widely distributed in the environment and are potentially useful for biocatalysis and bioremediation . The catabolic pathway of alkene assimilation in mycobacteria is not well characterized . It is clear that the initial step is a monooxygenase-mediated epoxidation that produces epoxyethane from ethene and chlorooxirane from VC, but the enzymes involved in subsequent transformation of the epoxides have not been identified . We investigated epoxyethane metabolism in Mycobacterium strain JS60 and discovered a coenzyme M (CoM)-dependent enzyme activity in extracts from VC- and ethene-grown cells . PCR amplifications using primers targeted at epoxyalkane:CoM transferase (EaCoMT) genes yielded part of the JS60 EaCoMT gene, which was used to clone an 8.4-kb genomic DNA fragment . The complete EaCoMT gene (etnE) was recovered, along with genes (etnABCD) encoding a four-component monooxygenase and two genes possibly involved in acyl-CoA ester metabolism . Reverse transcription-PCR indicated that the etnE and etnA genes were cotranscribed and inducible by ethene and VC . Heterologous expression of the etnE gene in Mycobacterium smegmatis mc2155 using the pMV261 vector gave a recombinant strain capable of transforming epoxyethane, epoxypropane, and chlorooxirane . A metabolite identified by mass spectrometry as 2-hydroxyethyl-CoM was produced from epoxyethane . The results indicate that the EaCoMT and monooxygenase enzymes encoded by a single operon (etnEABCD) catalyze the initial reactions in both the VC and ethene assimilation pathways . CoM-mediated reactions appear to be more widespread in bacteria than was previously believed . Purification and In Vitro Characterization of the Serratia marcescens NucC Protein, a Zinc-Binding Transcription Factor Homologous to P2 Ogr. Victor McAlister, 2003.NucC is structurally and functionally homologous to a family of prokaryotic zinc finger transcription factors required for late gene expression in P2- and P4-related bacteriophages . Characterization of these proteins in vitro has been hampered by their relative insolubility and tendency to aggregate . We report here the successful purification of soluble, active, wild-type NucC protein . Purified NucC exhibits site-specific binding to a conserved DNA sequence that is located upstream of NucC-dependent Serratia marcescens promoters and the late promoters of P2-related phages . This sequence is sufficient for binding of NucC in vitro . NucC binding to the S . marcescens nuclease promoter PnucA and to the sequence upstream of the P2 late promoter PF is accompanied by DNA bending . NucC protects about 25 nucleotides of the PF upstream region from DNase I digestion, and RNA polymerase protects the promoter region only in the presence of NucC . Template DNA, RNA polymerase holoenzyme, and purified NucC are the only macromolecular components required for transcription from PF in vitro . Community Structure of Ammonia-Oxidizing Bacteria within Anoxic Marine Sediments. Thomas E. Freitag, 2003.The potential for oxidation of ammonia in anoxic marine sediments exists through anaerobic oxidation by Nitrosomonas-like organisms, utilizing nitrogen dioxide, coupling of nitrification, manganese reduction, and anaerobic oxidation of ammonium by planctomycetes (the Anammox process) . Here we describe the presence of microbial communities with the potential to carry out these processes in a natural marine sediment system (Loch Duich, Scotland) . Natural microbial communities of Planctomycetales-Verrucomicrobia and ß- and  -proteobacterial ammonia-oxidizing bacteria were characterized by analysis of 16S rRNA genes amplified using group-specific primers by PCR- and reverse transcription-PCR amplification of 16S rDNA and RNA, respectively . Amplification products were analyzed by sequencing of clones and by denaturant gradient gel electrophoresis (DGGE) . Amplification of primers specific for Planctomycetales-Verrucomicrobia and ß-proteobacterial ammonia-oxidizing bacteria generated products at all sampling sites and depths, but no product was generated using primers specific for
-proteobacterial ammonia-oxidizing bacteria . 16S rDNA DGGE banding patterns indicated complex communities of ß-proteobacterial ammonia-oxidizing bacteria in anoxic marine sediments . Phylogenetic analysis of sequences from clones and those excised from DGGE gels suggests dominance of Nitrosospira cluster 1-like organisms and of strains belonging to a novel cluster represented in dominant bands in 16S rRNA DGGE banding patterns . Their presence indicates a group of organisms closely related to recognized ß-proteobacterial ammonia-oxidizing bacteria that may be selected in anoxic environments and may be capable of anoxic ammonia oxidation . Sequence analysis of planctomycete clone libraries and sequences excised from DGGE gels also demonstrated a diverse microbial community and suggested the presence of new subdivisions, but no sequence related to recognized Anammox organisms was detected .
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