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Genetic Manipulation of Carotenoid Biosynthesis in the Green Sulfur Bacterium Chlorobium tepidum. Niels-Ulrik Frigaard, 2004.The green sulfur bacterium Chlorobium tepidum is a strict anaerobe and an obligate photoautotroph . On the basis of sequence similarity with known enzymes or sequence motifs, nine open reading frames encoding putative enzymes of carotenoid biosynthesis were identified in the genome sequence of C . tepidum, and all nine genes were inactivated . Analysis of the carotenoid composition in the resulting mutants allowed the genes encoding the following six enzymes to be identified: phytoene synthase (crtB/CT1386), phytoene desaturase (crtP/CT0807),  -carotene
desaturase (crtQ/CT1414),
 -carotene
desaturase (crtU/CT0323), carotenoid 1',2'-hydratase (crtC/CT0301),
and carotenoid cis-trans isomerase (crtH/CT0649) .
Three mutants (CT0180, CT1357, and CT1416 mutants)
did not exhibit a discernible phenotype . The carotenoid biosynthetic
pathway in C . tepidum is similar to that in cyanobacteria and
plants by converting phytoene into lycopene using two plant-like
desaturases (CrtP and CrtQ) and a plant-like cis-trans
isomerase (CrtH) and thus differs from the pathway known in all other
bacteria . In contrast to the situation in cyanobacteria and plants,
the construction of a crtB mutant completely lacking
carotenoids demonstrates that carotenoids are not essential for
photosynthetic growth of green sulfur bacteria . However, the
bacteriochlorophyll a contents of mutants lacking colored
carotenoids (crtB, crtP, and crtQ mutants) were
decreased from that of the wild type, and these mutants exhibited a
significant growth rate defect under all light intensities tested .
Therefore, colored carotenoids may have both structural and
photoprotection roles in green sulfur bacteria . The ability to
manipulate the carotenoid composition so dramatically in C .
tepidum offers excellent possibilities for studying the roles of
carotenoids in the light-harvesting chlorosome antenna and
iron-sulfur-type (photosystem I-like) reaction center . The phylogeny
of carotenogenic enzymes in green sulfur bacteria and green
filamentous bacteria is also discussed .
Response of Archaeal Communities in Beach Sediments to Spilled Oil and Bioremediation. Wilfred F. M. Röling, 2004.While the contribution of Bacteria to bioremediation of oil-contaminated shorelines is well established, the response of Archaea to spilled oil and bioremediation treatments is unknown . The relationship between archaeal community structure and oil spill bioremediation was examined in laboratory microcosms and in a bioremediation field trial . 16S rRNA gene-based PCR and denaturing gradient gel analysis revealed that the archaeal community in oil-free laboratory microcosms was stable for 26 days . In contrast, in oil-polluted microcosms a dramatic decrease in the ability to detect Archaea was observed, and it was not possible to amplify fragments of archaeal 16S rRNA genes from samples taken from microcosms treated with oil . This was the case irrespective of whether a bioremediation treatment (addition of inorganic nutrients) was applied . Since rapid oil biodegradation occurred in nutrient-treated microcosms, we concluded that Archaea are unlikely to play a role in oil degradation in beach ecosystems . A clear-cut relationship between the presence of oil and the absence of Archaea was not apparent in the field experiment . This may have been related to continuous inoculation of beach sediments in the field with Archaea from seawater or invertebrates and shows that the reestablishment of Archaea following bioremediation cannot be used as a determinant of ecosystem recovery following bioremediation . Comparative 16S rRNA sequence analysis showed that the majority of the Archaea detected (94%) belonged to a novel, distinct cluster of group II uncultured Euryarchaeota, which exhibited less than 87% identity to previously described sequences . A minor contribution of group I uncultured Crenarchaeota was observed . In Situ Identification of Intracellular Bacteria Related to Paenibacillus spp . in the Mycelium of the Ectomycorrhizal Fungus Laccaria bicolor S238N. J. Bertaux, 2003.Bacterial proliferations have recurrently been observed for the past 15 years in fermentor cultures of the ectomycorrhizal fungus Laccaria bicolor S238N, suggesting the presence of cryptic bacteria in the collection culture of this fungus . In this study, intracellular bacteria were detected by fluorescence in situ hybridization in combination with confocal laser scanning microscopy in several collection subcultures of L . bicolor S238N . They were small (0.5 µm in diameter), rare, and heterogeneously distributed in the mycelium and were identified as Paenibacillus spp . by using a 16S rRNA-directed oligonucleotide probe initially designed for bacteria isolated from a fermentor culture of L . bicolor S238N .
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