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      Zinc deprivation of methanol fed anaerobic granular sludge bioreactors

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          Abstract

          The effect of omitting zinc from the influent of mesophilic (30 °C) methanol fed upflow anaerobic sludge bed (UASB) reactors, and latter zinc supplementation to the influent to counteract the deprivation, was investigated by coupling the UASB reactor performance to the microbial ecology of the bioreactor sludge. Limitation of the specific methanogenic activity (SMA) on methanol due to the absence of zinc from the influent developed after 137 days of operation. At that day, the SMA in medium with a complete trace metal solution except Zn was 3.4 g CH 4-COD g VSS −1 day −1, compared to 4.2 g CH 4-COD g VSS −1 day −1 in a medium with a complete (including zinc) trace metal solution. The methanol removal capacity during these 137 days was 99% and no volatile fatty acids accumulated. Two UASB reactors, inoculated with the zinc-deprived sludge, were operated to study restoration of the zinc limitation by zinc supplementation to the bioreactor influent. In a first reactor, no changes to the operational conditions were made. This resulted in methanol accumulation in the reactor effluent after 12 days of operation, which subsequently induced acetogenic activity 5 days after the methanol accumulation started. Methanogenesis could not be recovered by the continuous addition of 0.5 μM ZnCl 2 to the reactor for 13 days. In the second reactor, 0.5 μM ZnCl 2 was added from its start-up. Although the reactor stayed 10 days longer methanogenically than the reactor operated without zinc, methanol accumulation was observed in this reactor (up to 1.1 g COD-MeOH L −1) as well. This study shows that zinc limitation can induce failure of methanol fed UASB reactors due to acidification, which cannot be restored by resuming the continuous supply of the deprived metal.

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          Most cited references39

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          Optimizing fluorescent in situ hybridization with rRNA-targeted oligonucleotide probes for flow cytometric identification of microorganisms.

          A combination of fluorescent rRNA-targeted oligonucleotide probes ("phylogenetic stains") and flow cytometry was used for a high resolution automated analysis of mixed microbial populations. Fixed cells of bacteria and yeasts were hybridized in suspension with fluorescein- or tetramethylrhodamine-labeled oligonucleotide probes complementary to group-specific regions of the 16S ribosomal RNA (rRNA) molecules. Quantifying probe-conferred cell fluorescence by flow cytometry, we could discriminate between target and nontarget cell populations. We critically examined changes of the hybridization conditions, kinetics of the hybridization, and posthybridization treatments. Intermediate probe concentrations, addition of detergent to the hybridization buffer, and a posthybridization washing step were found to increase the signal to noise ratio. We could demonstrate a linear correlation between growth rate and probe-conferred fluorescence of Escherichia coli and Pseudomonas cepacia cells. Oligonucleotides labeled with multiple fluorochromes showed elevated levels of nonspecific binding and therefore could not be used to lower the detection limits, which still restrict studies with fluorescing rRNA-targeted oligonucleotide probes to well-growing microbial cells. Two probes of different specificities--one labeled with fluorescein, the other with tetramethylrhodamine--could be applied simultaneously for dual color analysis.
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            Nickel uptake and utilization by microorganisms.

            Nickel is an essential nutrient for selected microorganisms where it participates in a variety of cellular processes. Many microbes are capable of sensing cellular nickel ion concentrations and taking up this nutrient via nickel-specific permeases or ATP-binding cassette-type transport systems. The metal ion is specifically incorporated into nickel-dependent enzymes, often via complex assembly processes requiring accessory proteins and additional non-protein components, in some cases accompanied by nucleotide triphosphate hydrolysis. To date, nine nickel-containing enzymes are known: urease, NiFe-hydrogenase, carbon monoxide dehydrogenase, acetyl-CoA decarbonylase/synthase, methyl coenzyme M reductase, certain superoxide dismutases, some glyoxylases, aci-reductone dioxygenase, and methylenediurease. Seven of these enzymes have been structurally characterized, revealing distinct metallocenter environments in each case.
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              Fluorescence in situ hybridization using 16S rRNA-targeted oligonucleotides reveals localization of methanogens and selected uncultured bacteria in mesophilic and thermophilic sludge granules.

              16S rRNA-targeted in situ hybridization combined with confocal laser scanning microscopy was used to elucidate the spatial distribution of microbes within two types of methanogenic granular sludge, mesophilic (35 degrees C) and thermophilic (55 degrees C), in upflow anaerobic sludge blanket reactors fed with sucrose-, acetate-, and propionate-based artificial wastewater. The spatial organization of the microbes was visualized in thin sections of the granules by using fluorescent oligonucleotide probes specific to several phylogenetic groups of microbes. In situ hybridization with archaeal- and bacterial-domain probes within granule sections clearly showed that both mesophilic and thermophilic granules had layered structures and that the outer layer harbored mainly bacterial cells while the inner layer consisted mainly of archaeal cells. Methanosaeta-, Methanobacterium-, Methanospirillum-, and Methanosarcina-like cells were detected with oligonucleotide probes specific for the different groups of methanogens, and they were found to be localized inside the granules, in both types of which dominant methanogens were members of the genus Methanosaeta. For specific detection of bacteria which were previously detected by whole-microbial-community 16S ribosomal DNA (rDNA)-cloning analysis (Y. Sekiguchi, Y. Kamagata, K. Syutsubo, A. Ohashi, H. Harada, and K. Nakamura, Microbiology 144:2655-2665, 1998) we designed probes specific for clonal 16S rDNAs related to unidentified green nonsulfur bacteria and clones related to Syntrophobacter species. The probe designed for the cluster closely related to Syntrophobacter species hybridized with coccoid cells in the inner layer of the mesophilic granule sections. The probe for the unidentified bacteria which were clustered with the green nonsulfur bacteria detected filamentous cells in the outermost layer of the thermophilic sludge granule sections. These results revealed the spatial organizations of methanogens and uncultivated bacteria and their in situ morphologies and metabolic functions in both mesophilic and thermophilic granular sludges.
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                Author and article information

                Contributors
                +31-317-483851 , +31-317-482108 , piet.lens@wur.nl
                Journal
                J Ind Microbiol Biotechnol
                Journal of Industrial Microbiology & Biotechnology
                Springer-Verlag (Berlin/Heidelberg )
                1367-5435
                1476-5535
                19 February 2008
                June 2008
                : 35
                : 6
                : 543-557
                Affiliations
                [1 ]Sub-department of Environmental Technology, Wageningen University, “Biotechnion”-Bomenweg 2, P.O. Box 8129, 6700 EV Wageningen, The Netherlands
                [2 ]Microbial Ecology Laboratory, Department of Microbiology and Environmental Change Institute (ECI), National University of Ireland, Galway (NUI, Galway), University Road, Galway, Ireland
                [3 ]Pollution Prevention and Control Core, UNESCO-IHE, P.O. Box 3015, 2601 DA Delft, The Netherlands
                Article
                315
                10.1007/s10295-008-0315-z
                2668640
                18283507
                3f5b0150-ecb7-4d44-9271-781940c1d47c
                © The Author(s) 2008
                History
                : 14 September 2007
                : 10 January 2008
                Categories
                Original Paper
                Custom metadata
                © Society for Industrial Microbiology 2008

                Biotechnology
                trace metal limitation,methanogenic activity,nutrients,methanol,uasb reactor,fish,zinc
                Biotechnology
                trace metal limitation, methanogenic activity, nutrients, methanol, uasb reactor, fish, zinc

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