Methane-oxidizing bacteria in a Finnish raised mire complex: effects of site fertility and drainage.

Methane-oxidizing bacteria (MOB) are the only biological sinks for methane (CH4). Drainage of peatlands is known to decrease overall CH4 emission, but the effect on MOB is unknown. The objective of this work was to characterize the MOB community and activity in two ecohydrologically different pristine peatland ecosystems, a fen and a bog, and their counterparts that were drained in 1961. Oligotrophic fens are groundwater-fed peatlands, but ombrotrophic bogs receive additional water and nutrients only from rainwater. The sites were sampled in August 2003 down to 10 cm below the water table (WT), and cores were divided into 10-cm subsamples. CH4 oxidation was measured by gas chromatography (GC) to characterize MOB activity. The MOB community structure was characterized by polymerase chain reaction-denaturing gradient gel electrophoresis (DGGE) and sequencing methods using partial pmoA and mmoX genes. The highest CH4 oxidation rates were measured from the subsamples 20-30 and 30-40 cm above WT at the pristine oligotrophic fen (12.7 and 10.5 micromol CH4 dm-3 h-1, respectively), but the rates decreased to almost zero in the vicinity of WT. In the pristine ombrotrophic bog, the highest oxidation rate at 0-10 cm was lower than in the fen (8.10 micromol CH4 dm-3 h-1), but in contrast to the fen, oxidation rates of 4.5 micromol CH4 dm-3 h-1 were observed at WT and 10 cm below WT. Drainage reduced the CH4 oxidation rates to maximum values of 1.67 and 5.77 micromol CH4 dm-3 h-1 at 30-40 and 20-30 cm of the fen and bog site, respectively. From the total of 13 pmoA-derived DGGE bands found in the study, 11, 3, 6, and 2 were observed in the pristine fen and bog and their drained counterparts, respectively. According to the nonmetric multidimensional scaling of the DGGE banding pattern, the MOB community of the pristine fen differed from the other sites. The majority of partial pmoA sequences belonged to type I MOB, whereas the partial mmoX bands that were observed only in the bog sites formed a distinct group relating more to type II MOB. This study indicates that fen and bog ecosystems differ in MOB activity and community structure, and both these factors are affected by drainage.

Methanogen communities in a drained bog: effect of ash fertilization.

Forestry practises such has drainage have been shown to decrease emissions of the greenhouse gas methane (CH(4)) from peatlands. The aim of the study was to examine the methanogen populations in a drained bog in northern Finland, and to assess the possible effect of ash fertilization on potential methane production and methanogen communities. Peat samples were collected from control and ash fertilized (15,000 kg/ha) plots 5 years after ash application, and potential CH(4) production was measured. The methanogen community structure was studied by DNA isolation, PCR amplification of the methyl coenzyme-M reductase (mcr) gene, denaturing gradient gel electrophoresis (DGGE), and restriction fragment length polymorphism (RFLP) analysis. The drained peatland showed low potential methane production and methanogen diversity in both control and ash-fertilized plots. Samples from both upper and deeper layers of peat were dominated by three groups of sequences related to Rice cluster-I hydrogenotroph methanogens. Even though pH was marginally greater in the ash-treated site, the occurrence of those sequences was not affected by ash fertilization. Interestingly, a less common group of sequences, related to the Fen cluster, were found only in the fertilized plots. The study confirmed the depth related change of methanogen populations in peatland.

Pathways for methanogenesis and diversity of methanogenic archaea in three boreal peatland ecosystems.

The main objectives of this study were to uncover the pathways used for methanogenesis in three different boreal peatland ecosystems and to describe the methanogenic populations involved. The mesotrophic fen had the lowest proportion of CH4 produced from H2-CO2. The oligotrophic fen was the most hydrogenotrophic, followed by the ombrotrophic bog. Each site was characterized by a specific group of methanogenic sequences belonging to Methanosaeta spp. (mesotrophic fen), rice cluster-I (oligotrophic fen), and fen cluster (ombrotrophic bog).

Effect of Cd-containing wood ash on the microflora of coniferous forest humus.

The use of wood ash in forestry has been questioned because the cadmium (Cd) concentration of ash, which varies between 1 and 20 mg kg(-1) ash, exceeds the level allowed for fertilizers (3 mg kg(-1)) used in agriculture. To investigate the combined and separated effects of Cd and ash on the forest humus microflora, pumice or wood ash, spiked with a water-soluble (CdCl(2)) or -insoluble (CdO) form of Cd at three levels (0, 400 and 1000 mg kg(-1)), were applied at a fertilization level of 5000 kg ha(-1) in a laboratory microcosm study. The trial consisted of 60 microcosms (five replications per treatment), which were incubated in darkness at +20 degrees C and a constant relative air humidity of 60%. After two months the humus in the microcosms was sampled. Analyses of CO(2) evolution to measure the overall microbial activity and of phospholipid fatty acid (PLFA) pattern to measure microbial community structure were performed. The substrate-use patterns of Biolog EcoPlates were analyzed as a measure of bacterial functionality. Finally the bacterial (3)H-thymidine incorporation in the presence of different concentrations of Cd and the number of colony forming units (cfu) of bacteria on nutrient agar in the presence of 0, 5 and 20 mg Cd l(-1) agar were applied to measure Cd tolerance. The use of pumice (pH of humus under the pumice 4.0) did not induce any changes in the above variables compared to two untreated microcosms (humus pH 3.9). Pumice was therefore used to distribute the Cd evenly over the humus surface in order to estimate the possible effect of Cd without ash (pH of humus under the ash 7.0). The application of ash increased the microbial activity, changed the PLFA and substrate-use patterns and increased cfu compared to the humus under pumice. The form and level of Cd in the ash had no further effect on this result. In the humus under pumice the level, but not the form of Cd decreased the microbial activity and changed the PLFA pattern compared to the unspiked pumice. None of the treatments induced bacterial tolerance to Cd. Ash thus protected the humus microflora from the harmful effects of Cd.

Means to improve the effect of in situ bioremediation of contaminated soil: an overview of novel approaches.

Different aspects of bacterial degradation of organic contaminants in soil, and how to improve the efficiency and reproducibility is discussed in this review. Although bioremediation in principle includes the use of any type of organism in improving the condition of a contaminated site, most commonly bacteria are the degraders and other organisms, such as soil animals or plant roots, play a role in dissemination of bacteria and, indirectly, plasmids between bacteria, and in providing nutrients and co-substrates for the bacteria active in the degradation process. There are a number of different procedures that have been tested more-or-less successfully in attempts to improve reliability, cost efficiency and speed of bioremediation. The methods range from minimal intervention, such as mere monitoring of intrinsic bioremediation, through in situ introduction of nutrients and/or bacterial inocula or improvement of physico-chemical conditions, all the way to excavation followed by on site or ex situ composting in its different varieties. In the past the rule has been that more intervention (leading to higher costs) has been more reliable, but novel ideas are continuously tried out, both as a means to come up with new truly functional applications and also as a line of studies in basic soil microbial ecology. Both approaches generate valuable information needed when predicting outcome of remediation activities, evaluating environmental risks, deciding on cleaning-up approaches, etc. The emphasis of this review is to discuss some of the novel methods for which the value has not been clearly shown, but that in our view merit continued studies and efforts to make them work, separately or in combination.

Characterization and reclassification of an aromatic- and chloroaromatic-degrading Pseudomonas sp., strain HV3, as Sphingomonas sp. HV3.

Phylogenetic analyses of 16S rRNA gene sequences showed that the Gram-negative aromatic- and chloroaromatic-degrading Pseudomonas sp. strain HV3 carrying the mega-plasmid pSKY4 belongs to the genus Sphingomonas. The 16SrRNA sequence is most related to Sphingomonas chlorophenolica strains ATCC 33790(T) (98.5%) and SR3 (98.4%) and Sphingomonas sp. SS86 (98.4%). The G+C content was 64 mol%, and the DNA-DNA hybridization-based relative homology of strain HV3 to the S. chlorophenolica ATCC 33790(T) and S. chlorophenolica RA2 was 59.6% and 35.9%, respectively. The results showed that although strain HV3 is related to S. chlorophenolica it differs in certain characteristics. It is therefore proposed to reclassify Pseudomonas sp. strain HV3 as Sphingomonas sp. HV3.

Novel organization of catechol meta-pathway genes in Sphingomonas sp. HV3 pSKY4 plasmid.

Sphingomonas sp. strain HV3 (formerly Pseudomonas sp. HV3), which degrades aromatics and chloroaromatics, harbors a mega-plasmid, pSKY4. A sequenced 4 kb fragment of the plasmid reveals a novel gene organization for catechol meta-pathway genes. The putative meta operon starts with the cmpF gene encoding a 2-hydroxymuconic semialdehyde hydrolase. The gene has a 6 bp overlap with the previously characterized ring-cleavage gene, catechol 2,3-dioxygenase, cmpE. Downstream of cmpE is a 429 bp open reading frame of unknown function. Gene cmpC, encoding a 2-hydroxymuconic semialdehyde dehydrogenase, starts 44 bp further downstream. It has the highest homology to 2-hydroxymuconic semialdehyde dehydrogenases of dmp and xyl pathways and to XylC from the marine oligotroph Cycloclasticus oligotrophus. The gene organization is different from other known meta pathways. This is the first report of organization of plasmid-encoded meta-pathway genes in the genus Sphingomonas.

Cloning of cmpE, a plasmid-borne catechol 2,3-dioxygenase-encoding gene from the aromatic- and chloroaromatic-degrading Pseudomonas sp. HV3.

Pseudomonas sp. strain HV3 degrades aromatics and chloroaromatics. It harbours a mega-plasmid, designated pSKY4, from which the gene cmpE, encoding a catechol 2,3-dioxygenase (C23O) catalyzing the conversion of catechol to 2-hydroxymuconic semialdehyde, was cloned and sequenced. The deduced amino acid (aa) sequence shows the highest homology, 52%, to the deduced aa sequences of xylE1 and dmpB. The deduced 307-aa sequence of cmpE contains the extradiol ring-cleavage signature in the same position as other 307-aa C23O-encoding genes.