Microbial activity and community structure in a lake sediment used for psychrophilic anaerobic wastewater treatment

Aims: The aim of the study was to investigate the feasibility of a continuous reactor for psychrophilic anaerobic wastewater treatment by using the sludge from cold natural environment. Methods and Results: Six sludge samples (S1–S6) were collected from different cold natural locations to select sludge with high anaerobic microbial activity under low temperatures. After a 225‐day incubation, the maximum specific methane production rate of a waterfowl lake sediment (S1) at 15°C (70·5 mLCH₄ gVSS^?1 day^?1) was much higher than all other samples. S1 was thus chosen as the seed sludge for the reactor treating synthetic brewery wastewater at 15°C, by immobilizing the micro‐organisms on polyurethane foam carriers. The chemical oxygen demand (COD) removal efficiency reached over 80% after 240‐day operation at an organic loading rate of 5·3 kg m^?3 day^?1, and significant enrichment of biomass was observed. Clone libraries of the microbial communities in the inoculum had high diversities for both archaea and bacteria. Along with a decrease in microbial community diversities, the dominant bacteria (79·5%) at the end of the operation represented the phylum Firmicutes, while the dominant archaeon (41·5%) showed a similarity of 98% with the psychrotolerant methanogen Methanosarcina lacustris. Conclusions: The possibility of using anaerobic micro‐organisms from cold environments in anaerobic wastewater treatment under psychrophilic conditions is supported by these findings. Significance and Impact of the Study: This study enriches the theory on microbial community and the application on anaerobic treatment of sludge from cold natural environments.     

Characterization of microbial community and kinetics for spent sulfidic caustic applied autotrophic denitrification

Spent sulfidic caustic was applied to sulfur utilizing autotrophic denitrification as the simultaneous source of electron donor and alkalinity. The two experiment set-up of upflow anoxic hybrid growth reactor (UAHGR) and upflow anoxic suspended growth reactor (UASGR) was adopted and nitrate removals were similar in both reactors. Approximately 90% of the initial nitrate was denitrified at nitrate loading rate of 0.15∼0.40 kgNO₃ ⁻/m³·d. The experimental stoichiometric ratio of sulfate production to nitrate removal was ranged from 1.5 to 2.1 mgSO₄ ²⁻/mgNO₃ ⁻. During the operation period, denaturing gradient gel electrophoresis (DGGE) analysis of polymerase chain reaction (PCR)-amplified 16S rDNA fragments for the sludge sample of both reactors showed the change of microbial communities. Thiobacillus denitrificans-like microorganism occupied 28.5% (18 clones) of the 63 clones by cloning the PCR products from the sludge sample of UAHGR. Acidovorax avenae, which can reduce nitrate to nitrogen gas while oxidizing phenol (heterotrophic denitrifier), was also found in 7 clones (11.1%). Although an organic carbon source was not added to the medium, a microorganism (Kaistella koreensis) capable of oxidizing organic compounds was found in 7 clones (11.1%). Therefore, the microbial community of spent sulfidic caustic applied autotrophic denitrification process well corresponds to the substrate components of spent sulfidic caustic. Through the batch cultivation of microorganisms in UAHGR, the microbial kinetic coefficients of spent sulfidic caustic applied autotrophic denitrification were estimated to be μ _max = 0.097 h⁻¹, k _d = 0.0021 h⁻¹, K _s = 200 mgNO₃ ⁻/L, and Y = 0.31 mgMLVSS/mgNO₃ ⁻.     

Long-term effects of metals in sewage sludge on soils,microorganisms and plants

Summary This paper reviews the evidence for impacts of metals on the growth of selected plants and on the effects of metals on soil microbial activity and soil fertility in the long-term. Less is known about adverse long-term effects of metals on soil microorganisms than on crop yields and metal uptake. This is not surprising, since the effects of metals added to soils in sewage sludge are difficult to assess, and few long-term experiments exist. Controlled field experiments with sewage sludges exist in the UK, Sweden, Germany and the USA and the data presented here are from these long-term field experiments only. Microbial activity and populations of cyanobacteria,Rhizobium leguminosarum bv.trifolii, mycorrhizae and the total microbial biomass have been adversely affected by metal concentrations which, in some cases, are below the European Community's maximum allowable concentration limits for metals in sludge-treated soils. For example, N₂-fixation by free living heterotrophic bacteria was found to be inhibited at soil metal concentrations of (mg kg^–1): 127 Zn, 37 Cu, 21 Ni, 3.4 Cd, 52 Cr and 71 Pb. N₂-fixation by free-living cyanobacteria was reduced by 50% at metal concentrations of (mg kg^–1): 114 Zn, 33 Cu, 17 Ni, 2.9 Cd, 80 Cr and 40 Pb.Rhizobium leguminosarum bv.trifolii numbers decreased by several orders of magnitude at soil metal concentrations of (mg kg^–1): 130–200 Zn, 27–48 Cu, 11–15 Ni, and 0.8–1.0 Cd. Soil texture and pH were found to influence the concentrations at which toxicity occurred to both microorganisms and plants. Higher pH, and increased contents of clay and organic carbon reduced metal toxicity considerably. The evidence suggests that adverse effects on soil microbial parameters were generally found at surpringly modest concentrations of metals in soils. It is concluded that prevention of adverse effects on soil microbial processes and ultimately soil fertility, should be a factor which influences soil protection legislation.     

Monitoring of specific methanogenic activity of granular sludge by confocal laser scanning microscopy during start-up of thermophilic upflow anaerobic sludge blanket reactor

Confocal, laser-scanning microscopy was applied to acquire coenzyme F₄₂₀-based autofluorescence images of middle sections of sludge granules during start-up of a thermophilic reactor that were seeded with mesophilically-grown microorganisms of granular sludge. Digital images were analyzed to calculate weighted averages of autofluorescence. The values were related (r ²=0.97) to specific methanogenic activities of granular sludge as the granules developed to steady state.     

Distribution and Physiological State of Microorganisms in Petrochemical Oily Sludge

The occurrence, vertical distribution, and physiological state of microorganisms in a petrochemical oily sludge deposit were studied. The total number and the number of viable microbial cells at depths of 0.2 and 3 m were about 10¹⁰ and 10⁸ cells/g dry wt sludge. Most microbial cells taken from the middle (1 m deep) and the bottom (3 m deep) sludge horizons showed a delayed colony-forming ability, which suggested that the cells occurred in a hypometabolic state. The relative number of microaerobic denitrifying microorganisms steeply increased with depth. The amount of microorganisms tolerant to 3, 5, and 10% NaCl and capable of growing at 7 and 40°C varied from 10² to 10⁸ CFU/g dry wt sludge. Petrochemical oily sludge was found to maintain the growth of heterotrophs, among which the degraders of oily sludge and ten different individual polycyclic aromatic hydrocarbons were detected. The occurrence of highly adaptable microorganisms with an adequate metabolic potential in the petrochemical oily sludge deposit implies that its bioremediation is possible without introducing special microorganisms.     

Biodegradation of oily sludge in Kuwait soil

Soil microorganisms were not inhibited by mixing oily sludge in soil up to 8.7% (w/w) oil (15% sludge). Adding NH ₄ ⁺ and phosphate increased microbial activity. Microbial activity was also affected by seasonal variation. Thermotolerant microorganisms were more predominant during the summer. After 29 months, 72%, 84%, and 83% of the soil was degraded in fertilized soils dosed with 2.9, 5.8 and 8.7% oil, respectively.     

A new kinetic approach to microbial storage process

In this work, a new kinetic approach was proposed to describe the microbial growth, substrate consumption, and formation and utilization of the intracellular storage products (X _STO) in activated sludge. It was found that the formation of X _STO was coupled with energy generation and respiration and that the X _STO formation rate was proportional to the substrate utilization rate. A high amount of external substrate resulted in a relatively rapid storage process with a large fraction of substrate electrons for X _STO formation. The maximum growth rate of active biomass on X _STO and the yield coefficient for growth on the storage polymers were estimated as 0.12 h⁻¹ and 0.60 g chemical oxygen demand (COD) _X g⁻¹ COD_STO, respectively. This established model was verified with the experimental results from two different case studies with pure and mixed cultures. Results showed that this kinetic model was able to accurately and mechanistically describe the microbial storage processes.     

Biosorption of Copper (II) by the Microorganisms Isolated from the Crude-Oil-Contaminated Soil

The objective of this study was to isolate and screen the highly efficient copper-removing microorganisms from the petroleum hydrocarbon (PH)-contaminated sites in the Amazonian rain forest in Ecuador. Two bacterial strains (strain UEAB3 and UEAB6) have shown 100% microbial resistance on the nutrient medium containing 100 mM of MgCl₂, FeCl₃, and Al₂(SO₄)₃ separately. Though these two strains were less tolerant of ZnCl₂ and CuSO₄.5H₂O, they have proven 100% resistance at the lower concentrations of these two metals. According to atomic absorption spectroscopy (AAS) analysis, the filamentous fungi (strains UEAFr and UEAFg) were significantly (p<0.05) effective at bacteria in the biosorption (97–100%) of copper (5 mg L^?1) over 7 d. As per 16/18S rDNA sequences, UEAB3, UEAB6, UEAFr, and UEAFg were Bacillus thuringiensis, Bacillus cereus, Geomyces pannorum, and Geomyces sp., respectively. From these results, it can be comprehensively concluded that the isolated microbial cultures had a capacity to remove the copper metal from the liquid medium.     

The use of polyurethane foam for microbial retention in methanogenic fermentation of propionate

Summary The use of polyurethane foam (PUF) as a microbial support carrier was evaluated with a mesophilic propionate-acclimatized sludge. The acclimatized sludge could be immobilized rapidly and stably in PUF of smaller pore size under shaking conditions. The sludge retained in PUF could maintain a high propionate metabolic activity for a long period. High conversion rates of propionate to methane of 23–65 g chemical oxygen demand (COD)·1^–1 · day^–1 could be achieved in reactors packed with PUF-retained sludge. A dense sludge of 0.08–0.25 g mixed-liquor volatile suspended solids (MLVSS)·cm^–3 was retained in PUF. Microscopic analysis suggested that filamentous microorganisms, e.g., Methanothrix spp. could play an important role in the efficient retention of acclimatized sludge in PUF. Offprint requests to: Shiro Nagai     

Kinetics of soluble microbial product formation in substrate-sufficient batch culture of activated sludge

The kinetics of soluble microbial product (SMP) formation under substrate-sufficient conditions appear to exhibit different patterns from substrate-limited cultures. However, energy spilling-associated SMP formation is not taken into account in the existing kinetic models and classification of SMP. Based on the concepts of growth yield and energy uncoupling, a kinetic model describing energy spilling-associated SMP formation in relation to the ratio of initial substrate concentration to initial biomass concentration (S ₀/X ₀) was developed for substrate-sufficient batch culture of activated sludge, and was verified by experimental data. The specific rate of energy spilling-associated SMP formation showed an increasing trend with the S ₀/X ₀ ratio up to its maximum value. The SMP productivity coefficient (α _p/e) was defined from the model on the basis of energy spilling-associated substrate consumption. Results revealed that less than 5% of energy spilling-associated substrate consumption was converted into SMP. Electronic Publication     

Inhibition of plant and microbial ureases by phosphoroamides

Enzyme kinetic studies of inhibition of plant (jackbean) and microbial (Bacillus pasteurii) ureases by eight phosphoroamides [phenylphosphorodiamidate, 4-chlorophenylphosphorodiamidate, phosphoric triamide, N-(diaminophosphinyl)benzamide, N-(diaminophosphinyl)benzeneacetamide, 4-chloro-N-(diaminophosphinyl)benzamide, N-(4-nitrophenyl)phosphoric triamide, N-(diaminophosphinyl)-3-pyridinecarboxamide] demonstrated that these compounds are slow, tight-binding inhibitors of urease enzymes. Measurement of the dissociation constants (Ki^*) of the enzyme-inhibitor complexes (E · I^*) formed by interaction of the ureases and phosphoroamide inhibitors studied showed that these inhibitors had a much higher affinity (i.e., a lower Ki^*) for plant urease than for microbial urease. Measurement of rate constants for formation (k_on) and decay (k_off) of E · I^* showed that, whereas k_on varied greatly with the different inhibitors and ureases, k_off was constant for the phosphoroamides tested and had a characteristic value for each urease. The half-life of E · I^* (30°C; pH 7 THAM buffer) for the plant urease was much longer than that for the microbial urease, and this difference largely accounted for the much higher values of Ki^* (k_off/k_on) observed with microbial urease.     

Engineering the productivity of recombinant Escherichia coli for limonene formation from glycerol in minimal media

The efficiency and productivity of cellular biocatalysts play a key role in the industrial synthesis of fine and bulk chemicals. This study focuses on optimizing the synthesis of (S)‐limonene from glycerol and glucose as carbon sources using recombinant Escherichia coli. The cyclic monoterpene limonene is extensively used in the fragrance, food, and cosmetic industries. Recently, limonene also gained interest as alternative jet fuel of biological origin. Key parameters that limit the (S)‐limonene yield, related to genetics, physiology, and reaction engineering, were identified. The growth‐dependent production of (S)‐limonene was shown for the first time in minimal media. E. coli BL21 (DE3) was chosen as the preferred host strain, as it showed low acetate formation, fast growth, and high productivity. A two‐liquid phase fed‐batch fermentation with glucose as the sole carbon and energy source resulted in the formation of 700 mg L_org^–1 (S)‐limonene. Specific activities of 75 mU g_cdw^–1 were reached, but decreased relatively quickly. The use of glycerol as a carbon source resulted in a prolonged growth and production phase (specific activities of ≥50 mU g_cdw^–1) leading to a final (S)‐limonene concentration of 2,700 mg L_org^–1. Although geranyl diphosphate (GPP) synthase had a low solubility, its availability appeared not to limit (S)‐limonene formation in vivo under the conditions investigated. GPP rerouting towards endogenous farnesyl diphosphate (FPP) formation also did not limit (S)‐limonene production. The two‐liquid phase fed‐batch setup led to the highest monoterpene concentration obtained with a recombinant microbial biocatalyst to date.     

Nitrate-induced ethylene biosynthesis and the control of nodulation in alfalfa

We previously reported that inhibition of ethylene biosynthesis with aminoethoxyvinylglycine (AVG) eliminated the inhibitory effect of NO₃^– on nodulation of alfalfa (Medicago sativa L. cv. Aragon) plants grown aeroponically. In this work, the effect of Ag⁺, as an inhibitor of ethylene action, has been studied in plants growing aeroponically or in darkened tubes with vermiculite, and low-nitrate or high-nitrate solution. Vermiculite-grown plants developed up to 3 times as many nodules as did those growing aeroponically. Nodule formation was mirrored by dry-matter accumulation. High (10 mol m^–3) NO₃^– applied from planting inhibited nodulation to an equal extent (c. 50%) in the two growth conditions. In contrast, Ag⁺ treatment increased nodule formation at all NO₃^– concentrations assayed under the two growth conditions, with the stimulation being higher in plants grown aeroponically. Finally, no effect of Ag⁺ (10 mmol m^–3) on plant growth was observed in either of the growth conditions. The effectiveness of NO₃^– as a nodulation inhibitor and enhancer of ethylene biosynthesis in roots of alfalfa was also studied. Within 24 h after inoculation, 10 mol m^–3 NO₃^– exerted most of its inhibitory effect on nodulation. At the same time, both 1-aminocyclopropane-1-carboxylic acid (ACC) oxidase activity and ethylene evolution rates markedly increased in inoculated and uninoculated alfalfa roots treated with NO₃^–. Support for a role of endogenous ethylene in the control of nodule formation in legumes is discussed.     

蚯蚓作用下污泥重金属形态变化及其与化学生物学性质变化的关系

城市污泥处理是一项世界性难题,污泥农业利用是其最简单有效的资源化利用方式之一,但污泥中较高的重金属含量限制了其实际推广应用,利用蚯蚓-超富集植物联合修复污泥重金属的方法已引起国内外研究者的关注。以新鲜城市脱水污泥为研究对象,接种赤子爱胜蚓(Eisenia fetida)进行室内培养试验,系统研究蚯蚓作用下污泥重金属形态的变化,及其与污泥氧化还原条件、化学和微生物性质变化的关系,以期为蚯蚓-超富集植物联合修复技术在污泥重金属处理中的应用提供理论依据。结果表明,试验前期蚯蚓在污泥中能正常生长和存活,前20 d总生物量增加了52%。蚯蚓可以显著促进污泥中的Cu、Zn、Cd、Ni等重金属从残渣态和铁锰态等稳定形态向交换态和水溶态等有效形态转化。还可以显著降低污泥中还原性物质的含量,减缓p H值下降速度,降低总有机碳含量,促进铵态氮向硝态氮转化,减少污泥微生物的数量并增加其种群活性。蚯蚓作用下,污泥中重金属的活化程度与还原性物质的含量呈显著负相关,而与微生物种群的活性呈显著正相关(P0.05)。综上所述,蚯蚓可以促进污泥重金属的活化,并改善污泥的肥力条件,为修复植物在污泥中的正常生长和对重金属离子的快速吸收提供有利条件。     

Simulating a cyclic activated sludge system by employing a modified ASM3 model for wastewater treatment

To interpret the biological nutrient removal in a cyclic activated sludge system (CAS), a modified model was developed by combining the process of simultaneous storage and growth, and the kinetics of soluble microbial product (S _SMP) and extracellular polymeric substance (X _EPS) with activated sludge model no. 3 (ASM3). These most sensitive parameters were initially selected whilst parameters with low sensitivity were given values from literature. The selected parameters were then calibrated on an oxygen uptake rate test and a batch CAS reactor on an operational cycle. The calibrated model was validated using a combination of the measurements from a batch CAS reactor operated for 1 month and the average deviation method. The simulations demonstrated that the modified model was capable of predicting higher effluent concentrations compared to outputs of the ASM3 model. Additionally, it was also shown that the average deviation of effluent S _COD, S _NH, S _SMP and X _EPS simulated with the modified model was all less than 1 mg L⁻¹. In summary, the model could effectively describe biological processes in a CAS reactor and provide a wonderful tool for operation.

     

P450<Subscript>BM-3</Subscript>-catalyzed whole-cell biotransformation of α-pinene with recombinant <Emphasis Type="Italic">Escherichia coli</Emphasis> in an aqueous–organic two-phase system

A recombinant Escherichia coli BL21 (DE3) strain overexpressing a variant of P450_BM-3 (V26T/R47F/A74G/F87V/L188K; abbreviated: BL21 (P450_BM-3 QM)) oxyfunctionalizes the bicyclic monoterpene α-pinene to α-pinene oxide, verbenol, and myrtenol. To address the low water solubility and the toxicity of terpenoids, an aqueous–organic two-phase bioprocess was developed. Diisononyl phthalate was selected as a biocompatible organic carrier solvent capable of masking the toxic effects mediated by α-pinene and of efficiently extracting the products enabling scale-up to the bioreactor. With an aqueous to organic phase ratio of 3:2 and 30% (v/v) of α-pinene in the organic phase, a biocatalytic product formation period of more than 4 h was achieved. A comparison of the biotransformation performance of BL21 (P450_BM-3 QM) and a strain with an additional heterologous NADPH regeneration system comprising glucose facilitator and dehydrogenase, but only expressing half the amount of P450_BM-3 QM, shows comparable product concentrations of 1,020 ± 144 and 800 ± 61 mg l_Aq⁻¹, respectively. The total product yields Y _P/P450 (μmol μmol_P450⁻¹) were 80% higher when the strain with the cofactor regeneration system was used. A total product concentration of over 1 g l_Aq⁻¹, corresponding to the highest value reported for microbial α-pinene oxyfunctionalization so far, marks a promising step forward toward a future application of recombinant microorganisms for the selective oxidation of terpenoids to value-added products.     

Degradation of Phthalate and Di-(2-Ethylhexyl)phthalate by Indigenous and Inoculated Microorganisms in Sludge-Amended Soil

The metabolism of phthalic acid (PA) and di-(2-ethylhexyl)phthalate (DEHP) in sludge-amended agricultural soil was studied with radiotracer techniques. The initial rates of metabolism of PA and DEHP (4.1 nmol/g [dry weight]) were estimated to be 731.8 and 25.6 pmol/g (dry weight) per day, respectively. Indigenous microorganisms assimilated 28 and 17% of the carbon in [¹⁴C]PA and [¹⁴C]DEHP, respectively, into microbial biomass. The rates of DEHP metabolism were much greater in sludge assays without soil than in assays with sludge-amended soil. Mineralization of [¹⁴C]DEHP to ¹⁴CO₂ increased fourfold after inoculation of sludge and soil samples with DEHP-degrading strain SDE 2. The elevated mineralization potential was maintained for more than 27 days. Experiments performed with strain SDE 2 suggested that the bioavailability and mineralization of DEHP decreased substantially in the presence of soil and sludge components. The microorganisms metabolizing PA and DEHP in sludge and sludge-amended soil were characterized by substrate-specific radiolabelling, followed by analysis of ¹⁴C-labelled phospholipid ester-linked fatty acids (¹⁴C-PLFAs). This assay provided a radioactive fingerprint of the organisms actively metabolizing [¹⁴C]PA and [¹⁴C]DEHP. The ¹⁴C-PLFA fingerprints showed that organisms with different PLFA compositions metabolized PA and DEHP in sludge-amended soil. In contrast, microorganisms with comparable ¹⁴C-PLFA fingerprints were found to dominate DEHP metabolism in sludge and sludge-amended soil. Our results suggested that indigenous sludge microorganisms dominated DEHP degradation in sludge-amended soil. Mineralization of DEHP and PA followed complex kinetics that could not be described by simple first-order equations. The initial mineralization activity was described by an exponential function; this was followed by a second phase that was described best by a fractional power function. In the initial phase, the half times for PA and DEHP in sludge-amended soil were 2 and 58 days, respectively. In the late phase of incubation, the apparent half times for PA and DEHP increased to 15 and 147 days, respectively. In the second phase (after more than 28 days), the half time for DEHP was 2.9 times longer in sludge-amended soil assays than in sludge assays without soil. Experiments with radiolabelled DEHP degraders suggested that a significant fraction of the ¹⁴CO₂ produced in long-term degradation assays may have originated from turnover of labelled microbial biomass rather than mineralization of [¹⁴C]PA or [¹⁴C]DEHP. It was estimated that a significant amount of DEHP with poor biodegradability and extractability remains in sludge-amended soil for extended periods of time despite the presence of microorganisms capable of degrading the compound (e.g., more than 40% of the DEHP added is not mineralized after 1 year).     

Mixed culture hydrogenotrophic nitrate reduction in drinking water

Isolation and identification of the bacteria from a hydrogenotrophic reactor for the denitrification of drinking water revealed that several microorganisms are involved. Acinetobacter sp., Aeromonas sp., Pseudomonas sp. and Shewanella putrefaciens were repeatedly isolated from the hydrogenotrophic sludge and postulated to be of primary importance in the process. Nitrate reduction to nitrite appears to be a property of a diverse group of organisms. Nitrite reduction was found to be stimulated by the presence of organic growth factors. Thus, in a mixed culture, hydrogenotrophic denitrification reactor, NO _inf2 ^sup– formed by NO _inf3 ^sup– -reducers can be converted by true denitrifiers thriving on organic growth factors either present in the raw water, or excreted by the microbial community. Mixotrophic growth also contributes to NO _inf2 ^sup– reduction. Finally, chemolithotrophic bacteria participate in the nitrite to nitrogen gas conversion.Offprint requests to: W. Verstraete.     

Linking Belowground Carbon Allocation to Anaerobic CH4 and CO2 Production in a Forested Peatland,New York State

Heterotrophic soil microorganisms rely on carbon (C) allocated belowground in plant production, but belowground C allocation (BCA) by plants is a poorly quantified part of ecosystem C cycling, especially, in peat soil. We applied a C balance approach to quantify BCA in a mixed conifer-red maple (Acer rubrum) forest on deep peat soil. Direct measurements of CH₄ and CO₂ fluxes across the soil surface (soil respiration), production of fine and small plant roots, and aboveground litterfall were used to estimate respiration by roots, by mycorrhizae and by free-living soil microorganisms. Measurements occurred in two consecutive years. Soil respiration rates averaged 1.2 bm μmol m^{? 2} s^{? 1} for CO₂ and 0.58 nmol m^{? 2} s^{? 1} for CH₄ (371 to 403 g C m^{? 2} year^{? 1}). Carbon in aboveground litter (144 g C m^{? 2} year^{? 1}) was 84% greater than C in root production (78 g C m^{? 2} year^{? 1}). Complementary in vitro assays located high rates of anaerobic microbial activity, including methanogenesis, in a dense layer of roots overlying the peat soil and in large-sized fragments within the peat matrix. Large-sized fragments were decomposing roots and aboveground leaf and twig litter, indicating that relatively fresh plant production supported most of the anaerobic microbial activity. Respiration by free-living soil microorganisms in deep peat accounted for, at most, 29 to 38 g C m^{? 2} year^{? 1}. These data emphasize the close coupling between plant production, ecosystem-level C cycling and soil microbial ecology, which BCA can help reveal.