Methane turnover and temperature response of methane-oxidizing bacteria in permafrost-affected soils of northeast Siberia

The abundance, activity, and temperature response of aerobic methane-oxidizing bacteria were studied in permafrost-affected tundra soils of northeast Siberia. The soils were characterized by both a high accumulation of organic matter at the surface and high methane concentrations in the water-saturated soils. The methane oxidation rates of up to 835 nmol CH₄ h⁻¹ g⁻¹ in the surface soils were similar to the highest values reported so far for natural wetland soils worldwide. The temperature response of methane oxidation was measured during short incubations and revealed maximum rates between 22 °C and 28 °C. The active methanotrophic community was characterized by its phospholipid fatty acid (PLFA) concentrations and with stable isotope probing (SIP). Concentrations of 16:1ω8 and 18:1ω8 PLFAs, specific to methanotrophic bacteria, correlated significantly with the potential methane oxidation rates. In all soils, distinct 16:1 PLFAs were dominant, indicating a predominance of type I methanotrophs. However, long-term incubation of soil samples at 0 °C and 22 °C demonstrated a shift in the composition of the active community with rising temperatures. At 0 °C, only the concentrations of 16:1 PLFAs increased and those of 18:1 PLFAs decreased, whereas the opposite was true at 22 °C. Similarly, SIP with ¹³CH₄ showed a temperature-dependent pattern. When the soils were incubated at 0 °C, most of the incorporated label (83%) was found in 16:1 PLFAs and only 2% in 18:1 PLFAs. In soils incubated at 22 °C, almost equal amounts of ¹³C label were incorporated into 16:1 PLFAs and 18:1 PLFAs (33% and 36%, respectively). We concluded that the highly active methane-oxidizing community in cold permafrost-affected soils was dominated by type I methanotrophs under in situ conditions. However, rising temperatures, as predicted for the future, seem to increase the importance of type II methanotrophs, which may affect methane cycling in northern wetlands.     

Varying atmospheric methane concentrations affect soil methane oxidation rates and methanotroph populations in pasture,an adjacent pine forest,and a landfill

We describe experiments to better understand how CH₄ oxidation rates by different methanotroph communities respond to changing CH₄ concentrations. We used a novel system of automatically monitored chambers to investigate the response of CH₄ oxidation rates in a New Zealand pasture and adjacent pine forest soil exposed to varying atmospheric CH₄ concentrations.Type II methanotrophs that dominate CH₄ oxidation in the forest soil became progressively saturated as CH₄ concentrations rose from ambient (1.8 ppmv) to 570 ppmv, as shown by a decrease in uptake efficiency from 20% to 2% removal. By contrast, CH₄ oxidation in the pasture soil where Type I methanotrophs dominate increased in proportion to the increase in CH₄ inlet concentration, oxidising about 2% of the inlet CH₄ flux throughout. Modelling based on Michaelis-Menten kinetics revealed that low-affinity (Type I) methanotrophs were solely responsible for CH₄ oxidation in pasture soils, whereas high affinity (Type II) methanotrophs only contributed about 10% of the CH₄ oxidation in the forest soil. Increased aeration status using a soil–perlite (1:1) mixture doubled CH₄ oxidation rates at both ambient (1.8 ppmv) and 40 ppmv atmospheric CH₄. A similar volcanic soil previously exposed for 8 y to high CH₄ fluxes from a landfill had removal efficiencies consistently above 95% for atmospheric CH₄ concentrations up to 7500 ppmv when the CH₄ oxidation rate was7000 μg CH₄ kg⁻¹soil h⁻¹.     

Soil methane oxidation and methanotroph responses to afforestation of pastures with Pinus radiata stands

Afforestation of pastures in New Zealand reduces methane (CH₄) production from soil, while also stimulating oxidation of atmospheric CH₄ by soil methanotrophs. However, neither the mechanisms by which soil CH₄ oxidation is enhanced by afforestation, nor how long after forest planting tree-dependent responses in CH₄ oxidation become detectable are fully known. Here, we investigated the effects of different-aged stands (5-20 y) of the exotic pine (Pinus radiata (D. Don)) on CH₄ oxidation and methanotrophic community structure in soils, compared with adjacent, long-established pastures. Two of the pastures were on volcanic soils and two were on non-volcanic soils. Although the CH₄ fluxes in soils from these young stands were not significantly different from those in the associated pastures, the rate of oxidation of added ¹³CH₄ was higher in the pine soils. Both fluxes and ¹³CH₄ oxidation rates were higher in the volcanic than the non-volcanic soils. Combined phospholipid fatty acid (PLFA) and stable isotope probe (SIP) analyses suggested that type II methanotrophs (PLFA C18:1ω7) were most active in all soils followed by uncultivable bacteria (C17:0ai). Molecular analysis of the methanotrophic community structure using pmoA (particulate methane monooxygenase) genes suggested that a particular type II methanotroph (TRF 35) was dominant in all soils, but more so in the pine than in pasture soils. A type I methanotroph (TRF 245) was more prevalent in the pasture than in associated pine soils, whereas TRF 128 (a type II methanotroph) was slightly more dominant in soils under pine. Cloning and sequencing data suggest TRFs 35 and 128, which differ from one another, belong to distant relatives of Methylocapsa sp; TRF 245 is related to Methylococcus capsulatus. Land-use change resulted in changes in soil bulk density, porosity, moisture contents and in methanotrophic community structure. Methane oxidation rates were most closely related to soil moisture, as well as to the methanotrophic community structure, and nitrate-N, extractable C and total C concentrations. Stepwise multiple regression also suggested a weak effect (P = 0.06) of stand age on CH₄ oxidation rate. By contrast, the responses of the methanotrophic community structure to this land-use change were more readily detected by the specific molecular analyses, and indicated a predominance of type II methanotrophs in pine soils.     

Assessing effects of earthworm cast on methanotrophic community in a soil biocover by concurrent use of microarray and quantitative real-time PCR

Effects of earthworm cast (EC) on the methanotrophic community in a soil biocover were evaluated using microarray and quantitative real-time PCR (qRT-PCR). Soil was collected from a biocover with either soil alone or a mixture of soil and EC (3:1, w/w). The microarray results showed a more diverse methanotrophic community in the EC biocover than that in the soil biocover (p < 0.05). A principal component analysis result confirmed a substantial change in the methanotrophic community structure due to the added EC. Type I methanotrophs dominated both biocovers, with Methylobacter being most abundant. The qRT-PCR results showed that EC greatly increased the methanotrophic population levels (p < 0.05) up to 100 fold. In conclusion, EC can increase the population density of methanotrophs as well as their diversity, resulting in a substantial shift in the community structure. The results confirmed the promising potential of EC as a bed material for enhancing the biocover performance.     

Naphthalene addition to soil surfaces: A feasible method to reduce soil micro-arthropods with negligible direct effects on soil C dynamics

Soil fauna are a key component of soil biodiversity and a driver of soil functioning. While the importance of soil fauna is well recognized, quantitative estimates of the role of soil fauna on soil biogeochemical processes, such as plant litter decomposition, are limited by methodological constraints. The addition of naphthalene, a polycyclic aromatic hydrocarbon (C₁₀H₈), to suppress soil fauna has been used for decades in decomposition experiments, but its efficacy remains questioned. In fact, we lack a rigorous field assessment of the efficacy of naphthalene additions for soil fauna suppression and potential non-target effects on the soil microbial community and carbon cycling. We added naphthalene at a high rate (477 g m⁻²) monthly for 23 months on the bare soil surface of a tallgrass prairie. We determined the effect of such additions on the abundance of nematodes and micro-arthropods along the soil profile to a depth of 20 cm at 11, 16 and 23 months after initiating naphthalene application. We used the variation in the natural ¹³C abundance of the naphthalene (δ¹³C – 25.5‰) as compared to the native soil (δ¹³C ∼ −17‰) to quantify naphthalene contribution to soil CO₂ efflux and microbial biomarkers (PLFA). Naphthalene addition significantly reduced the abundance of oribatid mites (−45%), predatory mites (−52%) and springtails (−49%), but did not affect nematode abundance. The ¹³C abundance of a few Gram-negative (cy17:0, 18:1ω7c, 16:1ω7c), Gram-positive (a15:0, i15:0) and Actinobacteria (10Me-16:0, 10Me-18:0) PLFA markers decreased significantly in naphthalene treated plots, indicating bacterial utilization of naphthalene-derived C. Mixing models showed this contribution to be highly variable, with the highest naphthalene-C incorporation for Gram negative bacteria. Naphthalene-C was not incorporated in fungal PLFAs. This microbial utilization did not affect overall microbial abundance, community structure or activity, estimated as soil respiration. This experiment proves that naphthalene addition is a feasible method to reduce soil micro-arthropods in the field, with negligible direct effects on soil nematodes, microbial abundance and C dynamics.     

Soil organic matter in soil depth profiles: Distinct carbon preferences of microbial groups during carbon transformation

This study investigates how carbon sources of soil microbial communities vary with soil depth. Microbial phospholipid fatty acids (PLFA) were extracted from 0–20, 20–40 and 40–60 cm depth intervals from agricultural soils and analysed for their stable carbon isotopes (δ¹³C values). The soils had been subjected to a vegetation change from C3 (δ¹³C≈?29.3‰) to C4 plants (δ¹³C≈?12.5‰) 40 years previously, which allowed us to trace the carbon flow from plant-derived input (litter, roots, and root exudates) into microbial PLFA. While bulk soil organic matter (SOM) reflected ≈12% of the C4-derived carbon in top soil (0–20 cm) and 3% in deeper soil (40–60 cm), the PLFA had a much higher contribution of C4 carbon of about 64% in 0–20 cm and 34% in 40–60 cm. This implies a much faster turnover time of carbon in the microbial biomass compared to bulk SOM. The isotopic signature of bulk SOM and PLFA from C4 cultivated soil decreases with increasing soil depth (?23.7‰ to ?25.0‰ for bulk SOM and ?18.3‰ to ?23.3‰ for PLFA), which demonstrates decreasing influence of the isotopic signature of the new C4 vegetation with soil depth. In terms of soil microbial carbon sources this clearly shows a high percentage of C4 labelled and thus young plant carbon as microbial carbon source in topsoils. With increasing soil depth this percentage decreases and SOM is increasingly used as microbial carbon source. Among all PLFA that were associated to different microbial groups it could be observed that (a) depended on availability, Gram-negative and Gram-positive bacteria prefer plant-derived carbon as carbon source, however, (b) Gram-positive bacteria use more SOM-derived carbon sources while Gram-negative bacteria use more plant biomass. This tendency was observed in all three-depth intervals. However, our results also show that microorganisms maintain their preferred carbon sources independent on soil depth with an isotopic shift of 3–4‰ from 0–20 to 40–60 cm soil depth.     

Methane oxidation activity and bacterial community composition in a simulated landfill cover soil is influenced by the growth of Chenopodium album L.

Oxygen availability in landfill cover soil is a major limitation to the growth and activity of methanotrophs as methane oxidation is an aerobic microbial process. Plants tolerant to high concentrations of landfill gas (LFG) may play an important role in improving methane oxidation within landfill cover soil and reducing emission of methane, a greenhouse gas, from it. In this study, the effect of an LFG tolerant plant Chenopodium album L. on methane oxidation activity (MOA) and bacterial community composition in landfill cover soil was investigated. Soil samples from four simulated lysimeters with and without LFG and plant vegetation were taken at 4 stages during the plant's development cycle. Results showed that the total number of culturable bacteria in soil could be significantly increased (P < 0.05) by the growth of C. album. The total number of methanotrophs and MOA in soils with LFG was significantly higher (P < 0.05) than in soils without LFG on sampling days 90, 150 and 210. The total number of methanotrophs and MOA in lysimeters with LFG added increased in the presence of C. album when the plant entered the seed setting stage. Polymerase chain reaction and denaturing gradient gel electrophoresis (PCR-DGGE) gel patterns of 16S rDNA gene fragment and band sequencing analyses showed apparent differences in soil bacterial communities in the presence of LFG and plant vegetation. Members of the genus Methylosarcina were found to be the active and dominant methanotrophs in rhizosphere soil of C. album with LFG, while Methylococcus, Methylocystis, and Methylosinus were the primary methanotroph genera in LFG soil without C. album. Thus, C. album appears to select for specific methanotrophic bacteria in the presence of LFG. Soil MOA and microbial diversity can also be significantly affected by the presence of this plant.     

Investigation of the effect of ammonium sulfate on populations of ambient methane oxidising bacteria by C-labelling and GC/C/IRMS analysis of phospholipid fatty acids

The oxidation of atmospheric methane by methanotrophic bacteria residing in soils constitutes an important terrestrial methane sink with previous studies having revealed the inhibition of microbially mediated methane oxidation in the presence of salt ions. The bacteria responsible for ambient methane oxidation are not amenable to currently available methods of culturing, resulting in the need for a method of in situ analysis. A combination of phospholipid fatty acid (PLFA) analysis and stable isotopic labelling has been employed in this investigation as a means of cultivation-independent bacterial analysis. Soil samples were treated with an ammonium sulfate solution at a concentration that was known to inhibit methane oxidation or with distilled water, serving as a control, and incubated with ¹³C-labelled methane. PLFAs were analysed by GC/C/IRMS in order to determine their ¹³C content and, hence, the PLFA distribution of the methane oxidising bacteria. Ammonium sulfate treatment reduced the amount of ¹³C incorporated into the majority of PLFAs except the i17:0 PLFA in the presence of high concentrations of methane. These results implied a shift in the composition of the methane oxidising bacterial community in the soils treated with ammonium ions, with the treatment appearing to suppress one group of organisms more than another.     

Effect of soil moisture and bovine urine on microbial stress

While many studies have examined the cycling of urinary nutrients, few have focused on the effects ruminant urine might have on the soil microbial community. Urine application can cause microbial communities to become stressed, potentially changing community composition and microbial function with subsequent effects on nutrient dynamics. Identification of the factors that stress microbes may assist in explaining ruminant urine effects on nutrient cycling. In this laboratory study bovine urine, with either a high (15.0 g K⁺ l^?1) or low (10.4 g K⁺ l^?1) salt concentration, was added to repacked soil cores maintained at high or low soil moisture contents (70 or 35% water-filled pore space, respectively). Control cores did not receive urine. Microbial stress was measured using phospholipid fatty acid (PLFA) biomarker ratios. Urine addition increased stress as indicated by a decrease in the iso15:0/anteiso15:0 PLFA ratio from >1.35 to <0.95 in both wet and dry soils and by an increase in the 18:1ω9trans/18:1ω9cis PLFA ratio from 1.4 to 1.9 from day 8 onwards in wet soils. Higher stress was indicated by a lower Gram-positive/Gram-negative PLFA ratio in the urine treatments than in the control treatments on day 29 and this may have been a response to the reduction in substrate availability as the experiment progressed. The PLFA biomarkers showed that the salt treatments did not induce stress. Stress induced by urine addition and wet soil treatments was also indicated by principal component analyses and the metabolic quotient for CO₂, respectively. Thus microbial stress was induced by both urine addition and high soil moisture content, but not specifically by increasing the urinary salt concentration.     

Impact of the antibiotic sulfadiazine and pig manure on the microbial community structure in agricultural soils

Large amounts of veterinary antibiotics enter soil via manure of treated animals. The effects on soil microbial community structure are not well investigated. In particular, the impact of antibiotics in the presence of manure is poorly understood. In this study, two agricultural soils, a sandy Cambisol (KS) and a loamy Luvisol (ML), were spiked with manure and sulfadiazine (SDZ; 0, 10 and 100 μg g^?1) and incubated for 1, 4, 32 and 61 days. Untreated controls received only water. The microbial community structure was characterised by investigating phospholipid fatty acids (PLFA) and using PCR–denaturing gradient gel electrophoresis (DGGE) of 16S rDNA. The total concentration of PLFA increased with addition of manure and was reduced by both SDZ concentrations at incubation times >4 days. The SDZ addition decreased the bacteria:fungi ratio. The largest stress level, measured as ratio of PLFA (cyc17:0 + cyc19:0)/(16:1ω7c + 18:1ω7c), was found for the controls, followed by the manure treatments and the SDZ treatments. A discriminant analysis of the PLFA clearly separated treatments and incubation times. Both soils differed in total PLFA concentrations and Gram^?:Gram⁺ ratios, but showed similar changes in PLFA pattern upon soil treatment. Effects of manure and SDZ on the bacterial community structure were also revealed by DGGE analysis. Effects on pseudomonads and β-proteobacteria were less pronounced. While community structure remained altered even after two months, the extractable concentrations of SDZ decreased exponentially and the remaining solution concentrations after 32 days were ≤27% of the spiking concentration. Our results demonstrate that a single addition of SDZ has prolonged effects on the microbial community structure in soils.     

The differential effects of ammonium and nitrate on methanotrophs in rice field soil

It has been known that nitrogenous fertilizers can either stimulate or inhibit methane oxidation in soils. The mechanism, however, remains unclear. Here we conducted laboratory incubation experiments to evaluate the effects of ammonium versus nitrate amendment on CH₄ oxidation in a rice field soil. The results showed that both N forms stimulated CH₄ oxidation. But nitrate stimulated CH₄ oxidation to a greater extent than ammonium per unit N base. The 16S rRNA genes and the pmoA genes were analyzed to determine the dynamics of total bacterial and methanotrophic populations, respectively. The methanotrophic community consisted of type I and type II methanotrophs and was dominated by type I group after two weeks of incubation. Nitrate promoted both types of methanotrophs, but ammonium promoted only type I. DNA-based stable isotope probing confirmed that ammonium stimulated the incorporation of ¹³CH₄ into type I methanotrophs but not type II, while nitrate caused almost homogenous distribution of ¹³CH₄ in type I and type II methanotrophs. Our study suggests that nitrate can promote CH₄ oxidation more significantly than ammonium and is probably a better N source for both types of methanotrophs in rice field soil. More investigations, e.g. using ¹⁵N labeling, are necessary to elucidate this possibility.     

Responses of soil microbial biomass and community composition to biological soil crusts in the revegetated areas of the Tengger Desert

As a key component of desert ecosystems, biological soil crusts (BSCs) play an important role in dune fixation and maintaining soil biota. Soil microbial properties associated with the colonization and development of BSCs may indicate soil quality changes, particularly following dune stabilization. However, very little is known about the influence of BSCs on soil microbes in sand dunes. We examined the influence of BSCs on soil microbial biomass and community composition in revegetated areas of the Tengger Desert. BSCs increased soil microbial biomass (biomass C and N), microbial phospholipid fatty acid (PLFA) concentrations and the ratio of fungal to bacterial PLFAs. The effects varied with crust type and crust age. Moss crusts had higher microbial biomass and microbial PLFA concentrations than cyanobacteria-lichen crusts. Crust age was positively correlated with microbial biomass C and N, microbial PLFA concentrations, bacterial PLFA concentrations, fungal PLFA concentrations and the ratio of fungal to bacterial PLFAs. BSCs significantly affected microbial biomass C and N in the 0–20 cm soil layers, showing a significant negative correlation with soil depth. The study demonstrated that the colonization and development of BSCs was beneficial for soil microbial properties and soil quality in the revegetated areas. This can be attributed to BSCs increasing topsoil thickness after dunes have been stabilized, creating suitable habitats and providing an essential food source for soil microbes.     

Distinct microbial and faunal communities and translocated carbon in Lumbricus terrestris drilospheres

Lumbricus terrestris is a deep-burrowing anecic earthworm that builds permanent, vertical burrows with linings (e.g., drilosphere) that are stable and long-lived microhabitats for bacteria, fungi, micro- and mesofauna. We conducted the first non-culture based field study to assess simultaneously the drilosphere (here sampled as 0–2 mm burrow lining) composition of microbial and micro/mesofaunal communities relative to bulk soil. Our study also included a treatment of surface-applied ¹³C- and ¹⁵N-labeled plant residue to trace the short-term (40 d) translocation of residue C and N into the drilosphere, and potentially the assimilation of residue C into drilosphere microbial phospholipid fatty acids (PLFAs). Total C concentration was 23%, microbial PLFA biomass was 58%, and PLFAs associated with protozoa, nematodes, Collembola and other fauna were 200-to-300% greater in the drilosphere than in nearby bulk soil. Principal components analysis of community PLFAs revealed that distributions of Gram-negative bacteria and actinomycetes and other Gram-positive bacteria were highly variable among drilosphere samples, and that drilosphere communities were distinct from bulk soil communities due to the atypical distribution of PLFA biomarkers for micro- and mesofauna. The degree of microbial PLFA ¹³C enrichment in drilosphere soils receiving ¹³C-labeled residue was highly variable, and only one PLFA, 18:1ω9c, was significantly enriched. In contrast, 11 PLFAs from diverse microbial groups where enriched in response to residue amendment in bulk soil 0–5 cm deep. Among control soils, however, a significant δ¹³C shift between drilosphere and bulk soil at the same depth (5–15 cm) revealed the importance of L. terrestris for translocating perennial ryegrass-derived C into the soil at depth, where we estimated the contribution of the recent grass C (8 years) to be at least 26% of the drilosphere soil C. We conclude that L. terrestris facilitates the translocation of plant C into soil at depth and promotes the maintenance of distinct soil microbial and faunal communities that are unlike those found in the bulk soil.     

Evaluating effects of sewage sludge and household compost on soil physical,chemical and microbiological properties

Recycling of organic wastes within agriculture may help maintain soil fertility via effects on physical, chemical and biological properties. Efficient use, however, requires an individual assessment of waste products, and effects should be compared with natural variations due to climate and soil type. An 11-month incubation experiment was conducted between April 1998 and March 1999, in which a sandy loam without or with anerobically digested sewage sludge (4.2 t dry matter (DM) ha⁻¹) or household compost (17 t DM ha⁻¹) was incubated under constant laboratory conditions at 10 °C, as well as in the field. The following properties were monitored: wet-stability of soil aggregates, clay dispersibility, hot-water extractable carbohydrates, resin-extractable P_i, inorganic N, biomass C and N, PLFA profiles, FDA hydrolysis activity, β-glucosidase activity and CO₂ evolution. In general, effects of waste amendment were positive, but moderate compared to the dynamics observed in unamended soil, and mainly occurred in the first several weeks after amendment. The temporal dynamics of inorganic N, FDA hydrolysis activity, biomass C and PLFA composition appeared to be faster under the fluctuating climatic conditions in the field. To evaluate accumulated effects of repeated waste applications, soil was also sampled from a field trial, in which the sewage sludge and household compost had been applied at the same rates as in the incubation study for three consecutive years. Sampling took place after the final harvest, i.e. 5 months after the final waste application. Compost amendment had increased potentially mineralizable N by a factor of 1.8, and sludge amendment had increased the amount of resin-extractable P_i by a factor of 1.6. However, there were no accumulated effects of waste amendment on the fraction of soil in wet-stable aggregates, or on the microbiological properties tested, which supported the observation from the incubation study that effects of organic wastes were transient.     

Application of 13C-labeled litter and root materials for in situ decomposition studies using phospholipid fatty acids

Microorganisms play a central role in litter decomposition and partitioning C between CO₂ evolution and sequestration of C into semi-permanent pools in soils. At the ecosystem level, forest stand age influences rates of litter accumulation and quality, and micro-climatology which could affect the microbial community structure and C sequestration processes. Although numerous laboratory experiments have studied the decomposition of model ¹³C-labeled compounds, few studies have verified these findings under field conditions. The objective of this study was to track decomposition of ¹³C-labeled Douglas-fir (Pseudotsuga menziesii (Mirb.) Franco) materials into the soil microbial community using ¹³C-phospholipids fatty acid (PLFA) analysis in three different aged forest stands. A field experiment was conducted that had three forest stand age treatments: old-growth (>500 yrs); 8-year-old clear-cut (CC8); and 25-year-old clear-cut (CC25) (landscape reps of n = 2). Each stand age had in situ microcosms that were amended with either ¹³C-labeled surface litter or root material. Microcosms were destructively sampled seven times over a 22-month period and the soil was analyzed for the relative amounts of ¹³C incorporated (¹³C%_INCORP) into PLFAs and the proportional distribution of ¹³C incorporated into PLFAs. The ¹³C%_INCORP was affected by stand age and ¹³C source with greater ¹³C%_INCORP in samples from CC8 than OG or CC25. Also, the level of ¹³C%_INCORP was greater for labeled litter than root material in five out of the seven sample dates. In general, 18:1ω9 and 18:2ω6,9 (common fungal biomarkers) had the greatest amount of ¹³C incorporation throughout the study period in both clear-cut and old-growth sites, especially in plots with ¹³C-labeled litter. Our data showed a low fungal ¹³C-PLFA: bacterial ¹³C-PLFA ratio (0.45) 1 month after incubation was initiated compared to 5, 7 and 9 months after incubation (two of these dates were >1.0). This suggests that initially bacteria played a greater role in the decomposition of the added needles with fungi playing a more important role in subsequent sample dates. Our results illustrate that the use of ¹³C-labeled materials in field studies coupled with¹³C-PLFA profiling is a powerful tool for determining microbial dynamics during decomposition – enabling statistically significant detection of land management treatment effects on C acquisition by microbial functional groups.     

Soil microbial community dynamics and phenanthrene degradation as affected by rape oil application

The effects of rape oil application on soil microbial communities and phenanthrene degradation were characterized by examining phenanthrene concentrations, changes in microbial composition and incorporation of [¹³C] phenanthrene-derived carbon into phospholipid fatty acids (PLFAs). A Haplic Chernozem was incubated with and without rape oil in combination with and without phenanthrene over 60 days. High-performance liquid chromatography (HPLC) analysis showed a net reduction in extractable phenanthrene in the soils treated with rape oil but no net reduction in the soils without rape oil. Rape oil application increased the total PLFA content and changed microbial community composition predominantly due to growth of fungal groups and Gram-positive bacterial groups. Under rape oil and phenanthrene amendment all detected microbial groups grew until day 24 of incubation. The ¹³C PLFA profiles showed ¹³C enrichment for the PLFAs i14:0, 15:0, 18:0, 18:1ω5 and the fungal biomarker 18:2ω6,9 under rape oil application. Fungal PLFA growth was highest among detected all PLFAs, but its ¹³C incorporation was lower compared to the Gram-positive and Gram-negative bacteria PLFAs. Our results demonstrate the effect of rape oil application on the abundance of microbial groups in soil treated with phenanthrene and its impact on phenanthrene degradation.     

Tree species affect atmospheric CH4 oxidation without altering community composition of soil methanotrophs

Plant species exert strong effects on ecosystem functions and one of the emerging, and difficult to test hypotheses, is that plants alter soil functions through changing the community structure of soil microorganisms. We tested the hypothesis for atmospheric CH₄ oxidation by using soil samples from a Siberian afforestation experiment and exposing them to ¹³C-CH₄. We determined the activity of the soil methanotrophs under different tree species at three levels of initial CH₄ concentration (30, 200 and 1000 ppm) thus distinguishing the activities of low- and high-affinity methanotrophs. Half of the samples were incubated with ¹³C-enriched CH₄ (99.9%) and half with ¹²C-CH₄. This allowed an estimation of the amount of ¹³C incorporated into individual PLFAs and determination of PLFAs of methanotrophs involved in CH₄ oxidation at the different CH₄ concentrations. Tree species strongly altered the activity of atmospheric CH₄ oxidation without appearing to change the composition of high-affinity methanotrophs as evidenced by PLFA ¹³C labeling. The low diversity of atmospheric CH₄ oxidizers, presumably belonging to the UCSα group, may explain the lack of tree species effects on the composition of soil methanotrophs. We submit that the observed tree species effects on atmospheric CH₄ oxidation indicate an effect on biomass or cell-specific activities rather than by a community change and this may be related to the impact of the tree species on soil N cycling.     

Abundance and community composition of methanotrophs in a Chinese paddy soil under long-term fertilization practices

Background, aim, and scope  As the second most important greenhouse gas, methane (CH₄) is produced from many sources such as paddy fields. Methane-oxidizing bacteria (methanotrophs) consume CH₄ in paddy soil and, therefore, reduce CH₄ emission to the atmosphere. In order to estimate the contribution of paddy fields as a source of CH₄, it is important to monitor the effects of fertilizer applications on the shifts of soil methanotrophs, which are targets in strategies to combat global climate change. In this study, real-time polymerase chain reaction (PCR) and denaturing gradient gel electrophoresis (DGGE) based on 16S rRNA and pmoA genes, respectively, were used to analyze the soil methanotrophic abundance and community diversity under four fertilization treatments: urea (N), urea and potassium chloride (NK), urea, superphosphate, and potassium chloride (NPK), and urea, superphosphate, potassium chloride, and crop residues (NPK+C), compared to an untreated control (CON). The objective of this study was to examine whether soil methanotrophs responded to the long-term, different fertilizer regimes by using a combination of quantitative and qualitative molecular approaches. Materials and methods  Soil samples were collected from the Taoyuan Experimental Station of Agro-ecosystem Observation at Changde (28°55′ N, 111°26′ E), central Hunan Province of China, in July 2006. Soil DNAs were extracted from the samples, then the 16S rRNA genes were quantified by real-time PCR and the pmoA genes were amplified via general PCR followed by DGGE, cloning, sequencing, and phylogenetic analysis. The community diversity indices were assessed through the DGGE profile. Results  Except for NPK, other treatments of N, NK, and NPK+C showed significantly higher copy numbers of type I methanotrophs (7.0–9.6 × 10⁷) than CON (5.1 × 10⁷). The copy numbers of type II methanotrophs were significantly higher in NPK+C (2.8 × 10⁸) and NK (2.5 × 10⁸) treatments than in CON (1.4 × 10⁸). Moreover, the ratio of type II to type I methanotrophic copy numbers ranged from 1.88 to 3.32, indicating that the type II methanotrophs dominated in all treatments. Cluster analyses based on the DGGE profile showed that the methanotrophic community in NPK+C might respond more sensitively to the environmental variation. Phylogenetic analysis showed that 81% of the obtained pmoA sequences were classified as type I methanotrophs. Furthermore, the type I-affiliated sequences were related to Methylobacter, Methylomicrobium, Methylomonas, and some uncultured methanotrophic clones, and those type II-like sequences were affiliated with Methylocystis and Methylosinus genera. Discussion  There was an inhibitory effect on the methanotrophic abundance in the N and a stimulating effect in the NK and NPK+C treatments, respectively. During the rice-growing season, the type II methanotrophs might be more profited from such a coexistence of low O₂ and high CH₄ concentration environment than the type I methanotrophs. However, type I methanotrophs seemed to be more frequently detected. The relatively complex diversity pattern in the NPK+C treatment might result from the strong CH₄ production. Conclusions  Long-term fertilization regimes can both affect the abundance and the composition of the type I and type II methanotrophs. The inhibited effects on methanotrophic abundance were found in the N treatment, compared to the stimulated effects from the NK and NPK+C treatments. The fertilizers of nitrogen, potassium, and the crop residues could be important factors controlling the abundance and community composition of the methanotrophs in the paddy soil. Recommendations and perspectives  Methanotrophs are a fascinating group of microorganisms playing an important role in the biogeochemical carbon cycle and in the control of global climate change. However, it is still a challenge for the cultivation of the methanotrophs, although three isolates were obtained in the extreme environments very recently. Therefore, future studies will be undoubtedly conducted via molecular techniques just like the applications in this study.     

Effects of vegetation on soil microbial C,N, and P dynamics in a tropical forest and savanna of Central Africa

The forest–savanna transition zone is widely distributed on nutrient-poor oxisols in Central Africa. To reveal and compare the nutrient cycle in relation to soil microbes for forest and savanna vegetation in this area, we evaluated seasonal fluctuations in microbial biomass carbon (MBC), nitrogen (MBN), and phosphorus (MBP) for 13 months as well as soil moisture, temperature, soil pH levels, and nutrients for both vegetation types in eastern Cameroon. Soil pH was significantly lower in forest (4.3) than in savanna (5.6), and soil N availability was greater in forest (87.1 mg N kg⁻¹ soil) than in savanna (32.9 mg N kg⁻¹ soil). We found a significant positive correlation between soil moisture and MBP in forest, indicating the importance of organic P mineralization for MBP, whereas in savanna, we found a significant positive correlation between soil N availability and MBP, indicating N limitation for MBP. These results suggest that for soil microbes, forest is an N-saturated and P-limited ecosystem, whereas savanna is an N-limited ecosystem. Additionally, we observed a significantly lower MBN and larger MB C:N ratio in forest (50.7 mg N kg⁻¹ soil and 8.6, respectively) than in savanna (60.0 mg N kg⁻¹ soil and 6.5, respectively) during the experimental period, despite the rich soil N condition in forest. This may be due to the significantly lower soil pH in forest, which influences the different soil microbial communities (fungi-to-bacteria ratio) in forest versus savanna, and therefore, our results indicate that, in terms of microbial N dynamics, soil pH rather than soil substrate conditions controls the soil microbial communities in this area. Further studies should be focused on soil microbial community, such as PLFA, which was not evaluated in the present study.     

Transient elevation of carbon dioxide modifies the microbial community composition in a semi-arid grassland

Using open-top chambers (OTC) on the shortgrass steppe in northern Colorado, changes of microbial community composition were followed over the latter 3 years of a 5-year study of elevated atmospheric CO₂ as well as during 12 months after CO₂ amendment ended. The experiment was composed of nine experimental plots: three chambered plots maintained at ambient CO₂ levels of 360±20 μmol mol^?1 (ambient treatment), three chambered plots maintained at 720±20 μmol mol^?1 CO₂ (elevated treatment) and three unchambered plots. The abundance of fungal phospholipid fatty acids (PLFAs) shifted in the shortgrass steppe under the influence of elevation of CO₂ over the period of 3 years. Whereas the content of the fungal signature molecule (18:2ω6) was similar in soils of the ambient and elevated treatments in the third year of the experiment, CO₂ treatment increased the content of 18:2ω6 by around 60% during the two subsequent years. The shift of microbial community composition towards a more fungal dominated community was likely due to slowly changing substrate quality; plant community forage quality declined under elevated CO₂ because of a decline of N in all tested species as well as shift in species composition towards greater abundance of the low forage quality species (Stipa comata). In the year after which CO₂ enrichment had ceased, abundances of fungal and bacterial PLFAs in the post-CO₂ treatment plots shifted slowly back towards the control plots. Therefore, quantity and quality of available substrates had not changed sufficiently to shift the microbial community permanently to a fungal dominated community. We conclude from PLFA composition of soil microorganisms during the CO₂ elevation experiment and during the subsequent year after cessation of CO₂ treatment that a shift towards a fungal dominated system under higher CO₂ concentrations may slow down C cycling in soils and therefore enhance C sequestration in the shortgrass steppe in future CO₂-enriched atmospheres.