QMEC: a tool for high-throughput quantitative assessment of microbial functional potential in C,N, P,and S biogeochemical cycling

Microorganisms are major drivers of elemental cycling in the biosphere. Determining the abundance of microbial functional traits involved in the transformation of nutrients, including carbon(C), nitrogen(N), phosphorus(P) and sulfur(S), is critical for assessing microbial functionality in elemental cycling. We developed a high-throughput quantitative-PCR-based chip, Quantitative microbial element cycling(QMEC), for assessing and quantifying the genetic potential of microbiota to mineralize soil organic matter and to release C, N, P and S. QMEC contains 72 primer pairs targeting 64 microbial functional genes for C, N, P, S and methane metabolism. These primer pairs were characterized by high coverage(average of 18–20 phyla covered per gene)and sufficient specificity(70% match rate) with a relatively low detection limit(7–102 copies per run). QMEC was successfully applied to soil and sediment samples, identifying significantly different structures, abundances and diversities of the functional genes(P0.05). QMEC was also able to determine absolute gene abundance. QMEC enabled the simultaneous qualitative and quantitative determination of 72 genes from 72 samples in one run, which is promising for comprehensively investigating microbially mediated ecological processes and biogeochemical cycles in various environmental contexts including those of the current global change.     

Dissolved phosphorus concentrations and sediment interactions in effluent–dominated Ozark streams

Phosphorus (P) loads from point sources have a significant influence on dissolved P concentrations in streams and sediment-water column dynamics. The goal of this study was to quantify dissolved P concentrations and sediment-P interactions in Ozark (USA) headwater streams with high point source P loads. Specifically, the objectives were to: (1) compare soluble reactive P (SRP) upstream and downstream from wastewater treatment plant (WWTP) effluent discharges; (2) examine longitudinal gradients in SRP downstream from WWTPs; (3) evaluate the effect of WTTP P inputs on sediment-water column P equilibrium and sediment exchangeable P. Water and sediment samples were collected, extracted and analyzed from July 2002 through June 2003 at these Ozark streams. Mean SRP concentrations in the select Ozark streams were significantly greater downstream from effluent discharges (0.08–2.10 mg L⁻¹) compared to upstream (0.02–0.12 mg L⁻¹). Effluent discharge from the WWTPs increased equilibrium concentrations between stream sediments and the water column; mean sediment equilibrium P concentration (EPC₀) was between 0.01–0.07 mg L⁻¹ upstream from WWTP and the increase downstream was proportional to that observed in water column SRP. Sediment exchangeable P (EXP) was greater downstream from the effluent discharges (0.3–6.8 mg kg⁻¹) compared to upstream (0.03–1.4 mg kg⁻¹), representing a substantial transient storage of P inputs from WWTPs. Furthermore, P was generally not retained in these stream reaches when dilution was considered using a hydrologic tracer and was released in one stream reach where effluent P concentrations decreased over the study period. Thus, the effect of the WWTPs was profound in these streams increasing water column and sediment-bound P, and also reducing the ability of these stream reaches to retain P. In P-enriched streams, effluent P discharges likely regulate sediment and aqueous phase P equilibrium and sediment bioavailable P, not the sediments.     

Holm oak decline triggers changes in plant succession and microbial communities,with implications for ecosystem C and N cycling

Background and aims

The occurrence of drought-induced forest die-off events is projected to increase in the future, but we still lack complete understanding of its impact on plant-soil interactions, soil microbial diversity and function. We investigated the effects of holm oak (Quercus ilex) decline (HOD) on soil microbial community and functioning, and how these effects relate to changes in the herbaceous community.

Methods

We selected 30 holm oak trees with different defoliation degrees (healthy, affected and dead) and analyzed soil samples collected under the canopy (holm oak ecotype) and out of the influence (grassland ecotype) of each tree.

Results

HOD increased potential nitrogen (N) mineralization and decreased inorganic N concentrations. These results could be partially explained by changes in the herbaceous composition, an increased herbaceous abundance and changes in soil microbial functional diversity and structure, with HOD favoring bacteria against fungi. Moreover, herbaceous abundance and microbial functional diversity of holm oak and grassland ecotypes converged with HOD.

Conclusions

Our results show that HOD triggers a cascade effect on plant understory and soil microbial communities, as well as a plant succession (savannization) process, where understory species colonize the gaps left by dead holm oaks, with important implications for ecosystem C and N budgets.

     

Ethanol, BTEX and microbial community interactions in E-blend contaminated soil slurry

Degradation of benzene, toluene, ethylbenzene, m-, p- and o-xylenes (BTEX) and microbial community shifts in soil slurries contaminated with ethanol–gasoline blends (E-blends), containing 10, 50 or 90% (v/v) ethanol (E10, E50 and E90) were studied in soil slurries previously uncontaminated, contaminated by E-blends or ethanol. BTEX originating from E50 degraded fastest whereas from E10 slowest. Among the individual compounds, ethylbenzene degraded fastest (max 30% d⁻¹), and o-xylene slowest (min 1% d⁻¹) during aerobic conditions in previously not contaminated soils. Previous contamination by E-blends increased BTEX degradation significantly (3–19 times) compared with previously uncontaminated soils, whereas previous contamination with ethanol did not show significant difference in BTEX degradation. At least one type of the E-blends during aerobic conditions had a positive effect on total PLFAs (phospholipid fatty acids) and specific PLFAs, i.e. 10Me18:0, 16:1ω6 and cy17:0, but had a negative effect on cy19:0 and 18:2ω6,9c. The effects on total PLFAs, as well as the individual PLFAs, were particularly strong after repeated contamination. The single most affected PLFA was 16:1ω6, which increased 23 times during E10 treatment in soil slurries previously contaminated by E-blends. Altogether, the various E-blends had significantly different effects on BTEX degradation and also on individual PLFAs under aerobic conditions.     

Dynamics of C, N, P and microbial community composition in particulate soil organic matter during residue decomposition

Decomposing residues can be an important source of nutrients for plants, especially of N and P, but the relationship between N and P release and microbial community dynamics have rarely been studied. Two pea (Pisum sativum L.) residues with contrasting chemical composition, shoots from flowering pea (Pea-Y) with 2.9 mg P and 36 mg N kg⁻¹ and from mature pea (Pea-M) with 0.3 mg P and 13 mg N kg⁻¹, were added at a rate of 20 g kg soil⁻¹ to a sandy soil low in nutrients. Particulate organic matter (POM) was isolated on days (d) 0, 5, 15, 28, 42 and 61 after residue addition and analysed for C, N, P and microbial community structure (fatty acid methyl ester analysis). The recovery of POM from residue-amended soils decreased over time to 30–40% of added amounts for both residues. Apart from d 0, the N concentration in POM was lower in residue-amended soil than in the control. Due to a rapid decrease in P concentration during the first 5 days in Pea-Y and a slow increase over the whole experiment in Pea-M, P concentrations in POM on d 61 were similar in all treatments. In Pea-Y, the dynamics of C, N and P were coupled, with amounts of C, N and P decreasing during the first 15 days and remaining stable thereafter. In Pea-M, a steady loss of C from POM was contrasted by a slight increase in P. As a result, the C/P ratio decreased from 1,330 on d 0 to 390 on d 61. The C/N ratio of Pea-M decreased only during the second phase of decomposition. The different nutrient dynamics in Pea-Y and Pea-M led to similar amounts of N and P in POM towards the end of the incubation. Microbial community composition in the POM in Pea-Y and Pea-M remained distinct from the control, even though it changed over time. POM was shown to be an important source of potentially available nutrients after addition of plant residues. In the unamended soil, stable nutrient amounts in POM suggested very low net nutrient release from native POM compared to POM after residue addition.     

Seasonal patterns of sediment loading and benthic invertebrate community dynamics in Lake Tanganyika, Africa

1. This study investigated the spatiotemporal dynamics of benthic invertebrate communities located close to the mouths of the Kalambo and Lunzua Rivers, at the southern end of Lake Tanganyika, and subject to varying degrees of sediment load. 2. Metrics of exposure to riverine sediments were associated inversely with abundance and diversity of benthic organisms at sampling locations adjacent to both river mouths, indicating a major effect of riverine sediments on the structure of near‐shore benthic communities. 3. A lower abundance of benthos was found at the mouth of the Lunzua River, which exports significantly higher sediment loads than the Kalambo. 4. Seasonal cycles of abundance and diversity in the benthos varied with distance from river mouths. This has important implications for monitoring programmes investigating benthic communities close to the mouths of rivers. 5. Our findings suggest that recent increases in the sediment loading of Lake Tanganyika, owing to anthropogenic catchment disturbance, impact significantly on biological diversity throughout the lake littoral, which provides habitat for the majority of species in the lake.     

Cladoceran community responses to biomanipulation and re‐oligotrophication in Lake Vesijärvi,Finland, as inferred from remains in annually laminated sediment

1. We studied the role of zooplankton in biomanipulation and the subsequent recovery phase in the Enonselkä basin of Lake Vesijärvi, using subfossil cladocerans in annually laminated sediment. Measures to restore the Enonselkä basin included reduction in external nutrient loading and mass removal of plankti‐ and benthivorous fish. Water clarity increased and the lake changed from a eutrophic to a mesotrophic state. However, some signs of increased turbidity were observed after 5–10 years of successful recovery. 2. Annual laminae in a freeze core sample were identified and sliced, based on the seasonal succession of diatoms. Cladoceran remains and rotifer eggs were counted, and Daphnia ephippia and Eubosmina and Bosmina ephippia and carapaces were measured. Annual changes in pelagic species composition were studied with principal component analysis. Individual species abundance, size measurements and various cladoceran‐based indices or ratios (commonly used to reconstruct changes in trophic state and fish predation) were tested for change between four distinct periods: I (1985–1988) dense fish stocks, poor water quality; II (1989–1992) fish removal; III (1993–1997) low fish density, improved water quality; IV (1998–2002) slightly increased fish density and poorer water quality. 3. After the removal of fish, the mean size of Daphnia ephippia and Eubosmina crassicornis ephippia and carapaces increased significantly. In contrast, the percentage of Daphnia did not increase. When based on ephippia, the ratio Daphnia/(Daphnia + E. crassicornis) increased, but the interpretation was obscured by the tolerance of fish predation by small Daphnia and by the fact that bosminids were the preferred food of roach. Moreover, ephippial production by E. crassicornis decreased in recent years. 4. The abundance of Diaphanosoma brachyurum and Limnosida frontosa increased significantly after the fish population was reduced, while that of Ceriodaphnia and rotifers decreased. 5. The expanding littoral vegetation along with improved water clarity was clearly reflected in the concentration of littoral species in the deep sediment core. The species diversity index for the entire subfossil community also increased. 6. The period of faltering recovery was characterised by greater interannual variability and an increased percentage of rotifers. Nevertheless, the mean sizes of Daphnia ephippia and E. crassicornis ephippia and carapaces indicated a low density of fish. The deteriorating water quality was apparently related to multiple stressors in the catchment after rehabilitation, such as intensified lakeshore building, as well as to exceptional weather conditions, challenging the management methods in use.     

铜锈环棱螺生物扰动对"蓝藻水华"水体底泥及其间隙水中碳、氮、磷含量的影响

实验室内建立小型模拟生态系统,根据铜锈环棱螺(Bellamya aeruginosa)的密度设置了3个处理组和1个对照组.结果显示:铜锈环棱螺对底泥0～0.5 cm及0.5～2 cm有机质影响较明显,显著降低了底泥中的C:P.处理组3和处理组2间隙水NH4+-N含量分别在底泥0～0.5cm及0.5～2 cm深度和对照组之间存在显著差异(p＜0.05).间隙水中NO2--N+NO3--N的变化较复杂,处理组NO2--N+NO3--N含量在0～0.5 cm,0.5～2cm及2～4cm与对照组相比差异显著(p＜0.05).间隙水中DIP含量随底泥深度先增后减,在2～4 cm处含量达最大,DIP含量在0.5～2 cm深度处理组与对照组之间有显著差异(p＜0.05).铜锈环棱螺生物干扰增加了底泥表层有机质的含量,同时降低了其稳定性.     

The impact of bottlenecks on microbial survival,adaptation, and phenotypic switching in host–pathogen interactions

Microbial pathogens and viruses can often maintain sufficient population diversity to evade a wide range of host immune responses. However, when populations experience bottlenecks, as occurs frequently during initiation of new infections, pathogens require specialized mechanisms to regenerate diversity. We address the evolution of such mechanisms, known as stochastic phenotype switches, which are prevalent in pathogenic bacteria. We analyze a model of pathogen diversification in a changing host environment that accounts for selective bottlenecks, wherein different phenotypes have distinct transmission probabilities between hosts. We show that under stringent bottlenecks, such that only one phenotype can initiate new infections, there exists a threshold stochastic switching rate below which all pathogen lineages go extinct, and above which survival is a near certainty. We determine how quickly stochastic switching rates can evolve by computing a fitness landscape for the evolutionary dynamics of switching rates, and analyzing its dependence on both the stringency of bottlenecks and the duration of within‐host growth periods. We show that increasing the stringency of bottlenecks or decreasing the period of growth results in faster adaptation of switching rates. Our model provides strong theoretical evidence that bottlenecks play a critical role in accelerating the evolutionary dynamics of pathogens.     

Effects of suspended sediment and desiccation on the benthic tailwater community in the Colorado River, USA

We demonstrated that differences in habitatrequirements by C. glomerata and Oscillatoria havea profound bottom-up influence on the foodweb in thetailwaters below Glen Canyon Dam in the Colorado Riverthrough Grand Canyon National Park, USA. We examinedthe effects of suspended sediment and desiccation onthe colonization sequence of Cladophora glomerata andOscillatoria spp. and the consequent effects onmacroinvertebrates in each algal community in a seriesof reciprocal transplants in the regulated ColoradoRiver, AZ. Our experiments showed that C. glomeratagrows best in continuously submerged, clear-water,stable habitats, whereas Oscillatoria forms densemat-like matrices of trichomes and sand in varialzones and habitats with high suspended sedimentstypical of many southwestern USA streams. Varial zoneconditions have a stronger influence on communitystructure than habitats with high suspended sediments. Recruitment by chironomid larvae was less dependent onC. glomerata and less affected by suspended sedimentand periodic desiccation than Gammarus lacustris. Weestimated the energy from macroinvertebrate biomassassociated with tufts of C. glomerata to be an orderof magnitude higher than that in Oscillatoriamatrices. Therefore, loss of C. glomerata andreplacement of habitat more suitable for Oscillatoriaas a result of regulated flows indirectly reducespotential energy flow in the Colorado Riverfoodweb.     

Vermicompost derived from mushroom residues improves soil C/P cycling,bacterial community, and fungal abundance

The utilization of agricultural waste organic materials through composting technology has gained significant traction in agricultural production as an effective means of crop nutrient management. However, the differences in the impact of organic amendments prepared by traditional composting and vermicomposting on soil properties still deserve further research. Based on field experiments conducted in greenhouse, compared to chemical fertilizer treatments as control, we utilized traditional compost (OF) and vermicompost (VcF) derived from agricultural organic waste edible mushroom bran and cow manure (2:8). Variations in soil physiochemical properties, activities of soil enzymes related C and P cycling, abundances and diversities of bacterial 16S rRNA and fungal ITS gene at total DNA level were analyzed. Both compost treatments enhanced soil organic carbon, soil total phosphorus, and soil available P content significantly and also increased the activities of soil α-glucosidase, β-glucosidase, acid phosphomonoesterase, and alkaline phosphomonoesterase significantly. The above results suggested that soil C and P transformations were stimulated effectively by both organic amendments. OF and VcF increased the fungal ITS absolute abundances significantly while diversity indices of soil bacterial community increased significantly under both treatments. Correlation analysis indicated that bacterial community composition was strongly correlated with several soil property indexes while fungal community composition was only significantly correlated with soil total phosphorous content. In conclusion, similar to traditional compost, vermicompost significantly improved soil nutrient cycling (especially C and P aspects). In terms of soil microbes, bacteria and fungi showed different responding mechanism to vermicompost: bacteria adjust microbial structure, while fungi tend to proliferated. In consideration of the advantages of vermicompost in technology and economic cost, it could be applied in the subsequent agricultural production more frequently.     

Elemental formula annotation of polar and lipophilic metabolites using (13) C, (15) N and (34) S isotope labelling, in combination with high-resolution mass spectrometry

The unbiased and comprehensive analysis of metabolites in any organism presents a major challenge if proper peak annotation and unambiguous assignment of the biological origin of the peaks are required. Here we provide a comprehensive multi-isotope labelling-based strategy using fully labelled (13) C, (15) N and (34) S plant tissues, in combination with a fractionated metabolite extraction protocol. The extraction procedure allows for the simultaneous extraction of polar, semi-polar and hydrophobic metabolites, as well as for the extraction of proteins and starch. After labelling and extraction, the metabolites and lipids were analysed using a high-resolution mass spectrometer providing accurate MS and all-ion fragmentation data, providing an unambiguous readout for every detectable isotope-labelled peak. The isotope labelling assisted peak annotation process employed can be applied in either an automated database-dependent or a database-independent analysis of the plant polar metabolome and lipidome. As a proof of concept, the developed methods and technologies were applied and validated using Arabidopsis thaliana leaf and root extracts. Along with a large repository of assigned elemental compositions, which is provided, we show, using selected examples, the accuracy and reliability of the developed workflow.     

Sub-fossils of cladocerans in the surface sediment of 135 lakes as proxies for community structure of zooplankton,fish abundance and lake temperature

To elucidate the possibilities of using zooplankton remains in the surface sediment to describe present-days community structure and population dynamics of zooplankton, fish abundance and temperature, we compared contemporary data sampled in the pelagial during summer with the sediment record from the upper 1 cm of the sediment in 135 lakes covering a latitude gradient from Greenland in the north to New Zealand in the south. The abundance of three genera Bosmina, Daphnia and Ceriodaphnia of the total pool of ephippia was significantly related to the total abundance of the same taxa in the pelagic zone. However, in most lakes the abundance of Ceriodaphnia was higher in the sediment than in the water, which may be attributed to the overall preference by this genus for the littoral habitat. Using contemporary data from 27 Danish lakes sampled fortnightly during summer for 10 years, we found substantial inter-annual variations in the abundance of Daphnia spp., Ceriodaphnia spp., B. longirostris and B. coregoni. Yet, the sediment record mimicked the medium level well for most of the lakes, which suggests that the sediment record provides an integrated picture of the pelagic cladoceran community, which otherwise can be obtained only by long-term frequent contemporary sampling for several years. The contribution of Daphnia to the sum of Daphnia and Bosmina ephippia was negatively correlated with the abundance of fish expressed as catch per night in multi-mesh sized gill nets (CPUE). Yet, region-specific differences occurred, which partly could be eliminated by including nutrient state expressed as total phosphorus (TP) in a multiple regression. The average ratio of ephippia to the sum of ephippia and carapaces of Bosmina varied 40-fold between the sampling regions and was significantly negatively related to summer mean air temperature, and for Danish lakes also, albeit weakly, to fish CPUE but not to chlorophyll a. Apparently, temperature is the most important factor determining the ratio of parthenogenetic to ephippia producing specimens of Bosmina. We conclude that the sediment record of cladocerans is a useful indicator of community structure of pelagic cladocerans and the abundance of fish and temperature.     

The effect of single tree species on soil microbial activities related to C and N cycling in the Siberian artificial afforestation experiment

The effects of grassland conversion to forest vegetation and of individual tree species on microbial activity in Siberia are largely unstudied. Here, we examined the effects of the six most commonly dominant tree species in Siberian forests (Scots pine, spruce, Arolla pine, larch, aspen and birch) on soil C and N mineralization, N₂O-reduction and N₂O production during denitrification 30 years after planting. We also documented the effect of grassland conversion to different tree species on microbial activities at different soil depths and their relationships to soil chemical properties. The effects of tree species and grassland conversion were more pronounced on N than on C transformations. Tree species and grassland conversion did significantly alter substrate-induced respiration (SIR) and basal respiration, but the differences were not as large as those observed for N transformations. Variances in SIR and basal respiration within species were markedly lower than those in N transformations. Net N mineralization, net nitrification, and denitrification potential were highest under Arolla pine and larch, intermediate under deciduous aspen and birch, and lowest beneath spruce and Scots pine. Tree species caused similar effects on denitrification potential, net N mineralization, and net nitrification, but effects on N₂O reduction rate were idiosyncratic, indicating a decoupling of N₂O production and reduction. We predict that deciduous species should produce more N₂O in the field than conifers, and that Siberian forests will produce more N₂O if global climate change alters tree species composition. Basal respiration and SIR showed inverse responses to tree species: when basal respiration increased in response to a given tree species, SIR declined. SIR may have been controlled by NH₄ ⁺ availability and related therefore to N mineralization, which was negatively affected by grassland conversion. Basal respiration appeared to be less limited by NH₄ ⁺ and controlled mostly by readily available organic C (DOC), which was higher in concentration under forests than in grassland and therefore basal respiration was higher in forested soils. We conclude that in the Siberian artificial afforestation experiment, soil C mineralization was not limited by N.     

The impact of Chironomus plumosus larvae on organic matter decay and nutrient (N, P) exchange in a shallow eutrophic lake sediment following a phytoplankton sedimentation

The importance of Chironomus plumosus larvae onbenthic metabolism and nutrient exchange across thesediment–water interface was evaluated in a shalloweutrophic lake (Lake Arreskov, Denmark) following aphytoplankton sedimentation. Chironomus plumosuslarvae were added to laboratory sediment microcosms,corresponding to a density of 2825 larvae m⁻².Non-inhabited microcosms served as controls. Asedimentation pulse of organic matter was simulated byadding fresh algal material (Chlamydomonasreinhardii) to sediment cores (36 g dryweight m⁻²). The mineralization was followed bymeasuring fluxes of O₂, CO₂, dissolvedinorganic nitrogen and phosphate. A rapid clearance ofalgae from the water column in faunated microcosmssuggested that chironomids may be of major importancein controlling phytoplankton concentrations in shalloweutrophic lakes. Chironomids increased the sedimentO₂ uptake ≈ 3 times more than what wouldbe expected from their own respiration, indicating astimulation of microbial activity and decomposition oforganic matter in the sediment. Addition of algaeenhanced the release of CO₂, NH⁺ ₄ ando-P. The excess inorganic C, N and P released inamended non-inhabited sediment after 36 dayscorresponded to 65, 31 and 58% of the C, N and P inthe added algae. In sediment inhabited by Chironomus plumosus the corresponding numbers were147, 45 and 73%, indicating that mineralization oforganic matter also from the indigenous sediment poolwas stimulated by chironomids. This revised version was published online in July 2006 with corrections to the Cover Date.     

Bacterial,archaeal and eukaryotic diversity in Arctic sediment as revealed by 16S rRNA and 18S rRNA gene clone libraries analysis

We studied the microbial diversity in the sediment from the Kongsfjorden, Svalbard, Arctic, in the summer of 2005 based on the analysis of 16S rRNA and 18S rRNA gene clone libraries. The sequences of the cloned 16S rRNA and 18S rRNA gene inserts were used to determine the species identity or closest relatives by comparison with sequences of known species. Compared to the other samples acquired in Arctic and Antarctic, which are different from that of ours, the microbial diversity in our sediment is much higher. The bacterial sequences were grouped into 11 major lineages of the domain Bacteria: Proteobacteria (include α-, β-, γ-, δ-, and ε-Proteobacteria); Bacteroidetes; Fusobacteria; Firmicutes; Chloroflexi; Chlamydiae; Acidobacteria; Actinobacteria; Planctomycetes; Verrucomicrobiae and Lentisphaerae. Crenarchaeota were dominant in the archaeal clones containing inserts. In addition, six groups from eukaryotes including Cercozoa, Fungi, Telonema, Stramenopiles, Alveolata, and Metazoa were identified. Remarkably, the novel group Lentisphaerae was reported in Arctic sediment at the first time. Our study suggested that Arctic sediment as a unique habitat may contain substantial microbial diversity and novel species will be discovered.