Methane production potential and methanogenic archaeal community structure in tropical irrigated Indian paddy soils

Soil characteristics regulate various belowground microbial processes including methanogenesis and, consequently, affect the structure and function of methanogenic archaeal communities due to change in soil type which in turn influences the CH₄ production potential of soils. Thus, five different soil orders (Alfisol, Entisol, Inceptisol, Podzol and Vertisol) were studied to assess their CH₄ production potential and also the methanogenic archaeal community structure in dryland irrigated Indian paddy soils. Soil incubation experiments revealed CH₄ production to range from 178.4 to 431.2 μg CH₄ g^-1 dws in all soil orders as: Vertisol<Inceptisol<Entisol<Podzol<Alfisol. The numbers of methanogens as quantified using real-time quantitative polymerase chain reaction (qPCR) targeting mcrA genes varied between 0.06 and 72.97 (×10⁶ copies g^-1 dws) and were the highest in Vertisol soil and the least in Alfisol soil. PCR-denaturing gradient gel electrophoresis (DGGE)-based approach targeting 16S rRNA genes revealed diverse methanogenic archaeal communities across all soils. A total of 43 DGGE bands sequenced showed the closely related groups to Methanomicrobiaceae, Methanobacteriaceae, Methanocellales, Methanosarcinaceae, Methanosaetaceae and Crenarchaeota. The composition of methanogenic groups differed among all soils and only the Methanocellales group was common and dominant in all types of soils. The highest diversity of methanogens was found in Inceptisol and Vertisol soils. Methane production potential varied significantly in different soil orders with a positive relationship (p?<?0.05) with methanogens population size, permanganate oxidizable C (POXC) and CO₂ production. The present study suggested that CH₄ production potential of different soils depends on physicochemical properties, methanogenic archaeal community composition and the population size.     

Comparative diversity and abundance of ammonia monooxygenase genes in mulched and vegetated soils

Soil microbial habitats are altered by mulching, a common practice in urban areas during which vegetation is removed and soils covered to suppress weeds and retain moisture. Soil microorganisms drive nitrogen-cycling processes in mulched soils, because living plants no longer take up ammonium-N released during decomposition of residual organic matter. Because ammonia oxidizers carry out the first, rate-limiting step of nitrification, we compared ammonia oxidizers in experimental, unfertilized plots of mulched and vegetated soils. We hypothesized that mulched and vegetated soils would support contrasting communities of bacterial and archaeal ammonia oxidizers, as determined by quantitative PCR and primers specific for genes encoding ammonia monooxygenase subunit A (amoA). Clone libraries of archaeal amoA also were constructed to compare diversity in soil cores, duplicate blocked plots, and treatments (bark-mulched, gravel-mulched, and unmanaged old field vegetation). Gene copies from ammonia-oxidizing bacteria (AOB) ranged from 2.2 × 10⁶ to 2.7 × 10⁷ gene copies per gram dry soil and did not differ across treatments. In contrast, gene copies from ammonia-oxidizing archaea (AOA) ranged from 9.1 × 10⁵ to 1.0 × 10⁸ copies per gram dry soil, with bark-mulched soils having significantly lower abundance. Community structure of AOA in gravel-mulched soils was distinct from the other two treatments. At 97% amino acid similarity, 22 operational taxonomic units, or OTUs, were identified, with only one OTU found in all 18 clone libraries. This ubiquitous OTU-1, which was highly similar to published amoA sequences recovered from soils, comprised 55% of all 482 translated sequences. Greater variability in OTU richness was observed among cores from mulched soils than from vegetated soils. Our observations supported our hypothesis that AOA communities differ in mulched and vegetated soils, with mulched soils providing altered and variable microniches for these N cycling microorganisms.     

Dung application increases CH<Subscript>4</Subscript> production potential and alters the composition and abundance of methanogen community in restored peatland soils from Europe

Peatland restoration via rewetting aims to recover biological communities and biogeochemical processes typical to pristine peatlands. While rewetting promotes recovery of C accumulation favorable for climate mitigation, it also promotes methane (CH₄) emissions. The potential for exceptionally high emissions after rewetting has been measured for Central European peatland sites previously grazed by cattle. We addressed the hypothesis that these exceptionally high CH₄ emissions result from the previous land use. We analyzed the effects of cattle dung application to peat soils in a short- (2 weeks), a medium- (1 year) and a long-term (grazing) approach. We measured the CH₄ production potentials, determined the numbers of methanogens by mcrA qPCR, and analyzed the methanogen community by mcrA T-RFLP-cloning-sequencing. Dung application significantly increased the CH₄ production potential in the short- and the medium-term approach and non-significantly at the cattle-grazed site. The number of methanogens correlated with the CH₄ production in the short- and the long-term approach. At all three time horizons, we found a shift in methanogen community due to dung application and a transfer of rumen methanogen sequences (Methanobrevibacter spp.) to the peatland soil that seemed related to increased CH₄ production potential. Our findings indicate that cattle grazing of drained peatlands changes their methanogenic microbial community, may introduce rumen-associated methanogens and leads to increased CH₄ production. Consequently, rewetting of previously cattle-grazed peatlands has the potential to lead to increased CH₄ emissions. Careful consideration of land use history is crucial for successful climate mitigation with peatland rewetting.     

Long-term submergence of non-methanogenic oxic upland field soils helps to develop the methanogenic archaeal community as revealed by pot and field experiments

The community structure of methanogenic archaea is relatively stable,i.e.,it is sustained at a high abundance with minimal changes in composition,in paddy field soils irrespective of submergence and drainage.In contrast,the abundance in non-methanogenic oxic soils is much lower than that in paddy field soils.This study aimed to describe methanogenic archaeal community development following the long-term submergence of non-methanogenic oxic upland field soils in pot and field experiments.In the pot experiment,a soil sample obtained from an upland field was incubated under submerged conditions for 275 d.Soil samples periodically collected were subjected to culture-dependent most probable number(MPN)enumeration,polymerase chain reaction-denaturing gradient gel electrophoresis(PCR-DGGE)analysis of archaeal 16 S r RNA gene,and quantitative PCR analysis of the methyl-coenzyme M reductase alpha subunit gene(mcr A)of methanogenic archaea.The abundance of methanogenic archaea increased from 102 to 103 cells g-1 dry soil and 104 to 107 copies of mcr A gene g-1 dry soil after submergence.Although no methanogenic archaeon was detected prior to incubation by the DGGE analysis,members from Methanocellales,Methanosarcinaceae,and Methanosaetaceae proliferated in the soils,and the community structure was relatively stable once established.In the field experiment,the number of viable methanogenic archaea in a rice paddy field converted from meadow(reclaimed paddy field)was monitored by MPN enumeration over five annual cycles of field operations.Viability was also determined simultaneously in a paddy field where the plow layer soil from a farmer’s paddy field was dressed onto the meadow(dressed paddy field)and an upland crop field converted from the meadow(reclaimed upland field).The number of viable methanogenic archaea in the reclaimed paddy field was below the detection limit before the first cultivation of rice and in the reclaimed upland field.Then,the number gradually increased over five years and finally reached 103–104 cells g-1 dry soil,which was comparable to that in the dressed paddy field.These findings showed that the low abundance of autochthonous methanogenic archaea in the non-methanogenic oxic upland field soils steadily proliferated,and the community structure was developed following repeated and long-term submergence.These results suggest that habitats suitable for methanogenic archaea were established in soil following repeated and long-term submergence.     

Comparative analysis of soil microbial communities and their responses to the short-term drought in bog,fen, and riparian wetlands

The frequency of drought is anticipated to increase in wetland ecosystems as global warming intensifies. However, information on microbial communities involved in greenhouse gas emissions and their responses to drought remains sparse. We compared the gene abundance of eubacterial 16S rRNA, nitrite reductase (nirS) and methyl coenzyme M reductase (mcrA), and the diversity and composition of eubacteria, methanogens and denitrifiers among bog, fen and riparian wetlands. The gene abundance, diversity and composition significantly differed among wetlands (p < 0.01) with the exception of the diversity of methanogens. The gene abundance was ranked in the order of the bog = fen > riparian wetland, whereas the diversity was in the riparian wetland ≥ fen > bog. In addition, we conducted a short-term drought experiment and compared microbial communities between control (water-logged) and drought (?15 cm) treatments. Drought led to significant decline in the gene abundance in the bog (16S rRNA, nirS, mcrA) (p < 0.01) and fen (16S rRNA, nirS) (p < 0.05), but not in the riparian wetland. There were no differences in the diversity and composition of denitrifiers and methanogens at all sites following drought. Our results imply that denitrifiers and methanogens inhabiting bogs and fens would suffer from short-term droughts, but remain unchanged in riparian wetlands.     

Changes in methane emission and methanogenic and methanotrophic communities in restored wetland with introduction of <Emphasis Type="Italic">Alnus trabeculosa</Emphasis>

Purpose

The dynamics and uncertainties in wetland methane budgets affected by the introduction of Alnus trabeculosa H. necessitate research on production of methane by methanogenic archaea and consumption by methane-oxidizing microorganisms simultaneously.

Materials and methods

This study investigated methane emission in situ by the closed chamber method, and methanogenic and methanotrophic communities using denatured gradient gel electrophoresis (DGGE) and quantitative PCR based on mcrA (methyl coenzyme M reductase), pmoA (particulate methane monooxygenase) genes in the rhizosphere and non-rhizosphere soils in the indigenous pure Phragmites australis T., and A. trabeculosa–P. australis mixed communities in Chongxi wetland.

Results and discussion

Methane flux rate from the pure P. australis community was 2.4 times larger than that of A. trabeculosa–P. australis mixed community in the rhizosphere and 1.7 times larger in the non-rhizosphere, respectively. The abundance of methanogens was lower in the mixed community soils (3.56?×?10³–6.90?×?10³ copies g^?1 dry soil) compared with the P. australis community (1.47?×?10⁴–1.89?×?10⁴ copies g^?1 dry soil), whereas the methanotrophs showed an opposite trend (2.08?×?10⁶–1.39?×?10⁶ copies g^?1 dry soil for P. australis and 6.20?×?10⁶–1.99?×?10⁶ copies g^?1 dry soil for mixed community soil). A liner relationship between methane emission rates against pmoA/mcrA ratios (R ²?=?0.5818, p?<?0.05, n?=?15) was observed. The community structures of the methane-cycling microorganism based on mcrA and pmoA suggested that acetoclastic methanogens belonging to Methanosarcinaceae and a particular type II methanotroph, Methylocystis, were dominant in these two plant communities.

Conclusions

The introduction of A. trabeculosa would promote the proliferation of methanotrophs, especially the dominant Methylocystis, but not methanogens, ultimately diminishing methane emission in the wetland.

     

Bacterial community structure of nirK-bearing denitrifiers and the development of properties of soils in created mitigation wetlands

We investigated the abundance and genetic heterogeneity of bacterial nitrite reductase genes (nir) and soil structural properties in created and natural freshwater wetlands in the Virginia piedmont. Soil attributes included soil organic matter (SOM), total organic carbon (TOC), total nitrogen (TN), pH, gravimetric soil moisture (GSM), and bulk density (D_b). A subset of soil attributes were analyzed across the sites, using euclidean cluster analysis, resulting in three soil condition (SC) groups of increasing wetland soil development (i.e., SC1 < SC2 < SC3; less to more developed or matured) as measured by accumulation of TOC, TN, the increase of GSM, and the decrease of D_b. There were no difference found in the bacterial community diversity between the groups (p = 0.4). NirK gene copies detected ranged between 3.6 × 10⁴ and 3.4 × 10⁷ copies g⁻¹ soil and were significantly higher in the most developed soil group, SC3, than in the least developed soil group, SC1. However, the gene copies were lowest in SC2 that had a significantly higher soil pH (~6.6) than the other two SC groups (~5.3). The same pattern was found in denitrifying enzyme activity (DEA) on a companion study where DEA was found negatively correlated with soil pH. Gene fragments were amplified and products were screened by terminal restriction fragment length polymorphism (T-RFLP) analysis. Among 146 different T-RFs identified, fourteen were dominant and together made up more than 65% of all detected fragments. While SC groups did not relate to whole nirK communities, most soil properties that identified SC groups did significantly correlate to dominant members of the community.     

Depth-specific distribution and importance of nitrite-dependent anaerobic ammonium and methane-oxidising bacteria in an urban wetland

Anaerobic ammonium oxidation (anammox) and nitrite-dependent anaerobic methane oxidation (n-damo) are two recently discovered processes in the nitrogen cycle that are catalysed by anammox bacteria and n-damo bacteria, respectively. Here, the depth-specific distribution and importance of anammox bacteria and n-damo bacteria were studied in an urban wetland, Xixi Wetland, Zhejiang Province (China). Anammox bacteria related to Candidatus Brocadia, Candidatus Kuenenia and Candidatus Anammoxoglobus, and n-damo bacteria related to “Candidatus Methylomirabilis oxyfera” were present in the collected soil samples. The abundance of anammox bacteria (2.6–8.6 × 10⁶ copies g⁻¹ dry soil) in the shallow soils (0–10 cm and 20–30 cm) was higher than that (2.5–9.8 × 10⁵ copies g⁻¹ dry soil) in the deep soils, whereas the abundance of n-damo bacteria (0.6–1.3 × 10⁷ copies g⁻¹ dry soil) in the deep soils (50–60 cm and 90–100 cm) was higher than that (3.4–4.5 × 10⁶ copies g⁻¹ dry soil) in the shallow soils. Anammox activity was detected at all depths, and higher potential rates (12.1–21.4 nmol N₂ g⁻¹ dry soil d⁻¹) were observed at depths of 0–10 cm and 20–30 cm compared with the rates (3.5–8.7 nmol N₂ g⁻¹ dry soil d⁻¹) measured at depths of 50–60 and 90–100 cm. In contrast, n-damo was mainly occurred at depths of 50–60 cm and 90–100 cm with potential rates of 0.7–5.0 nmol CO₂ g⁻¹ dry soil d⁻¹. This study suggested the niche segregation of the anammox bacteria and n-damo bacteria in wetland soils, with anammox bacteria being active primarily in deep soils and n-damo bacteria being active primarily in shallow soils.     

The impact of sludge amendment on methanogen community structure in an upland soil

In the EU, municipal sewage sludge application to agricultural land has increased dramatically since the ban on dumping at sea came into effect in 1998. There are many concerns related to potential contamination and reduction in plant productivity. In this study, the aim was to assess the impact of repeated long-term soil amendment with anaerobically digested sewage sludge on methanogen diversity in an upland soil ecosystem. Sludge-treated and untreated upland soil samples as well as samples of the sludge used, were analysed for the diversity of methanogens using TGGE, PCR-RFLP and DNA sequence analysis of approximately 490 bp of the mcrA operon. PCR analysis using mcrA specific oligonucleotide primers confirmed the presence of methanogen DNA in treated and untreated soil samples and in sewage sludge. TGGE was used to describe the diversity of methanogen mcrA sequences and the differences in community structure between samples. Ninety-six mcrA gene PCR products were screened using RFLP analysis representing methanogen DNA amplified from anaerobically digested sewage sludge, control soils and sludge treated soils. Fourteen RFP's were detected in all treatments, five of which were common to all three treatments. Thirty-eight cloned amplimers were selected for sequencing and phylogenetic analysis. These included representatives of each RFP. From control soils, sludge and sludged soil samples 15, 16 and 7 clones were sequenced, respectively. Phylogenetic analysis suggested that they represented hitherto uncharacterised mcr genes; 35 of the clones fell into 7 clusters supported by moderate to high bootstrap values. The diversity of methanogens in an upland soil (treated and untreated) and sludge was evaluated and marked differences in the diversity of the methanogen communities was observed between the treatments. Our results indicate that sludge application may reduce soil methanogen community diversity.     

Temporal variation in methanogenic community structure and methane production potential of tropical rice ecosystem

Temporal variations in diversity of methanogenic community and CH₄ production potential were analyzed in an Indian tropical rice ecosystem. Laboratory incubations showed that methane production varied from 20.86 to 134.11 μg CH₄ g⁻¹ d.w.s. during the two consecutive years, 2009 and 2010. CH₄ production potential was high at the flowering stage of the rice crop followed by ripening, tillering, post-harvest and pre-plantation stage. Phylogenetic analysis of 16S rRNA genes of methanogenic community indicated that flowering and ripening stages comprised of Methanomicrobiaceae, Methanosarcinaceae, Methanosaetaceae and RC I methanogenic groups, while only the members of Methanomicrobiaceae and RC I were present in the remaining stages. Further, the dominance of RC I was observed in all stages. This study demonstrates that flowering and ripening stages of rice crop offer relatively favorable ecological niche for methanogenic community. The overall analyses suggest that the temporal change in diversity of methanogens regulates CH₄ production potential in rice field soils.     

Soil–methane sink increases with soil age in forefields of Alpine glaciers

In upland soils aerobic methane-oxidizing bacteria (MOB) catalyze methane (CH₄) oxidation, and thus regulate the sole terrestrial sink for atmospheric CH₄. While confirmed in mature upland soils, little is known about this important function in young mountainous soils in glacier forefields, which are progressively formed as a result of glacier recession. We assessed four attributes of the soil CH₄ sink, i.e., soil-atmosphere CH₄ flux (J_atm), CH₄ oxidation activity (k), MOB abundance and variation in community composition along the 6–120-yr soil chronosequence in two Alpine glacier forefields on siliceous and calcareous bedrock. At most sampling locations soil CH₄ profiles showed stable uptake of atmospheric CH₄, with J_atm in the range of −0.082 to −2.2 mg CH₄ m⁻² d⁻¹. Multiple-linear-regression analyses indicated that J_atm significantly increased with soil age, whereas k did not. Instead, water content and CH₄ profiles in the youngest soils often indicated dry, inactive top layers with k < 0.1 h⁻¹, and active deeper layers (0.2 h⁻¹ ≤ k ≤ 11 h⁻¹) with more favorable water content. With increasing soil age the zone of highest CH₄ oxidation activity gradually moved upwards and eventually focused in the 10–40-cm layer (0.2 h⁻¹ ≤ k ≤ 16 h⁻¹). Copy numbers of pmoA genes significantly increased with soil age at both sites, ranging from 2.4 × 10³ to 5.5 × 10⁵ copies (g soil w.w.)⁻¹, but also correlated with mineral nitrogen content. Terminal restriction-fragment-length-polymorphism and cluster analyses showed differences in MOB community composition apparently related to bedrock type rather than soil age. Yet, regardless of bedrock type, the soil CH₄ sink established within a few years of soil development, and J_atm increased to values comparable to mature soils within decades. Thus, young mountainous soils have the potential to consume substantial amounts of atmospheric CH₄, and should be incorporated into future estimates of global soil CH₄ uptake.     

Parent material and vegetation influence bacterial community structure and nitrogen functional genes along deep tropical soil profiles at the Luquillo Critical Zone Observatory

Microbial communities mediate every step of the soil nitrogen cycle, yet the structure and associated nitrogen cycle functions of soil microbial communities remain poorly studied in tropical forests. Moreover, tropical forest soils are often many meters deep, but most studies of microbial nitrogen cycling have focused exclusively on surface soils. The objective of our study was to evaluate changes in bacterial community structure and nitrogen functional genes with depth in soils developed on two contrasting geological parent materials and two forest types that occur at different elevations at the Luquillo Critical Zone Observatory in northeast Puerto Rico. We excavated three soil pits to 140 cm at four different sites representing the four soil × forest combinations (n = 12), and collected samples at ten-centimeter increments from the surface to 140 cm. We used bacterial 16S rRNA gene-DGGE (denaturant gradient gel electrophoresis) to fingerprint microbial community structures, and quantitative PCR to measure the abundance of five functional genes involved in various soil nitrogen transformations: nifH (nitrogen fixation), chiA (organic nitrogen decomposition), amoA (ammonia oxidation), nirS (nitrite reduction) and nosZ (nitrous oxide reduction). Multivariate analyses of DGGE fingerprinting patterns revealed differences in bacterial community structure across the four soil × forest types that were strongly correlated with soil pH (r = 0.69, P < 0.01) and nutrient stoichiometry (r² ≥ 0.36, P < 0.05). Across all soil and forest types, nitrogen functional genes declined significantly with soil depth (P < 0.001). Denitrification genes (nirS and nosZ) accounted for the largest proportion of measured nitrogen functional genes. Measured nitrogen functional genes were positively correlated with soil carbon, nitrogen and phosphorus concentrations (P < 0.001) and all genes except amoA were significantly more abundant in the Inceptisol soil type compared with the Oxisol soil type (P < 0.03). Greater abundances and a stronger vertical zonation of nitrogen functional genes in Inceptisols suggest more dynamic nitrogen transformation processes in this soil type. As the first study to examine bacterial nitrogen functional gene abundances below the surface 20 cm in tropical forest soils, our work provides insight into how pedogenically-driven vertical gradients control the nitrogen-cycling capacity of soil microbial communities. While previous studies have shown evidence for redox-driven hotspots in tropical nitrogen cycling on a watershed scale, our study corroborates this finding on a molecular scale.     

Survival and persistence of genetically modified Sinorhizobium meliloti in soil

Studies were conducted to evaluate the survival and persistence of Sinorhizobium meliloti 104A14 and two acid phosphatase-negative mutants in Kirkland (fine, mixed, thermic Udertic Paleustolls) silt loam soils with various fertility levels, and to assess the impact of inoculation on nodule occupancy and soil microbial community structure in the inoculated alfalfa (Medicago sativa L.) rhizosphere. Recovery of the inoculated strains was 100% (in the order of 10⁸ cells g⁻¹ soil) immediately following inoculation to soils, but decreased from 10⁸ cells g⁻¹ soil to undetectable levels in a nutrient-poor soil within 32 days. In a nutrient-rich soil, approximately 2–3% (4.7–7.43×10⁶ cells g⁻¹ soil) of the mutants and 23% (5.84×10⁷ cells g⁻¹ soil) of the wild-type inocula persisted for more than 64 days. Survivability and persistence of the wild-type S. meliloti were significantly greater than that of the genetically modified acid phosphatase negative mutants in all the soils tested. The persistence and nodule occupancy of the introduced S. meliloti in sterile and non-sterile soils were also tested for two repeated alfalfa growth periods in the same plant growth units, with a 1 month interval in between and no additional inoculation for the second period. Nodule occupancy of the introduced S. meliloti in non-sterile soils ranged from 30 to 60% for the first period and 85 to 100% for the second period. Our results suggest that survival and persistence of S. meliloti was enhanced by alfalfa cultivation and increased soil fertility, but impaired by mutation of acid phosphatase genes regardless of phosphorus nutritional levels. Moreover, inoculation with genetically modified S. meliloti strain 104A14 promoted indigenous bacterial growth in soil (increased bacterial population from 1.4×10⁶ to 4.3×10⁶ cells g⁻¹ soil), but not the growth of fungi and yeast. However, inoculation of the wild-type S. meliloti or genetically modified mutants did not result in significant changes in microbial community structure as indicated by EP indices and ratios of r/K strategists.     

The community structure of soil Sarcodina in Baiyun Mountain,Guangzhou, China

Community structures of soil Sarcodina in 7 different habitats within Baiyun Mountain in Guangzhou, China were investigated with qualitative and quantitative analyses. The abundance, dominance, species diversity and community similarity index of soil sarcodina with different physicochemical parameters were comparatively analyzed. A total 67 species of sarcodina belonging to 4 Super-groups, 6 First ranks and 14 Second ranks were identified. The first dominant group was Tubulinea, followed by Flabellinea, with dominance of 59.7% and 13.4%, respectively. The highest abundance of sarcodina appeared in autumn of Site 5, reaching 1.20 × 10⁵ ind g^?1; the lowest in spring of Site 2 with 1.73 × 10³ ind g^?1. Margalef's biodiversity index ranged from 1.26 (winter of Site 6) to 2.51 (summer of Site 1). Statistical analyses showed the sarcodina abundance was positively correlated with organic matter, soil moisture, soil pH, ammonia nitrogen and total nitrogen, but the correlation coefficient of total potassium was negative. Total phosphorus, nitrate nitrogen and sulphate showed no significant effect on sarcodina abundance in the present study.     

The fate and toxicity of the flavonoids naringenin and formononetin in soil

The flavonoid class of plant secondary metabolites play a multifunctional role in below-ground plant–microbe interactions with their best known function as signals in the nitrogen fixing legume–rhizobia symbiosis. Flavonoids enter rhizosphere soil as a result of root exudation and senescence but little is known about their subsequent fate or impacts on microbial activity. Therefore, the present study examined the sorptive behaviour, biodegradation and impact on dehydrogenase activity (as determined by iodonitrotetrazolium chloride reduction) of the flavonoids naringenin and formononetin in soil. Organic carbon normalised partition coefficients, log K_oc, of 3.12 (formononetin) and 3.19 (naringenin) were estimated from sorption isotherms and, after comparison with literature log K_oc values for compounds whose soil behaviour is better characterised, the test flavonoids were deemed to be moderately sorbed. Naringenin (spiked at 50 μg g^?1) was biodegraded without a detectable lag phase with concentrations reduced to 0.13±0.01 μg g^?1 at the end of the 96 h time course. Biodegradation of formononetin proceeded after a lag phase of ～24 h with concentrations reduced to 4.5±1% of the sterile control after 72 h. Most probable number (MPN) analysis revealed that prior to the addition of flavonoids, the soil contained 5.4×10⁶ MPN g^?1 (naringenin) and 7.9×10⁵ MPN g^?1 (formononetin) catabolic microbes. Formononetin concentration had no significant (p>0.05) effect on soil dehydrogenase activity, whereas naringenin concentration had an overall but non-systematic impact (p=0.045). These results are discussed with reference to likely total and bioavailable concentrations of flavonoids experienced by microbes in the rhizosphere.     

Control of cotton <Emphasis Type="Italic">Verticillium</Emphasis> wilt and fungal diversity of rhizosphere soils by bio-organic fertilizer

Cotton Verticillium wilt is a destructive soil-borne disease affecting cotton production. In this study, application of bio-organic fertilizer (BIO) at the beginning of nursery growth and/or at the beginning of transplanting was evaluated for its ability to control Verticillium dahliae Kleb. The most efficient control of cotton Verticillium wilt was achieved when the nursery application of BIO was combined with a second application in transplanted soil, resulting in a wilt disease incidence of only 4.4%, compared with 90.0% in the control. Denaturing gradient gel electrophoresis patterns showed that the consecutive applications of BIO at nursery and transplanting stage resulted in the presence of a unique group of fungi not found in any other treatments. Humicola sp., Metarhizium anisopliae, and Chaetomium sp., which were considered to be beneficial fungi, were found in the BIO treatment, whereas some harmful fungi, such as Alternaria alternate, Coniochaeta velutina, and Chaetothyriales sp. were detected in the control. After the consecutive applications of BIO at nursery and transplanting stage, the V. dahliae population in the rhizosphere soil in the budding period, flowering and boll-forming stage, boll-opening stage, and at harvest time were 8.5 × 10², 3.1 × 10², 4.6 × 10², and 1.7 × 10² colony-forming units per gram of soil (cfu g⁻¹), respectively, which were significantly lower than in the control (6.1 × 10³, 3.4 × 10³, 5.2 × 10³, and 7.0 × 10³ cfu g⁻¹, respectively). These results indicate that the suggested application mode of BIO could effectively control cotton Verticillium wilt by significantly changing the fungal community structure and reducing the V. dahliae population in the rhizosphere soil.     

Structural and functional diversity of bacterial community in soil treated with the herbicide napropamide estimated by the DGGE,CLPP and r/K-strategy approaches

Napropamide is one of the most commonly used herbicide in agricultural practice and can exhibit toxic effect to soil microorganisms. Therefore, the main objective of this study was to examine the genetic and functional diversity of microbial communities in soil treated with napropamide at field rate (FR, 2.25 mg kg⁻¹ of soil) and 10 times the FR (10 × FR, 22.5 mg kg⁻¹ of soil) by the denaturing gradient gel electrophoresis (DGGE) and the community level physiological profile (CLPP) methods. In addition, the r/K-strategy approach was used to evaluate the effect of this herbicide on the community structure of the culturable soil bacteria. DGGE patterns revealed that napropamide affected the structure of microbial community; however, the richness (S) and genetic diversity (H) values indicated that the FR dosage of napropamide experienced non-significant changes. In turn, the 10 × FR dosage of herbicide caused significant changes in the S and H values of dominant soil bacteria. DGGE profiles suggest an evolution of bacteria capable of degrading napropamide among indigenous microflora. Analysis of the CLPPs indicated that the catabolic activity of microbial community expressed as AWCD (average well-color development) was temporary positively affected after napropamide application and resulted in an increase of the substrate richness (S_R) as well as functional biodiversity (H) values. Analysis of the bacterial growth strategy revealed that napropamide affected the r- or K-type bacterial classes (ecotypes). In treated-soil samples K-strategists dominated the population, as indicated by the decreased ecophysiological (EP) index. Napropamide significantly affected the physiological state of culturable bacteria and caused a reduction in the rate of colony formation as well as a prolonged time of growth rate. Obtained results indicate that application of napropamide may poses a potential risk for soil functioning.     

Shifts of archaeal community structure in soil along an elevation gradient in a reservoir water level fluctuation zone

Purpose

Although archaea play an important role in nutrients cycling, the archaeal community in a reservoir water-level fluctuation zone (WLFZ) remains unclear. An elucidation of archaeal community responding to the environmental variables is essential to understand the nutrients dynamics in WLFZ. This study focused on the response of the archaeal community structure and abundance to the periodic water flooding along an elevation gradient in the WLFZ of the Three Gorges Reservoir.

Materials and methods

Along the elevation gradient (152–175 m) of the study area, soil samples in the beginning and late stages of water flooding were collected to investigate the influence of water flooding on the archaeal community in soil, using quantitative PCR and Illumina high-throughput sequencing approaches.

Results and discussion

An increase of archaeal abundance from 3.8?×?10⁸ to 3.8?×?10⁹ copies (g d.w.s)^?1 on average was observed after water flooding. The archaeal abundance was positively correlated with the contents of ammonium, organic matter, and moisture in soil and with the accumulated flooding time. Higher diversity was observed in dry samples (non-flooded soil samples) rather than wet samples (flooded soil samples). The Thaumarchaeota were predominant in most of the dry samples. Interestingly, high proportions of Candidatus Nitrososphaera were observed in the transition zone, while euryarchaeotal methanogens dominated the wet samples. The proportion of methanogens decreased dramatically in the dry samples at higher elevations, which was associated with the decrease of the moisture content and the probably increase of available oxygen in soil.

Conclusions

Archaeal abundance, diversity, and community composition shifted along an elevation gradient and were influenced by water flooding. The increased archaea abundance after water flooding and elevation related community composition and diversity indicated that water flooding was a key dynamic environmental variable in the WLFZ.