Shift in abundance and structure of soil ammonia-oxidizing bacteria and archaea communities associated with four typical forest vegetations in subtropical region

Purpose

Nitrogen (N) is one of the most important elements that can limit plant growth in forest ecosystems. Ammonia-oxidizing bacteria (AOB) and archaea (AOA) are considered as the key drivers of global N biogeochemical cycling. Soil ammonia-oxidizing microbial communities associated with subtropical vegetation remain poorly characterized. The aim of this study was to determine how AOA and AOB abundance and community structure shift in response to four typical forest vegetations in subtropical region.

Materials and methods

Broad-leaved forest (BF), Chinese fir forest (CF), Pinus massoniana forest (PF), and moso bamboo forest (MB) were widely distributed in the subtropical area of southern China and represented typical vegetation types. Four types of forest stands of more than 30 years grew adjacent to each other on the same soil type, slope, and elevation, were chosen for this experiment. The abundance and community structure of AOA and AOB were characterized by using real-time PCR and denaturing gradient gel electrophoresis (DGGE). The impact of soil properties on communities of AOA and AOB was tested by canonical correspondence analysis (CCA).

Results and discussion

The results indicated that AOB dominated in numbers over AOA in both BF and MB soils, while the AOA/AOB ratio shifted with different forest stands. The highest archaeal and bacterial amoA gene copy numbers were detected in CF and BF soils, respectively. The AOA abundance showed a negative correlation with soil pH and organic C but a positive correlation with NO₃ ^??N concentration. The structures of AOA communities changed with vegetation types, but vegetation types alone would not suffice for shaping AOB community structure among four forest soils. CCA results revealed that NO₃ ^??N concentration and soil pH were the most important environmental gradients on the distribution of AOA community except vegetation type, while NO₃ ^??N concentration, soil pH, and organic C significantly affected the distribution of the AOB communities.

Conclusions

These results revealed the differences in the abundance and structure of AOA and AOB community associated with different tree species, and AOA was more sensitive to vegetation and soil chemical properties than AOB. N bioavailability could be directly linked to AOA and AOB community, and these results are useful for management activities, including forest tree species selection in areas managed to minimize N export to aquatic systems.     

Preliminary study on the distribution of ammonia oxidizers and their contribution to potential ammonia oxidation in the plant-bed/ditch system of a constructed wetland

Purpose

Ammonia oxidation—as the rate-limiting step of nitrification—has been found to be performed by both ammonia-oxidizing archaea (AOA) and bacteria (AOB). However, how ammonium content and oxidation–reduction status regulate the distribution of ammonia oxidizers in constructed wetlands and their contribution to potential ammonia oxidation rate are still in dispute. This study aimed to explore the effects of ammonium content and oxidation–reduction status on the abundances of AOA/AOB and examine the contributions of AOA and AOB populations to ammonia oxidation rates in the plant-bed/ditch system of a constructed wetland.

Materials and methods

Sampling was carried out in the plant-bed/ditch system of the Shijiuyang Constructed Wetland, China. Three plant-bed soil cores were collected using a soil auger and sampled at depths of 0, 20, and 50 cm in 5-cm increments. Five ditch surface sediments (0–5 cm) were collected along the water flow direction. The abundances of AOA and AOB were investigated by quantitative polymerase chain reaction based on amoA genes. The potential ammonia oxidation rate was determined using the chlorate inhibition method.

Results and discussion

The results showed that AOA outnumbered AOB in the plant-bed surface soil which had lower ammonium content (4.67–7.63 mg kg^?1), but that AOB outnumbered AOA in the ditch surface sediment which had higher ammonium content (14.0–22.9 mg kg^?1). Ammonium content was found to be the crucial factor influencing the relative abundances of AOA and AOB in the surface samples of the plant-bed/ditch system. In the deep layers of the plant bed, AOA abundance outnumbered AOB, though much lower oxidation–reduction potential occurred along the water flow direction. Thus, the oxidation–reduction potential may be another factor influencing the distributions of AOA and AOB in the deep layers of the plant bed without significant difference in ammonium content (p?<?0.05). Moreover, the potential ammonia oxidation rate was significantly dominated by AOB rather than AOA in the plant-bed/ditch system.

Conclusions

The high ammonium content in the ditch sediment likely favored AOB. AOA seemed to persist more readily even under low oxidation–reduction potential in the deep layers of the plant bed. Ammonium content and the oxidation–reduction potential were important parameters influencing the distribution of AOA and AOB in the plant-bed/ditch system of Shijiuyang Constructed Wetland. AOB contributed more to ammonia oxidation than AOA, both in the plant-bed soils (r?=?0.592, p?=?0.0096) and in the ditch sediments (r?=?0.873, p?=?0.0002).     

Contrasting response of nitrification capacity in three agricultural soils to N addition during short-term incubation

Purpose

Human disturbance is a major culprit driving imbalances in the biological transformation of nitrogen from the nonreactive to the reactive pool and is therefore one of the greatest concerns for nitrogen (N) cycling. The objective of this study was to compare potential nitrification rates and the abundance of ammonia oxidizers responsible for nitrification, with the amendment of external N in different agricultural soils.

Materials and methods

Three typical Chinese agricultural soils, QiYang (QY) acid soil, ShenYang (SY) neutral soil, and FengQiu (FQ) alkaline soil, were amended with 0, 20, 150, and 300 μg NH₄ ⁺-N g^?1 soil and incubated for 40 days. The abundance of ammonia oxidizing bacteria (AOB) and archaea (AOA) at the end of incubation in the soil microcosms was determined using the real-time PCR.

Results and discussion

There was a significant decrease in ammonium concentration in the QY soil from the highest to the lowest N-loading treatments, while no significant difference in ammonium concentrations was detected among the different N-loading treatments for the SY and FQ soils. A significantly higher potential nitrification rate (PNR) was observed in the FQ soil while lowest PNR was found in the QY soil. Quantitative PCR analysis of AOB amoA genes demonstrated that AOB abundance was significantly higher in the high N-loading treatments than in the control for the QY soil only, while no significant difference among treatments in the SY and FQ soils. A significant positive correlation between PNR and AOB amoA abundance, however, was found for the SY and FQ soils, but not for the QY soil. Little difference in AOA amoA abundance between different N-loading treatments was observed for all the soils.

Conclusions

This study suggested that ammonia oxidation capacity in the FQ and SY soils was higher than those in the QY soil with the addition of ammonium fertilizer for a short-term. These findings indicated that understanding the differential responses of biological nitrification to varying input levels of ammonium fertilizer is important for maximizing N use efficiency and thereby improving agricultural fertilization management.     

pH-dependent distribution of soil ammonia oxidizers across a large geographical scale as revealed by high-throughput pyrosequencing

Purpose

Ammonia-oxidizing archaea (AOA) and bacteria (AOB) are ubiquitous and important for nitrogen transformations in terrestrial ecosystems. However, the distribution patterns of these microorganisms as affected by the terrestrial environments across a large geographical scale are not well understood. This study was designed to gain insights into the ecological characteristics of AOA and AOB in 65 soils, collected from a wide range of soil and ecosystem types.

Materials and methods

Barcoded pyrosequencing in combination with quantitative PCR was employed to characterize the relative abundance, diversity, and community composition of archaeal 16S rRNA gene, and AOA and AOB amoA genes in 65 soil samples.

Results and discussion

The operational taxonomic unit richness and Shannon diversity of Thaumarchaeota, AOA, and AOB were highly variable among different soils, but their variations were best explained by soil pH. Soil pH was strongly correlated with the overall community composition of ammonia oxidizers, as measured by the pairwise Bray–Curtis dissimilarity across all sites. These findings were further corroborated by the evident pH-dependent distribution patterns of four thaumarchaeal groups (I.1a-associated, I.1b, I.1c, and I.1c-associated) and four AOB clusters (2, 3a.1, 10, and 12). The ratios of AOA to AOB amoA gene copy numbers significantly decreased with increasing pH, suggesting a competitive advantage of AOA over AOB in acidic soils.

Conclusions

These results suggest that the distribution of ammonia oxidizers across large-scale biogeographical settings can be largely predicted along the soil pH gradient, thus providing important indications for the ecological characteristics of AOA and AOB in different soils.     

Prevalence of ammonia-oxidizing bacteria over ammonia-oxidizing archaea in sediments as related to nutrient loading in Chinese aquaculture ponds

Purpose

Nitrogen (N) application in excess of assimilatory capacity for aquaculture ponds can lead to water-quality deterioration through ammonia accumulation with toxicity to fish. Ammonia-oxidizing archaea (AOA) and bacteria (AOB) potentially process extra ammonium, so their abundance and diversity are of great ecological significance. This study aimed to reveal variations in communities of AOA and AOB as affected by aquaculture activities.

Materials and methods

From June to September 2012, water and sediments were sampled monthly in three ponds feeding Mandarin fish in a suburb of Wuhan City, China. Molecular methods based on ammonia monooxygenase (amoA) gene were used to determine abundance and diversity of AOA and AOB in the sediments.

Results and discussion

The pond with the highest fish stock had the highest nutrient loadings in terms of different forms of N and carbon (C) in both sediment and water. The abundance and diversity of AOB were significantly higher than those of AOA in the sediment. The AOB abundance showed a significantly positive relationship to concentration of soluble reactive phosphorus (SRP) in interstitial water, and both abundance and diversity of AOA were significantly negative to concentration of ammonium in interstitial water. Furthermore, AOA species affiliated to Nitrososphaera-like and Nitrosophaera Cluster was distinguishable from those observed in other aquaculture environments.

Conclusions

Nutrients in sediment were enriched by intensive aquaculture activity, among which organic N and C, together with ammonium and SRP, shaped the communities of ammonia oxidizers, with AOB dominating over AOA in terms of abundance and diversity.

     

Abundance and community structure of ammonia oxidizing bacteria and archaea in a Sweden boreal forest soil under 19-year fertilization and 12-year warming

Purpose

Boreal forests are considered to be more sensitive to global climate change compared with other terrestrial ecosystems, but the long-term impact of climate change and forest management on soil microbial functional diversity is not well understood. Ammonia-oxidizing bacteria (AOB) and archaea (AOA) are the most important players in nitrogen (N) cycling-associated processes in terrestrial ecosystems. This study investigated the separate and combined impacts of long-term soil warming and fertilization on soil AOB and AOA community structures and abundances in a Norway spruce stand in northern Sweden.

Materials and methods

The soil-warming experiment was established in the buffer zones of two irrigated plots (I) and complete nutrient solution plots (IL) since 1995. The warming treatment started in April each year by maintaining soil temperature on warmed plots at 5°C above the temperature in unwarmed plots using heating cables. In August 2006, soil samples were collected from eight subplots for molecular analysis. The abundance of bacterial and archaeal amoA genes was determined by quantitative polymerase chain reaction. Similarly, total bacterial and archaeal population sizes have also been determined. The diversity of AOB and AOA was assessed by constructing amoA gene clone libraries, and different genotypes were screened with restriction fragment length polymorphism.

Results and discussion

Results showed that fertilization did not significantly affect the abundance of the bacterial amoA gene under either warming or non-warming conditions; however, warming decreased the abundance under fertilization treatments. No significant effects of fertilization and soil warming were observed on the number of thaumarchaeal amoA gene copies across all treatments. In this study, amoA gene abundance of AOB was significantly higher than that of AOA across all treatments. The community structure of both AOB and AOA was strongly influenced by fertilization. For bacterial amoA genes, Nitrosospira cluster 2 was present across all treatments, but the only genotype was observed in the fertilization treatments while, for thaumarchaeal amoA genes, the relative abundance of soil cluster 5 increased in fertilization treatments. By comparison, soil-warming effects on AOB and AOA community structure were not significant. Canonical correspondence analysis showed a positive correlation between fertilization and both dominant genotypes of AOB and AOA.

Conclusions

These results indicated that the abundance of AOA and AOB was not affected by fertilization or warming alone, but the interaction of fertilization and warming reduced the abundance of AOB. The community composition of ammonia-oxidizers was more affected by the nutrient-optimized fertilization than the soil warming.     

Ammonia oxidizers and denitrifiers in response to reciprocal elevation translocation in an alpine meadow on the Tibetan Plateau

Purpose

Global climate change, in particular temperature variation, is likely to alter soil microbial abundance and composition, with consequent impacts on soil biogeochemical cycling and ecosystem functioning. However, responses of belowground nitrogen transformation microorganisms to temperature changes in high-elevation terrestrial ecosystems are not well understood.

Materials and methods

Here, the effects of simulated cooling and warming on the abundance and community composition of ammonia-oxidizing archaea (AOA) and bacteria (AOB), as well as the abundance of denitrifiers, were investigated using quantitative polymerase chain reaction and clone library approaches, on the basis of a 2-year reciprocal elevation translocation experiment along an elevation gradient from 3,200 to 3,800 m above sea level on the Tibetan Plateau.

Results and discussion

We found that, compared with the temperature variations caused by elevation translocation, the soil origin exerted a much stronger influence on AOA abundance. There were significant effects of both soil origin and elevation translocation on AOB abundance, which was particularly decreased by elevation-enhanced (simulated cooling) and increased by elevation-decreased (simulated warming) treatments. Altered temperature affected the abundance of nirK rather than nirS and nosZ genes, and the latter two seemed to be associated tightly with the soil origin. Furthermore, the results showed that temperature changes had obvious influences on the community structure and diversity of AOB, but not AOA. More apparent response of AOB to warming than in other studies on grassland and forest ecosystems may be attributed to higher elevation and lower mean annual temperature in this study.

Conclusions

Our findings thus suggest that, in comparison with AOA and denitrifying populations, AOB may respond more sensitively to natural temperature variation caused by elevation translocation in this alpine grassland ecosystem on the Tibetan Plateau.     

The effect of soil pH and dicyandiamide (DCD) on N2O emissions and ammonia oxidiser abundance in a stimulated grazed pasture soil

Purpose

Nitrous oxide (N₂O) is a potent greenhouse gas which is mainly produced from agricultural soils through the processes of nitrification and denitrification. Although denitrification is usually the major process responsible for N₂O emissions, N₂O production from nitrification can increase under some soil conditions. Soil pH can affect N₂O emissions by altering N transformations and microbial communities. Bacterial (AOB) and archaeal (AOA) ammonia oxidisers are important for N₂O production as they carry out the rate-limiting step of the nitrification process.

Material and methods

A field study was conducted to investigate the effect of soil pH changes on N₂O emissions, AOB and AOA community abundance, and the efficacy of a nitrification inhibitor, dicyandiamide (DCD), at reducing N₂O emissions from animal urine applied to soil. The effect of three pH treatments, namely alkaline treatment (CaO/NaOH), acid treatment (HCl) and native (water) and four urine and DCD treatments as control (no urine or DCD), urine-only, DCD-only and urine + DCD were assessed in terms of their effect on N₂O emissions and ammonia oxidiser community growth.

Results and discussion

Results showed that total N₂O emissions were increased when the soil was acidified by the acid treatment. This was probably due to incomplete denitrification caused by the inhibition of the assembly of the N₂O reductase enzyme under acidic conditions. AOB population abundance increased when the pH was increased in the alkaline treatment, particularly when animal urine was applied. In contrast, AOA grew in the acid treatment, once the initial inhibitory effect of the urine had subsided. The addition of DCD decreased total N₂O emissions significantly in the acid treatment and decreased peak N₂O emissions in all pH treatments. DCD also inhibited AOB growth in both the alkaline and native pH treatments and inhibited AOA growth in the acid treatment.

Conclusions

These results show that N₂O emissions increase when soil pH decreases. AOB and AOA prefer different soil pH environments to grow: AOB growth is favoured in an alkaline pH and AOA growth favoured in more acidic soils. DCD was effective in inhibiting AOB and AOA when they were actively growing under the different soil pH conditions.     

Management practices have a major impact on nitrifier and denitrifier communities in a semiarid grassland ecosystem

Purpose

Nitrification and denitrification, two of the key nitrogen (N) transformation processes in the soil, are carried out by a diverse range of microorganisms and catalyzed by a series of enzymes. Different management practices, such as continuous grazing, mowing, and periodic fencing off from grazing, dramatically influenced grassland ecosystems. This study aimed to examine the effects of management practices on the abundance and community structure of nitrifier and denitrifier communities in grassland ecosystems.

Materials and methods

Soil samples were collected from a semiarid grassland ecosystem in Xilingol region, Inner Mongolia, where long-term management practices including free-grazing, different periods of enclosure from grazing, and different frequencies of mowing were conducted. Real-time quantitative polymerase chain reaction (Q-PCR), denaturing gradient gel electrophoresis (DGGE), sequencing, and phylogenetic analysis were applied to estimate the abundance and composition of amoA, nirS, nirK, and nosZ genes.

Results and discussion

The ammonia-oxidizing archaea (AOA) amoA copies were in the range 5.99?×?10⁸ to 8.60?×?10⁸, while those of ammonia-oxidizing bacteria (AOB) varied from 3.02?×?10⁷ to 4.61?×?10⁷. The abundance of AOA was substantially higher in the light grazing treatment (LG) than in the mowing treatments. The quantity and intensity of DGGE bands of AOA varied with pasture management. In stark contrast, AOB population abundance and community structure remained largely unchanged in all the soils irrespective of the management practices. All these results suggested that ammonia oxidizers were dominated by AOA. The higher gene abundance and greater intensity of DGGE bands of nirS and nosZ under the enclosure treatments would suggest greater stimulated denitrification. The ratio of nosZ/(nirS?+?nirK) was higher in mowing treatments than in the free-grazing and enclosure treatments, possibly leading to more complete denitrification. Correlation analysis indicated that soil moisture and inorganic nitrogen content were the two main soil environmental variables that influence the community structure of nitrifiers and denitrifiers.

Conclusions

In this semiarid neutral to alkaline grassland ecosystem under low temperature conditions, AOA mainly affiliated with Nitrososphaera dominated nitrification. These results clearly demonstrate that grassland management practices can have a major impact on nitrifier and denitrifier communities in this semiarid grassland ecosystem, under low temperature conditions.

     

Effect of application rate of a nitrification inhibitor,dicyandiamide (DCD), on nitrification rate,and ammonia-oxidizing bacteria and archaea growth in a grazed pasture soil: An incubation study

Purpose

Dicyandiamide (DCD) has been used commercially in New Zealand to reduce nitrate leaching and N₂O emissions in grazed pastures. However, there is a lack of information in the literature on the optimum rate of DCD to achieve the environmental benefits while at the same time reducing the cost of the technology. The objective of this study was to determine the effect of DCD application rate on its effectiveness to inhibit ammonia oxidizer growth and nitrification rate in a grazed pasture soil.

Materials and methods

The soil was a Templeton silt loam (Immature Pallic Soil; Udic Haplustepts) collected from Lincoln University Research Dairy Farm with a mixed pasture consisting of perennial ryegrass (Lolium perenne L.) and white clover (Trifolium repens L.) and was incubated alone (control) or with cow urine at 700 kg N/ha with 6 rates of DCD [0, 2.5, 5, 7.5, 10 (applied twice), 15 and 20 kg/ha] in incubation vessels. The incubation vessels were placed randomly in an incubator with a constant temperature of 12 °C. During 112 days of incubation, soil subsamples were taken at different time intervals to measure the concentrations of NO₃ ^?-N and NH₄ ⁺-N and the amoA gene copy numbers of ammonia-oxidizing bacteria (AOB) and ammonia-oxidizing archaea (AOA).

Results and discussion

DCD applied at all the different rates inhibited nitrification in urine-treated soils, but the effectiveness increased with DCD application rate. In addition, AOB growth and the amounts of nitrate-N in the soil were significantly related to the application rate of DCD. However, AOA population abundance showed no relationship to the application rate of DCD. The DCD rate at which the AOB growth rate and nitrate-N concentration were halved (effective dosage that causes 50 % reduction in nitrification rate, or ED₅₀) was about 10 kg DCD/ha.

Conclusions

These results suggest that DCD applied at relatively low rates still slowed down the nitrification rate, and the current recommended rate of 10 kg DCD/ha for DCD use in New Zealand grazed pastures would result in a 50 % reduction in nitrification rate in this soil. The actual rate of DCD application used would depend on the cost of the product and the environmental and agronomic benefits that would result from its use.     

Impacts of continuous excessive fertilization on soil potential nitrification activity and nitrifying microbial community dynamics in greenhouse system

Purpose

Sampling and analysis of greenhouse soils were conducted in Shouguang, China, to study continuous excessive fertilization effect on nitrifying microbial community dynamics in greenhouse environment.

Materials and methods

Potential nitrification activity (PNA), abundance, and structure of nitrifying microbial communities as well as the correlations with soil properties were investigated.

Results and discussion

Short-term excessive fertilization increased soil nutrient contents and the diversity of nitrifying microbial communities under greenhouse cultivation. However, the abundance and diversity of nitrifying communities decreased greatly due to the increase of soil acidity and salinity after 14 years of high fertilization in greenhouse. There was a significant positive correlation between soil PNA and the abundance of ammonia-oxidizing bacteria (AOB) but not that of ammonia-oxidizing archaea (AOA) in topsoil (0–20 cm) when pH ≥7. Soil PNA and AOB were strongly influenced by soil pH. The groups of Nitrososphaeraceae, Nitrosomonadaceae, and Nitrospiraceae were predominant in the AOA, AOB, and nitrite-oxidizing bacteria (NOB) communities, respectively. Nitrifying community structure was significantly correlated with soil electrical salinity (EC), organic carbon (OC), and nitrate nitrogen (NO₃ ^?–N) content by redundancy analysis (RDA).

Conclusions

Nitrification was predominated by AOB in greenhouse topsoil with high fertilizer loads. Soil salinity, OC, NO₃ ^?–N content, and pH affected by continuous excessive fertilization were the major edaphic factors in shaping nitrifying community structure in greenhouse soils.

     

Comammox—a newly discovered nitrification process in the terrestrial nitrogen cycle

Purpose

Nitrification, the microbial oxidation of ammonia to nitrate via nitrite, is a pivotal component of the biogeochemical nitrogen cycle. Nitrification was conventionally assumed as a two-step process in which ammonia oxidation was thought to be catalyzed by ammonia-oxidizing archaea (AOA) and bacteria (AOB), as well as nitrite oxidation by nitrite-oxidizing bacteria (NOB). This long-held assumption of labour division between the two functional groups, however, was challenged by the recent unexpected discovery of complete ammonia oxidizers within the Nitrospira genus that are capable of converting ammonia to nitrate in a single organism (comammox). This breakthrough raised fundamental questions on the niche specialization and differentiation of comammox organisms with other canonical nitrifying prokaryotes in terrestrial ecosystems.

Materials and methods

This article provides an overview of the recent insights into the genomic analysis, physiological characterization and environmental investigation of the comammox organisms, which have dramatically changed our perspective on the aerobic nitrification process. By using quantitative PCR analysis, we also compared the abundances of comammox Nitrospira clade A and clade B, AOA, AOB and NOB in 300 forest soil samples from China spanning a wide range of soil pH.

Results and discussion

Comammox Nitrospira are environmentally widespread and numerically abundant in natural and engineered habitats. Physiological data, including ammonia oxidation kinetics and metabolic versatility, and comparative genomic analysis revealed that comammox organisms might functionally outcompete other canonical nitrifiers under highly oligotrophic conditions. These findings highlight the necessity in future studies to re-evaluate the niche differentiation between ammonia oxidizers and their relative contribution to nitrification in various terrestrial ecosystems by including comammox Nitrospira in such comparisons.

Conclusions

The discovery of comammox and their broad environmental distribution added a new dimension to our knowledge of the biochemistry and physiology of nitrification and has far-reaching implications for refined strategies to manipulate nitrification in terrestrial ecosystems and to maximize agricultural productivity and sustainability.

     

Effect of agricultural land use change on community composition of bacteria and ammonia oxidizers

Purpose

Soil microbial communities can be strongly influenced by agricultural practices, but little is known about bacterial community successions as land use changes. The objective of this study was to determine microbial community shifts following major land use changes in order to improve our understanding of land use impacts on microbial community composition and functions.

Materials and methods

Four agricultural land use patterns were selected for the study, including old rice paddy fields (ORP), Magnolia nursery planting (MNP), short-term vegetable (STV), and long-term vegetable (LTV) cultivation. All four systems are located in the same region with same soil parent material (alluvium), and the MNP, STV, and LTV systems had been converted from ORP for 10, 3, and 30 years, respectively. Soil bacteria and ammonia oxidizer community compositions were analyzed by 454 pyrosequencing and terminal restriction fragment length polymorphism, respectively. Quantitative PCR was used to determine 16S rRNA and amoA gene copy numbers.

Results and discussion

The results showed that when land use was changed from rice paddy to upland systems, the relative abundance of Chloroflexi increased whereas Acidobacteria decreased significantly. While LTV induced significant shifts of bacterial composition, MNP had the highest relative abundance of genera GP1, GP2, and GP3, which were mainly related to the development of soil acidity. The community composition of ammonia-oxidizing bacteria (AOB) but not ammonia-oxidizing archaea was strongly impacted by the agricultural land use patterns, with LTV inducing the growth of a single super predominant AOB group. The land use changes also induced significant shifts in the abundance of 16S rRNA and bacterial amoA genes, but no significant differences in the abundance of archaea amoA was detected among the four land use patterns. Soil total phosphorous, available phosphorous, NO₃ ^?, and soil organic carbon contents and pH were the main determinants in driving the composition of both bacteria and AOB communities.

Conclusions

These results clearly show the significant impact of land use change on soil microbial community composition and abundance and this will have major implications on the microbial ecology and nutrient cycling in these systems, some of which is unknown. Further research should be directed to studying the impacts of these microbial community shifts on nutrient dynamics in these agroecosystems so that improved nutrient management systems can be developed.     

Responses of soil microbial community to nitrogen fertilizer and precipitation regimes in a semi-arid steppe

Purpose

Changes of nitrogen (N) cycle caused by N fertilization and precipitation regimes have affected the key ecosystem structure and functions in temperate steppe, which may modify the structure of soil microbial communities involved in N transformation. This paper was designated to examine the response of soil ammonia oxidizers and denitrifiers to the N fertilization and precipitation regimes in a semi-arid steppe where N and water contents are major limiting factors of the grassland productivity.

Materials and methods

This study was based on a long-term N fertilization and precipitation regimes experiment in Inner Mongolia (116° 17′ 20″ E, 42° 2′ 29″ N). The treatments including CK (control), R (reduced precipitation), W (30% increase in precipitation), N (10 g N m^?2 y^?1), RN (reduced precipitation and 10 g N m^?2 y^?1), and WN (30% increase in precipitation and 10 g N m^?2 y^?1). Soil basic chemical properties and microbial activities were analyzed. Molecular methods were applied to determine the abundance, structure and diversity of ammonia oxidizers and denitrifiers. Statistical analysis detected the main and interactive effect of treatments on soil microbial communities and revealed the relationship between soil microbial community structures and environmental factors.

Results and discussion

N fertilization significantly increased ammonia-oxidizing bacteria (AOB) abundance. Ammonia-oxidizing archaea (AOA) community structure was markedly changed in N fertilizer treatment and strongly affected by soil pH, while soil nitrate and water content correlated with AOB community structure. Soil nitrate was the key factor influencing nirK gene community structure, while soil pH and water content explained much of the variations of nosZ gene community. AOB-amoA and nosZ gene community diversities were influenced by precipitation regimes and interaction of N fertilization and precipitation regimes, respectively.

Conclusions

N fertilization and precipitation regimes had significant influences on the changes of soil properties and microbial functional communities. Soil nitrification was mainly driven by AOB in the semi-arid grassland. Changes of substrate content and soil pH were the key factors in shifting functional microbial communities. The non-synergistic effects of N fertilization and precipitation regimes on the microbial functional groups indicated that the negative effect of lower pH induced by N fertilization would be alleviated by precipitation regimes, which should be well considered in grassland restoration.

     

The short-term effects of nitrification inhibitors on the abundance and expression of ammonia and nitrite oxidizers in a long-term field experiment comparing land management

Microcosms were set up to evaluate the effect of nitrification inhibitors (DCD, c-PTiO, and NaClO₃) on the abundance and expression of ammonia-oxidizing bacteria (AOB) and archaea (AOA), as well as the nitrite-oxidizing bacteria (NOB) Nitrospira and Nitrobacter. Both DCD and NaClO₃ inhibited the net nitrification rate, while c-PTiO had no significant effects, and NaClO₃ had a much greater inhibitory effect (> 60%) in all soils than DCD. No significant changes in total microbial abundance were observed with DCD and NaClO₃. DCD limited only the growth of AOB; however, NaClO₃ inhibited growth of both AOA and Nitrospira-NOB with no significant effects on AOB and Nitrobacter-NOB. Probably NaClO₃ inhibited both ammonia oxidation and nitrite oxidation. This is the first report to reveal the inhibitory effects of NaClO₃ on a specific nitrification process, helping to clarify the ecological niche of nitrifiers and the potential of nitrification inhibitors applied to soil.     

Litterbag decomposition of residues from Bacillus thuringiensis (Bt) rice hybrids and the parental lines under multiple field conditions

Purpose

The cultivation of genetically modified (GM) crops has raised environmental concerns, since large amounts of plant materials remain in the field after harvesting. Specific proteins of GM crops might negatively impact soil ecosystem by changing residue decomposition dynamics. Particularly, the residue decomposition of crop-wild hybrids, which were formed through transgene escape to wild population, remains unexplored.

Materials and methods

We used litter bags to assess residue (leaves, stems and roots) decomposition dynamics of two stacked genes from Bacillus thuringiensis (Bt) Cry1Ac and the sck (a modified CpTI gene encoding a cowpea trypsin-inhibitor) (Bt/CpTI) rice lines (Kefeng-6 and Kefeng-8), a non-transgenic rice near isoline (Minghui86), wild rice (Oryza rufipogon) and Bt wild rice at three sites. The enzyme-linked immunosorbent assay (ELISA) was used to monitor the changes of the Cry1Ac protein in Bt rice residues.

Results and discussion

Mass remaining, total N and total C concentrations of rice residues declined over time and varied among plant tissues, with significant differences among cultivar, crop-wild hybrids and wild rice, but no differences between Bt and non-Bt rice cultivars. The initial concentration of Cry1Ac was higher in leaves and stems than in roots and was different between rice types. The degradation dynamics of Cry1Ac fitted best to a first-order kinetics model and correlated with the level of total nitrogen in residues but did not correlate with the mass decomposition rate. The predicted DT50 (50 % degradation time) of the protein ranged from 10.7 to 63.6 days, depending on plant types, parts and burial sites. By the end of the study (~170 days), the protein was present in low concentration in the remaining residues.

Conclusions

Our results suggest that the impacts of the stacked Bt/CpTI gene inserts on the decomposition dynamics of rice residues are insignificant.     

Bacterial community incorporating carbon derived from plant residue in an anoxic non-rhizosphere soil estimated by DNA-SIP analysis

Purpose

Plant residues are one of the main sources of soil organic matter in paddy fields, and elucidation of the bacterial communities decomposing plant residues was important to understand their function and roles, as the microbial decomposition of plant residues is linked to soil fertility. We conducted a DNA stable isotope probing (SIP) experiment to elucidate the bacterial community assimilating 13-carbon (¹³C) derived from plant residue under an anoxic soil condition. In addition, we compared the bacterial community with that under the oxic soil condition, which was elucidated in our previous study (Lee et al. in Soil Biol Biochem 43:814–822, 2011).

Materials and methods

We used the ¹³C-labeled dried rice callus cells as a model of rice plant residue. A paddy field soil was incubated with unlabeled and ¹³C-labeled callus cells. DNA extracted from the soils was subjected to buoyant density gradient centrifugation to fractionate ¹³C-enriched DNA. Then, polymerase chain reaction (PCR) and denaturing gradient gel electrophoresis (DGGE) analysis of bacterial 16S rDNA band patterns and band sequencing method were used to evaluate bacterial community.

Results and discussion

DGGE analysis showed that the band patterns in the ¹³C-enriched fractions were distinctly changed over time, while the changes in the community structure before fractionation were minor. Sequencing of the ¹³C-labeled DGGE bands revealed that Clostridia were a major group in the bacterial communities incorporating the callus-derived carbon although Gram-negative bacteria, and Actinobacteria also participated in the carbon flow from the callus under the anoxic condition. The proportion of Gram-negative bacteria and Actinobacteria increased on 14 days after the onset of incubation, suggesting that the callus was decomposed by diverse bacterial members on this phase. When the bacterial groups incorporating the ¹³C were compared between under anoxic and oxic soil conditions, the composition was largely different under the two opposite conditions. However, some members of Gram-negative bacteria were commonly found under the anoxic and oxic soil conditions.

Conclusions

The majority of bacterial members assimilating the callus carbon was Clostridia in the soil under anoxic conditions. However, several Gram-negative bacterial members, such as Acidobacteria, Bacteroidetes, and Proteobacteria, also participated in the decomposition of callus under anoxic soil conditions. Our study showed that carbon flow into the diverse bacterial members during the callus decomposition and the distinctiveness of the bacterial communities was formed under the anoxic and oxic soil conditions.

     

Carbon flux from decomposing 13C-labeled transgenic and nontransgenic parental rice straw in paddy soil

Purpose

Genetic modification of Bt rice may affect straw decomposition and soil carbon pool under flood conditions. This study aims to assess the effects of cry gene transformation in rice on the residue decomposition and fate of C from residues under flooded conditions.

Materials and methods

A decomposition experiment was set up using ¹³C-enriched rice straws from transgenic and nontransgenic Bt rice to evaluate the soil C dynamics and CH₄ or CO₂ emission rates in the root and non-root zones. The concentrations and stable carbon isotope compositions of the soil organic carbon (SOC), dissolved organic carbon (DOC), microbial biomass carbon (MBC), CH₄, and CO₂ of the root and non-root zones were determined from 7 to 110 days after rice straw incorporation.

Results and discussion

Rice straw incorporation into soil significantly increased the SOC, DOC, and MBC concentrations and the CH₄ and CO₂ emission rates. The percentage of ¹³C-SOC remaining in the root zone was significantly lower than that in the non-root zone with rice straw decomposition. The DOC and MBC concentrations significantly increased in both the root and non-root zones between 0 and 80 days after rice straw incorporation. However, no significant differences were found after Bts (Bt rice straw added into soil) and Cks (nontransgenic Bt rice straw added into soil) incorporation in the root and non-root zones. This result may be attributed to the priming effects of sufficient oxygen and nutrients on straw degradation in the root zone.

Conclusions

Bt gene insertion did not affect the SOC, DOC, and MBC concentrations and the CH₄ and CO₂ emission rates in both the root and non-root zones. However, rice straw incorporation and root exudation significantly increased the SOC, DOC, and MBC concentrations and the CH₄ and CO₂ emission rates.     

The effects of different irrigation regimes on nitrous oxide emissions and influencing factors in greenhouse tomato fields

Purpose

Intensive agricultural practices have enhanced problems associated with the competing use of limited water resources. Nitrous oxide (N₂O) is a major contributor to global warming. It is important for researchers to ascertain the relationship between irrigation and soil N₂O emissions in order to identify mitigation strategies to reduce nitrous oxide emissions. Different irrigation amounts affect soil water dynamics and nitrogen turnover. The effect of three lower limits of irrigation on soil N₂O emissions, influencing factors, and abundance of genes involved in nitrification and denitrification were investigated in tomato irrigated in a greenhouse.

Materials and methods

Observations were performed between April and August 2015 in a long-term irrigated field subjected to different lower limits of irrigation: 20 kPa (D₂₀), 30 kPa (D₃₀), and 40 kPa (D₄₀) from greenhouse soil during the tomato crop season. Soil N₂O fluxes were monitored using the static chamber-gas chromatograph method. Copy numbers of genes were determined using the real-time quantitative polymerase chain reaction (real-time PCR) technique. Characteristics of soil N₂O emissions were analyzed, and differences between irrigation regimes were determined. The effects of influencing factors on soil N₂O emissions were analyzed, including soil temperature, soil moisture, soil pH, and soil mineral nitrogen, as well as changes in the abundance of soil ammonia-oxidizing bacteria (AOB) and ammonia-oxidizing archaea (AOA) based on amoA genes and denitrifier genes (nosZ, nirK, and cnorB).

Results and discussion

Our results showed that peaks in N₂O emissions occurred 1–5 days after each irrigation. During the whole tomato growth period, soil N₂O fluxes were lowest under D₃₀ treatment compared with those under D₂₀ and D₄₀ treatments. Soil NO₃ ^?-N concentrations were significantly higher than NH₄ ⁺-N concentrations. Soil N₂O fluxes were significantly related to soil moisture, NH₄ ⁺-N concentrations (P < 0.01), soil pH, and AOA copy numbers (P < 0.05). There was no consistent correlation between soil N₂O emissions, soil temperature, and soil NO₃ ^?-N concentrations. Different irrigation regimes significantly affected AOA copy numbers but did not affect the expression of other genes. AOA copy numbers were higher than those of AOB. Soil N₂O fluxes significantly affected the AOA copy numbers and potential nitrification rates (P < 0.05).

Conclusions

Soil moisture, pH, and NH₄ ⁺-N concentration were important factors affecting soil N₂O emissions. Compared with other genes associated with nitrification and denitrification, AOA plays an important role in N₂O emissions from greenhouse soils. Selecting a lower limit of irrigation of 30 kPa could effectively reduce N₂O emissions from vegetable soils.

     

Soil pH and plant diversity drive co-occurrence patterns of ammonia and nitrite oxidizer in soils from forest ecosystems

In this study, we investigated how co-occurrence patters of ammonia and nitrite oxidizers, which drive autotrophic nitrification, are influenced by tree species composition as well as soil pH in different forest soils. We expected that a decline of ammonia oxidizers in coniferous forests, as a result of excreted nitrification inhibitors and at acidic sites with low availability of ammonia, would reduce the abundance of nitrite-oxidizing bacteria (NOB). To detect shifts in co-occurrence patterns, the abundance of key players was measured at 50 forest plots with coniferous respectively deciduous vegetation and different soil pH levels in the region Schwäbische Alb (Germany). We found ammonia-oxidizing archaea (AOA) and Nitrospira-like NOB (NS) to be dominating in numbers over their counterparts across all forest types. AOA co-occurred mostly with NS, while bacterial ammonia oxidizers (AOB) were correlated with Nitrobacter-like NOB (NB). Co-occurrence patterns changed from tight significant relationships of all ammonia and nitrite oxidizers in deciduous forests to a significant relationship of AOB and NB in coniferous forests, where AOA abundance was reduced. Surprisingly, no co-occurrence structures between ammonia and nitrite oxidizers could be determined at acidic sites, although abundances were correlated to the respective nitrogen pools. This raises the question whether interactions with heterotrophic nitrifiers may occur, which needs to be addressed in future studies.