Hang-Wei Hu

Ammonia-oxidizing archaea have more important role than ammonia-oxidizing bacteria in ammonia oxidation of strongly acidic soils

Increasing evidence demonstrated the involvement of ammonia-oxidizing archaea (AOA) in the global nitrogen cycle, but the relative contributions of AOA and ammonia-oxidizing bacteria (AOB) to ammonia oxidation are still in debate. Previous studies suggest that AOA would be more adapted to ammonia-limited oligotrophic conditions, which seems to be favored by protonation of ammonia, turning into ammonium in low-pH environments. Here, we investigated the autotrophic nitrification activity of AOA and AOB in five strongly acidic soils (pH<4.50) during microcosm incubation for 30 days. Significantly positive correlations between nitrate concentration and amoA gene abundance of AOA, but not of AOB, were observed during the active nitrification. 13CO2-DNA-stable isotope probing results showed significant assimilation of 13C-labeled carbon source into the amoA gene of AOA, but not of AOB, in one of the selected soil samples. High levels of thaumarchaeal amoA gene abundance were observed during the active nitrification, coupled with increasing intensity of two denaturing gradient gel electrophoresis bands for specific thaumarchaeal community. Addition of the nitrification inhibitor dicyandiamide (DCD) completely inhibited the nitrification activity and CO2 fixation by AOA, accompanied by decreasing thaumarchaeal amoA gene abundance. Bacterial amoA gene abundance decreased in all microcosms irrespective of DCD addition, and mostly showed no correlation with nitrate concentrations. Phylogenetic analysis of thaumarchaeal amoA gene and 16S rRNA gene revealed active 13CO2-labeled AOA belonged to groups 1.1a-associated and 1.1b. Taken together, these results provided strong evidence that AOA have a more important role than AOB in autotrophic ammonia oxidation in strongly acidic soils.

Microbial regulation of terrestrial nitrous oxide formation: understanding the biological pathways for prediction of emission rates

The continuous increase of the greenhouse gas nitrous oxide (N2O) in the atmosphere due to increasing anthropogenic nitrogen input in agriculture has become a global concern. In recent years, identification of the microbial assemblages responsible for soil N2O production has substantially advanced with the development of molecular technologies and the discoveries of novel functional guilds and new types of metabolism. However, few practical tools are available to effectively reduce in situ soil N2O flux. Combating the negative impacts of increasing N2O fluxes poses considerable challenges and will be ineffective without successfully incorporating microbially regulated N2O processes into ecosystem modeling and mitigation strategies. Here, we synthesize the latest knowledge of (i) the key microbial pathways regulating N2O production and consumption processes in terrestrial ecosystems and the critical environmental factors influencing their occurrence, and (ii) the relative contributions of major biological pathways to soil N2O emissions by analyzing available natural isotopic signatures of N2O and by using stable isotope enrichment and inhibition techniques. We argue that it is urgently necessary to incorporate microbial traits into biogeochemical ecosystem modeling in order to increase the estimation reliability of N2O emissions. We further propose a molecular methodology oriented framework from gene to ecosystem scales for more robust prediction and mitigation of future N2O emissions.

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

PurposeAmmonia-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 methodsBarcoded 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 discussionThe 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.ConclusionsThese 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.

Water addition regulates the metabolic activity of ammonia oxidizers responding to environmental perturbations in dry subhumid ecosystems.

Terrestrial arid and semi-arid ecosystems (drylands) constitute about 41% of the Earths land surface and are predicted to experience increasing fluctuations in water and nitrogen availability. Mounting evidence has confirmed the significant importance of ammonia-oxidizing archaea (AOA) and bacteria (AOB) in nitrification, plant nitrogen availability and atmospheric N2 O emissions, but their responses to environmental perturbations in drylands remain largely unknown. Here we evaluate how the factorial combinations of irrigation and fertilization in forests and land-use change from grassland to forest affects the dynamics of AOA and AOB following a 6-year dryland field study. Potential nitrification rates and AOA and AOB abundances were significantly higher in the irrigated plots, accompanied by considerable changes in community compositions, but their responses to fertilization alone were not significant. DNA-stable isotope probing results showed increased (13) CO2 incorporation into the amoA gene of AOA, but not of AOB, in plots receiving water addition, coupled with significantly higher net mineralization and nitrification rates. High-throughput microarray analysis revealed that active AOA assemblages belonging to Nitrosopumilus and Nitrosotalea were increasingly labelled by (13) CO2 following irrigation. However, no obvious effects of land-use changes on nitrification rates or metabolic activity of AOA and AOB could be observed under dry conditions. We provide evidence that water addition had more important roles than nitrogen fertilization in influencing the autotrophic nitrification in dryland ecosystems, and AOA are increasingly involved in ammonia oxidation when dry soils become wetted.

Ammonia-Oxidizing Archaea Play a Predominant Role in Acid Soil Nitrification

Acid soils, extensively used for nitrogen-fertilized agriculture and agroforestry, have important roles in maintaining global biogeochemical cycling and ecosystem functions. Huge inputs of nitrogen-based fertilizers into terrestrial ecosystems accelerate soil acidification, concomitantly altering the nitrogen transformation processes. Nitrification, as a critical component of the nitrogen cycle, is a microbially mediated process from ammonia to nitrate via nitrite, contributing to enormous losses of fertilizers through atmospheric emissions of greenhouse gas N2O and nitrate leaching to groundwater. However, the functionally dominant nitrifiers and underlying mechanisms for the acid soil nitrification are a long-standing mystery, which have confused scientists for more than 100 years. This century-long paradox originated from the early observations of active nitrification activity in acid soils, intensified by the failure of ammonia-oxidizing bacteria (AOB) cultures to sustain the nitrification in liquid batch under acid conditions, might be resolved by the recent progress in metagenomic, isotopic probing studies, and isolation of acidophilic ammonia-oxidizing archaea (AOA). Emerging evidence led to the supposition of the predominant role of AOA in controlling the autotrophic ammonia oxidation of acid soils, which has radically revised the previous perception that this oxidative reaction was exclusively regulated by autotrophic AOB and occasionally by heterotrophic nitrifiers. In this chapter, we review the recent progress in our understanding of the pH-impacted distribution of ammonia oxidizers, the niche differentiation of AOA and AOB shaped by acid stress, and the possible mechanisms of autotrophic nitrification in acid soils. The unveiling of this key process in widely distributed acid soils would help to identify effective biological strategies for better management of terrestrial nitrogen turnover and balance.

Field‐based evidence for copper contamination induced changes of antibiotic resistance in agricultural soils

Bacterial resistance to antibiotics and heavy metals are frequently linked, suggesting that exposure to heavy metals might select for bacterial assemblages conferring resistance to antibiotics. However, there is a lack of clear evidence for the heavy metal-induced changes of antibiotic resistance in a long-term basis. Here, we used high-capacity quantitative PCR array to investigate the responses of a broad spectrum of antibiotic resistance genes (ARGs) to 4-5 year copper contamination (0-800 mg kg-1 ) in two contrasting agricultural soils. In total, 157 and 149 unique ARGs were detected in the red and fluvo-aquic soil, respectively, with multidrug and β-lactam as the most dominant ARG types. The highest diversity and abundance of ARGs were observed in medium copper concentrations (100-200 mg kg-1 ) of the red soil and in high copper concentrations (400-800 mg kg-1 ) of the fluvo-aquic soil. The abundances of total ARGs and several ARG types had significantly positive correlations with mobile genetic elements (MGEs), suggesting mobility potential of ARGs in copper-contaminated soils. Network analysis revealed significant co-occurrence patterns between ARGs and microbial taxa, indicating strong associations between ARGs and bacterial communities. Structural equation models showed that the significant impacts of copper contamination on ARG patterns were mainly driven by changes in bacterial community compositions and MGEs. Our results provide field-based evidence that long-term Cu contamination significantly changed the diversity, abundance and mobility potential of environmental antibiotic resistance, and caution the un-perceived risk of the ARG dissemination in heavy metal polluted environments.

Soil pH determines the alpha diversity but not beta diversity of soil fungal community along altitude in a typical Tibetan forest ecosystem

PurposeDespite their symbiotic relationship with trees and the vital role as decomposer in forest, soil fungi received limited attention regarding their changes with altitude in forest ecosystems. This study aimed to determine the diversity patterns of soil fungi along an altitudinal gradient on Mt. Shegyla, a typical forest ecosystem on the Tibetan Plateau.Materials and methodsHigh-throughput barcoded pyrosequencing and quantitative PCR approaches were employed to measure the community composition, diversity, and abundance patterns of soil fungal 18S ribosomal RNA (rRNA) gene in 20 samples collected along the altitudinal gradient of Mt. Shegyla.Results and discussionAbundant taxa in the fungal community were Agaricomycetes and Leotiomyceta on Mt. Shegyla. Fungal abundance decreased significantly with increasing altitude. Beta diversity of the fungal community, as measured using weighted UniFrac distance, was significantly related to altitude. Significant correlation was observed between altitude and alpha diversity including richness and phylodiversity, but not with evenness. Network analysis revealed that Ceramothyrium and Clavulina were two important hubs in the community, and an uncultured fungal taxon that previously detected in glacier forefront dominated this network. Distance-based linear model identified soil pH as the dominant driver which significantly related with fungal alpha diversity including richness, phylodiversity, and evenness. However, fungal abundance and the first component of PCoA on weighted UniFrac matrix (beta diversity) did not change significantly with pH.ConclusionsThese results provided strong evidence that soil pH was the dominant driver for structuring altitudinal alpha diversity pattern but not beta diversity pattern or community abundance of soil fungi in this typical forest on the Tibetan Plateau.

Long-Term Nickel Contamination Increases the Occurrence of Antibiotic Resistance Genes in Agricultural Soils

Heavy metal contamination is assumed to be a selection pressure on antibiotic resistance, but to our knowledge, evidence of the heavy metal-induced changes of antibiotic resistance is lacking on a long-term basis. Using quantitative PCR array and Illumina sequencing, we investigated the changes of a wide spectrum of soil antibiotic resistance genes (ARGs) following 4-5 year nickel exposure (0-800 mg kg-1) in two long-term experimental sites. A total of 149 unique ARGs were detected, with multidrug and β-lactam resistance as the most prevailing ARG types. The frequencies and abundance of ARGs tended to increase along the gradient of increasing nickel concentrations, with the highest values recorded in the treatments amended with 400 mg nickel kg-1 soil. The abundance of mobile genetic elements (MGEs) was significantly associated with ARGs, suggesting that nickel exposure might enhance the potential for horizontal transfer of ARGs. Network analysis demonstrated significant associations between ARGs and MGEs, with the integrase intI1 gene having the most frequent interactions with other co-occurring ARGs. The changes of ARGs were mainly driven by nickel bioavailability and MGEs as revealed by structural equation models. Taken together, long-term nickel exposure significantly increased the diversity, abundance, and horizontal transfer potential of soil ARGs.

Abundance and community structure of ammonia-oxidizing Archaea and Bacteria in response to fertilization and mowing in a temperate steppe in Inner Mongolia

Based on a 6-year field trial in a temperate steppe in Inner Mongolia, we investigated the effects of nitrogen (N) and phosphorus (P) fertilization and mowing on the abundance and community compositions of ammonia-oxidizing Bacteria (AOB) and Archaea (AOA) upon early (May) and peak (August) plant growth using quantitative PCR (qPCR), terminal-restriction fragment length polymorphism (T-RFLP), cloning and sequencing. The results showed that N fertilization changed AOB community composition and increased AOB abundance in both May and August, but significantly decreased AOA abundance in May. By contrast, P fertilization significantly influenced AOB abundance only in August. Mowing significantly decreased AOA abundance and had little effect on AOA community compositions in May, while significantly influencing AOB abundance in both May and August, Moreover, AOA and AOB community structures showed obvious seasonal variations between May and August. Phylogenetic analysis showed that all AOA sequences fell into the Nitrososphaera cluster, and the AOB community was dominated by Nitrosospira Cluster 3. The results suggest that fertilization and mowing play important roles in affecting the abundance and community compositions of AOA and AOB.

Temporal changes of antibiotic-resistance genes and bacterial communities in two contrasting soils treated with cattle manure

The emerging environmental spread of antibiotic-resistance genes (ARGs) and their subsequent acquisition by clinically relevant microorganisms is a major threat to public health. Animal manure has been recognized as an important reservoir of ARGs; however, the dissemination of manure-derived ARGs and the impacts of manure application on the soil resistome remain obscure. Here, we conducted a microcosm study to assess the temporal succession of total bacteria and a broad spectrum of ARGs in two contrasting soils following manure application from cattle that had not been treated with antibiotics. High-capacity quantitative PCR detected 52 unique ARGs across all the samples, with β-lactamase as the most dominant ARG type. Several genes of soil indigenous bacteria conferring resistance to β-lactam, which could not be detected in manure, were found to be highly enriched in manure-treated soils, and the level of enrichment was maintained over the entire course of 140 days. The enriched β-lactam resistance genes had significantly positive relationships with the relative abundance of the integrase intI1 gene, suggesting an increasing mobility potential in manure-treated soils. The changes in ARG patterns were accompanied by a significant effect of cattle manure on the total bacterial community compositions. Our study indicates that even in the absence of selective pressure imposed by agricultural use of antibiotics, manure application could still strongly impact the abundance, diversity and mobility potential of a broad spectrum of soil ARGs. Our findings are important for reliable prediction of ARG behaviors in soil environment and development of appropriate strategies to minimize their dissemination.