Unveiling the culturable and non-culturable actinobacterial diversity in two macroalgae species from the northern Portuguese coast.

Actinomycetota, associated with macroalgae, remains one of the least explored marine niches. The secondary metabolism of Actinomycetota, the primary microbial source of compounds relevant to biotechnology, continues to drive research into the distribution, dynamics, and metabolome of these microorganisms. In this study, we employed a combination of traditional cultivation and metagenomic analysis to investigate the diversity of Actinomycetota in two native macroalgae species from the Portuguese coast. We obtained and taxonomically identified a collection of 380 strains, which were distributed across 12 orders, 15 families, and 25 genera affiliated with the Actinomycetia class, with Streptomyces making up approximately 60% of the composition. Metagenomic results revealed the presence of Actinomycetota in both Chondrus crispus and Codium tomentosum datasets, with relative abundances of 11% and 2%, respectively. This approach identified 12 orders, 16 families, and 17 genera affiliated with Actinomycetota, with minimal overlap with the cultivation results. Acidimicrobiales emerged as the dominant actinobacterial order in both macroalgae, although no strain affiliated with this taxonomic group was successfully isolated. Our findings suggest that macroalgae represent a hotspot for Actinomycetota. The synergistic use of both culture-dependent and independent approaches proved beneficial, enabling the identification and recovery of not only abundant but also rare taxonomic members.

Distinct Assembly Patterns of Soil Antibiotic Resistome Revealed by Land-Use Changes over 30 Years.

Compared with the ever-growing information about the anthropogenic discharge of nutrients, metals, and antibiotics on the disturbance of antibiotic resistance genes (ARGs), less is known about how the potential natural stressors drive the evolutionary processes of antibiotic resistance. This study examined how soil resistomes evolved and differentiated over 30 years in various land use settings with spatiotemporal homogeneity and minimal human impact. We found that the contents of soil organic carbon, nitrogen, soil microbial biomass, and bioavailable heavy metals, as well as related changes in the antibiotic resistome prevalence including diversity and abundance, declined in the order of grassland > cropland > bareland. Sixty-nine remaining ARGs and 14 mobile genetic elements (MGEs) were shared among three land uses. Multiple factors (i.e., soil properties, heavy metals, bacterial community, and MGEs) contributed to the evolutionary changes of the antibiotic resistome, wherein the resistome profile was dominantly driven by MGEs from both direct and indirect pathways, supported by a partial least-squares path model analysis. Our results suggest that pathways to mitigate ARGs in soils can coincide with land degradation processes, posing a challenge to the common goal of managing our environment sustainably.

Selection of rhizosphere communities of diverse rotation crops reveals unique core microbiome associated with reduced banana Fusarium wilt disease.

Crop rotation can assemble distinct core microbiota as functionally specific barriers against the invasion of banana Fusarium oxysporum pathogens. However, the taxonomic identity of rotation-unique core taxa and their legacy effects are poorly understood under field conditions. Pepper and eggplant rotations were employed to reveal rotation crop- and banana-unique antagonistic core taxa by in situ tracking of the soil microbiome assembly patterns for 2 yr. The rotation crop-unique antagonistic taxa were isolated and functionally verified by culture-dependent techniques, high-throughput sequencing, and pot experiments. Pepper and eggplant rotations resulted in eight and one rotation-unique antagonistic core taxa out of 12 507 microbial taxa, respectively. These nine antagonistic taxa were retained the following year and significantly decreased banana wilt disease incidence via legacy effects, although the cultivated strains were exclusively of the genera Bacillus and Pseudomonas. The fermentation broth and volatiles of these two taxa showed strong antagonistic activity, and pot experiments demonstrated high suppression of wilt disease and significant promotion of banana growth. Our study provides a mechanistic understanding of the identification of rotation crop-unique antagonistic taxa and highlights the importance of targeted cultivation of beneficial microorganisms for optimizing crop rotation-based scenarios in support of banana agriculture sustainability.

Revealing the community and metabolic potential of active methanotrophs by targeted metagenomics in the Zoige wetland of the Tibetan Plateau.

The Zoige wetland of the Tibetan Plateau is one of the largest alpine wetlands in the world and a major emission source of methane. Methane oxidation by methanotrophs can counteract the global warming effect of methane released in the wetlands. Understanding methanotroph activity, diversity and metabolism at the molecular level can guide the isolation of the uncultured microorganisms and inform strategy-making decisions and policies to counteract global warming in this unique ecosystem. Here we applied DNA stable isotope probing using 13 C-labelled methane to label the genomes of active methanotrophs, examine the methane oxidation potential and recover metagenome-assembled genomes (MAGs) of active methanotrophs. We found that gammaproteobacteria of type I methanotrophs are responsible for methane oxidation in the wetland. We recovered two phylogenetically novel methanotroph MAGs distantly related to extant Methylobacter and Methylovulum. They belong to type I methanotrophs of gammaproteobacteria, contain both mxaF and xoxF types of methanol dehydrogenase coding genes, and participate in methane oxidation via H4 MPT and RuMP pathways. Overall, the community structure of active methanotrophs and their methanotrophic pathways revealed by DNA-SIP metagenomics and retrieved methanotroph MAGs highlight the importance of methanotrophs in suppressing methane emission in the wetland under the scenario of global warming.

Active Soil Nitrifying Communities Revealed by In Situ Transcriptomics and Microcosm-Based Stable-Isotope Probing.

Long-term nitrogen field fertilization often results in significant changes in nitrifying communities that catalyze a key step in the global N cycle. However, whether microcosm studies are able to inform the dynamic changes in communities of ammonia-oxidizing bacteria (AOB) and archaea (AOA) under field conditions remains poorly understood. This study aimed to evaluate the transcriptional activities of nitrifying communities under in situ conditions, and we found that they were largely similar to those of 13C-labeled nitrifying communities in the urea-amended microcosms of soils that had received different N fertilization regimens for 22 years. High-throughput sequencing of 16S rRNA genes and transcripts suggested that Nitrosospira cluster 3-like AOB and Nitrososphaera viennensis-like AOA were significantly stimulated in N-fertilized fresh soils. Real-time quantitative PCR demonstrated that the significant increase of AOA and AOB in fresh soils upon nitrogen fertilization could be preserved in the air-dried soils. DNA-based stable-isotope probing (SIP) further revealed the greatest labeling of Nitrosospira cluster 3-like AOB and Nitrosospira viennensis-like AOA, despite the strong advantage of AOB over AOA in the N-fertilized soils. Nitrobacter-like nitrite-oxidizing bacteria (NOB) played more important roles than Nitrospira-like NOB in urea-amended SIP microcosms, while the situation was the opposite under field conditions. Our results suggest that long-term fertilization selected for physiologically versatile AOB and AOA that could have been adapted to a wide range of substrate ammonium concentrations. It also provides compelling evidence that the dominant communities of transcriptionally active nitrifiers under field conditions were largely similar to those revealed in 13C-labeled microcosms.IMPORTANCE The role of manipulated microcosms in microbial ecology has been much debated, because they cannot entirely represent the in situ situation. We collected soil samples from 20 field plots, including 5 different treatments with and without nitrogen fertilizers for 22 years, in order to assess active nitrifying communities by in situ transcriptomics and microcosm-based stable-isotope probing. The results showed that chronic N enrichment led to competitive advantages of Nitrosospira cluster 3-like AOB over N. viennensis-like AOA in soils under field conditions. Microcosm labeling revealed similar results for active AOA and AOB, although an apparent discrepancy was observed for nitrite-oxidizing bacteria. This study suggests that the soil microbiome represents a relatively stable community resulting from complex evolutionary processes over a large time scale, and microcosms can serve as powerful tools to test the theory of environmental filtering on the key functional microbial guilds.

Atmospheric Methane Oxidizers Are Dominated by Upland Soil Cluster Alpha in 20 Forest Soils of China.

Upland soil clusters alpha and gamma (USCα and USCγ) are considered a major biological sink of atmospheric methane and are often detected in forest and grassland soils. These clusters are phylogenetically classified using the particulate methane monooxygenase gene pmoA because of the difficulty of cultivation. Recent studies have established a direct link of pmoA genes to 16S rRNA genes based on their isolated strain or draft genomes. However, whether the results of pmoA-based assays could be largely represented by 16S rRNA gene sequencing in upland soils remains unclear. In this study, we collected 20 forest soils across China and compared methane-oxidizing bacterial (MOB) communities by high-throughput sequencing of 16S rRNA and pmoA genes using different primer sets. The results showed that 16S rRNA gene sequencing and the semi-nested polymerase chain reaction (PCR) of the pmoA gene (A189/A682r nested with a mixture of mb661 and A650) consistently revealed the dominance of USCα (accounting for more than 50% of the total MOB) in 12 forest soils. A189f/A682r successfully amplified pmoA genes (mainly RA14 of USCα) in only three forest soils. A189f/mb661 could amplify USCα (mainly JR1) in several forest soils but showed a strong preferential amplification of Methylocystis and many other type I MOB groups. A189f/A650 almost exclusively amplified USCα (mainly JR1) and largely discriminated against Methylocystis and most of the other MOB groups. The semi-nested PCR approach weakened the bias of A189f/mb661 and A189f/A650 for JR1 and balanced the coverage of all USCα members. The canonical correspondence analysis indicated that soil NH4+-N and pH were the main environmental factors affecting the MOB community of Chinese forest soils. The RA14 of the USCα group prefers to live in soils with low pH, low temperature, low elevation, high precipitation, and rich in nitrogen. JR1's preferences for temperature and elevation were opposite to RA14. Our study suggests that combining the deep sequencing of 16S rRNA and pmoA genes to characterize MOB in forest soils is the best choice.

Co-occurrence patterns among prokaryotes across an age gradient in pit mud of Chinese strong-flavor liquor.

It is widely believed that the quality and characteristics of Chinese strong-flavor liquor (CSFL) are closely related to the age of the pit mud; CSFL produced from older pit mud tastes better. This study aimed to investigate the alteration and interaction of prokaryotic communities across an age gradient in pit mud. Prokaryotic microbes in different-aged pit mud (1, 6, and 10 years old) were analyzed by Illumina MiSeq sequencing of the 16S rRNA gene. Analysis of the 16S rRNA gene indicated that the prokaryotic community was significantly altered with pit mud age. There was a significant increase in the genera Methanosarcina, Methanobacterium, and Aminobacterium with increased age of pit mud, while the genus Lactobacillus showed a significant decreasing trend. Network analysis demonstrated that both synergetic co-occurrence and niche competition were dominated by 68 prokaryotic genera. These genera formed 10 hubs of co-occurrence patterns, mainly under the phyla Firmicutes, Euryarchaeota, and Bacteroidetes, playing important roles on ecosystem stability of the pit mud. Environmental variables (pH, NH4+, available P, available K, and Ca2+) correlated significantly with prokaryotic community assembly. The interaction of prokaryotic communities in the pit mud ecosystem and the relationship among prokaryotic communities and environmental factors contribute to the higher quality of the pit mud in older fermentation pits.

Multifunctional Periphytic Biofilms: Polyethylene Degradation and Cd2+ and Pb2+ Bioremediation under High Methane Scenario.

Priority pollutants such as polyethylene (PE) microplastic, lead (Pb2+), and cadmium (Cd2+) have attracted the interest of environmentalists due to their ubiquitous nature and toxicity to all forms of life. In this study, periphytic biofilms (epiphyton and epixylon) were used to bioremediate heavy metals (HMs) and to biodegrade PE under high (120,000 ppm) methane (CH4) doses. Both periphytic biofilms were actively involved in methane oxidation, HMs accumulation and PE degradation. Epiphyton and epixylon both completely removed Pb2+ and Cd2+ at concentrations of 2 mg L-1 and 50 mg L-1, respectively, but only partially removed these HMs at a relatively higher concentration (100 mg L-1). Treatment containing 12% 13CH4 proved to be most effective for biodegradation of PE. A synergistic effect of HMs and PE drastically changed microbial biota and methanotrophic communities. High-throughput 16S rRNA gene sequencing revealed that Cyanobacteria was the most abundant class, followed by Gammaproteobacteria and Alphaproteobacteria in all high-methane-dose treatments. DNA stable-isotope probing was used to label 13C in a methanotrophic community. A biomarker for methane-oxidizing bacteria, pmoA gene sequence of a 13C-labeled fraction, revealed that Methylobacter was most abundant in all high-methane-dose treatments compared to near atmospheric methane (NAM) treatment, followed by Methylococcus. Methylomonas, Methylocystis, Methylosinus, and Methylocella were also found to be increased by high doses of methane compared to NAM treatment. Overall, Cd+2 had a more determinantal effect on methanotrophic activity than Pb2+. Epiphyton proved to be more effective than epixylon in HMs removal and PE biodegradation. The findings proved that both epiphyton and epixylon can be used to bioremediate HMs and biodegrade PE as an efficient ecofriendly technique under high methane concentrations.

Glucose-induced changes in the bacterial communities of mine tailings at different acidification stages.

Ecological restoration technologies applied to tailings can influence the associated bacterial communities. However, it is unknown if the shifts in these bacterial communities are caused by increased organic carbon. Glucose-induced respiration and high-throughput sequencing were used to assess the microbial activity and bacterial communities, respectively. Glucose addition increased the microbial activity, and glucose + ammonium nitrate addition resulted in slightly higher CO2 emission than did glucose addition alone, suggesting that carbon and nitrogen limited microbial community growth. In neutral pH tailings, the bacterial taxa that increased by glucose addition were assigned to the phyla Proteobacteria, Acidobacteria, Actinobacteria, Bacteroidetes, Firmicutes, and Planctomycetes. However, the bacterial taxa that increased by glucose addition in acidic tailings only belonged to the phylum Actinobacteria (maximum increase of 43.78%). In addition, the abundances of the total nitrogen-fixing genera and of the genus Arthrobacter (representing approximately 97.89% of the total nitrogen-fixing genera) increased by glucose addition in acidic tailings (maximum increase of 46.98%). In contrast, the relative abundances of the total iron- and (or) sulfur-oxidizing bacteria decreased (maximum decrease of 10.41%) in response to the addition of glucose. These findings indicate that the addition of organic carbon is beneficial to the development of bacterial communities in mine tailings.

Stable-isotope probing of bacterial community for dissolved inorganic carbon utilization in Microcystis aeruginosa-dominated eutrophic water.

Dissolved inorganic carbon (DIC) is an important source of carbon in aquatic ecosystems, especially under conditions of increased frequency of cyanobacterial bloom. However, the importance of bacteria in direct or indirect utilization of DIC has been widely overlooked in eutrophic freshwater. To identify the functional bacteria that can actively utilize DIC in eutrophic freshwater during cyanobacterial bloom, stable-isotope probing (SIP) experiments were conducted on eutrophic river water with or without inoculation with cyanobacteria (Microcystis aeruginosa). Our 16S rRNA sequencing results revealed the significance of Betaproteobacteria, with similar relative abundance as Alphaproteobacteria, in the active assimilation of H13CO3- into their DNA directly or indirectly, which include autotrophic genera Betaproteobacterial ammonia-oxidizing bacteria. Other bacterial groups containing autotrophic members, e.g. Planctomycetes and Nitrospira, also presented higher abundance among free-living bacteria in water without cyanobacteria. Microcystis aggregates showed a preference for some specific bacterial members that may utilize H13CO3- metabolized by Microcystis as organic matter, e.g. Bacteroidetes (Cytophagales, Sphingobacteriales), and microcystin-degrading bacteria Betaproteobacteria (Paucibacter/Burkholderiaceae). This study provides some valuable information regarding the functional bacteria that can actively utilize DIC in eutrophic freshwater.

Community Structure of Active Aerobic Methanotrophs in Red Mangrove (Kandelia obovata) Soils Under Different Frequency of Tides.

Methanotrophs are important microbial communities in coastal ecosystems. They reduce CH4 emission in situ, which is influenced by soil conditions. This study aimed to understand the differences in active aerobic methanotrophic communities in mangrove forest soils experiencing different inundation frequency, i.e., in soils from tidal mangroves, distributed at lower elevations, and from dwarf mangroves, distributed at higher elevations. Labeling of pmoA gene of active methanotrophs using DNA-based stable isotope probing (DNA-SIP) revealed that methanotrophic activity was higher in the dwarf mangrove soils than in the tidal mangrove soils, possibly because of the more aerobic soil conditions. Methanotrophs affiliated with the cluster deep-sea-5 belonging to type Ib methanotrophs were the most dominant methanotrophs in the fresh mangrove soils, whereas type II methanotrophs also appeared in the fresh dwarf mangrove soils. Furthermore, Methylobacter and Methylosarcina were the most important active methanotrophs in the dwarf mangrove soils, whereas Methylomonas and Methylosarcina were more active in the tidal mangrove soils. High-throughput sequencing of the 16S ribosomal RNA (rRNA) gene also confirmed similar differences in methanotrophic communities at the different locations. However, several unclassified methanotrophic bacteria were found by 16S rRNA MiSeq sequencing in both fresh and incubated mangrove soils, implying that methanotrophic communities in mangrove forests may significantly differ from the methanotrophic communities documented in previous studies. Overall, this study showed the feasibility of 13CH4 DNA-SIP to study the active methanotrophic communities in mangrove forest soils and revealed differences in the methanotrophic community structure between coastal mangrove forests experiencing different tide frequencies.

[Impact of periodical flooding-drying on nitrification and ammonia oxidizers in hydro-fluctuation belt of the Three Gorges Reservoir].

Objective: This study was aimed to elucidate the effect of periodic flooding-drying to ecological processes of ammonia oxidizers in the hydro-fluctuation belt of the Three Gorges Reservoir. Methods: Soil samples were collected at thee altitudes in regions of Wanzhou, Fengdu and Changshou, representing 8, 5 and 0 times floodingdrying management, respectively. Soil physiochemical properties were analyzed and microcosms were constructed to monitor nitrification activity by fertilizing soils with ammonium substrate. Real-time PCR was used to quantify the population size of ammonia-oxidizing archaea (AOA) and bacteria (AOB). DGGE fingerprints and clone libraries were conducted to study the shift of AOA and AOB compositions in nitrifying soils. Results: Among the physiochemical characteristics of the soils, soil organic matter and total phosphates increased along with cycle increasing. After incubation for 13 days, the net nitrification rates of the samples with 8 cycles exceeded those with 5 cycles. The quantities of both AOA and AOB have increased during the incubation. Phylogenetic analysis showed that AOA were placed within the soil group 1.1b and soil group 1.1a, while bacterial ammonia oxidizers were closely related to Nitrosospira and Cluster 0. Conclusion: Periodical flooding-drying increased soil organic matter, enhanced soil nitrification activity and likely played important roles in shaping community structures of soil ammonia oxidizers.

Phylogenetically distinct phylotypes modulate nitrification in a paddy soil.

Paddy fields represent a unique ecosystem in which regular flooding occurs, allowing for rice cultivation. However, the taxonomic identity of the microbial functional guilds that catalyze soil nitrification remains poorly understood. In this study, we provide molecular evidence for distinctly different phylotypes of nitrifying communities in a neutral paddy soil using high-throughput pyrosequencing and DNA-based stable isotope probing (SIP). Following urea addition, the levels of soil nitrate increased significantly, accompanied by an increase in the abundance of the bacterial and archaeal amoA gene in microcosms subjected to SIP (SIP microcosms) during a 56-day incubation period. High-throughput fingerprints of the total 16S rRNA genes in SIP microcosms indicated that nitrification activity positively correlated with the abundance of Nitrosospira-like ammonia-oxidizing bacteria (AOB), soil group 1.1b-like ammonia-oxidizing archaea (AOA), and Nitrospira-like nitrite-oxidizing bacteria (NOB). Pyrosequencing of 13C-labeled DNA further revealed that 13CO2 was assimilated by these functional groups to a much greater extent than by marine group 1.1a-associated AOA and Nitrobacter-like NOB. Phylogenetic analysis demonstrated that active AOB communities were closely affiliated with Nitrosospira sp. strain L115 and the Nitrosospira multiformis lineage and that the 13C-labeled AOA were related to phylogenetically distinct groups, including the moderately thermophilic "Candidatus Nitrososphaera gargensis," uncultured fosmid 29i4, and acidophilic "Candidatus Nitrosotalea devanaterra" lineages. These results suggest that a wide variety of microorganisms were involved in soil nitrification, implying physiological diversification of soil nitrifying communities that are constantly exposed to environmental fluctuations in paddy fields.

Active ammonia oxidizers in an acidic soil are phylogenetically closely related to neutrophilic archaeon.

All cultivated ammonia-oxidizing archaea (AOA) within the Nitrososphaera cluster (former soil group 1.1b) are neutrophilic. Molecular surveys also indicate the existence of Nitrososphaera-like phylotypes in acidic soil, but their ecological roles are poorly understood. In this study, we present molecular evidence for the chemolithoautotrophic growth of Nitrososphaera-like AOA in an acidic soil with pH 4.92 using DNA-based stable isotope probing (SIP). Soil microcosm incubations demonstrated that nitrification was stimulated by urea fertilization and accompanied by a significant increase in the abundance of AOA rather than ammonia-oxidizing bacteria (AOB). Real-time PCR analysis of amoA genes as a function of the buoyant density of the DNA gradient following the ultracentrifugation of the total DNA extracted from SIP microcosms indicated a substantial growth of soil AOA during nitrification. Pyrosequencing of the total 16S rRNA genes in the "heavy" DNA fractions suggested that archaeal communities were labeled to a much greater extent than soil AOB. Acetylene inhibition further showed that (13)CO2 assimilation by nitrifying communities depended solely on ammonia oxidation activity, suggesting a chemolithoautotrophic lifestyle. Phylogenetic analysis of both (13)C-labeled amoA and 16S rRNA genes revealed that most of the active AOA were phylogenetically closely related to the neutrophilic strains Nitrososphaera viennensis EN76 and JG1 within the Nitrososphaera cluster. Our results provide strong evidence for the adaptive growth of Nitrososphaera-like AOA in acidic soil, suggesting a greater metabolic versatility of soil AOA than previously appreciated.

Response of phyllosphere bacterial communities to elevated CO2 during rice growing season.

The phyllosphere, the aerial parts of terrestrial plants, represents the largest biological interface on Earth. This habitat is colonized by diverse microorganisms that affect plant health and growth. However, the community structure of these phyllosphere microorganisms and their responses to environmental changes, such as rising atmospheric CO2, are poorly understood. Using a massive parallel pyrosequencing technique, we investigated the feedback of a phyllosphere bacterial community in rice to elevated CO2 (eCO2) at the tillering, filling, and maturity stages under nitrogen fertilization with low (LN) and high application rates (HN). The results revealed 9,406 distinct operational taxonomic units that could be classified into 8 phyla, 13 classes, 26 orders, 59 families, and 120 genera. The family Enterobacteriaceae within Gammaproteobacteria was the most dominant phylotype during the rice growing season, accounting for 61.0-97.2 % of the total microbial communities. A statistical analysis indicated that the shift in structure and composition of phyllosphere bacterial communities was largely dependent on the rice growing stage. eCO2 showed a distinct effect on the structure of bacterial communities at different growth stages, and the most evident response of the community structure to eCO2 was observed at the filling stage. eCO2 significantly increased the relative abundance of the most dominant phylotype (Enterobacteriaceae) from 88.6 % at aCO2 (ambient CO2) to 97.2 % at eCO2 under LN fertilization at the filling stage, while it significantly decreased the total relative abundance of other phylotypes from 7.48 to 1.35 %. Similarly, higher value for the relative abundance of the most dominant family (Enterobacteriaceae) and lower value for the total relative abundance of other families were observed under eCO2 condition at other growth stages and under different N fertilizations, but the difference was not statistically significant. No consistent response pattern was observed along growth stages that could be attributed to N treatments. These results provide useful insights into our understanding of the response of a phyllosphere bacterial community to eCO2 with regards to the diversity, composition, and structure during rice growing seasons.

[Research progress of atmospheric methane oxidizers in soil].

Microbial oxidation in soil is the only biological sink for atmospheric methane (about 1.8 ppmv). Two groups of atmospheric methane oxidizing bacteria are existed in aerobic soils: obligate and alternative atmospheric methane oxidizing bacteria. The former, such as upland soil cluster alpha (USCalpha) and upland soil cluster gamma (USCgamma), are widely distributed in a variety of aerobic upland soils, and their particulate methane monooxygenase (pMMO) have very high affinity for methane in low concentration. Bacteria in this group are probably genuine oligotrophs. However, so far, there is still no cultivated strain of this group. The latter (Methylocystis/Methylosinus) belongs to traditional methane-oxidizing bacteria, and are widely distributed in soil environments with periodic high methane emission. Most strains of these two genera are known to possess two pMMO isozymes with low and high affinity to methane respectively, and these strains can keep atmospheric methane oxidizing activity for relative long periods ( > 3 months) relying on the high affinity pMMO (pMMO2). However, the growth and reproduction of bacteria in this group are still dependent on endogenous high-concentration methane which is periodically produced within the soils. We reviewed the research progress of these two groups of atmospheric methane-oxidizing bacteria, their possible living strategies, and the effects of several key environmental factors (e. g. soil temperature and moisture, soil pH, vegetation, land use, nitrogen input) on their community composition and methane oxidizing activity. Several important research directions of atmospheric methane oxidizing bacteria have also been proposed.

[Comparative analysis of soil microbial communities by pyrosequencing and DGGE].

OBJECTIVE: We aimed to assess the advantage and disadvantage of next-generation pyrosequencing and traditional Denaturing Gradient Gel Electrophoresis (DGGE) in fingerprinting analysis of soil microbial communities. METHODS: We analyzed microbial compositions, abundance and diversity of typical grassland and forest soils by 16S rRNA gene-based pyrosequencing and DGGE to compare the accuracy and reproducibility of the two techniques on soil microbial communities. RESULTS: For grassland soils, pyrosequencing technique revealed 22 phyla, 54 classes, 60 orders, 131 families and 350 genera; DGGE only detected 6 phyla, 9 classes, 8.orders, 10 families and 10 genera. The results show that DGGE greatly underestimated soil community compositions. Similar results were obtained for forest soils, and the detection sensitivity of pyrosequencing of forest soils was 3.8, 6.7, 6.4, 19.2 and 39.4 times higher than that of DGGE at the taxonomic levels of phylum, class, order, family and genera respectively. Furthermore, DGGE overwhelmingly overestimated the relative abundance of dominant microorganisms represented by the high-intensity bands, leading up to a 2000-fold difference. Both DGGE and pyrosequencing showed consistent results of microbial diversity changing patterns, although the DGGE-based diversity index was much lower than pyrosequencing. CONCLUSION: Pyrosequencing thus provides more comprehensive and accurate fingerprints of soil microbial community structure than DGGE. DGGE only can represent a few numerically dominant phylotypes with apparent overestimation of their relative abundance in soil microbial communities.

[Restoration of microbial ammonia oxidizers in air-dried forest soils upon wetting].

OBJECTIVE: This study was aimed to investigate the abundance and community shift of ammonia-oxidizing archaea (AOA) and bacteria (AOB) in air-dried forest soils in response to water addition, to explore the applicability of air-dried soil for microbial ecology study, and to elucidate whether AOA within the marine group 1. 1a dominate ammonia oxidizers communities in the acidic forest soils in China. METHODS: Soil samples were collected from 10 forest sites of the China Ecosystem Research Network (CERN) and kept under air-drying conditions in 2010. In 2013 the air-dried soil samples were adjusted to 60% of soil maximum water holding capacity for a 28-day incubation at 28 degrees C in darkness. DGGE fingerprinting, clone library construction, pyrosequencing and quantitative PCR of amoA genes were performed to assess community change of ammonia oxidizers in air-dried and re-wetted soils. RESULTS: After incubation for 28 days, the abundance of bacteria and archaea increased significantly, up to 3,230 and 568 times, respectively. AOA increased significantly in 8 samples, and AOB increased significantly in 5 of 10 samples. However, pyrosequencing of amoA genes reveals insignificant changes in composition of AOA and AOB communities. Phylogenetic analysis of amoA genes indicates that archaeal ammonia oxidizers were predominated by AOA within the soil group 1. 1b lineage, while the Nitrosospira-like AOB dominate bacteria ammonia oxidizer communities. There was a significantly positive correlation between AOA/AOB ratio and total nitrogen (r2 = 0.54, P < 0.05), implying that soil ammonia oxidation might be dominated by AOA in association with ammonium released from soil mineralization. CONCLUSION: Phylogenetic analysis suggest that AOA members within the soil group 1. 1b lineage were not restricted to non-acidic soils as previously thought. The abundance rather than composition of AOA and AOB changed in response to water addition. This indicates that air-dried soil could be of help for microbial biogeography study.

[Microbial metabolism in typical flooded paddy soils ].

[OBJECTIVE] The object of this study is to reveal the composition of active microorganism and their metabolic activities in flooded paddy soils with long-term fertilization ( Mineral nitrogen, phosphorus, and potassium, NPK) and without fertilizer (Control check, CK) by environmental transcriptomics. [METHODS] Flooded soil microcosms were incubated in the laboratory for two weeks, then total RNA were extracted from the soil for transcriptome sequencing. Resulting fastq files were uploaded to the Metagenomics Analysis Server (MG-RAST) for taxonomic analysis, gene annotation and function classification. [RESULTS] Transcripts from diverse active microorganism, including bacteria ( > 95% ) , archaea, eukaryotes and viruses, were detected in both flooded paddy soils of CK and NPK treatments. Most of the transcripts (active genes) of bacteria and archaea were derived from Proteobacteria (more than 50% of total bacterial transcripts) and Thaumarchaaeota (about 70% of total archaeal transcripts ) respectively in both treatments. Transcriptional activity of Acidobacteria in NPK treatment paddy soil was significantly higher than that in CK treatment paddy soil. As for other phyla of bacteria and archaea, there were no significant differences of transcriptional activity of them between CK and NPK treatment paddy soils. The highest expressed gene in both CK and NPK treatment paddy soils is ABC transporter encoding gene which related to the transmembrane transport of substances. Based on gene function category of COG (Clusters of Orthologous Genes), Subsystem and KEGG (Kyoto Encyclopedia of Genes and Genomes) database, we found that the main metabolic activities of microorganisms in both CK and NPK treatment paddy soils were related to energy production and conversion, carbohydrate metabolism, protein metabolism and amino acid metabolism, and the dominant KEGG pathways were oxidative phosphorylation and aminoacyl-tRNA biosynthesis. [ CONCLUSION] Composition of active microorganism in CK and NPK treatment paddy soils was generally similar, except Acidobacteria whose transcriptional activity was significantly different between these two treatment paddy soils. It was also very similar between CK and NPK treatment paddy soils considering the metabolic activities of microorganisms in them, for dominant metabolic processes in these two soils were both related to energy obtaining and protein metabolism. So, dominant metabolic activities of microorganism in flooded paddy soils used in this study were not altered significantly under long - term inorganic fertilization.

[Spatial shifts in methanogenic archaeal community composition in eight Chinese paddy soils].

OBJECTIVE: The methanogenic archaea plays many ecological roles in paddy ecosystems such as the regulation of CH4 emissions. However, knowledge gaps exist about the mechanisms of their spatial shift in population. METHODS: We collected 8 latitudinal paddy soil samples from 20.55 degrees N to 47.43 degrees N in China, and studied their methanogenic archaeal community compositions by PCR-DGGE fingerprinting and phylogenetic analyses. Mechanism of spatial shift in community composition was unraveled by canonical correspondence analysis and Venn diagram. RESULTS: Soil pH was one of the main environmental triggers to the composition of methanogenic archaea community in paddy soils. In addition, community composition shifted along latitudinal gradients. CONCLUSION: It is the first report on biogeography of methanogenic archaeal community in arable soils and its environmental driving factors. The information would contribute to understanding spatial shifts in the transformation of organic matters along Chinese latitudinal gradient.