Man-Young Jung

Enrichment and Characterization of an Autotrophic Ammonia-Oxidizing Archaeon of Mesophilic Crenarchaeal Group I.1a from an Agricultural Soil

ABSTRACT Soil nitrification is an important process for agricultural productivity and environmental pollution. Though one cultivated representative of ammonia-oxidizing Archaea from soil has been described, additional representatives warrant characterization. We describe an ammonia-oxidizing archaeon (strain MY1) in a highly enriched culture derived from agricultural soil. Fluorescence in situ hybridization microscopy showed that, after 2 years of enrichment, the culture was composed of >90% archaeal cells. Clone libraries of both 16S rRNA and archaeal amoA genes featured a single sequence each. No bacterial amoA genes could be detected by PCR. A [13C]bicarbonate assimilation assay showed stoichiometric incorporation of 13C into Archaea-specific glycerol dialkyl glycerol tetraethers. Strain MY1 falls phylogenetically within crenarchaeal group I.1a; sequence comparisons to “Candidatus Nitrosopumilus maritimus” revealed 96.9% 16S rRNA and 89.2% amoA gene similarities. Completed growth assays showed strain MY1 to be chemoautotrophic, mesophilic (optimum at 25°C), neutrophilic (optimum at pH 6.5 to 7.0), and nonhalophilic (optimum at 0.2 to 0.4% salinity). Kinetic respirometry assays showed that strain MY1s affinities for ammonia and oxygen were much higher than those of ammonia-oxidizing bacteria (AOB). The yield of the greenhouse gas N2O in the strain MY1 culture was lower but comparable to that of soil AOB. We propose that this new soil ammonia-oxidizing archaeon be designated “Candidatus Nitrosoarchaeum koreensis.”

Intact Polar and Core Glycerol Dibiphytanyl Glycerol Tetraether Lipids of Group I.1a and I.1b Thaumarchaeota in Soil

ABSTRACT Ecological studies of thaumarchaeota often apply glycerol dibiphytanyl glycerol tetraether (GDGT)-based intact membrane lipids. However, these components have only been characterized for thaumarchaeota from aquatic environments. Thaumarchaeota have been shown to play an important role in the nitrogen cycle in soil as ammonium oxidizers, and GDGTs are common lipids encountered in soil. We report the core and intact polar lipid (IPL) GDGTs produced by three newly available thaumarchaeota isolated from grassland soil in Austria (“Nitrososphaera viennensis,” group I.1b) and enriched from agricultural soils in South Korea (“Candidatus Nitrosoarchaeum koreensis” MY1, group I.1a; and “Candidatus Nitrososphaera” strain JG1, group I.1b). The soil thaumarchaeota all synthesize crenarchaeol as their major core GDGT, in agreement with the fact that crenarchaeol has also been detected in thaumarchaeota from aquatic environments. The crenarchaeol regioisomer apparently is produced in significant quantities only by soil thaumarchaeota of the I.1b subgroup. In addition, GDGTs with 0 to 4 cyclopentane moieties and GDGTs containing an additional hydroxyl group were detected. The IPL head groups of their membrane lipids comprised mainly monohexose, dihexose, trihexose, phosphohexose, and hexose-phosphohexose moieties. The hexose-phosphohexose head group bound to crenarchaeol occurred in all soil thaumarchaeota, and this IPL is at present the only lipid that is detected in all thaumarchaeota analyzed so far. This specificity and its lability indicate that it is the most suitable biomarker lipid to trace living thaumarchaeota. This study, in combination with previous studies, also suggests that hydroxylated GDGTs occur in the I.1a, but not in the I.1b, subgroup of the thaumarchaeota.

Isotopic signatures of N2O produced by ammonia-oxidizing archaea from soils

N2O gas is involved in global warming and ozone depletion. The major sources of N2O are soil microbial processes. Anthropogenic inputs into the nitrogen cycle have exacerbated these microbial processes, including nitrification. Ammonia-oxidizing archaea (AOA) are major members of the pool of soil ammonia-oxidizing microorganisms. This study investigated the isotopic signatures of N2O produced by soil AOA and associated N2O production processes. All five AOA strains (I.1a, I.1a-associated and I.1b clades of Thaumarchaeota) from soil produced N2O and their yields were comparable to those of ammonia-oxidizing bacteria (AOB). The levels of site preference (SP), δ15Nbulk and δ18O -N2O of soil AOA strains were 13–30%, −13 to −35% and 22–36%, respectively, and strains MY1–3 and other soil AOA strains had distinct isotopic signatures. A 15N-NH4+-labeling experiment indicated that N2O originated from two different production pathways (that is, ammonia oxidation and nitrifier denitrification), which suggests that the isotopic signatures of N2O from AOA may be attributable to the relative contributions of these two processes. The highest N2O production yield and lowest site preference of acidophilic strain CS may be related to enhanced nitrifier denitrification for detoxifying nitrite. Previously, it was not possible to detect N2O from soil AOA because of similarities between its isotopic signatures and those from AOB. Given the predominance of AOA over AOB in most soils, a significant proportion of the total N2O emissions from soil nitrification may be attributable to AOA.

Hydrogen peroxide detoxification is a key mechanism for growth of ammonia-oxidizing archaea

Significance Ammonia-oxidizing archaea (AOA) are major players in global nitrogen cycling, but the AOA carbon-nutrition paradigm is poorly understood. Once considered strict autotrophs, AOA also have been reported to assimilate organic carbon. We used a marine AOA isolate to test hypotheses about the role of fixed carbon in AOA nutrition. Results were confirmed with tests with four additional marine and terrestrial AOA. We discovered that α-keto acids (pyruvate, oxaloacetate) were not directly incorporated into AOA cells. Instead, the α-keto acids functioned as chemical scavengers that detoxified intracellularly produced H2O2 during ammonia oxidation. H2O2 toxicity was also counteracted by co-inoculating the AOA with bacteria harboring catalases. Thus, H2O2 toxicity in AOA may be an evolutionary force controlling AOA communities and global ammonia cycling. Ammonia-oxidizing archaea (AOA), that is, members of the Thaumarchaeota phylum, occur ubiquitously in the environment and are of major significance for global nitrogen cycling. However, controls on cell growth and organic carbon assimilation by AOA are poorly understood. We isolated an ammonia-oxidizing archaeon (designated strain DDS1) from seawater and used this organism to study the physiology of ammonia oxidation. These findings were confirmed using four additional Thaumarchaeota strains from both marine and terrestrial habitats. Ammonia oxidation by strain DDS1 was enhanced in coculture with other bacteria, as well as in artificial seawater media supplemented with α-keto acids (e.g., pyruvate, oxaloacetate). α-Keto acid-enhanced activity of AOA has previously been interpreted as evidence of mixotrophy. However, assays for heterotrophic growth indicated that incorporation of pyruvate into archaeal membrane lipids was negligible. Lipid carbon atoms were, instead, derived from dissolved inorganic carbon, indicating strict autotrophic growth. α-Keto acids spontaneously detoxify H2O2 via a nonenzymatic decarboxylation reaction, suggesting a role of α-keto acids as H2O2 scavengers. Indeed, agents that also scavenge H2O2, such as dimethylthiourea and catalase, replaced the α-keto acid requirement, enhancing growth of strain DDS1. In fact, in the absence of α-keto acids, strain DDS1 and other AOA isolates were shown to endogenously produce H2O2 (up to ∼4.5 μM), which was inhibitory to growth. Genomic analyses indicated catalase genes are largely absent in the AOA. Our results indicate that AOA broadly feature strict autotrophic nutrition and implicate H2O2 as an important factor determining the activity, evolution, and community ecology of AOA ecotypes.

A Mesophilic, Autotrophic, Ammonia-Oxidizing Archaeon of Thaumarchaeal Group I.1a Cultivated from a Deep Oligotrophic Soil Horizon

ABSTRACT Soil nitrification plays an important role in the reduction of soil fertility and in nitrate enrichment of groundwater. Various ammonia-oxidizing archaea (AOA) are considered to be members of the pool of ammonia-oxidizing microorganisms in soil. This study reports the discovery of a chemolithoautotrophic ammonia oxidizer that belongs to a distinct clade of nonmarine thaumarchaeal group I.1a, which is widespread in terrestrial environments. The archaeal strain MY2 was cultivated from a deep oligotrophic soil horizon. The similarity of the 16S rRNA gene sequence of strain MY2 to those of other cultivated group I.1a thaumarchaeota members, i.e., Nitrosopumilus maritimus and “Candidatus Nitrosoarchaeum koreensis,” is 92.9% for both species. Extensive growth assays showed that strain MY2 is chemolithoautotrophic, mesophilic (optimum temperature, 30°C), and neutrophilic (optimum pH, 7 to 7.5). The accumulation of nitrite above 1 mM inhibited ammonia oxidation, while ammonia oxidation itself was not inhibited in the presence of up to 5 mM ammonia. The genome size of strain MY2 was 1.76 Mb, similar to those of N. maritimus and “Ca. Nitrosoarchaeum koreensis,” and the repertoire of genes required for ammonia oxidation and carbon fixation in thaumarchaeal group I.1a was conserved. A high level of representation of conserved orthologous genes for signal transduction and motility in the noncore genome might be implicated in niche adaptation by strain MY2. On the basis of phenotypic, phylogenetic, and genomic characteristics, we propose the name “Candidatus Nitrosotenuis chungbukensis” for the ammonia-oxidizing archaeal strain MY2.

Unveiling abundance and distribution of planktonic Bacteria and Archaea in a polynya in Amundsen Sea, Antarctica.

Polynyas, areas of open water surrounded by sea ice, are sites of intense primary production and ecological hotspots in the Antarctic Ocean. This study determined the spatial variation in communities of prokaryotes in a polynya in the Amundsen Sea using 454 pyrosequencing technology, and the results were compared with biotic and abiotic environmental factors. The bacterial abundance was correlated with that of phytoplankton, Phaeocystis spp. and diatoms. A cluster analysis indicated that the bacterial communities in the surface waters of the polynya were distinct from those under the sea ice. Overall, two bacterial clades, Polaribacter (20-64%) and uncultivated Oceanospirillaceae (7-34%), dominated the surface water in the polynya while the Pelagibacter clade was abundant at all depths (7-42%). The archaeal communities were not as diverse as the bacterial communities in the polynya, and marine group I was dominant (> 80%). Canonical correspondence analysis indicated that the oceanographic properties facilitated the development of distinct prokaryotic assemblages in the polynya. This analysis of the diversity and composition of the psychrophilic prokaryotes associated with high phytoplankton production provides new insights into the roles of prokaryotes in biogeochemical cycles in high-latitude polynyas.

Thioalbus denitrificans gen. nov., sp. nov., a chemolithoautotrophic sulfur-oxidizing gammaproteobacterium, isolated from marine sediment

A mesophilic, facultatively anaerobic, autotrophic bacterium, designated strain Su4(T), was isolated from marine sediment. The isolate was able to utilize reduced sulfur compounds including thiosulfate, tetrathionate, sulfur and sulfide but not sulfite as the energy source. Growth occurred under aerobic and denitrifying chemolithoautotrophic conditions in the presence of thiosulfate as an electron donor and bicarbonate as a carbon source. The G+C content of the genomic DNA was 64.5 mol%. Comparative 16S rRNA gene sequence studies showed that strain Su4(T) was clearly affiliated with the class Gammaproteobacteria. The isolate was Gram-negative-staining and rod-shaped, lacked flagella and grew in artificial seawater medium at 10-40 °C (optimum 28-32 °C) and in 1-5 % (w/v) NaCl (optimum 3 % NaCl). Strain Su4(T) possessed C₁₆:₀, C₁₆:₁ω7c/iso-C₁₅:₀ 2-OH and C₁₈:₁ω7c/ω9t/ω12t as the major fatty acids. On the basis of phenotypic and phylogenetic analysis, the isolate represents a novel species of a novel genus, for which the name Thioalbus denitrificans is proposed. The type strain is Su4(T) ( = KCTC 5699(T)  = JCM 15568(T)).

An uncultivated nitrate-reducing member of the genus Herminiimonas degrades toluene

ABSTRACT Stable isotope probing (SIP) is a cultivation-free methodology that provides information about the identity of microorganisms participating in assimilatory processes in complex communities. In this study, a Herminiimonas-related bacterium was identified as the dominant member of a denitrifying microcosm fed [13C]toluene. The genome of the uncultivated toluene-degrading bacterium was obtained by applying pyrosequencing to the heavy DNA fraction. The draft genome comprised ∼3.8 Mb, in 131 assembled contigs. Metabolic reconstruction of aromatic hydrocarbon (toluene, benzoate, p-cresol, 4-hydroxybenzoate, phenylacetate, and cyclohexane carboxylate) degradation indicated that the bacterium might specialize in anaerobic hydrocarbon degradation. This characteristic is novel for the order Burkholderiales within the class Betaproteobacteria. Under aerobic conditions, the benzoate oxidation gene cluster (BOX) system is likely involved in the degradation of benzoate via benzoyl coenzyme A. Many putative genes for aromatic hydrocarbon degradation were closely related to those in the Rhodocyclaceae (particularly Aromatoleum aromaticum EbN1) with respect to organization and sequence similarity. Putative mobile genetic elements associated with these catabolic genes were highly abundant, suggesting gene acquisition by Herminiimonas via horizontal gene transfer.

Influence of Deglaciation on Microbial Communities in Marine Sediments Off the Coast of Svalbard, Arctic Circle

Increases in global temperatures have been shown to enhance glacier melting in the Arctic region. Here, we have evaluated the effects of meltwater runoff on the microbial communities of coastal marine sediment located along a transect of Temelfjorden, in Svalbard. As close to the glacier front, the sediment properties were clearly influenced by deglaciation. Denaturing gradient gel electrophoresis profiles showed that the sediment microbial communities of the stations of glacier front (stations 188–178) were distinguishable from that of outer fjord region (station 176). Canonical correspondence analysis indicated that total carbon and calcium carbonate in sediment and chlorophyll a in bottom water were key factors driving the change of microbial communities. Analysis of 16S rRNA gene clone libraries suggested that microbial diversity was higher within the glacier–proximal zone (station 188) directly affected by the runoffs than in the outer fjord region. While the crenarchaeotal group I.1a dominated at station 176 (62%), Marine Benthic Group-B and other Crenarchaeota groups were proportionally abundant. With regard to the bacterial community, alpha-Proteobacteria and Flavobacteria lineages prevailed (60%) at station 188, whereas delta-Proteobacteria (largely sulfate-reducers) predominated (32%) at station 176. Considering no clone sequences related to sulfate-reducers, station 188 may be more oxic compared to station 176. The distance-wise compositional variation in the microbial communities is attributable to their adaptations to the sediment environments which are differentially affected by melting glaciers.

Ammonia‐oxidising archaea living at low pH: Insights from comparative genomics

Summary Obligate acidophilic members of the thaumarchaeotal genus Candidatus Nitrosotalea play an important role in nitrification in acidic soils, but their evolutionary and physiological adaptations to acidic environments are still poorly understood, with only a single member of this genus (Ca. N. devanaterra) having its genome sequenced. In this study, we sequenced the genomes of two additional cultured Ca. Nitrosotalea strains, extracted an almost complete Ca. Nitrosotalea metagenome‐assembled genome from an acidic fen, and performed comparative genomics of the four Ca. Nitrosotalea genomes with 19 other archaeal ammonia oxidiser genomes. Average nucleotide and amino acid identities revealed that the four Ca. Nitrosotalea strains represent separate species within the genus. The four Ca. Nitrosotalea genomes contained a core set of 103 orthologous gene families absent from all other ammonia‐oxidizing archaea and, for most of these gene families, expression could be demonstrated in laboratory culture or the environment via proteomic or metatranscriptomic analyses respectively. Phylogenetic analyses indicated that four of these core gene families were acquired by the Ca. Nitrosotalea common ancestor via horizontal gene transfer from acidophilic representatives of Euryarchaeota. We hypothesize that gene exchange with these acidophiles contributed to the competitive success of the Ca. Nitrosotalea lineage in acidic environments.