Nonmarine crenarchaeol in Nevada hot springs

Glycerol dialkyl glycerol tetraethers (GDGTs) are core membrane lipids of the Crenarchaeota. The structurally unusual GDGT crenarchaeol has been proposed as a taxonomically specific biomarker for the marine planktonic group I archaea. It is found ubiquitously in the marine water column and in sediments. In this work, samples of microbial community biomass were obtained from several alkaline and neutral-pH hot springs in Nevada, United States. Lipid extracts of these samples were analyzed by high-performance liquid chromatography-mass spectrometry and by gas chromatography-mass spectrometry. Each sample contained GDGTs, and among these compounds was crenarchaeol. The distribution of archaeal lipids in Nevada hot springs did not appear to correlate with temperature, as has been observed in the marine environment. Instead, a significant correlation with the concentration of bicarbonate was observed. Archaeal DNA was analyzed by denaturing gradient gel electrophoresis. All samples contained 16S rRNA gene sequences which were more strongly related to thermophilic crenarchaeota than to Cenarchaeum symbiosum, a marine nonthermophilic crenarchaeon. The occurrence of crenarchaeol in environments containing sequences affiliated with thermophilic crenarchaeota suggests a wide phenotypic distribution of this compound. The results also indicate that crenarchaeol can no longer be considered an exclusive biomarker for marine species.     

Nonmarine Crenarchaeol in Nevada Hot Springs

Glycerol dialkyl glycerol tetraethers (GDGTs) are core membrane lipids of the Crenarchaeota. The structurally unusual GDGT crenarchaeol has been proposed as a taxonomically specific biomarker for the marine planktonic group I archaea. It is found ubiquitously in the marine water column and in sediments. In this work, samples of microbial community biomass were obtained from several alkaline and neutral-pH hot springs in Nevada, United States. Lipid extracts of these samples were analyzed by high-performance liquid chromatography-mass spectrometry and by gas chromatography-mass spectrometry. Each sample contained GDGTs, and among these compounds was crenarchaeol. The distribution of archaeal lipids in Nevada hot springs did not appear to correlate with temperature, as has been observed in the marine environment. Instead, a significant correlation with the concentration of bicarbonate was observed. Archaeal DNA was analyzed by denaturing gradient gel electrophoresis. All samples contained 16S rRNA gene sequences which were more strongly related to thermophilic crenarchaeota than to Cenarchaeum symbiosum, a marine nonthermophilic crenarchaeon. The occurrence of crenarchaeol in environments containing sequences affiliated with thermophilic crenarchaeota suggests a wide phenotypic distribution of this compound. The results also indicate that crenarchaeol can no longer be considered an exclusive biomarker for marine species.     

Distribution of ether lipids and composition of the archaeal community in terrestrial geothermal springs: impact of environmental variables

Archaea can respond to changes in the environment by altering the composition of their membrane lipids, for example, by modification of the abundance and composition of glycerol dialkyl glycerol tetraethers (GDGTs). Here, we investigated the abundance and proportions of polar GDGTs (P‐GDGTs) and core GDGTs (C‐GDGTs) sampled in different seasons from Tengchong hot springs (Yunnan, China), which encompassed a pH range of 2.5–10.1 and a temperature range of 43.7–93.6°C. The phylogenetic composition of the archaeal community (reanalysed from published work) divided the Archaea in spring sediment samples into three major groups that corresponded with spring pH: acidic, circumneutral and alkaline. Cluster analysis showed correlation between spring pH and the composition of P‐ and C‐GDGTs and archaeal 16S rRNA genes, indicating an intimate link between resident Archaea and the distribution of P‐ and C‐GDGTs in Tengchong hot springs. The distribution of GDGTs in Tengchong springs was also significantly affected by temperature; however, the relationship was weaker than with pH. Analysis of published datasets including samples from Tibet, Yellowstone and the US Great Basin hot springs revealed a similar relationship between pH and GDGT content. Specifically, low pH springs had higher concentrations of GDGTs with high numbers of cyclopentyl rings than neutral and alkaline springs, which is consistent with the predominance of high cyclopentyl ring‐characterized Sulfolobales and Thermoplasmatales present in some of the low pH springs. Our study suggests that the resident Archaea in these hot springs are acclimated if not adapted to low pH by their genetic capacity to effect the packing density of their membranes by increasing cyclopentyl rings in GDGTs at the rank of community.     

Fossilization and degradation of archaeal intact polar tetraether lipids in deeply buried marine sediments (Peru Margin)

Glycerol dibiphytanyl glycerol tetraether (GDGT) lipids are part of the cellular membranes of Thaumarchaeota, an archaeal phylum composed of aerobic ammonia oxidizers, and are used in the paleotemperature proxy TEX₈₆. GDGTs in live cells possess polar head groups and are called intact polar lipids (IPL‐GDGTs). Their transformation to core lipids (CL) by cleavage of the head group was assumed to proceed rapidly after cell death, but it has been suggested that some of these IPL‐GDGTs can, just like the CL‐GDGTs, be preserved over geological timescales. Here, we examined IPL‐GDGTs in deeply buried (0.2–186 mbsf, ~2.5 Myr) sediments from the Peru Margin. Direct measurements of the most abundant IPL‐GDGT, IPL‐crenarchaeol, specific for Thaumarchaeota, revealed depth profiles, which differed per head group. Shallow sediments (<1 mbsf) contained IPL‐crenarchaeol with both glycosidic and phosphate head groups, as also observed in thaumarchaeal enrichment cultures, marine suspended particulate matter and marine surface sediments. However, hexose, phosphohexose‐crenarchaeol is not detected anymore below 6 mbsf (~7 kyr), suggesting a high lability. In contrast, IPL‐crenarchaeol with glycosidic head groups is preserved over timescales of Myr. This agrees with previous analyses of deeply buried (>1 m) marine sediments, which only reported glycosidic and no phosphate‐containing IPL‐GDGTs. TEX₈₆ values of CL‐GDGTs did not markedly change with depth, and the TEX₈₆ of IPL‐derived GDGTs decreased only when the proportions of monohexose‐ to dihexose‐GDGTs changed, likely due to the enhanced preservation of the monohexose GDGTs. Our results support the hypothesis that in situ GDGT production and differential IPL degradation in sediments is not substantially affecting TEX₈₆ paleotemperature estimations based on CL–GDGTs and indicates that likely only a small amount of IPL‐GDGTs present in deeply buried sediments is part of cell membranes of active archaea. The amount of archaeal biomass in the deep biosphere based on these IPLs may have been substantially overestimated.     

Archaeal and bacterial glycerol dialkyl glycerol tetraether lipids in hot springs of yellowstone national park

Glycerol dialkyl glycerol tetraethers (GDGTs) are core membrane lipids originally thought to be produced mainly by (hyper)thermophilic archaea. Environmental screening of low-temperature environments showed, however, the abundant presence of structurally diverse GDGTs from both bacterial and archaeal sources. In this study, we examined the occurrences and distribution of GDGTs in hot spring environments in Yellowstone National Park with high temperatures (47 to 83 degrees C) and mostly neutral to alkaline pHs. GDGTs with 0 to 4 cyclopentane moieties were dominant in all samples and are likely derived from both (hyper)thermophilic Crenarchaeota and Euryarchaeota. GDGTs with 4 to 8 cyclopentane moieties, likely derived from the crenarchaeotal order Sulfolobales and the euryarchaeotal order Thermoplasmatales, are usually present in much lower abundance, consistent with the relatively high pH values of the hot springs. The relative abundances of cyclopentane-containing GDGTs did not correlate with in situ temperature and pH, suggesting that other environmental and possibly genetic factors play a role as well. Crenarchaeol, a biomarker thought to be specific for nonthermophilic group I Crenarchaeota, was also found in most hot springs, though in relatively low concentrations, i.e., <5% of total GDGTs. Its abundance did not correlate with temperature, as has been reported previously. Instead, the cooccurrence of relatively abundant nonisoprenoid GDGTs thought to be derived from soil bacteria suggests a predominantly allochthonous source for crenarchaeol in these hot spring environments. Finally, the distribution of bacterial branched GDGTs suggests that they may be derived from the geothermally heated soils surrounding the hot springs.     

A combined lipidomic and 16S rRNA gene amplicon sequencing approach reveals archaeal sources of intact polar lipids in the stratified Black Sea water column

Archaea are important players in marine biogeochemical cycles, and their membrane lipids are useful biomarkers in environmental and geobiological studies. However, many archaeal groups remain uncultured and their lipid composition unknown. Here, we aim to expand the knowledge on archaeal lipid biomarkers and determine the potential sources of those lipids in the water column of the euxinic Black Sea. The archaeal community was evaluated by 16S rRNA gene amplicon sequencing and by quantitative PCR. The archaeal intact polar lipids (IPLs) were investigated by ultra‐high‐pressure liquid chromatography coupled to high‐resolution mass spectrometry. Our study revealed both a complex archaeal community and large changes with water depth in the IPL assemblages. In the oxic/upper suboxic waters (<105 m), the archaeal community was dominated by marine group (MG) I Thaumarchaeota, coinciding with a higher relative abundance of hexose phosphohexose crenarchaeol, a known marker for Thaumarchaeota. In the suboxic waters (80–110 m), MGI Nitrosopumilus sp. dominated and produced predominantly monohexose glycerol dibiphytanyl glycerol tetraethers (GDGTs) and hydroxy‐GDGTs. Two clades of MGII Euryarchaeota were present in the oxic and upper suboxic zones in much lower abundances, preventing the detection of their specific IPLs. In the deep sulfidic waters (>110 m), archaea belonging to the DPANN Woesearchaeota, Bathyarchaeota, and ANME‐1b clades dominated. Correlation analyses suggest that the IPLs GDGT‐0, GDGT‐1, and GDGT‐2 with two phosphatidylglycerol (PG) head groups and archaeol with a PG, phosphatidylethanolamine, and phosphatidylserine head groups were produced by ANME‐1b archaea. Bathyarchaeota represented 55% of the archaea in the deeper part of the euxinic zone and likely produces archaeol with phospho‐dihexose and hexose‐glucuronic acid head groups.     

Archaeal and Bacterial Glycerol Dialkyl Glycerol Tetraether Lipids in Hot Springs of Yellowstone National Park

Glycerol dialkyl glycerol tetraethers (GDGTs) are core membrane lipids originally thought to be produced mainly by (hyper)thermophilic archaea. Environmental screening of low-temperature environments showed, however, the abundant presence of structurally diverse GDGTs from both bacterial and archaeal sources. In this study, we examined the occurrences and distribution of GDGTs in hot spring environments in Yellowstone National Park with high temperatures (47 to 83°C) and mostly neutral to alkaline pHs. GDGTs with 0 to 4 cyclopentane moieties were dominant in all samples and are likely derived from both (hyper)thermophilic Crenarchaeota and Euryarchaeota. GDGTs with 4 to 8 cyclopentane moieties, likely derived from the crenarchaeotal order Sulfolobales and the euryarchaeotal order Thermoplasmatales, are usually present in much lower abundance, consistent with the relatively high pH values of the hot springs. The relative abundances of cyclopentane-containing GDGTs did not correlate with in situ temperature and pH, suggesting that other environmental and possibly genetic factors play a role as well. Crenarchaeol, a biomarker thought to be specific for nonthermophilic group I Crenarchaeota, was also found in most hot springs, though in relatively low concentrations, i.e., <5% of total GDGTs. Its abundance did not correlate with temperature, as has been reported previously. Instead, the cooccurrence of relatively abundant nonisoprenoid GDGTs thought to be derived from soil bacteria suggests a predominantly allochthonous source for crenarchaeol in these hot spring environments. Finally, the distribution of bacterial branched GDGTs suggests that they may be derived from the geothermally heated soils surrounding the hot springs.     

Distribution of intact and core membrane lipids of archaeal glycerol dialkyl glycerol tetraethers among size-fractionated particulate organic matter in hood canal, puget sound

There is great interest in the membrane lipids of archaea (glycerol dialkyl glycerol tetraethers [GDGTs]) as tracers of archaeal biomass because of their utility as paleoproxies and because of the biogeochemical importance of archaea. While core GDGTs (formed by hydrolysis of polar head groups of intact GDGTs after cell death) are appropriate for paleostudies, they have also been used to trace archaeal populations. Also, despite the small size (0.2 by 0.7 μm) of cultivated marine archaea, 0.7-μm glass-fiber filters (GFFs) are typically used to collect GDGTs from natural waters. We quantified both core and intact GDGTs in free-living (0.2- to 0.7-μm), suspended (0.7- to 60-μm), and aggregate (>60-μm) particle size fractions in Puget Sound (Washington State). On average, the free-living fraction contained 36% of total GDGTs, 90% of which were intact. The intermediate-size fraction contained 62% of GDGTs, and 29% of these were intact. The aggregate fraction contained 2% of the total GDGT pool, and 29% of these were intact. Our results demonstrate that intact GDGTs are largely in the free-living fraction. Because only intact GDGTs are present in living cells, protocols that target this size fraction and analyze the intact GDGT pool are necessary to track living populations in marine waters. Core GDGT enrichment in larger-size fractions indicates that archaeal biomass may quickly become attached or entrained in particles once the archaea are dead or dying. While the concentrations of the two pools were generally not correlated, the similar sizes of the core and intact GDGT pools suggest that core GDGTs are removed from the water column on timescales similar to those of cell replication, on timescales of days to weeks.     

In Situ Production of Crenarchaeol in Two California Hot Springs

Crenarchaeol, a membrane-spanning glycerol dialkyl glycerol tetraether (GDGT) containing a cyclohexane moiety in addition to four cyclopentane moieties, was originally hypothesized to be synthesized exclusively by the mesophilic Crenarchaeota. Recent studies reporting the occurrence of crenarchaeol in hot springs and as a membrane constituent of the recently isolated thermophilic crenarchaeote “Candidatus Nitrosocaldus yellowstonii,” however, have raised questions regarding its taxonomic distribution and function. To determine whether crenarchaeol in hot springs is indeed synthesized by members of the Archaea in situ or is of allochthonous origin, we quantified crenarchaeol present in the form of both intact polar lipids (IPLs) and core lipids in sediments of two California hot springs and in nearby soils. IPL-derived crenarchaeol (IPL-crenarchaeol) was found in both hot springs and soils, suggesting in situ production of this GDGT over a wide temperature range (12°C to 89°C). Quantification of archaeal amoA gene abundance by quantitative PCR showed a good correspondence with IPL-crenarchaeol, suggesting that it was indeed derived from living cells and that crenarchaeol-synthesizing members of the Archaea in our samples may also be ammonia oxidizers.Numerous groups of the Archaea synthesize isoprenoid glycerol dialkyl glycerol tetraethers (GDGTs) as a major component of their core membrane lipids, which can contain up to eight cyclopentane moieties (e.g., see reference 7) (Fig. ​(Fig.1).1). An increase in the number of cyclopentane moieties results in denser packing of membrane lipids, allowing for the maintenance of both cellular membrane integrity at high temperatures and stable proton gradients under low-pH conditions (8). This biophysical characteristic is hypothesized to be among those traits essential for the survival and persistence of the Archaea in the “extreme” environments in which they are commonly found (42). GDGTs are synthesized by a large number of cultivated members of the Archaea (see overviews in references 20 and 34), and in nature, they are abundant in hot springs (24, 25, 34, 46), for example, where members of the Archaea are known to thrive at high temperatures and over a wide pH range (3, 21).Open in a separate windowFIG. 1.Structures of GDGTs referred to in the text. “IS,” C₄₆ internal standard.Crenarchaeol is unique among the GDGTs in that it contains a cyclohexane moiety in addition to four cyclopentane moieties (Fig. ​(Fig.1).1). It was first reported in large abundances from Holocene and ancient sediments collected from various marine settings as supporting evidence for the widespread distribution of low-temperature relatives of the hyperthermophilic Archaea (31). It was later proposed that crenarchaeol was synthesized exclusively by marine group I Crenarchaeota (36), a hypothesis further supported by core lipid analysis of the mesophilic marine group I.1a crenarchaeotes “Cenarchaeum symbiosum” (38) and “Candidatus Nitrosopumilus maritimus” SCM1 (30), which showed that both of these organisms synthesize crenarchaeol at moderate temperatures. In addition to this, the apparent absence of crenarchaeol in cultures of (hyper)thermophilic members of the Archaea (see overviews in references 20 and 34) and molecular modeling (8, 37) led to the hypothesis that crenarchaeol decreases lipid density, effectively allowing archaeal membranes composed of membrane-spanning GDGTs to function at mesophilic temperatures (37). Hence, crenarchaeol synthesis was thought to be instrumental in the evolution and radiation of mesophilic Crenarchaeota from thermophilic habitats (17).Recent studies, however, have reported the occurrence of crenarchaeol in hot springs with temperatures of up to 86.5°C (24, 25, 34, 46). That work has been debated to some extent, as there exists the potential for the allochtonous input of fossilized lipid material from weathering of nearby soils where mesophilic Crenarchaeota may thrive: Schouten et al. (34) previously found large relative amounts of specific soil bacterium biomarkers in tandem with crenarchaeol in Yellowstone hot springs. In contrast, Reigstad et al. (28) reported the occurrence of crenarchaeol in the absence of soil-specific biomarkers in Icelandic hot springs. Furthermore, the recently isolated thermophilic crenarchaeote “Candidatus Nitrosocaldus yellowstonii” was shown to synthesize crenarchaeol at a growth temperature of 72°C (6).Core lipids (CLs) that occur in biological membranes generally contain polar head groups such as sugars and phosphates, which are rapidly cleaved upon cell senescence (10, 44). The loss of head groups from intact polar lipids (IPLs) leaves relatively recalcitrant CLs to accumulate in the environment over time as fossil biomarkers. Therefore, depending on the extraction and/or analytical protocols, CLs present in environmental lipid extracts may be derived from both living cells and fossil biomass, including a mixture of both CL-derived GDGTs (CL-GDGTs) and IPL-derived GDGTs (IPL-GDGTs). Most studies of the presence of crenarchaeol in hot springs reported to date have analyzed directly extracted CL-crenarchaeol or CL-crenarchaeol released by the acid hydrolysis of Bligh-Dyer IPL lipid extracts, i.e., without prior separation of CL-GDGTs from IPL-GDGTs (24, 25, 28, 34, 46). In these cases, the reported GDGT distributions represent an integrated signal of both “living” and fossilized material, rendering it impossible to distinguish what proportion (if any) of the observed crenarchaeol was derived from local living archaeal communities. Thus, the in situ production of crenarchaeol in hot springs and its importance relative to that of the in situ production of other archaeal GDGTs remain uncertain.Here we have used a recently described chromatographic method (22, 26) to separately quantify the potential contributions of both in situ-produced and fossilized crenarchaeol (as well as other archaeal GDGTs) in two Californian hot springs and their surrounding soils. In addition, we have quantified the amounts of archaeal amoA and archaeal 16S rRNA gene copies from one site to make quantitative comparisons between gene abundance and IPL-GDGT concentrations.     

Intact polar and core glycerol dibiphytanyl glycerol tetraether lipids of group I.1a and I.1b thaumarchaeota in soil

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.     

Hydrogen production potential of fermentative microorganisms from the Sargasso Sea

Mounting evidence suggests that ammonia-oxidizing archaea (AOA) may play important roles in nitrogen cycling in geothermal environments. In this study, the diversity, distribution and ecological significance of AOA in terrestrial hot springs in Kamchatka (Far East Russia) were explored using amoA genes complemented by analysis of glycerol dialkyl glycerol tetraethers (GDGTs) of archaea. PCR amplification of functional genes (amoA) from AOA and ammonia-oxidizing bacteria (AOB) was performed on microbial mats/streamers and sediments collected from three hot springs (42°C to 87°C and pH 5.5-7.0). No amoA genes of AOB were detected. The amoA genes of AOA formed three distinct phylogenetic clusters with Cluster 3 representing the majority (～59%) of OTUs. Some of the sequences from Cluster 3 were closely related to those from acidic soil environments, which is consistent with the predominance of low pH (<7.0) in these hot springs. Species richness (estimated by Chao1) was more frequently higher at temperatures below 75°C than above it, indicating that AOA may be favored in the moderately high temperature environments. Quantitative PCR of 16S rRNA genes showed that crenarchaeota counted for up to 80% of total archaea. S-LIBSHUFF separated all samples into two phylogenetic groups. The profiles of GDGTs were well separated among the studied springs, suggesting a spatial patterning of archaeal lipid biomarkers. However, this patterning did not correlate significantly with variation in archaeal amoA, suggesting that AOA are not the predominant archaeal group in these springs producing the observed GDGTs.     

Core and intact polar glycerol dibiphytanyl glycerol tetraether lipids of ammonia-oxidizing archaea enriched from marine and estuarine sediments

Glycerol dibiphytanyl glycerol tetraether (GDGT)-based intact membrane lipids are increasingly being used as complements to conventional molecular methods in ecological studies of ammonia-oxidizing archaea (AOA) in the marine environment. However, the few studies that have been done on the detailed lipid structures synthesized by AOA in (enrichment) culture are based on species enriched from nonmarine environments, i.e., a hot spring, an aquarium filter, and a sponge. Here we have analyzed core and intact polar lipid (IPL)-GDGTs synthesized by three newly available AOA enriched directly from marine sediments taken from the San Francisco Bay estuary ("Candidatus Nitrosoarchaeum limnia"), and coastal marine sediments from Svalbard, Norway, and South Korea. Like previously screened AOA, the sedimentary AOA all synthesize crenarchaeol (a GDGT containing a cyclohexane moiety and four cyclopentane moieties) as a major core GDGT, thereby supporting the hypothesis that crenarchaeol is a biomarker lipid for AOA. The IPL headgroups synthesized by sedimentary AOA comprised mainly monohexose, dihexose, phosphohexose, and hexose-phosphohexose moieties. The hexose-phosphohexose headgroup bound to crenarchaeol was common to all enrichments and, in fact, the only IPL common to every AOA enrichment analyzed to date. This apparent specificity, in combination with its inferred lability, suggests that it may be the most suitable biomarker lipid to trace living AOA. GDGTs bound to headgroups with a mass of 180 Da of unknown structure appear to be specific to the marine group I.1a AOA: they were synthesized by all three sedimentary AOA and "Candidatus Nitrosopumilus maritimus"; however, they were absent in the group I.1b AOA "Candidatus Nitrososphaera gargensis."     

Thermophilic archaeon orchestrates temporal expression of GDGT ring synthases in response to temperature and acidity stress

Glycerol dibiphytanyl glycerol tetraethers (GDGTs) are unique archaeal membrane-spanning lipids with 0–8 cyclopentane rings on the biphytanyl chains. The cyclization pattern of GDGTs is affected by many environmental factors, such as temperature and pH, but the underlying molecular mechanism remains elusive. Here, we find that the expression regulation of GDGT ring synthase genes grsA and grsB in thermophilic archaeon Sulfolobus acidocaldarius is temperature- and pH-dependent. Moreover, the presence of functional GrsA protein, or more likely its products cyclic GDGTs rather than the accumulation of GrsA protein itself, is required to induce grsB expression, resulting in temporal regulation of grsA and grsB expression. Our findings establish a molecular model of GDGT cyclization regulated by environment factors in a thermophilic ecosystem, which could be also relevant to that in mesophilic marine archaea. Our study will help better understand the biological basis for GDGT-based paleoclimate proxies. Archaea inhabit a wide range of terrestrial and marine environments. In response to environment fluctuations, archaea modulate their unique membrane GDGTs lipid composition with different strategies, in particular GDGTs cyclization significantly alters membrane permeability. However, the regulation details of archaeal GDGTs cyclization in response to different environmental factor changes remain unknown. We demonstrated, for the first time, thermophilic archaea orchestrate the temporal expression of GDGT ring synthases, leading to delicate control of GDGTs cyclization to respond environmental temperature and acidity stress. Our study provides insight into the regulation of archaea membrane plasticity, and the survival strategy of archaea in fluctuating environments.     

Community Structure of Archaea in the Water Column above Gas Hydrates in the Gulf of Mexico

Microorganisms play fundamental roles in the ecosystem of the Gulf of Mexico (GOM), yet their vertical distributions along the depth continuum of water column are not well known. In this study, we presented the 16S rDNA sequences and lipid profiles in the context of water chemistry to characterize the archaeal community structure above a gas hydrate mound (MC 118) in GOM. Our results showed that all archaeal sequences were related to unknown species of Crenarchaeota or Euryarchaeota. Phylogenetically, group II –β Euryarchaeota dominated the surface water and mid-depth (400-m) water (74% and 58% of total archaeal species, respectively) whereas the marine group I-γ Crenarchaeota dominated the bottom (869 m) water (61% of total archaeal species). Estimates of the Shannon index showed the highest diversity of planktonic Archaea at the 400 m depth. Glycerol dialkyl glycerol tetraether (GDGT) lipids were detected from the 400- and 869-m depths only and characterized by relatively high abundances of GDGT-5 (crenarchaeol) and GDGT-0. Our studies suggested a possible zonation of archaeal community in the water column, which did not seem to be affected by the possible venting of hydrocarbons from the hydrate location in GOM.     

Temperature and pH controls on glycerol dibiphytanyl glycerol tetraether lipid composition in the hyperthermophilic crenarchaeon <Emphasis Type="Italic">Acidilobus sulfurireducens</Emphasis>

Cyclization in glycerol dibiphytanyl glycerol tetraethers (GDGTs) results in internal cyclopentane moieties which are believed to confer thermal stability to crenarchaeal membranes. While the average number of rings per GDGT lipid (ring index) is positively correlated with temperature in many temperate environments, poor correlations are often observed in geothermal environments, suggesting that additional parameters may influence GDGT core lipid composition in these systems. However, the physical and chemical parameters likely to influence GDGT cyclization which are often difficult to decouple in geothermal systems, making it challenging to assess their influence on lipid composition. In the present study, the influence of temperature (range 65–81°C), pH (range 3.0–5.0), and ionic strength (range 10.1–55.7 mM) on GDGT core lipid composition was examined in the hyperthermoacidophile Acidilobus sulfurireducens, a crenarchaeon originally isolated from a geothermal spring in Yellowstone National Park, Wyoming. When cultivated under defined laboratory conditions, the composition of individual and total GDGTs varied significantly with temperature and to a lesser extent with the pH of the growth medium. Ionic strength over the range of values tested did not influence GDGT composition. The GDGT core lipid ring index was positively correlated with temperature and negatively correlated with pH, suggesting that A. sulfurireducens responds to increasing temperature and acidity by increasing the number of cyclopentyl rings in GDGT core membrane lipids.     

Intact membrane lipids of "Candidatus Nitrosopumilus maritimus," a cultivated representative of the cosmopolitan mesophilic group I Crenarchaeota

In this study we analyzed the membrane lipid composition of "Candidatus Nitrosopumilus maritimus," the only cultivated representative of the cosmopolitan group I crenarchaeota and the only mesophilic isolate of the phylum Crenarchaeota. The core lipids of "Ca. Nitrosopumilus maritimus" consisted of glycerol dialkyl glycerol tetraethers (GDGTs) with zero to four cyclopentyl moieties. Crenarchaeol, a unique GDGT containing a cyclohexyl moiety in addition to four cyclopentyl moieties, was the most abundant GDGT. This confirms unambiguously that crenarchaeol is synthesized by species belonging to the group I.1a crenarchaeota. Intact polar lipid analysis revealed that the GDGTs have hexose, dihexose, and/or phosphohexose head groups. Similar polar lipids were previously found in deeply buried sediments from the Peru margin, suggesting that they were in part synthesized by group I crenarchaeota.     

Archaeal and bacterial diversity in hot springs on the Tibetan Plateau, China

The diversity of archaea and bacteria was investigated in ten hot springs (elevation >4600 m above sea level) in Central and Central-Eastern Tibet using 16S rRNA gene phylogenetic analysis. The temperature and pH of these hot springs were 26-81°C and close to neutral, respectively. A total of 959 (415 and 544 for bacteria and archaea, respectively) clone sequences were obtained. Phylogenetic analysis showed that bacteria were more diverse than archaea and that these clone sequences were classified into 82 bacterial and 41 archaeal operational taxonomic units (OTUs), respectively. The retrieved bacterial clones were mainly affiliated with four known groups (i.e., Firmicutes, Proteobacteria, Cyanobacteria, Chloroflexi), which were similar to those in other neutral-pH hot springs at low elevations. In contrast, most of the archaeal clones from the Tibetan hot springs were affiliated with Thaumarchaeota, a newly proposed archaeal phylum. The dominance of Thaumarchaeota in the archaeal community of the Tibetan hot springs appears to be unique, although the exact reasons are not yet known. Statistical analysis showed that diversity indices of both archaea and bacteria were not statistically correlated with temperature, which is consistent with previous studies.     

Energy flux controls tetraether lipid cyclization in Sulfolobus acidocaldarius

Microorganisms regulate the composition of their membranes in response to environmental cues. Many Archaea maintain the fluidity and permeability of their membranes by adjusting the number of cyclic moieties within the cores of their glycerol dibiphytanyl glycerol tetraether (GDGT) lipids. Cyclized GDGTs increase membrane packing and stability, which has been shown to help cells survive shifts in temperature and pH. However, the extent of this cyclization also varies with growth phase and electron acceptor or donor limitation. These observations indicate a relationship between energy metabolism and membrane composition. Here we show that the average degree of GDGT cyclization increases with doubling time in continuous cultures of the thermoacidophile Sulfolobus acidocaldarius (DSM 639). This is consistent with the behavior of a mesoneutrophile, Nitrosopumilus maritimus SCM1. Together, these results demonstrate that archaeal GDGT distributions can shift in response to electron donor flux and energy availability, independent of pH or temperature. Paleoenvironmental reconstructions based on GDGTs thus capture the energy available to microbes, which encompasses fluctuations in temperature and pH, as well as electron donor and acceptor availability. The ability of Archaea to adjust membrane composition and packing may be an important strategy that enables survival during episodes of energy stress.     

Constraints on the Biological Source(s) of the Orphan Branched Tetraether Membrane Lipids

A soil profile from the Saxnäs Mosse peat bog, Sweden, has been analysed for glycerol dialkyl glycerol tetraether (GDGT) membrane lipids and 16S rRNA genes in order to constrain the source of the yet ‘orphan,’ but supposedly bacterial, branched GDGTs. Branched GDGT lipids dominate over archaeal membrane lipids. The Acidobacteria comprise the dominant bacterial group, accounting for the majority of total Bacteria, and are generally more abundant than methanogenic archaea. Analysed acidobacterial strains did not contain branched GDGT lipids. Thus, the source organism must likely be searched for in other acidobacterial phyla or in another abundant group within the remaining bacteria.