Archaeal Biomass Research Articles

Integrative Genomics Sheds Light on Global Deep Terrestrial Biosphere Communities

Active microbial lineages inhabit the deep terrestrial subsurface (Fry et al. 1997, Lopez-Fernandez et al. 2018) with estimates suggesting deep subsurface ecosystems to host ca. 90% of the total bacterial and archaeal biomass on Earth and about 10-20% of all terrestrial biomass (McMahon and Parnell 2014, Bar-On et al. 2018, Magnabosco et al. 2018, Drake and Reiners 2021). This highlights deep subsurface ecosystems as a vast reservoir of genetic innovation that remains poorly characterized. The biological diversity of deep subsurface ecosystems seem to be influenced by parameters such as temperature, pressure, water residence times, and geochemistry (Drake and Reiners 2021). The deep subsurface environment remains underrepresented in metagenomic databases, and yet the publicly available datasets are not synoptically analyzed in an integrated way. In this project, we collected a large set of publicly available and published metagenomes, published metagenome assembled genomes (MAGs) as well as single cell amplified genomes (SAGs) originating from global deep oligotrophic terrestrial subsurface samples (>70 m). Reconstructed MAGs/SAGs from this survey were integrated in to our own existing Fennoscandian Shield Genomic Database (FSGD) (Holmfeldt et al. 2021, Mehrshad et al. 2021). This expanded global database of the deep oligotrophic terrestrial subsurface contains >4000 MAGs/SAGs. In this dataset, representatives of phyla Firmicutes, Desulfobacterota, Patescibacteria, and Chlroflexota were the most prevalent taxa. The presence of Candidatus Desulforudis audaxviator across different continents and samples highlighted its adaptation to different deep biosphere environments. In addition, novel bacterial lineages at the class level were identified in several locations, pointing to the importance and necessity of further studies to characterize deep biosphere microbial diversity. This extensive study of the global deep oligotrophic terrestrial subsurface will reveal whether deep terrestrial populations are ubiquitous and how their distribution patterns are affected by different physicochemical variables such as water types and geology. By Identifying key populations present across multiple global groundwaters, our study will advance our understanding of the global diversity hosted in deep oligotrophic terrestrial subsurface ecosystems.

Deterministic Development of Soil Microbial Communities in Disturbed Soils Depends on Microbial Biomass of the Bioinoculum.

Despite its enormous importance for ecosystem services, factors driving microbial recolonization of soils after disturbance are still poorly understood. Here, we compared the microbial recolonization patterns of a disturbed, autoclaved soil using different amounts of the original non-disturbed soil as inoculum. By using this approach, we manipulated microbial biomass, but did not change microbial diversity of the inoculum. We followed the development of a new soil microbiome after reinoculation over a period of 4 weeks using a molecular barcoding approach as well as qPCR. Focus was given on the assessment of bacteria and archaea. We could show that 1 week after inoculation in all inoculated treatments bacterial biomass exceeded the values from the original soil as a consequence of high dissolved organic carbon (DOC) concentrations in the disturbed soil resulting from the disturbance. This high biomass was persistent over the complete experimental period. In line with the high DOC concentrations, in the first 2 weeks of incubation, copiotrophic bacteria dominated the community, which derived from the inoculum used. Only in the disturbed control soils which did not receive a microbial inoculum, recolonization pattern differed. In contrast, archaeal biomass did not recover over the experimental period and recolonization was strongly triggered by amount of inoculated original soil added. Interestingly, the variability between replicates of the same inoculation density decreased with increasing biomass in the inoculum, indicating a deterministic development of soil microbiomes if higher numbers of cells are used for reinoculation.

Diverse Bathyarchaeotal Lineages Dominate Archaeal Communities in the Acidic Dajiuhu Peatland, Central China.

Bathyarchaeota are believed to have roles in the carbon cycle in marine systems. However, the ecological knowledge of Bathyarchaeota is limited in peatland ecosystems. Here, we investigated the vertical distribution of Bathyarchaeota community structure using quantitative PCR and high-throughput sequencing technology of ribosomal 16S rRNA gene integrated with detailed chemical profiling in the Dajiuhu Peatland, central China. Eight archaeal phyla were observed in peat samples, which mainly composed of Bathyarchaeota with a mean relative abundance about 88%, followed by Thaumarchaeota (9%). Bathyarchaeota were further split into 17 subgroups, and some subgroups showed habitat specificity to peat horizons with distinct lithological and physicochemical properties, for example, Bathy-6 and Bathy-15 had preference for the acrotelm, Bathy-5b, Bathy-16, and Bathy-19 were enriched in the catotelm, Bathy-5a, Bathy-8, and Bathy-11 were specific for the clay horizon. This spatial distribution pattern of archaeal communities along peat profile was mainly influenced by water content as indicated by RDA ordination and permutational MANOVA, whereas organic matter content exclusively affected Bathyarchaeota distribution along the peat profile significantly. The abundance of archaeal 16S rRNA genes ranged from 105 to 107 copies per gram dry sediment, and the highest archaeal biomass was observed in the periodically oxic mesotelm horizon with more dynamic archaeal interaction relationship as indicated by the network analysis. Bathyarchaeota dominated the archaeal interaction network with 82% nodes, 96% edges, and 71% keystone species. Our results provide an overview of the archaeal population, community structure, and relationship with environmental factors that affect the vertical distribution of archaeal communities and emphasize the ecology of bathyarchaeotal lineages in terrestrial peatland ecosystems.

Effects of low pH conditions on decay of methanogenic biomass

Acidic failure is relatively common in anaerobic digesters that receive readily biodegradable food wastes at high loading. Under low pH conditions, the activity of methanogenic biomass decreases resulting in complete failure of the digestion process. In this experimental study, we demonstrated that one of the causes for the digester failure under low pH conditions is due to accelerated decay of methanogenic biomass. When enriched acetate degrading methanogens were exposed to a low pH environment (pH = 5.1 with phosphoric acid) in a batch experiment without external substrate, the specific decay rate was observed to increase as much as 10 times of that at pH 7.0. The specific decay rate for formate degrader was also found to increase under low pH conditions whilst the fermentative microorganisms in the cultures appeared to be tolerant to low pH conditions. A Propidium Mono-Azide-quantitative Polymerase Chain Reaction (PMA-qPCR) analysis revealed that the archaeal biomass dominated by methanogens dropped by 71–79% from the initial concentration after 6 days of the acidic batch experiment whilst the bacterial biomass dominating acidogens decreased by only 25%. The decrease in the number of living cells in the batch experiments at different pH was monitored with time to determine a correlation between decay rate and incubation pH.

Recycling of archaeal biomass as a new strategy for extreme life in Dead Sea deep sediments

Archaea and Bacteria that inhabit the deep subsurface (known as the deep biosphere) play a prevalent role in the recycling of sedimentary organic carbon. In such environments, this process can occur over millions of years and requires microbial communities to cope with extremely limited sources of energy. Because of this scarcity, metabolic processes come at a high energetic cost, but the ways heterotrophic microbial communities develop to minimize energy expenses for a maximized yield remain unclear. Here, we report molecular biomarker evidence for the recycling of archaeal cell wall constituents in extreme evaporitic facies of Dead Sea deep sediments. Wax esters derived from the recombination of hydrolyzed products of archaeal membrane lipids were observed in gypsum and/or halite sedimentary deposits down to 243 m below the lake floor, implying the reutilization of archaeal necromass possibly by deep subsurface bacteria. By recycling the building blocks of putatively better-adapted archaea, heterotrophic bacteria may build up intracellular carbon stocks and mitigate osmotic stress in this energy-deprived environment. This mechanism illustrates a new pathway of carbon transformation in the subsurface and demonstrates how life can be maintained in extreme environments characterized by long-term isolation and minimal energetic resources.

Mapping anaerobic sludge bed community adaptations to manure supernatant in biogas reactors

In upflow anaerobic sludge bed (UASB) reactors, biomass present as granules allows for long solids retention time. Here, granules from a process treating pulp and paper industrial wastewater were successfully applied as inoculum in UASB reactors treating pig manure supernatant, despite high particle content and high ammonium concentrations in the influent. We did a detailed characterization of archaeal and bacterial communities associated with the inoculum and with the aggregated and dispersed fractions of the influent and the reactors after one year of operation. The granular communities underwent major changes and adapted to the highly distinct conditions without disintegration of the granules. Although the granules persisted in the reactors, non-granular aggregates accumulated, and partly replaced the granules. Particles introduced to the reactors by the pig manure influent apparently contributed both as food and biofilm growth support. Archaeal communities in the dispersed reactor phase were similar to those dispersed in the influents, implying successful retention and little loss of archaeal biomass due to detachment or disintegration of granules and other aggregates. Unique bacterial communities developed in the dispersed fraction of the reactors despite of low hydraulic retention times. They probably consisted of fast growing organisms consuming readily degradable organic matter.

Understanding microbial ecology can help improve biogas production in AD

454-Pyrosequencing and lipid fingerprinting were used to link anaerobic digestion (AD) process parameters (pH, alkalinity, volatile fatty acids (VFAs), biogas production and methane content) with the reactor microbial community structure and composition. AD microbial communities underwent stress conditions after changes in organic loading rate and digestion substrates. 454-Pyrosequencing analysis showed that, irrespectively of the substrate digested, methane content and pH were always significantly, and positively, correlated with community evenness. In AD, microbial communities with more even distributions of diversity are able to use parallel metabolic pathways and have greater functional stability; hence, they are capable of adapting and responding to disturbances. In all reactors, a decrease in methane content to <30% was always correlated with a 50% increase of Firmicutes sequences (particularly in operational taxonomic units (OTUs) related to Ruminococcaceae and Veillonellaceae). Whereas digesters producing higher methane content (above 60%), contained a high number of sequences related to Synergistetes and unidentified bacterial OTUs. Finally, lipid fingerprinting demonstrated that, under stress, the decrease in archaeal biomass was higher than the bacterial one, and that archaeal Phospholipid etherlipids (PLEL) levels were correlated to reactor performances. These results demonstrate that, across a number of parameters such as lipids, alpha and beta diversity, and OTUs, knowledge of the microbial community structure can be used to predict, monitor, or optimise AD performance.

An archaeal biomarker record of paleoenvironmental change across the onset of the Messinian salinity crisis in the absence of evaporites (Piedmont Basin, Italy)

A sudden change from normal marine to extreme paleoenvironmental conditions occurred approximately 6Ma ago in the Mediterranean Basin at the onset of the late Miocene Messinian salinity crisis, one of the most severe ecological crises in Earth history. Strong evaporation and tectonics led to hypersaline conditions, resulting in widespread deposition of evaporites and the apparent annihilation of the marine metazoan biosphere. In contrast to the prominent occurrence of evaporites elsewhere in the Mediterranean, evaporites did not form in the deeper part of some marginal basins at the onset of the crisis. The strata of the Pollenzo section (Piedmont Basin, NW Italy) studied here were deposited in such a paleogeographic setting. Instead of evaporites, a cyclic succession of organic-rich shales and carbonates formed during the early phase of the crisis in the study area. These sediments record a sharp increase in the contents of archaeal molecular fossils that are mostly represented by isoprenoidal dialkyl glycerol diethers (DGDs) and isoprenoidal glycerol dialkyl glycerol tetraethers (GDGTs). Such an expansion of archaeal biomass is best explained by a change of the archaeal communities at the onset of the Messinian salinity crisis. In particular, the appearance of molecular fossils of extremophilic archaea, mostly producing DGDs (archaeol and extended archaeol), suggests the emergence of halophilic archaea. At the same time, lipids of planktonic Thaumarchaeota, especially crenarchaeol, are present across the entire section, suggesting the local persistence of normal marine conditions. In agreement with the sudden appearance of tetrahymanol in the upper part of the section, the persistence of crenarchaeol indicates the establishment of water column stratification after the advent of the Messinian salinity crisis. To further investigate the Piedmont Basin paleoenvironmental conditions, we test the Archaeol Caldarchaeol Ecometric (ACE), a proxy developed for identifying high paleosalinities in waters and possibly in sediments. Despite high ACE values found for the Messinian salinity crisis samples, these values are in contrast with the absence of any lithological evidence for high salinities as well with other biomarker-independent information, indicating low to normal seawater salinities. This apparent contradiction is likely explained by the complexity of the sources of archaeal lipids, especially of caldarchaeol and archaeol, limiting the utility of the ACE for the reconstruction of paleosalinities for Messinian strata.

Efficacy of methanogenic biomass acclimation in mesophilic anaerobic digestion of ultrasound pretreated sludge

ABSTRACTMethanogenic biomass plays a key role with regard to methane production during anaerobic bioconversion of organic substrates. In this study, the effect of two different acclimated inocula on digestion performances was investigated by means of anaerobic batch tests on untreated and sonicated waste-activated sludge. Organics solubilization and removal kinetics, the abundance and physiological conditions of archaeal cells on ultimate methane yield were evaluated. The simultaneous presence of Methanosarcina and Methanosaeta in the archaeal biomass, the higher initial archaeal cells relative abundance and their occurrence in the aggregated forms were the main factors positively affecting the conversion into methane. The presence of the acclimated inoculum at the start-up influenced positively the methane improvement due to sonication, and the methane-specific production increased from 0.335 ± 0.03 to 0.420 ± 0.05 Nm3/kg VSfed. Moreover, the better physiological state of methanogens permitted to appreciate the effect of hydrolysis improvement by ultrasound pretreatment.

Kinetic analysis of biological degradation for tetramethylammonium hydroxide (TMAH) in the anaerobic activated sludge system at ambient temperature

With rapid development of thin-film transistor liquid crystal display industries, improvements of the wastewater treatments for spent developing fluid (Tetramethylammonium hydroxide (TMAH)) is essential. As an alternative to the conventional aerobic processes, an anaerobic process in ambient temperature composed of a suspended sludge reactor and subsequent two gravity thickeners with addition of flocculants was elaborated. Methylotrophic TMAH-degraders were enriched from an anaerobic digester at a municipal wastewater treatment plant, feeding TMAH as a sole substrate. In the system TMAH was converted to methane via methanol whilst trimethylamine, dimethylamine, monomethylamine and ammonium were sequentially produced with the biomass yield of 0.14 gCOD/gTMAH, maximum specific growth rate at 0.372d⿿1 and specific decay rate at 0.017d⿿1 under 23°C. The system could remove TMAH at 0.37⿿1.2kgTMAH/m3/d with 2000⿿6000mg/L of sludge concentration in the reactor, and achieved low effluent dissolved organics at about 2⿿5mgC/L. A kinetic model of TMAH degradation, including cryptic growth of ordinary acidogens, acetotrophic and hydrogenotrophic methanogens from the decayed TMAH-degraders was built, and validated using a quantitative PCR method. Since the calculated fractions of bacterial and archaeal biomass were consistent with those detected, the kinetic model based on the metabolic pathways could be used as a design platform to maximise the target species and reaction rates in the new system.

Archaeal rhizosphere communities differ between the native and invasive lineages of the wetland plant Phragmites australis (common reed) in a Chesapeake Bay subestuary

Phragmites australis is a common wetland plant species worldwide and best known in North America as a persistent invasive species. Only in recent decades was a native lineage, Phragmites australis subsp. americanus, confirmed in North American wetlands. This study investigated whether the two lineages support unique microbial communities in the rhizosphere. Paired stands of the native and invasive lineages were sampled at four locations across a Chesapeake Bay subestuary to assess differences in soil chemical parameters and the microbial community. Microbial characterization included quantification of genes coding for bacterial and archaeal 16S rRNA, archaeal ammonia-monooxygenase, methane co-enzyme A, and particulate methane-monooxygenase. These gene targets helped quantify bacterial biomass, archaeal biomass, ammonia-oxidizers, methanogens, and methane oxidizers, respectively. Bacterial and archaeal community composition was determined by creating terminal restriction fragment length polymorphism profiles of the 16S rRNA, and bacterial and fungal composition was assessed using phospholipid fatty acid profiles. Bacterial biomass and composition did not differ between the two lineages, but significant differences were observed in archaeal community composition. There were significantly more copies of archaeal 16S rRNA genes in invasive (1.8 × 108 g−1 soil) compared to native (7.2 × 107 g−1 soil) lineages (P = 0.04) and composition profiles were significantly different according to PerMANOVA (P = 0.006). Phospholipid fatty acid profiles also differed between invasive and native lineages (P = 0.023). These findings support other studies that have noted the ability of native and invasive Phragmites to differentially affect soil biogeochemistry and suggest that archaea in the rhizosphere of wetland plants may be more sensitive to plant variation than bacteria.

Towards a better quantitative assessment of the relevance of deep-sea viruses, Bacteria and Archaea in the functioning of the ocean seafloor

The deep-ocean interior contains the majority of microbes present on Earth. Most deep-sea microbes are concentrated in surface sediments, with abundances up to 4 orders of magnitude higher, per unit of volume, than in highly productive waters of the photic zone. To date, it has been shown that prokaryotic biomass largely dominates over all other biotic components, but the relative importance of Bacteria, Archaea and viruses to the global benthic biomass has not yet been quantified. Here, we report that the microbial abundance in the top 50 cm of deep-sea sediments of the world oceans is on the order of 1.5 ± 0.4 × 1029. This is largely represented by viruses (9.8 ± 2.5 × 1028), followed by Bacteria (3.5 ± 0.9 × 1028 cells) and Archaea (1.4 ± 0.4 × 1028 cells). The overall biomass in the top 50 cm of the deep-sea sediments is 1.7 ± 0.4 Pg C, largely represented by bacterial biomass (ca. 78%), followed by archaeal biomass (ca. 21%) and viruses (<1%). The bathymetric patterns of abundance and biomass of the 3 microbial components show differences: abundance and biomass of Bacteria decrease with increasing water depth, whereas those of Archaea and viruses remain constant. These results support the hypothesis that the role of Archaea and viruses could be more relevant in the deepest part of the ocean floor.

High-resolution chemostratigraphy of the late Aptian–early Albian oceanic anoxic event (OAE 1b) from the Poggio le Guaine section (Umbria–Marche Basin, central Italy)

Abstract An integrated study of multiple geochemical proxies (δ13Ccarb, δ13Corg, TOC, HI, CaCO3, trace elements/Al ratios) of the late Aptian–early Albian oceanic anoxic event (OAE) 1b has been performed on the pelagic sedimentary sequence of Poggio le Guaine (Umbria–Marche Basin, central Italy). A comparison of the newly collected stable isotope carbon curve with the records from the Vocontian Basin (SE France), DSDP Site 545 and Hole 1049C provided a reliable and precise identification of the four main prominent black shale levels (113/Jacob, Kilian, Urbino/Paquier and Leenhardt) that characterize the OAE 1b. The studied record shows an increase in the marine organic carbon accumulation rate, in particular in the 113/Jacob and Urbino/Paquier levels. In the other black shales, TOC values are Notably, the Kilian and Urbino/Paquier levels from the PLG section are characterized by the absence of correlative shifts in δ13Ccarb and δ13Corg. The increase in the δ13Corg values in these levels is explained by an increase in the relative contribution of 13C enriched marine planktonic archaeal biomass, while the concomitant negative excursions recorded in the δ13Ccarb could reflect a major contribution of isotopically light terrestrial carbonate ions from increased continental runoff during documented more humid conditions.

The anaerobic treatment of pharmaceutical industry wastewater in an anaerobic batch and upflow packed-bed reactor

The pharmaceutical industry constitutes an important place in terms of health and environment, both in our country and throughout the world. Various complex organic chemicals are present in the wastewater generated by pharmaceutical industries. The accumulation of toxic and persistent organic substances in wastewater is a serious problem for the environment. Therefore, it is important that pharmaceutical industry wastewater is treated before being discharged into receiving waters. Today, anaerobic treatment systems are commonly used for wastewater-containing high organic matter. The first stage of this study statistically investigates the optimization of anaerobic treatment conditions of pharmaceutical industry wastewater in a batch study. In the second stage, continuous treatment processes were planned using data obtained as a result of the batch study. For processing, an upflow anaerobic packed-bed reactor was used for treating substrate mixtures containing 10–100% pharmaceutical industry wastewater. The effects of operating parameters on the chemical oxygen demand (COD) removal efficiency and the methane production rate were evaluated. COD removal efficiencies of 93–97% were obtained for the pharmaceutical industry wastewater using a 2.5–4 d hydraulic retention time and a 0.6–2.2g COD d−1 organic loading rate. The overall results suggested that the mixed bacterial and archaeal biomass was able to efficiently treat pharmaceutical industry wastewater under determined anaerobic conditions.

Quantitative Analysis of Coenzyme F430 in Environmental Samples: A New Diagnostic Tool for Methanogenesis and Anaerobic Methane Oxidation

Coenzyme F430 is a nickel hydrocorphinoid and is the prosthetic group of methyl-coenzyme M reductase that catalyzes the last step of the methanogenic reaction sequence and its reversed reaction for anaerobic methane oxidation by ANME. As such, function-specific compound analysis has the potential to reveal the microbial distribution and activity associated with methane production and consumption in natural environments and, in particular, in deep subsurface sediments where microbiological and geochemical techniques are restricted. Herein, we report the development of a technique for high-sensitivity analysis of F430 in environmental samples, including paddy soils, marine sediments, microbial mats, and an anaerobic groundwater. The lower detection limit of F430 analysis by liquid chromatography/mass spectrometry is 0.1 femto mol, which corresponds to 6 × 10(2) to 1 × 10(4) cells of methanogens. F430 concentrations in these natural environmental samples range from 63 × 10(-6) to 44 nmol g(-1) and are consistent with the methanogenic archaeal biomass estimated by microbiological analyses.

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.

Molecular comparative assessment of the microbial ecosystem in rumen and faeces of goats fed alfalfa hay alone or combined with oats

The objective of this work was to compare the biomass and community structure of bacteria, protozoa and archaea communities in samples of rumen and faeces of goats and to what extent the diet (alfalfa hay with or without supplemented oats) offered to them exert an influence. Four cannulated adult goats fistulated in the rumen were used in a cross over design experiment in two experimental periods of 26 days, consisting in 14 days of adaptation, 7 days of sampling rumen contents and 5 days of digestibility measurement. Bacterial, protozoa and archaeal biomass and the communities' structure was assessed by real time PCR (qPCR) and denaturing gradient gel electrophoresis (DGGE), respectively. The numbers of archaea and bacteria in both rumen and faeces were higher and lower, respectively, in animals fed AH diet (P < 0.005). Contrary, protozoal numbers were not affected by the diet but were lower (P < 0.001) in faeces than in rumen. The analysis of the community structure revealed a consistently different population in structure in rumen and faeces for the three studied microbial groups and that supplementing alfalfa hay with oats led to a decrease in the similarity between sites in the rumen and faeces: similarity indexes for bacteria (57 and 27%), archaea (26 and 9%) and protozoa (62 and 22%) in animals fed AH and AHO diets, respectively.

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.

Distribution of intact and core GDGTs in marine sediments

We conducted a survey of archaeal GDGT (glycerol dibiphytanyl glycerol tetraether) distributions in marine sediments deposited in a range of depositional settings. The focus was comparison of two pools presumed to have distinct geobiological significance, i.e. intact polar GDGTs (IP GDGTs) and core GDGTs (C GDGTs). The former pool has been suggested to be related to living communities of benthic archaea in marine sediments, while the latter is commonly interpreted to consist of molecular fossils from past planktonic archaeal communities that inhabited the surface ocean. Understanding the link between these two pools is important for assessment of the validity of current molecular proxies for sedimentary archaeal biomass and past sea surface temperatures. The relative distributions of GDGTs in the two pools in a core at a CH 4 rich site in the Black Sea provide evidence for in situ production of glycosidic IP GDGTs and their subsequent degradation to corresponding C GDGTs on timescales that are short in geological terms. In addition, we monitored the relationship between the IP GDGT and C GDGT pools in a sample set from various ocean basins with subseafloor depth from a few cm to 320 m and 0 to 4 Myr in age. Notable differences between the two pools can be summarized as follows: the GDGT with acyclic biphytanes, GDGT-0, and its analogues with two and three cyclopentane moieties (GDGT-2 and -3) are generally more abundant in the pool of IP GDGTs, while crenarchaeol tends to be more abundant in the C GDGT pool. Consequently, the ring index is generally higher for the C GDGTs while TEX 86, a molecular proxy ratio not considering the two major GDGTs, tends to be higher in the IP GDGT pool. These differences in the proportion of individual GDGTs in the two pools are probably due to in situ production of IP GDGTs with distributions differing from those of C GDGTs. Despite these differences, we observed significant correlation of these two ratios between the two pools. Specifically, in both pools TEX 86 is high in sediments from warm oceanic regimes and low in cold regimes. We discuss these relationships and suggest that recycling of core GDGTs by benthic archaea is an important mechanism linking both molecular pools.

Changes in intact membrane lipid content of archaeal cells as an indication of metabolic status

Archaea build their cellular membrane with glycerol dialkyl glycerol tetraether (GDGT) core lipids, bound to polar head groups to form intact GDGTs. These lipids are commonly used as indicators of archaeal biomass in the environment. Here, we measured both intact and core GDGTs and archaeal 16S rRNA gene copies (a proxy for cell abundance) in order to determine cellular GDGT content in a coastal fjord at different locations, seasons and depths. We found that the abundance of both intact and core GDGTs relative to 16S rRNA gene copy numbers varied by two orders of magnitude, were generally higher in surface than deeper water and were elevated during periods associated with seasonal phytoplankton blooms. Core and intact GDGTs followed similar trends, but the ratio of core GDGT/16S rRNA gene copy number reached a maximum in October, following a peak in the ratio of intact GDGT/16S rRNA gene copy number that occurred in August. Our data suggest that spatial and temporal variability in these ratios must be considered when attempting to quantify cell numbers in the environment using either GDGTs or 16S rRNA genes. We used optimized extraction methods for both GDGTs and DNA. However, there is still potential for bias in qPCR methodology that could influence the GDGT/16S rRNA gene copy ratio. The apparent variability in cellular membrane lipid concentration could serve to evaluate the status of archaeal populations as it relates to: (i) varying cell size resulting from changes in either community structure or nutritional status, (ii) population turnover times and (iii) changing environmental factors that affect the lifetime of intact GDGTs after cell death.