Phosphorus (P) is an essential element for all living organisms, playing a critical role in various biological processes. Tracing the origin and cycling of P is vital to understand its fate and transformation as it relates to broader Earth system processes. Unlike other major elements with multiple stable isotopes, P only has one stable isotope (31P), making direct isotopic analysis unfeasible. However, phosphate oxygen isotopes (δ18OP) are a powerful tool to study P across various environments. Measuring phosphate oxygen isotopes requires the collection, purification, and isolation of inorganic phosphate (PO4). Here, we provide a cross-discipline evaluation of the range of pre-processing techniques and best practices required to obtain pure PO4 from diverse environmental matrices for δ18OP analysis. This critical evaluation is intended to increase efficiency and accessibility for new and existing practitioners by serving as a centralized source of information for the most common matrices analyzed for δ18OP, such as bioapatite, water, soil, sediment, plants, and rocks. New opportunities and methodological approaches are also identified to overcome current challenges and improve future applications of δ18OP, thereby informing our understanding of the behavior, fate, and distribution of P in the environment.

Phosphorus (P) is critical to modern biochemical functions and can control ecosystem growth. It was presumably important as a reagent in prebiotic chemistry. However, on the early Earth, P sources may have consisted primarily of poorly soluble calcium phosphates, which may have rendered phosphate as a minimally available nutrient or reagent if these minerals were the sole source. Here, we review aqueous P availability on the early Earth (>2.5 Gyr ago), considering both mineral sources and geochemical sinks relevant to its solvation, and activation by abiotic and biological pathways. Phosphorus on Earth’s early surface would have been present as a mixture of phosphate minerals, as a minor element in silicate minerals, and in reactive, reduced phases from accreted dust, meteorites and asteroids. These P sources would have weathered and plausibly furnished the prebiotic Earth with abundant and potentially reactive P. After the origin of a biosphere, life evolved to draw on not just reactive available P sources, but also insoluble and unreactive sources. The rise of an ecosystem dependent on this element at some point forged a P-limited biosphere, with evolutionary stress forcing the efficient extraction and recycling of P from both abiotic and biotic sources and sinks.

Although lakes dominated by macrophytes are conducive to ecological balance, this balance is easily disrupted by excessive nutrients flowing into the lake. However, knowledge of whether excessive nutrients lead to different microbial environmental vulnerabilities in the lake sediment between macrophyte-dominated areas and macrophyte-free areas is a prerequisite for the implementation of targeted protection measures. In this study, we investigated bacterial communities in sediments using high-throughput sequencing of 16S rRNA genes. Our results showed that the sources of total nitrogen (TN) and organic matter (OM) were related to the macrophytes. The structure, drivers, and interspecific associations of bacterial community, which were more susceptible to increased changes in TN and OM, differed significantly between macrophyte-dominated areas and macrophyte-free areas. More precisely, the lake edge, where was occupied by macrophytes, had a higher proportion of deterministic phylogenetic turnover (88.89%) than other sites, as well as a wider ecological niche and a tighter network structure. Further, as the difference in TN increased, the main assembly processes in surface sediments changed from stochastic to deterministic. However, the majority of phyla from the lake edge showed a greater correlation with excessive nutrients, and the selection of the community by excessive nutrients was more obvious at the edge of the lake. In addition, our results demonstrated that the stability of the bacterial community in macrophyte-free areas is greater than in macrophyte-dominated areas, while an excessively high deterministic process ratio and nutrient (TN and OM) concentration significantly reduced bacterial community stability at macrophyte-dominated areas. Taken together, these results provide a better understanding of the effects of excessive nutrients derived from macrophytes on bacterial community patterns, and highlight the importance of avoiding the accumulation of TN and OM in macrophyte-dominated areas to enhance the sustainability of the ecosystem after restoration of lakes with macrophytes.

Inorganic pyrophosphatase (PPase) is an enzyme that catalyzes the hydrolysis of the phosphoanhydride bond in pyrophosphate (PPi) to release inorganic phosphate (Pi) and simultaneously exchange oxygen isotopes between Pi and water. Here, we quantified the exchange kinetics of oxygen isotopes between five Pi isotopologues (P18O4, P18O316O, P18O216O2, P18O16O3, and P16O4) and water using Raman spectroscopy and 31P nuclear magnetic resonance (NMR) during the PPase-catalyzed 18O-16O isotope exchange reaction in Piwater and PPi-water systems. At a high PPi concentration (300 mM), hydrolysis of PPi by PPase was predominant, and only a small fraction of PPi (???1%) took part in the reversible hydrolysis-condensation reaction (PPi <-> Pi), leading to the oxygen isotope exchange between Pi and water. We demonstrated that Raman and NMR methods can be equally applied for monitoring the kinetics of the oxygen exchange between the Pi isotopologue and water. It was found that the isotope exchange determined by the spectroscopic methods was detectable as low as 0.2% 18O abundance, but the reliability below 1% was much lower. Given that high P concentrations (>= 1 mM) are required in these methods, environmental application of these methods is limited to rare high P conditions in engineered and agricultural environments.

Logging data are measurements of physical properties of the formation surrounding a borehole, acquired in situ after completion of coring (wireline logging) or during drilling (Logging-While-Drilling, LWD). The range of data (resistivity, gamma radiation, velocity, density, borehole images,…) in any hole depends on the scientific objectives and operational constraints.

Logging data are measurements of physical properties of the formation surrounding a borehole, acquired in situ after completion of coring (wireline logging) or during drilling (Logging-While-Drilling, LWD). The range of data (resistivity, gamma radiation, velocity, density, borehole images,…) in any hole depends on the scientific objectives and operational constraints.

Phosphorus (P) is essential for all known forms of life. The oxygen isotopic composition of phosphate can carry a strong imprint from the metabolic processes in Earth's surface environments, including the deep subseafloor biosphere extending to > 1 km beneath the seafloor. Here, we report the O-18/O-16 ratios (delta O-18) of dissolved inorganic phosphate (DIP) to identify different pathways of P cycling in deep-sea sediments sampled up to 200 meters below the seafloor during Ocean Drilling Program (ODP) Leg 201. Our results, along with a diagenetic model, indicate that the delta O-18 of DIP (delta(ODIP)-O-18) is mainly controlled by three pathways of P cycling at Site 1230: (1) release of DIP by extracellular enzymatic degradation of organic matter (i.e., organophosphate), (2) precipitation of authigenic apatite, (3) enzyme-catalyzed oxygen isotopic exchange between phosphate and water. Our diagenetic model quantitatively deconvolves the rates of the three P cycling pathways. In particular, a shift of delta(ODIP)-O-18 towards equilibrium around 140 m below the seafloor corresponds well with a shift in microbial communities, suggesting that delta(ODIP)-O-18 has potential as a proxy for microbial activities in the deep subseafloor biosphere. We also find that the rate of oxygen isotopic exchange can be scaled with the rate of organic matter degradation, suggesting microbially controlled oxygen isotopic exchange. Further, the pattern of variation between authigenic phosphate in sediments and DIP in porewaters bolsters the case that authigenic phosphate (authigenic apatite and Fe-bound phosphate) should reflect the oxygen isotopic composition of contemporaneous porewater DIP and thus can be used to track biogeochemical cycling of P. (C) 2021 Published by Elsevier Ltd.

It has been recently demonstrated that both temperature and intracellular/metabolic water are recorded by the PO4 moieties comprising the backbone of DNA in microbial cells as well as in total microbial biomass PO4 (Blake et al., 2016; Li et al., 2016). Temperature and intracellular water composition are reflected in the O-18/O-16 ratio of PO4 (delta O-18(P)) in DNA (Blake et al., 2016). To determine whether the reported temperature recording by microbial DNA-PO4 could be an artifact of variable microbial growth rate, which may also vary as a function of temperature, three strains of microorganisms having distinct and different growth rate patterns between 23 and 42 degrees C-Pseudomonas fluorescens, Acinetobacter ADP1 and Marinobacter aquaeolei were cultured over a range of temperatures and growth rate patterns. Growth curve patterns for the different strains were distinct and growth rates increased with temperature. However, as we show here, variations in delta O-18(P) values of DNA-PO4 did not correlate with changes in bacterial growth rates at a given temperature, and O isotope fractionations (i.e., partitioning of O isotopes) between PO4 in DNA and O in water were not significantly different between strains (< 1 parts per thousand). Results suggest that for some microbial strains, strain-specific growth rates could cause minor variation in delta O-18(P) values of microbial DNA independent of temperature effects. Thus, we interpret observed trends as strain-specific, intrinsic differences in growth patterns that do not significantly overprint the recording of temperature and intracellular water O isotopic composition by PO4 comprising the backbone of DNA.

This study reports on the compositional diversity of organic compounds in metal(loid)-bearing tailings samples from both active and abandoned tailings ponds. Tailings samples were qualitatively analyzed by comprehensive two-dimensional gas chromatography time-of-flight mass spectrometry (GC × GC-TOFMS). In addition, the priority PAHs (16), PAEs (6), and phenols (2) were quantitatively analyzed using gas chromatography-mass spectrometry (GC-MS). We attribute the presence of some of aromatic organics in studied tailings ponds to particular sources. Mineral floatation reagents are likely the major sources of small-ring aromatics in tailings ponds, and products from metallurgical processing and burning of fossil fuels in the mining area or further afield are also possible contributors and might be the main source of large-ring aromatics. We found that tailings ponds abandoned for decades can still have organics concentrations at levels of concern. Large-ring aromatics are generally more toxic than other contaminants, and these were more abundant in abandoned tailings ponds. This suggests that these large-ring organics do not readily decompose or biodegrade into less toxic byproducts, as do volatiles and many other organic compounds. Our aromatic contaminants database provides an important starting point for researchers to investigate and compare similar contaminants that might be also present in other tailings ponds and emphasizes the necessity of considering their transformations over time.

The ocean is an important source of methane, however, the sources of oceanic methane and mechanisms of its release to the atmosphere have only recently begun to be understood. Recent studies have identified methylphosphonate (MPn) as a previously unknown and likely source of methane in the aerobic ocean (Karl et al., 2008), as well as shown the biosynthesis of methylphosphonic acid to be a widespread trait in marine microbes (Metcalf et al., 2012). The mechanisms and reaction pathways from MPn to free methane, however, have not been well studied. Here we present results of laboratory studies on the photo-degradation of MPn, a likely mechanism of methane release to the atmosphere and phosphate release to the surface oceans. Phosphonoacetic acid was also studied as an additional model compound for comparison. We used the multi-labeled water isotope probing (MLWIP) approach, involving 18O-labeled waters to probe the photolytic mechanism of CP bond cleavage in phosphates through analysis of P released from MPn as PO4. These studies identified distinct reaction pathways involving phosphates compared with other common organophosphorus compounds (e.g., phosphoesters), as well as suggest the involvement of both ambient water and atmospheric oxygen in CP bond cleavage. There is only a small amount of water oxygen incorporated into product PO4 after cleavage of the CP bond in MPn, suggesting atmospheric O2 or radicals formed from O2 under Ultra Violet Radiation (UVR), as the primary source of O that replaces C in the CP bond of MPn. Model calculations suggest that the δ18OP signature of phosphate released via UV-degradation of phosphates is largely (75%) inherited from the original phosphate substrate. This opens up the possibility of tracing and differentiating specific phosphate sources of dissolved phosphate from other organophosphorus (Porg) sources (e.g., phosphoesters) used in primary production, as well as for tracing specific MPn sources of atmospheric methane.

The oxygen isotopic composition of phosphate (δ18OP) has been increasingly used as an effective tracer for the biogeochemical cycling of phosphorus (P) in soils and other environments. However, diverse pretreatment methods (e.g. storage, preparation and extraction) are being used for soil samples. For the uniformity of methods as well as for the comparison of results, it is important to understand if specific treatment methods can compromise original δ18OP values. Here, Ag3PO4 and KH2PO4 were used to test whether a modified Hedley sequential extraction and purification procedure can alter the δ18OP values of phosphate standards. Additionally, to test the effect of sample storage and drying conditions, two types of soils were first processed by using eight different pretreatment methods including sterilizing, storing, drying, and sieving and then the δ18OP values of each soil P pool were measured. Results indicate that the extraction and purification procedure, drying temperature (<0°C to 80°C) and sieving mesh (20 to 100) had no significant effect on the δ18OP values of Pi (inorganic P) pools, but storage at room temperature (without microbial growth inhibitor-HgCl2 added) can lead to significant changes in δ18OP values of almost all P pools. For the two soils studied, the δ18OP values of Pi pools decrease from H2O (H2O-Pi) to NaHCO3 (NaHCO3-Pi), NaOH (NaOH-Pi) and HCl (HCl-Pi), and organic P was also found in the extraction solution of HCl. Furthermore, the δ18OP values calculated from isotope mass balance were different from the measured values suggesting variable extraction of different P pools during single and sequential extraction methods. Collectively these results highlight the need for a unified and standard processing and extraction methods for soil samples to allow meaningful intercomparison of results.

Kinetics of oxygen isotope exchange between dissolved phosphate and water catalyzed by inorganic pyrophosphatase from 3-26 oC S.J. CHANG*, R.E. BLAKE AND A.S. COLMAN Korea Basic Science Institute, Chungbuk 28119, Republic of Korea (*correspondence: sjchang15@kbsi.re.kr) Dept. of Geology & Geophysics, Yale University, New Haven, CT 06511, USA (ruth.blake@yale.edu) Dept. of the Geophyscial Sciences, The University of Chicago, Chicago, IL 60637, USA (asc25@uchicago.edu)

Knowledge of the relative contributions of different water sources to intracellular fluids and body water is important for many fields of study, ranging from animal physiology to paleoclimate. The intracellular fluid environment of cells is challenging to study due to the difficulties of accessing and sampling the contents of intact cells. Previous studies of multicelled organisms, mostly mammals, have estimated body water composition-including metabolic water produced as a byproduct of metabolism-based on indirect measurements of fluids averaged over the whole organism (e.g., blood) combined with modeling calculations. In microbial cells and aquatic organisms, metabolic water is not generally considered to be a significant component of intracellular water, due to the assumed unimpeded diffusion of water across cell membranes. Here we show that the (18)O/(16)O ratio of PO4 in intracellular biomolecules (e.g., DNA) directly reflects the O isotopic composition of intracellular water and thus may serve as a probe allowing direct sampling of the intracellular environment. We present two independent lines of evidence showing a significant contribution of metabolic water to the intracellular water of three environmentally diverse strains of bacteria. Our results indicate that ∼30-40% of O in PO4 comprising DNA/biomass in early stationary phase cells is derived from metabolic water, which bolsters previous results and also further suggests a constant metabolic water value for cells grown under similar conditions. These results suggest that previous studies assuming identical isotopic compositions for intracellular/extracellular water may need to be reconsidered.

Oxygen isotope thermometry has been traditionally based on the ratio of ^18^O/^16^O in oxyanions of minerals such as carbonate (CaCO~3~) in shells/tests, and phosphate in bioapatites (Ca~5~(PO~4~)~3~OH) of marine invertebrates/vertebrates (for example, fish) and mammals. The requirement of mineral biomass, however, has restricted the application of oxygen isotope thermometry to only those organisms possessing biomineral hardparts. This has completely omitted from study, not only organisms lacking hard mineral tissues, but two entire Domains of life: Bacteria and Archaea. Prokaryotic organisms in the domains Bacteria and Archaea comprise the majority of earth9s biodiversity and also inhabit the most extreme environments on earth. This calls for a thermometry based on a more rudimentary component of biomass that is present in all organisms such as DNA. Our previous studies of the ubiquitous intracellular enzyme inorganic pyrophosphatase (PPase), which catalyzes oxygen isotope exchange between dissolved inorganic PO~4~ (P~i~) and water inside of cells, suggest that DNA may contain even more information than the blueprint for life. Here we show that PO~4~ moieties in DNA record the temperature at which life forms and evolves. Our results demonstrate that the ^18^O/^16^O ratio of PO~4~ (δ^18^O~P~) in DNA as well as in bulk biomass, reflects the temperature-dependent exchange of oxygen isotopes between PO~4~ and intracellular water. Thus, δ^18^O~P~ values of DNA may serve as both a new soft-tissue bio-thermometer and probe of intracellular PO~4~ or water during DNA synthesis and cellular growth. Results presented here of the first direct measurements of the oxygen isotopic composition of PO~4~ in microbial DNA, demonstrate systematic variation in δ^18^O~P~ of DNA from several different strains of microorganisms as a function of temperature, and also extend the PO~4~-water O-isotope thermometer to \>70 °C. A composite calibration curve based on several strains of bacteria is presented for DNA-PO~4~--water --temperature relations between 12 and 75 °C and suggests a "universal" DNA-based oxygen isotope thermometry for microorganisms that may further extend to all organisms. Our results open the possibility of connecting temperature with taxonomy and also expand the range of investigations of habitat temperatures and limits to life in extreme and diverse environments, from Earth9s subsurface deep biosphere to Antarctic ice sheets and extraterrestrial systems where life may have originated at extreme temperatures.

Oxygen isotope thermometry has been traditionally based on the ratio of O-18/O-16 in oxyanions of minerals such as carbonate (CaCO3) in shells/tests, and phosphate in bioapatites (Ca-5(PO4)(3)OH) of marine invertebrates/vertebrates (for example, fish) and mammals. The requirement of mineral biomass, however, has restricted the application of oxygen isotope thermometry to only those organisms possessing biomineral hardparts. This has completely omitted from study, not only organisms lacking hard mineral tissues, but two entire Domains of life: Bacteria and Archaea. Prokaryotic organisms in the domains Bacteria and Archaea comprise the majority of earth's biodiversity and also inhabit the most extreme environments on earth. This calls for a thermometry based on a more rudimentary component of biomass that is present in all organisms such as DNA. Our previous studies of the ubiquitous intracellular enzyme inorganic pyrophosphatase (PPase), which catalyzes oxygen isotope exchange between dissolved inorganic PO4 (P-i) and water inside of cells, suggest that DNA may contain even more information than the blueprint for life. Here we show that PO4 moieties in DNA record the temperature at which life forms and evolves. Our results demonstrate that the O-18/O-16 ratio of PO4 (delta O-18(P)) in DNA as well as in bulk biomass, reflects the temperature-dependent exchange of oxygen isotopes between PO4 and intracellular water. Thus, delta O-18(P) values of DNA may serve as both a new soft-tissue bio-thermometer and probe of intracellular PO4 or water during DNA synthesis and cellular growth.Results presented here of the first direct measurements of the oxygen isotopic composition of PO4 in microbial DNA, demonstrate systematic variation in delta O-18(P) of DNA from several different strains of microorganisms as a function of temperature, and also extend the PO4-water O-isotope thermometer to >70 degrees C. A composite calibration curve based on several strains of bacteria is presented for DNA-PO4-water -temperature relations between 12 and 75 degrees C and suggests a "universal" DNA-based oxygen isotope thermometry for microorganisms that may further extend to all organisms. Our results open the possibility of connecting temperature with taxonomy and also expand the range of investigations of habitat temperatures and limits to life in extreme and diverse environments, from Earth's subsurface deep biosphere to Antarctic ice sheets and extraterrestrial systems where life may have originated at extreme temperatures.