Energy flux couples sulfur isotope fractionation to proteomic and metabolite profiles in Desulfovibrio vulgaris.

Microbial sulfate reduction is central to the global carbon cycle and the redox evolution of Earth's surface. Tracking the activity of sulfate reducing microorganisms over space and time relies on a nuanced understanding of stable sulfur isotope fractionation in the context of the biochemical machinery of the metabolism. Here, we link the magnitude of stable sulfur isotopic fractionation to proteomic and metabolite profiles under different cellular energetic regimes. When energy availability is limited, cell-specific sulfate respiration rates and net sulfur isotope fractionation inversely covary. Beyond net S isotope fractionation values, we also quantified shifts in protein expression, abundances and isotopic composition of intracellular S metabolites, and lipid structures and lipid/water H isotope fractionation values. These coupled approaches reveal which protein abundances shift directly as a function of energy flux, those that vary minimally, and those that may vary independent of energy flux and likely do not contribute to shifts in S-isotope fractionation. By coupling the bulk S-isotope observations with quantitative proteomics, we provide novel constraints for metabolic isotope models. Together, these results lay the foundation for more predictive metabolic fractionation models, alongside interpretations of environmental sulfur and sulfate reducer lipid-H isotope data.

Sulfur isotope-based source apportionment and control mechanisms of PM2.5 sulfate in Seoul, South Korea during winter and early spring (2017-2020).

High level of particulate matter (PM) concentrations are a major environmental concern in Seoul, South Korea, especially during winter and early spring. Sulfate is a major component of PM and induces severe environmental pollution, such as acid precipitation. Previous studies have used numerical models to constrain the relative contributions of domestic and trans-boundary sources to PM2.5 sulfate concentration in South Korea. Because of the scarce measurement result of δ34S for PM2.5 sulfate in South Korea, poorly defined δ34S value of domestic sulfur sources, and no application of sulfur isotope fractionation during sulfate formation in previous observation-based studies, source apportionment results conducted by model studies have not been corroborated from independent chemical observations. Here, we examined the δ34S of PM2.5 in Seoul and domestic sulfur sources, and considered the sulfur isotope fractionation for accurate source apportionment constraint. Accordingly, domestic and trans-boundary sulfur sources accounted for approximately (16-32) % and (68-84) % of the sulfate aerosols in Seoul, respectively, throughout the winter and early spring of 2017-2020. Air masses passing through north-eastern China had relatively low sulfate concentrations, enriched δ34S, and a low domestic source contribution. Those passing through south-eastern China had relatively a high sulfate concentrations, depleted δ34S, and high domestic source contribution. Furthermore, elevated PM2.5 sulfate concentrations (>10 µg m-3) were exclusively associated with a weak westerly wind speed of <3 m s-1. From December 2019 to March 2020, Seoul experienced relatively low levels of PM2.5 sulfate, which might be attributed to favorable weather conditions rather than the effects of COVID-19 containment measures. Our results demonstrate the potential use of δ34S for accurate source apportionment and for identifying the crucial role of regional air mass transport and meteorological conditions in PM2.5 sulfate concentration. Furthermore, the data provided can be essential for relevant studies and policy-making in East Asia.

What Controls the Sulfur Isotope Fractionation during Dissimilatory Sulfate Reduction?

Sulfate often behaves conservatively in the oxygenated environments but serves as an electron acceptor for microbial respiration in a wide range of natural and engineered systems where oxygen is depleted. As a ubiquitous anaerobic dissimilatory pathway, therefore, microbial reduction of sulfate to sulfide has been of continuing interest in the field of microbiology, ecology, biochemistry, and geochemistry. Stable isotopes of sulfur are an effective tool for tracking this catabolic process as microorganisms discriminate strongly against heavy isotopes when cleaving the sulfur-oxygen bond. Along with its high preservation potential in environmental archives, a wide variation in the sulfur isotope effects can provide insights into the physiology of sulfate reducing microorganisms across temporal and spatial barriers. A vast array of parameters, including phylogeny, temperature, respiration rate, and availability of sulfate, electron donor, and other essential nutrients, has been explored as a possible determinant of the magnitude of isotope fractionation, and there is now a broad consensus that the relative availability of sulfate and electron donors primarily controls the magnitude of fractionation. As the ratio shifts toward sulfate, the sulfur isotope fractionation increases. The results of conceptual models, centered on the reversibility of each enzymatic step in the dissimilatory sulfate reduction pathway, are in qualitative agreement with the observations, although the underlying intracellular mechanisms that translate the external stimuli into the isotopic phenotype remain largely unexplored experimentally. This minireview offers a snapshot of our current understanding of the sulfur isotope effects during dissimilatory sulfate reduction as well as their potential quantitative applications. It emphasizes the importance of sulfate respiration as a model system for the isotopic investigation of other respiratory pathways that utilize oxyanions as terminal electron acceptors.

A Simple Bacteremia Score for Predicting Bacteremia in Patients with Suspected Infection in the Emergency Department: A Cohort Study.

Bacteremia is a life-threatening condition that has increased in prevalence over the past two decades. Prompt recognition of bacteremia is important; however, identification of bacteremia requires 1 to 2 days. This retrospective cohort study, conducted from 10 November 2014 to November 2019, among patients with suspected infection who visited the emergency department (ED), aimed to develop and validate a simple tool for predicting bacteremia. The study population was randomly divided into derivation and development cohorts. Predictors of bacteremia based on the literature and logistic regression were assessed. A weighted value was assigned to predictors to develop a prediction model for bacteremia using the derivation cohort; discrimination was then assessed using the area under the receiver operating characteristic curve (AUC). Among the 22,519 patients enrolled, 18,015 were assigned to the derivation group and 4504 to the validation group. Sixteen candidate variables were selected, and all sixteen were used as significant predictors of bacteremia (model 1). Among the sixteen variables, the top five with higher odds ratio, including procalcitonin, neutrophil-lymphocyte ratio (NLR), lactate level, platelet count, and body temperature, were used for the simple bacteremia score (model 2). The proportion of bacteremia increased according to the simple bacteremia score in both cohorts. The AUC for model 1 was 0.805 (95% confidence interval [CI] 0.785-0.824) and model 2 was 0.791 (95% CI 0.772-0.810). The simple bacteremia prediction score using only five variables demonstrated a comparable performance with the model including sixteen variables using all laboratory results and vital signs. This simple score is useful for predicting bacteremia-assisted clinical decisions.

A Cristae-Like Microcompartment in Desulfobacterota.

Some Alphaproteobacteria contain intracytoplasmic membranes (ICMs) and proteins homologous to those responsible for the mitochondrial cristae, an observation which has given rise to the hypothesis that the Alphaproteobacteria endosymbiont had already evolved cristae-like structures and functions. However, our knowledge of microbial fine structure is still limited, leaving open the possibility of structurally homologous ICMs outside the Alphaproteobacteria. Here, we report on the detailed characterization of lamellar cristae-like ICMs in environmental sulfate-reducing Desulfobacterota that form syntrophic partnerships with anaerobic methane-oxidizing (ANME) archaea. These structures are junction-bound to the cytoplasmic membrane and resemble the form seen in the lamellar cristae of opisthokont mitochondria. Extending these observations, we also characterized similar structures in Desulfovibrio carbinolicus, a close relative of the magnetotactic D. magneticus, which does not contain magnetosomes. Despite a remarkable structural similarity, the key proteins involved in cristae formation have not yet been identified in Desulfobacterota, suggesting that an analogous, but not a homologous, protein organization system developed during the evolution of some members of Desulfobacterota. IMPORTANCE Working with anaerobic consortia of methane oxidizing ANME archaea and their sulfate-reducing bacterial partners recovered from deep sea sediments and with the related sulfate-reducing bacterial isolate D. carbinolicus, we discovered that their intracytoplasmic membranes (ICMs) appear remarkably similar to lamellar cristae. Three-dimensional electron microscopy allowed for the novel analysis of the nanoscale attachment of ICMs to the cytoplasmic membrane, and these ICMs are structurally nearly identical to the crista junction architecture seen in metazoan mitochondria. However, the core junction-forming proteins must be different. The outer membrane vesicles were observed to bud from syntrophic Desulfobacterota, and darkly stained granules were prominent in both Desulfobacterota and D. carbinolicus. These findings expand the taxonomic breadth of ICM-producing microorganisms and add to our understanding of three-dimensional microbial fine structure in environmental microorganisms.

Extracorporeal Life-support for Out-of-hospital Cardiac Arrest: A Nationwide Multicenter Study.

BACKGROUND: Despite potential clinical roles of extracorporeal life support (ECLS) for out-of-hospital cardiac arrest (OHCA) compared to that of conventional cardiopulmonary resuscitation (CCPR), use of ECLS for OHCA is not strongly endorsed by current clinical guidelines. OBJECTIVE: The purpose of this study is to investigate the clinical roles of extracorporeal life support (ECLS) compared with that of conventional cardiopulmonary resuscitation (CCPR) for out-of-hospital cardiac arrest (OHCA) patients. METHODS: The outcomes of OHCA between 2015 and 2020, enrolled in the Korean Cardiac Arrest Research Consortium (KoCARC), a multicenter OHCA patient registry including 65 participating hospitals throughout the Republic of Korea (ClinicalTrials.gov, number NCT03222999). Differences in clinical features were adjusted by matching the propensity for ECLS. The primary outcome was 30-day neurologically favorable survival with cerebral performance category of 1 or 2. Restricted mean survival time (RMST) was used to compare outcomes between groups. RESULTS: Of 12,006 patients included, ECLS was applied to 272 patients (2.2%). The frequency of neurologically favorable survival was higher in the ECLS group than the CCPR group (RMST difference, 5.5 days [95% CI, 4.1-7.0 days], P < 0.001). In propensity score-matched 271 pairs, the clinical outcome of ECLS and CCPR did not differ to a statistically significant extent (RMST difference, 0.4 days [95% CI -1.6 to 2.5 days], P = 0.67). Subgroup analyses revealed that the clinical roles of ECLS was evident in patients with nonshockable rhythm or CPR time ≥20 min (RMST difference, 2.7 days [95% CI 0.5-4.8 days], P = 0.015), but not in patients without these features (RMST difference, -3.7 days [95% CI -7.6 to 0.2 days], P = 0.07). CONCLUSIONS: In this real-world data analysis, ECLS compared to CCPR did not result in better overall clinical outcomes of OHCA. The clinical efficacy of ECLS may be limited to a subgroup of high-risk patients.

Cardiac troponin I and the risk of cardiovascular or non-cardiovascular death in patients visiting the emergency department.

The prognostic implication of cardiac troponin I (cTnI) values for the determination of the magnitude or duration of cause-specific death risk is limited. We included consecutive patients with maximal cTnI values within 24 h of their emergency department visits. Multivariate analyses using variables selected by the Bayesian information criterion were performed to investigate the impact of cTnI on the event rate, time-dependent risk, and dose-dependent risk of cardiovascular or non-cardiovascular death within 360 days. There were 5472 (14.9%) all-cause deaths including 881 (2.4%) cardiovascular deaths and 4591 (12.5%) non-cardiovascular deaths. In patients with positive cTnI, defined as the ≥ 99th percentile of the upper normal limit, the cumulative risk of cardiac and non-cardiac death was 4.4- and 1.4-fold higher, respectively, than that of negative cTnI, respectively. In the competing risk analysis, positive cTnI was linked to 2.4- and 1.2-fold higher risks of cardiovascular and non-cardiovascular death, respectively. The cTnI value showed a positive relationship with the risk of both cardiovascular and non-cardiovascular deaths. In the time-dependent risk analysis, the excess risk of cardiovascular death was mostly evident in the first few weeks. Higher cTnI value was associated with an increased risk of both cardiovascular and non-cardiovascular death, especially which was in the early period.

Physiological, genomic, and sulfur isotopic characterization of methanol metabolism by Desulfovibrio carbinolicus.

Methanol is often considered as a non-competitive substrate for methanogenic archaea, but an increasing number of sulfate-reducing microorganisms (SRMs) have been reported to be capable of respiring with methanol as an electron donor. A better understanding of the fate of methanol in natural or artificial anaerobic systems thus requires knowledge of the methanol dissimilation by SRMs. In this study, we describe the growth kinetics and sulfur isotope effects of Desulfovibrio carbinolicus, a methanol-oxidizing sulfate-reducing deltaproteobacterium, together with its genome sequence and annotation. D. carbinolicus can grow with a series of alcohols from methanol to butanol. Compared to longer-chain alcohols, however, specific growth and respiration rates decrease by several fold with methanol as an electron donor. Larger sulfur isotope fractionation accompanies slowed growth kinetics, indicating low chemical potential at terminal reductive steps of respiration. In a medium containing both ethanol and methanol, D. carbinolicus does not consume methanol even after the cessation of growth on ethanol. Among the two known methanol dissimilatory systems, the genome of D. carbinolicus contains the genes coding for alcohol dehydrogenase but lacks enzymes analogous to methanol methyltransferase. We analyzed the genomes of 52 additional species of sulfate-reducing bacteria that have been tested for methanol oxidation. There is no apparent relationship between phylogeny and methanol metabolizing capacity, but most gram-negative methanol oxidizers grow poorly, and none carry homologs for methyltransferase (mtaB). Although the amount of available data is limited, it is notable that more than half of the known gram-positive methanol oxidizers have both enzymatic systems, showing enhanced growth relative to the SRMs containing only alcohol dehydrogenase genes. Thus, physiological, genomic, and sulfur isotopic results suggest that D. carbinolicus and close relatives have the ability to metabolize methanol but likely play a limited role in methanol degradation in most natural environments.

Role of APS reductase in biogeochemical sulfur isotope fractionation.

Sulfur isotope fractionation resulting from microbial sulfate reduction (MSR) provides some of the earliest evidence of life, and secular variations in fractionation values reflect changes in biogeochemical cycles. Here we determine the sulfur isotope effect of the enzyme adenosine phosphosulfate reductase (Apr), which is present in all known organisms conducting MSR and catalyzes the first reductive step in the pathway and reinterpret the sedimentary sulfur isotope record over geological time. Small fractionations may be attributed to low sulfate concentrations and/or high respiration rates, whereas fractionations greater than that of Apr require a low chemical potential at that metabolic step. Since Archean sediments lack fractionation exceeding the Apr value of 20‰, they are indicative of sulfate reducers having had access to ample electron donors to drive their metabolisms. Large fractionations in post-Archean sediments are congruent with a decline of favorable electron donors as aerobic and other high potential metabolic competitors evolved.

Fractionation of sulfur isotopes by Desulfovibrio vulgaris mutants lacking hydrogenases or type I tetraheme cytochrome c 3.

The sulfur isotope effect produced by sulfate reducing microbes is commonly used to trace biogeochemical cycles of sulfur and carbon in aquatic and sedimentary environments. To test the contribution of intracellular coupling between carbon and sulfur metabolisms to the overall magnitude of the sulfur isotope effect, this study compared sulfur isotope fractionations by mutants of Desulfovibrio vulgaris Hildenborough. We tested mutant strains lacking one or two periplasmic (Hyd, Hyn-1, Hyn-2, and Hys) or cytoplasmic hydrogenases (Ech and CooL), and a mutant lacking type I tetraheme cytochrome (TpI-c 3). In batch culture, wild-type D. vulgaris and its hydrogenase mutants had comparable growth kinetics and produced the same sulfur isotope effects. This is consistent with the reported redundancy of hydrogenases in D. vulgaris. However, the TpI-c 3 mutant (ΔcycA) exhibited slower growth and sulfate reduction rates in batch culture, and produced more H2 and an approximately 50% larger sulfur isotope effect, compared to the wild type. The magnitude of sulfur isotope fractionation in the CycA deletion strain, thus, increased due to the disrupted coupling of the carbon oxidation and sulfate reduction pathways. In continuous culture, wild-type D. vulgaris and the CycA mutant produced similar sulfur isotope effects, underscoring the influence of environmental conditions on the relative contribution of hydrogen cycling to the electron transport. The large sulfur isotope effects associated with the non-ideal stoichiometry of sulfate reduction in this study imply that simultaneous fermentation and sulfate reduction may be responsible for some of the large naturally-occurring sulfur isotope effects. Overall, mutant strains provide a powerful tool to test the effect of specific redox proteins and pathways on sulfur isotope fractionation.

Effects of iron and nitrogen limitation on sulfur isotope fractionation during microbial sulfate reduction.

Sulfate-reducing microbes utilize sulfate as an electron acceptor and produce sulfide that is depleted in heavy isotopes of sulfur relative to sulfate. Thus, the distribution of sulfur isotopes in sediments can trace microbial sulfate reduction (MSR), and it also has the potential to reflect the physiology of sulfate-reducing microbes. This study investigates the relationship between the availability of iron and reduced nitrogen and the magnitude of S-isotope fractionation during MSR by a marine sulfate-reducing bacterium, DMSS-1, a Desulfovibrio species, isolated from salt marsh in Cape Cod, MA. Submicromolar levels of iron increase sulfur isotope fractionation by about 50% relative to iron-replete cultures of DMSS-1. Iron-limited cultures also exhibit decreased cytochrome c-to-total protein ratios and cell-specific sulfate reduction rates (csSRR), implying changes in the electron transport chain that couples carbon and sulfur metabolisms. When DMSS-1 fixes nitrogen in ammonium-deficient medium, it also produces larger fractionation, but it occurs at faster csSRRs than in the ammonium-replete control cultures. The energy and reducing power required for nitrogen fixation may be responsible for the reverse trend between S-isotope fractionation and csSRR in this case. Iron deficiency and nitrogen fixation by sulfate-reducing microbes may lead to the large observed S-isotope effects in some euxinic basins and various anoxic sediments.

Large sulfur isotope fractionation does not require disproportionation.

The composition of sulfur isotopes in sedimentary sulfides and sulfates traces the sulfur cycle throughout Earth's history. In particular, depletions of sulfur-34 ((34)S) in sulfide relative to sulfate exceeding 47 per mil (‰) often serve as a proxy for the disproportionation of intermediate sulfur species in addition to sulfate reduction. Here, we demonstrate that a pure, actively growing culture of a marine sulfate-reducing bacterium can deplete (34)S by up to 66‰ during sulfate reduction alone and in the absence of an extracellular oxidative sulfur cycle. Therefore, similar magnitudes of sulfur isotope fractionation in sedimentary rocks do not unambiguously record the presence of other sulfur-based metabolisms or the stepwise oxygenation of Earth's surface environment during the Proterozoic.

Outcome of in-hospital adult cardiopulmonary resuscitation assisted with portable auto-priming percutaneous cardiopulmonary support.

BACKGROUND: Outcome from in-hospital cardiopulmonary resuscitation (CPR) is still unsatisfactory. CPR assisted with percutaneous cardiopulmonary support (PCPS) is expected to improve the outcome of in-hospital CPR. METHODS: We retrospectively analyzed 83 consecutive cases of adult in-hospital CPR assisted by a portable pre-assembled auto-priming PCPS system (EBS, Terumo, Japan) from January 2004 to December 2007. RESULTS: PCPS was successfully performed in 97.6% of the patients and could be weaned in 57.8% of the patients. The survival-to-discharge rate was 41.0% with an acceptable neurological status in 85.3% of the patients. The 6-month survival was 38.6%. Survival-to-discharge decreased about 1% for each 1 min increase in the duration of CPR. The probability of survival was about 65%, 45%, and 19% when the duration of CPR was 10, 30, or 60 min, respectively. Multivariate analysis adjusted with clinical factors including organ dysfunction severity scores revealed that defibrillation and CPR duration less than 35 min were independent predictors for both survival-to-discharge (odds ratio=8.0, 95% CI=2.8-23.0, p<0.001) and 6-month survival (hazard ratio=3.3, 95% CI=1.9-5.9, p<0.001). CONCLUSIONS: Our results showed that CPR assisted with PCPS results in an acceptable survival-to-discharge rate and mid-term prognosis.

Biophysical basis for the geometry of conical stromatolites.

Stromatolites may be Earth's oldest macroscopic fossils; however, it remains controversial what, if any, biological processes are recorded in their morphology. Although the biological interpretation of many stromatolite morphologies is confounded by the influence of sedimentation, conical stromatolites form in the absence of sedimentation and are, therefore, considered to be the most robust records of biophysical processes. A qualitative similarity between conical stromatolites and some modern microbial mats suggests a photosynthetic origin for ancient stromatolites. To better understand and interpret ancient fossils, we seek a quantitative relationship between the geometry of conical stromatolites and the biophysical processes that control their growth. We note that all modern conical stromatolites and many that formed in the last 2.8 billion years display a characteristic centimeter-scale spacing between neighboring structures. To understand this prominent-but hitherto uninterpreted-organization, we consider the role of diffusion in mediating competition between stromatolites. Having confirmed this model through laboratory experiments and field observation, we find that organization of a field of stromatolites is set by a diffusive time scale over which individual structures compete for nutrients, thus linking form to physiology. The centimeter-scale spacing between modern and ancient stromatolites corresponds to a rhythmically fluctuating metabolism with a period of approximately 20 hr. The correspondence between the observed spacing and the day length provides quantitative support for the photosynthetic origin of conical stromatolites throughout geologic time.

Morphological record of oxygenic photosynthesis in conical stromatolites.

Conical stromatolites are thought to be robust indicators of the presence of photosynthetic and phototactic microbes in aquatic environments as early as 3.5 billion years ago. However, phototaxis alone cannot explain the ubiquity of disrupted, curled, and contorted laminae in the crests of many Mesoproterozoic, Paleoproterozoic, and some Archean conical stromatolites. Here, we demonstrate that cyanobacterial production of oxygen in the tips of modern conical aggregates creates contorted laminae and submillimeter-to-millimeter-scale enmeshed bubbles. Similarly sized fossil bubbles and contorted laminae may be present only in the crestal zones of some conical stromatolites 2.7 billion years old or younger. This implies not only that cyanobacteria built Proterozoic conical stromatolites but also that fossil bubbles may constrain the timing of the evolution of oxygenic photosynthesis.