Po-Ge-Jiu-Xin decoction alleviate sepsis-induced cardiomyopathy via regulating phosphatase and tensin homolog-induced putative kinase 1 /parkin-mediated mitophagy.

ETHNOPHARMACOLOGICAL RELEVANCE: Sepsis is a life-threatening systemic syndrome usually accompanied by myocardial dysfunction. Po-Ge-Jiu-Xin decoction (PGJXD), a traditional Chinese prescription medicine, has been used clinically to treat cardiovascular disease including heart failure, sepsis-induced cardiomyopathy (SIC) and even septic shock. Previous clinical studies suggested PGJXD has shown promising results in improving cardiac function and treating heart failure in sepsis. However, more research is needed to elucidate the mechanisms underlying PGJXD's therapeutic effects in sepsis-induced cardiomyopathy. MATERIALS AND METHODS: Initially, we identified the major compounds of PGJXD through ultra-performance liquid chromatography-mass spectrometry technology analysis. We established in a SIC rat model using cecal ligation and puncture(CLP) and treated by PGJXD and levosimendan. We evaluated pathological damage by hematoxylin and eosin staining and measured serum myocardial injury biomarkers. Myocardial apoptosis was detected by Tunel staining and quantifying specific biomarker protein levels. Subsequently, we evaluated myocardium mitochondrial quality using Transmission electron microscope (TEM), antioxidant stress indexes and tissue adenosine triphosphate(ATP) content. We detected the expression of phosphatase and tensin homolog (PTEN)-induced putative kinase 1 (PINK1), parkin, LC3, and p62 using Western blotting and Quantitative real time polymerase chain reaction(qRT-PCR). (Lipopolysaccharides, LPS)-induced H9c2 cell model was established to further explore the mechanism of PGJXD on SIC. In addition to measuring cell viability, we measured mitochondrial membrane potential using JC-1 staining. Additionally, Parkin-siRNA transfected into H9c2 cells to validate whether PGJXD conducted protective effects against SIC through PINK1/Parkin-mediated mitophagy. RESULTS: It has been demonstrated that PGJXD reduced mortality in septic rat, contributed to ameliorating myocardium injury, suppressed inflammatory response and ameliorated the myocardial apoptosis. PGJXD could also alleviate mitochondrial structural abnormality, mitigated oxidative stress injury and promoted energy synthesis in CLP models. Western blotting and qRT-PCR have further confirmed that PGJXD can activate PINK1/parkin pathway-mediated mitophagy, resulting in preserving mitochondrial quality in the myocardium. Furthermore, Parkin siRNA partially reversed the beneficial effect of PGJXD on mitochondrial fission/fusion and mitophagy in vitro. Therefore, the cardioprotective effect of PGJXD is achieved by inducing PINK1/Parkin-mediated mitophagy in maintaining mitochondrial homeostasis. CONCLUSIONS: These results suggest that the potential therapeutic effect of PGJXD on cardiac dysfunction during sepsis and support its mechanism of targeted induction of PINK1-Parkin-mediated mitophagy.

Blood Transcriptomic and Inflammatory Protein Biomarkers Associated with Imminent Pulmonary Exacerbation Risk in Cystic Fibrosis.

RATIONALE: The factors that lead to poor pulmonary exacerbation (PEx) outcomes in individuals with cystic fibrosis (CF) are still being investigated; however, delayed diagnosis and treatment are likely contributory. Identifying individuals at imminent risk of PEx could enable closer monitoring and/or earlier initiation of therapies to improve outcomes. OBJECTIVE: The goal of this study was to develop blood-based biomarkers that associate with imminent PEx risk in CF individuals. METHODS: We examined the whole blood transcriptome and 55 inflammatory proteins from plasma and serum on 72 blood samples from 53 CF individuals. Biomarker candidate genes and proteins were selected from 14 CF individuals with paired stable and PEx visits (Cohort 1). The biomarker candidates were then estimated and tested to classify CF individuals who would experience a PEx within 4-months of a stable clinic visit or not (Cohort 2). RESULTS: A 16-gene panel and 9-protein panel were identified that could distinguish paired stable and PEx visits (AUC = 0.83 ± SE 0.28 and AUC = 0.92 ± SE 0.18, respectively). These two panels also demonstrated strong performance in classifying CF individuals who would experience a PEx within 4 months of a clinically stable visit or not (16-gene panel: AUC = 0.88; 9-protein panel: AUC = 0.83). In comparison, serum calprotectin and clinical variables (i.e. sex, ppFEV1, and the number of IV antibiotics in the preceding year) had AUCs of 0.75 and 0.71, respectively. CONCLUSION: Blood-based mRNA and protein biomarkers demonstrated strong performance in classifying CF individuals at risk of imminent PEx. If the findings from this study can be validated, there is the potential to use blood biomarkers to enable more personalized disease activity monitoring in CF.

Tandem transcription factors PpNAC1 and PpNAC5 synergistically activate the transcription of the PpPGF to regulate peach softening during fruit ripening.

Peach fruit rapidly soften after harvest, a significant challenge for producers and marketers as it results in rotting fruit and significantly reduces shelf life. In this study, we identified two tandem genes, PpNAC1 and PpNAC5, within the sr (slow ripening) locus. Phylogenetic analysis showed that NAC1 and NAC5 are highly conserved in dicots and that PpNAC1 is the orthologous gene of Non-ripening (NOR) in tomato. PpNAC1 and PpNAC5 were highly expressed in peach fruit, with their transcript levels up-regulated at the onset of ripening. Yeast two-hybrid and bimolecular fluorescence complementation assays showed PpNAC1 interacting with PpNAC5 and this interaction occurs with the tomato and apple orthologues. Transient gene silencing experiments showed that PpNAC1 and PpNAC5 positively regulate peach fruit softening. Yeast one-hybrid and dual luciferase assays and LUC bioluminescence imaging proved that PpNAC1 and PpNAC5 directly bind to the PpPGF promoter and activate its transcription. Co-expression of PpNAC1 and PpNAC5 showed higher levels of PpPGF activation than expression of PpNAC1 or PpNAC5 alone. In summary, our findings demonstrate that the tandem transcription factors PpNAC1 and PpNAC5 synergistically activate the transcription of PpPGF to regulate fruit softening during peach fruit ripening.

Effects of Chronic Stress from High Stocking Density in Mariculture: Evaluations of Growth Performance and Lipid Metabolism of Rainbow Trout (Oncorhychus mykiss).

(1) Background: In aquaculture, chronic stress due to high stocking density impairs animals' welfare and results in declined fishery production with low protein quality. However, most previous studies evaluated the effects of high stocking density on trout in freshwater rather than seawater. (2) Methods: Juvenile trout were reared for 84 days in circular tanks under three stocking densities, including low density ("LD", 9.15 kg/m3), moderate density ("MD", 13.65 kg/m3), and high density ("HD", 27.31 kg/m3) in seawater. The final densities of LD, MD, and HD were 22.00, 32.05 and 52.24 kg/m3, respectively. Growth performance and lipid metabolism were evaluated. (3) Results: Growth performance and feeding efficiency were significantly reduced due to chronic stress under high density in mariculture. The digestive activity of lipids was promoted in the gut of HD fish, while the concentration of triglycerides was decreased in the blood. Furthermore, decreased acetyl-CoA carboxylase (ACC) and fatty acid synthase (FAS), increased hormone-sensitive lipase (HSL) concentrations, and activated hepatic ß-oxidation processes were observed in trout under HD. Redundancy analysis showed that glycerol and HSL can be used as potential markers to evaluate the growth performance of trout in mariculture. (4) Conclusions: We showed that chronic high stocking density led to negative effects on growth performance, reduced de novo synthesis of fatty acids, and enhanced lipolysis.

Effects of Dietary Zinc on Growth Performance, Digestive Enzyme Activities, Antioxidant Status, and Immune Responses of Sea Cucumber Apostichopus japonicus.

Zinc is an essential micronutrient for organisms involved in regulating various biological processes. This study evaluated the effects of dietary zinc on growth performance, digestive enzyme activities, antioxidant status, and immune responses of sea cucumber Apostichopus japonicus. Five experimental diets were formulated with graded levels of zinc (0, 20, 40, 60, and 80 mg/kg, respectively), and the actual dietary zinc values were 31.4, 51.0, 68.2, 91.9, and 110.8 mg/kg diet, respectively. Sea cucumbers were fed with diets for 2 months. The results showed the growth performance, amylase, and trypsin activities of sea cucumber increased significantly with zinc supplementation, and the best growth performance and enzyme activities were observed at 40 mg/kg zinc diet. Zinc supplementation significantly increased activities of superoxide dismutase, catalase, anti-superoxide anion, and inhibiting hydroxyl radical, while significantly reduced the malondialdehyde content. Furthermore, the higher zinc supplementation levels resulted in significantly upregulated immune-related genes of hsp90, p105, rel, and lsz, suggesting that excessive zinc caused oxidative stress. The broken-line regression analysis of specific growth rate indicated dietary zinc requirement in juvenile sea cucumber was ~ 66.3 mg/kg diet. Overall, dietary zinc contributes to the growth and immune resistance of juvenile sea cucumber, and our study will provide insights into the rational use of dietary zinc in aquaculture.

One-Step Surface-to-Bulk Modification of High-Voltage and Long-Life LiCoO2 Cathode with Concentration Gradient Architecture.

Raising the charging cut-off voltage of layered oxide cathodes can improve their energy density. However, it inevitably introduces instabilities regarding both bulk structure and surface/interface. Herein, exploiting the unique characteristics of high-valence Nb5+ element, a synchronous surface-to-bulk-modified LiCoO2 featuring Li3 NbO4 surface coating layer, Nb-doped bulk, and the desired concentration gradient architecture through one-step calcination is achieved. Such a multifunctional structure facilitates the construction of high-quality cathode/electrolyte interface, enhances Li+ diffusion, and restrains lattice-O loss, Co migration, and associated layer-to-spinel phase distortion. Therefore, a stable operation of Nb-modified LiCoO2 half-cell is achieved at 4.6 V (90.9% capacity retention after 200 cycles). Long-life 250 Wh kg-1 and 4.7 V-class 550 Wh kg-1 pouch cells assembled with graphite and thin Li anodes are harvested (both beyond 87% after 1600 and 200 cycles). This multifunctional one-step modification strategy establishes a technological paradigm to pave the way for high-energy density and long-life lithium-ion cathode materials.

Ambient Pressure Drying of Freeze-Cast Ceramics from Aqueous Suspension.

Freeze-casting has been wildly exploited to construct porous ceramics but usually requires costly and demanding freeze-drying (high vacuum, size limit, and supercooled chamber), which can be avoided by the ambient pressure drying (APD) technique. However, applying APD to freeze-cast ceramic based on an aqueous suspension is still challenging due to inert surface chemistry. Herein, a modified APD strategy is developed to improve the drying process of freeze-cast ceramics by exploiting the simultaneous ice etching, ionic cross-linking, and solvent exchange under mild conditions (-10-0 °C, ambient pressure). This versatile strategy is applicable to various ceramic species, metal ions, and freezing techniques. The incorporated metal ions not only enhance liquid-phase sintering, producing ceramics with higher density and mechanical properties than freeze-cast counterparts, but also render customizable coloration and antibacterial property. The cost-/time-efficient APD is promising for mass production and even successive production of large-size freeze-cast ceramics that exceed the size of commercial freeze-dryers.

Unexplained Left Ventricular Hypertrophy Diagnosed as a Cardiac Variant of Late-Onset Fabry Disease: A Case Report.

The cardiac variant of Fabry disease (FD) has high rates of missed diagnosis and misdiagnosis due to the lack of systemic symptoms. Here, we report a case of a 68-year-old female with delayed-onset FD presenting as concentric left ventricular hypertrophy (LVH) with right bundle branch block, atrial fibrillation, and diastolic dysfunction, which was first presented with coronary artery spasm. Early cardiac-specific signs are crucial for diagnosing this disease due to the lack of extracardiac indications and the late onset of symptoms. This case raises a new red flag that patients with unexplained LVH and its atypical electrocardiographic (ECG) manifestations accompanied by diastolic dysfunction should be considered for FD. We also recommend further refinement of examinations associated with Fabry disease, which will contribute to the early diagnosis and treatment of the disease.

Metal-Free Late-Stage Alkylation of Tryptophan and Tryptophan-Containing Peptides with 1,3-Dithiane Derivatives.

Late-stage diversification of structurally complex peptides has enormous potential for drug discovery and molecular imaging. We report a simple, metal-free, late-stage reductive C2 alkylation of tryptophan and tryptophan-containing peptides using readily available 1,3-dithianes. This alkylation protocol has a wide substrate scope and an excellent tolerance for reactive functional groups.

Regioselective Alkenylation of Five-Membered Heteroarenes via a Dual 1,3-Sulfur Rearrangement.

Herein, we report a protocol for the stereoselective C-H alkenylation of five-membered heteroarenes including pyrroles (containing free NH pyrrole), thiophenes, and furans with 1,3-dithiane derivatives through dual 1,3-sulfur rearrangements. The site-selective and regioselective alkenylation of the five-membered heteroarenes proceeded in good yields via vinyl thionium ions to produce C2 or C5 Heck-type products, respectively.

Prevotella oris-caused meningitis and spinal canal infection: A case report.

BACKGROUND: Prevotella oris-induced meningitis and Prevotella oris-induced meningitis concomitant with spinal canal infection are extremely rare. To the best of our knowledge, only 1 case of Prevotella oris-induced central system infection has been reported. This is the second report on meningitis combined with spinal canal infection due to Prevotella oris. CASE SUMMARY: We report a case of a 9-year-old boy suffering from meningitis and spinal canal infection. The patient presented to the neurosurgery department with lumbosacral pain for 1 mo and headache and vomiting for 1 d. He had been treated with cephalosporin and nonsteroidal anti-inflammatory drugs for fever, otalgia and pharyngalgia in a local hospital 2 mo prior to this admission. During hospitalization, magnetic resonance imaging suggested meningitis and L3-S1 lumbosacral dural sac infection. The cerebrospinal fluid and blood cultures were negative, but the cerebrospinal fluid specimen indicated the presence of Prevotella oris by metagenomic next-generation sequencing. Previous cases of Prevotella oris infection were retrieved from PubMed to characterize the clinicopathological features and identify the prognostic factors and related antimicrobial treatment of infection due to Prevotella oris. CONCLUSION: This report shed light on the characteristics of Prevotella oris infection and highlighted the role of metagenomic next-generation sequencing in pathogen detection.

Pulmonary exacerbation inflammatory phenotypes in adults with cystic fibrosis.

BACKGROUND: Adults with cystic fibrosis (CF) develop exuberant inflammatory responses during pulmonary exacerbations (PEx) but whether distinct systemic inflammatory profiles can be identified and whether these associate with disparate treatment outcomes are unclear. We conducted a pilot study to address this question and hypothesized that CF adults with a pauci-inflammatory phenotype might derive less clinical benefit from intravenous (IV) antibiotic treatment than patients with other systemic inflammatory phenotypes. METHODS: Six proteins reflective of systemic inflammation were examined in 37 PEx from 28 unique CF subjects. We applied exploratory factor analysis and cluster analysis to identify biological clusters. Levels of blood proteins at PEx and clinical outcomes following IV antibiotic treatment were compared between clusters. RESULTS: Three clusters of PEx were identified. The pauci-inflammatory phenotype was characterized by lower levels of interleukin (IL)-1ß, IL-6, IL-10, tumor necrosis factor (TNF)-α, calprotectin, and C-reactive protein (CRP) (p < 0.05). Higher levels of IL-6 and IL-1ß were observed in the other 2 inflammatory clusters, but one of them was associated with higher calprotectin levels (p = 0.001) (neutrophil-predominant phenotype); whereas the other was associated with increased TNF-α and IL-10 levels (p < 0.001) (pro-inflammatory phenotype). A greater proportion of events from the neutrophil-predominant phenotype presented with acute respiratory symptoms and a larger decrease in ppFEV1 from baseline to hospital admission than the other two inflammatory phenotypes (p = 0.03). CONCLUSIONS: Three distinct inflammatory phenotypes were identified at PEx admission and each presented with unique clinical characteristics.

C-H Alkenylation of Indoles through a Dual 1,3-Sulfur Migration Process.

Methods for the modification of indole derivatives are powerful techniques in organic, medicinal, and biomolecular chemistry. Here, we report a protocol for the C-H alkenylation of N-H indoles with ß-chlorovinyl dithianes to furnish alkenyl indoles through dual 1,3-sulfur rearrangements. This alkenylation protocol could selectively prepare a variety of vinyl indoles in moderate to high yields with excellent functional group tolerance.

One-Pot Sequential Synthesis of 3,3'- or 2,3'-Bis(indolyl)methanes by Using 1,3-Dithiane as the Methylene Source.

A simple and efficient method for structurally diverse symmetrical and unsymmetrical 3,3'- and 2,3'-bisindolylmethanes has been developed through a one-pot sequential reaction using 1,3-dithiane as the methylene source. The important AhR agonists ICZ and malassezin were synthesized with excellent efficiency by this straightforward approach.

Understanding the Degradation of a Model Si Anode in a Li-Ion Battery at the Atomic Scale.

To advance the understanding of the degradation of the liquid electrolyte and Si electrode, and their interface, we exploit the latest developments in cryo-atom probe tomography. We evidence Si anode corrosion from the decomposition of the Li salt before charge-discharge cycles even begin. Volume shrinkage during delithiation leads to the development of nanograins from recrystallization in regions left amorphous by the lithiation. The newly created grain boundaries facilitate pulverization of nanoscale Si fragments, and one is found floating in the electrolyte. P is segregated to these grain boundaries, which confirms the decomposition of the electrolyte. As structural defects are bound to assist the nucleation of Li-rich phases in subsequent lithiations and accelerate the electrolyte's decomposition, these insights into the developed nanoscale microstructure interacting with the electrolyte contribute to understanding the self-catalyzed/accelerated degradation Si anodes and can inform new battery designs unaffected by these life-limiting factors.

Bridged Carbon Fabric Membrane with Boosted Performance in AC Line-Filtering Capacitors.

High-frequency responsive capacitors with lightweight, flexibility, and miniaturization are among the most vital circuit components because they can be readily incorporated into various portable devices to smooth out the ripples for circuits. Electrode materials no doubt are at the heart of such devices. Despite tremendous efforts and recent advances, the development of flexible and scalable high-frequency responsive capacitor electrodes with superior performance remains a great challenge. Herein, a straightforward and technologically relevant method is reported to manufacture a carbon fabric membrane "glued" by nitrogen-doped nanoporous carbons produced through a polyelectrolyte complexation-induced phase separation strategy. The as-obtained flexible carbon fabric bearing a unique hierarchical porous structure, and high conductivity as well as robust mechanical properties, serves as the free-standing electrode materials of electrochemical capacitors. It delivers an ultrahigh specific areal capacitance of 2632 µF cm-2 at 120 Hz with an excellent alternating current line filtering performance, fairly higher than the state-of-the-art commercial ones. Together, this system offers the potential electrode material to be scaled up for AC line-filtering capacitors at industrial levels.

Synchrotron X-Ray Tomography for Rechargeable Battery Research: Fundamentals, Setups and Applications.

Understanding the complicated interplay of the continuously evolving electrode materials in their inherent 3D states during the battery operating condition is of great importance for advancing rechargeable battery research. In this regard, the synchrotron X-ray tomography technique, which enables non-destructive, multi-scale, and 3D imaging of a variety of electrode components before/during/after battery operation, becomes an essential tool to deepen this understanding. The past few years have witnessed an increasingly growing interest in applying this technique in battery research. Hence, it is time to not only summarize the already obtained battery-related knowledge by using this technique, but also to present a fundamental elucidation of this technique to boost future studies in battery research. To this end, this review firstly introduces the fundamental principles and experimental setups of the synchrotron X-ray tomography technique. After that, a user guide to its application in battery research and examples of its applications in research of various types of batteries are presented. The current review ends with a discussion of the future opportunities of this technique for next-generation rechargeable batteries research. It is expected that this review can enhance the reader's understanding of the synchrotron X-ray tomography technique and stimulate new ideas and opportunities in battery research.

Isolated synchronous Virchow lymph node metastasis of sigmoid cancer: A case report.

BACKGROUND: Colorectal cancer (CRC) is one of the most common malignant tumors of the digestive tract. Lymphatic metastases of this tumor are mostly confined to the regional lymph nodes, and distant supraclavicular lymph node metastases are very rare. CASE SUMMARY: In this report, we describe a patient with sigmoid carcinoma and isolated synchronous supraclavicular lymph node metastases. A 56-year-old male presented with a left cervical mass that was confirmed as a lymph node metastasis from sigmoid cancer by several auxiliary examinations. After 6 cycles of chemotherapy with the 5-fluorouracil, leucovorin and oxaliplatin + cetuximab regimen, the sigmoid colon tumor and Virchow's lymph node metastasis were significantly smaller than before treatment, and no new metastatic sites were observed. Considering the effects of chemotherapy on quality of life, resection of the primary tumor was performed followed by 4 cycles of chemotherapy with the original chemotherapy regimen. Virchow's lymph node dissection was selected by mutual consultation between the patient and us. After the second surgery, the patient received capecitabine and cetuximab chemotherapy and did not experience recurrence or metastasis during follow-up. CONCLUSION: In conclusion, supraclavicular lymph node metastasis without any other solid organ metastasis is a potential metastatic pathway for CRC. In addition, after resection of the primary lesion, postoperative chemotherapy combined with supraclavicular lymph node dissection is feasible for the treatment of patients with CRC and isolated synchronous Virchow's lymph node metastases.

Differences in Gall Induction of Flower-like Galls on Haloxylon by Psyllids (Hemiptera: Aphalaridae), and the Emergence of Corresponding Parasitoids.

Saxaul is a kind of dominant perennial psammophyte that widely distributes in arid and semi-arid desert areas, and it has multiple functions in preventing desertification, especially in windbreak and sand fixation. Various gall inducers induce galls on the saxaul, including the flower-like gall. Parasitoids have great potentiality in controlling gall inducers. However, studies about gall inducers and parasitoids of flower-like galls on Haloxylon, as well as the parasitic efficacy of the parasitoids, are rarely reported. In this study, the flower-like galls were observed on Haloxylon ammodendron and H. persicum in Fukang, Xinjiang, China. Two types of flower-like galls were found on H. ammodendron, while only one type was found on H. persicum. In total, five species of gall inducers and three species of parasitoids were obtained from the galls mentioned above. All the galls were induced by Caillardia (Hemiptera: Aphalaridae), which were mostly bivoltine in Fukang. Besides, their parasitoids Psyllaephaguscaillardiae and P. longiventris could be observed on all the types of galls. Additionally, correlative studies on the parasitization indexes demonstrated that all the dominant parasitoids of diverse flower-like galls were P. caillardiae, which were slightly more in number than the P. ogazae discovered in the flower bud-like galls. In addition, the relevance between the emergence or lifespan of parasitoids and temperature was also investigated. The results showed that the number of parasitoids emerging decreased rapidly after a period of enhancement with the increase of temperature, including an optimum temperature, while the lifespan of wasps gradually shortened with the temperature rising. Our results highlight the importance of the biological investigation of parasitoids in the gall inducers lived in closed galls, which may provide critical evidence for us to understand its potential application in biological control.

Effects of temperature, dissolved oxygen, and their interaction on the growth performance and condition of rainbow trout (Oncorhynchus mykiss).

The individual effects of temperature and dissolved oxygen (DO) on rainbow trout (Oncorhynchus mykiss), an important aquaculture species, are clearly established; however, little is known about the interactive effects of these parameters. In this study, the effects of temperature, DO, and their interaction on the growth, antioxidant status, digestive enzyme activity, serum biochemical parameters, and liver IGF-1 expression in rainbow trout were evaluated. Fish (initial weight, 109.98 ± 3.28 g) were reared in a recirculating system for 4 weeks and subjected to 6 treatments at three temperatures (13 °C, 17 °C, and 21 °C) and two DO contents (4.2 mg L-1 and 9.6 mg L-1). Physiological parameters were determined at the end of the trial. Specific growth rate and feed consumption were the highest at 17 °C and the lowest at 21 °C. Additionally, lysozyme, trypsin, lipase, and amylase activities, serum glucose and serum triglyceride contents, and IGF-1 expression decreased significantly at 21 °C and total serum protein and albumin contents were significantly higher at 21 °C than at 13 °C and 17 °C, indicating that high temperature impaired the immunity, digestion, and growth of rainbow trout. However, the adverse effects of high temperature can be alleviated by a high DO content, as evidenced by the smaller increments and decrements of these parameters under hyperoxic conditions than under hypoxic conditions. In response to high temperature stress, an increase in antioxidant enzyme activity led to the removal of oxygen free radicals under hyperoxic conditions; however, this increase was inhibited under hypoxia. Our results indicated that high temperatures have adverse effects on rainbow trout, and these harmful effects can be reduced by a high DO content.