The effect of lysophosphatidic acid-supplemented culture medium on human immature oocytes matured in vitro.

BACKGROUND: Lysophosphatidic acid-supplemented culture medium significantly increases the oocyte maturation rate in vitro. However, potential targets and pathways involved remain unknown. METHODS: A total of 43 women, who underwent cesarean section and aged between 18 and 35 years with good health, were included in this study. Immature oocytes were obtained and cultured with 10 µM lysophosphatidic acid. After culture, oocyte maturation was assessed and oocytes and cumulus cells were collected for RNA sequencing. Hierarchical indexing for spliced alignment of transcripts 2 method was used to align clean reads to the human genome. The featureCounts and edgeR package were used to calculate gene expression and analyze differences between groups respectively. ClusterProfiler program was used to perform Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analysis. RESULTS: Oocyte maturation rate increased significantly following 48 h culture with lysophosphatidic acid. In cumulus cells, Gene Ontology analysis revealed the top 20 items enriched by upregulated genes and downregulated genes respectively; Kyoto Encyclopedia of Genes and Genomes analysis showed that upregulated genes in the treatment group were enriched in TNF signaling and insulin secretion pathways and downregulated genes were enriched in TNF signaling and cell adhesion molecules. In oocytes, Gene Ontology analysis revealed the top 20 items enriched by upregulated genes and downregulated genes respectively; Kyoto Encyclopedia of Genes and Genomes analysis showed that upregulated genes in the treatment group were enriched in MAPK signaling, gap junction, and cell cycle pathways and downregulated genes were enriched in MAPK signaling, estrogen signaling, RAP1 signaling, and gap junction pathways. CONCLUSIONS: Lysophosphatidic acid in culture medium enhances human oocyte maturation in vitro and the identified some potential pathways may associate with oocyte maturation.

Single-cell multiomic analysis of in vivo and in vitro matured human oocytes.

STUDY QUESTION: Are there any differences between in vivo (IVV) and in vitro (IVT) matured metaphase II (MII) oocytes at the molecular level? SUMMARY ANSWER: Between IVV and IVT oocytes, 507 differentially expressed genes (DEGs) were identified; the non-CpG methylomes were significantly different, but the CpG methylomes and genomic copy number variations (CNVs) were similar. WHAT IS KNOWN ALREADY: A previous study using microarray and single-cell RNA-seq analysis revealed that numerous genes were differentially expressed between IVV and IVT oocytes. Independent studies of DNA methylation profiling in human oocytes have revealed negative correlations between gene transcription and the DNA methylation level at gene promoter regions. No study has compared global CpG or non-CpG methylation between these two groups of oocytes. Although a high level of aneuploidy has been reported in MII oocytes, no direct comparison of IVV and IVT oocytes based on single-cell sequencing data has been performed. STUDY DESIGN, SIZE, DURATION: We collected eight IVV oocytes from six patients and seven IVT oocytes from seven patients and then analysed each oocyte using the previously established single-cell triple omics sequencing (scTrioseq) analysis to determine associations among the transcriptome, DNA methylome and chromosome ploidy in the oocytes. PARTICIPANTS/MATERIALS, SETTING, METHODS: All IVV oocytes were donated by patients who received 150 IU gonadotropin per day from the third day of their menstrual cycle, followed by GnRH antagonist after 5 days of gonadotropin stimulation. All IVT oocytes were from immature oocytes which were donated by volunteers undergoing delivery by caesarean section then cultured in oocyte maturation medium containing 75 mIU/ml hMG for 24 to 48 h. Every single oocyte was analysed using the previously established single-cell multiomic sequencing analysis. MAIN RESULTS AND THE ROLE OF CHANCE: There were 507 genes differentially expressed between the IVV (n = 8) and IVT (n = 7) oocytes, even though their global transcriptome profiles were similar. The enriched genes in IVV oocytes were related to the cell cycle process while those in IVT oocytes were related to mitochondrial respiration biogenesis. Although the global CpG methylation of the two groups of oocytes was similar, the non-CpG methylation level in IVV oocytes was higher than that in IVT oocytes. A high aneuploidy ratio was found in both groups, but the aneuploidy did not affect transcription according to the correlation analysis. LARGE-SCALE DATA: N/A. LIMITATIONS AND REASONS FOR CAUTION: Due to the difficulty in collecting MII oocytes, especially IVV matured oocytes, the sample size was limited. WIDER IMPLICATIONS OF THE FINDINGS: Our findings indicate that single-cell multiomic sequencing can be utilised to examine the similarity and differences between IVV and IVT matured MII oocytes. STUDY FUNDING/COMPETING INTEREST(S): This work was supported by the Ministry of Science and Technology of China, National Key R&D Program of China (No. 2017YFC1001601). The donated oocytes were collected by Shanghai Tenth People's Hospital. The authors declare no competing interests.

Nitrite-dependent microbial utilization for simultaneous removal of sulfide and methane in sewers.

Chemicals are commonly dosed in sewer systems to reduce the emission of hydrogen sulfide (H2S) and methane (CH4), incurring high costs and environmental concerns. Nitrite dosing is a promising approach as nitrite can be produced from urine wastewater, which is a feasible integrated water management strategy. However, nitrite dosing usually requires strict conditions, e.g., relatively high nitrite concentration (e.g., ∼200 mg N/L) and acidic environment, to inhibit microorganisms. In contrast to "microbial inhibition", this study proposes "microbial utilization" concept, i.e., utilizing nitrite as a substrate for H2S and CH4 consumption in sewer. In a laboratory-scale sewer reactor, nitrite at a relatively low concentrations of 25-48 mg N/L was continuously dosed. Two nitrite-dependent microbial utilization processes, i.e., nitrite-dependent anaerobic methane oxidation (n-DAMO) and microbial sulfide oxidation, successfully occurred in conjunction with nitrite reduction. The occurrence of both processes achieved a 58 % reduction in dissolved methane and over 90 % sulfide removal in the sewer reactor, with microbial activities measured as 15.6 mg CH4/(L·h) and 29.4 mg S/(L·h), respectively. High copy numbers of n-DAMO bacteria and sulfide-oxidizing bacteria (SOB) were detected in both sewer biofilms and sediments. Mechanism analysis confirmed that the dosed nitrite at a relatively low level did not cause the inhibition of sulfidogenic process due to the downward migration of activity zones in sewer sediments. Therefore, the proposed "microbial utilization" concept offers a new alternative for simultaneous removal of sulfide and methane in sewers.

Methane mitigation via the nitrite-DAMO process induced by nitrate dosing in sewers.

Nitrate or nitrite-dependent anaerobic methane oxidation (n-DAMO) is a microbial process that links carbon and nitrogen cycles as a methane sink in many natural environments. This study demonstrates, for the first time, that the nitrite-dependent anaerobic methane oxidation (nitrite-DAMO) process can be stimulated in sewer systems under continuous nitrate dosing for sulfide control. In a laboratory sewer system, continuous nitrate dosing not only achieved complete sulfide removal, but also significantly decreased dissolved methane concentration by ∼50 %. Independent batch tests confirmed the coupling of methane oxidation with nitrate and nitrite reduction, revealing similar methane oxidation rates of 3.68 ± 0.5 mg CH4 L-1 h-1 (with nitrate as electron acceptor) and 3.57 ± 0.4 mg CH4 L-1 h-1 (with nitrite as electron acceptor). Comprehensive microbial analysis unveiled the presence of a subgroup of the NC10 phylum, namely Candidatus Methylomirabilis (n-DAMO bacteria that couples nitrite reduction with methane oxidation), growing in sewer biofilms and surface sediments with relative abundances of 1.9 % and 1.6 %, respectively. In contrast, n-DAMO archaea that couple methane oxidation solely to nitrate reduction were not detected. Together these results indicated the successful enrichment of n-DAMO bacteria in sewerage systems, contributing to approx. 64 % of nitrite reduction and around 50 % of dissolved methane removal through the nitrite-DAMO process, as estimated by mass balance analysis. The occurrence of the nitrite-DAMO process in sewer systems opens a new path to sewer methane emissions.

Reducing sulfide and methane production in gravity sewer sediments through urine separation, collection and intermittent dosing.

Sulfide and methane production are a major concern in sewer management. Many solutions with the use of chemicals have been proposed yet incurring huge costs. Here, this study reports an alternative solution to reduce sulfide and methane production in sewer sediments. This is achieved through integration of urine source separation, rapid storage, and intermittent in situ re-dosing into a sewer. Based on a reasonable capacity of urine collection, an intermittent dosing strategy (i.e. 40 min per day) was designed and then experimentally tested using two laboratory sewer sediment reactors. The long-term operation showed that the proposed urine dosing in the experimental reactor effectively reduced sulfidogenic and methanogenic activities by 54% and 83%, compared to those in the control reactor. In-sediment chemical and microbial analyses revealed that the short-term exposure to urine wastewater was effective in suppressing sulfate-reducing bacteria and methanogenic archaea, particularly within a surface active zone of sediments (0-0.5 cm) likely attributed to the biocidal effect of urine free ammonia. Economic and environmental assessments indicated that the proposed urine approach can save 91% in total costs, 80% in energy consumption and 96% in greenhouse gas emissions compared to the conventional use of chemicals (including ferric salt, nitrate, sodium hydroxide, and magnesium hydroxide). These results collectively demonstrated a practical solution without chemical input to improve sewer management.

In-situ advanced oxidation of sediment iron for sulfide control in sewers.

Sulfide control is a significant problem in urban sewer management. Although in-sewer dosing of chemicals has been widely applied, it is prone to high chemical consumption and cost. A new approach is proposed in this study for effective sulfide control in sewers. It involves advanced oxidation of ferrous sulfide (FeS) in sewer sediment, to produce hydroxyl radical (·OH) in-situ, leading to simultaneous sulfide oxidation and reduction of microbial sulfate-reducing activity. Long-term operation of three laboratory sewer sediment reactors was used to test the effectiveness of sulfide control. The experimental reactor with the proposed in-situ advanced FeS oxidation substantially reduced sulfide concentration to 3.1 ± 1.8 mg S/L. This compares to 9.2 ± 2.7 mg S/L in a control reactor with sole oxygen supply, and 14.1 ± 4.2 mg S/L in the other control reactor without either iron or oxygen. Mechanistic investigations illustrated the critical role of ·OH, produced from the oxidation of sediment iron, in regulating microbial communities and the chemical sulfide oxidation reaction. Together these results demonstrate that incorporating the advanced FeS oxidation process in sewer sediment enable superior performance of sulfide control at a much lower iron dosage, thereby largely saving chemical use.

The advantage of a two-stage nitrification method for fertilizer recovery from human urine.

Recycling nutrients (nitrogen, phosphorus, and potassium) from human urine can potentially offset more than 13% of global agricultural fertilizer demand. Biological nitrification is a promising method for converting volatile ammonia in high-strength human urine into stable ammonium nitrate (a typical fertilizer), but it is usually terminated in the intermediate production of nitrite due to the inhibition of nitrite-oxidizing bacteria by free nitrous acid (FNA). This study aimed to develop a stable nitrification process in a unique two-stage bioreactor by removing critical barriers associated with FNA inhibition. Experimental results show that half of the ammonium in high-strength urine was successfully converted into nitrate, forming valuable ammonium nitrate (with a nitrogen concentration greater than 1500 mg N/L). The ammonium nitrate solution could retain most phosphorus (75% ± 3%) and potassium (96% ± 1%) in human urine, resulting in nearly full nutrient recovery. Once concentrated, the liquid compound fertilizer of ammonium nitrate was generated. Based on an assessment of economic and environmental impacts at the urban scale, urine diversion for nutrient recovery using a technical combination of nitrification and reverse osmosis could reduce total energy input by 43%, greenhouse gas emission by 40%, and cost by 33% compared to conventional wastewater management. Further research is needed to optimize the two-stage nitrification method on a larger scale.

Simultaneous control of sulfide and methane in sewers achieved by a physical approach targeting dominant active zone in sediments.

Sewer sediments not only induce sewer blockages, but also contributes to significant sulfide and methane productions in gravity sewer systems. Chemical control of sulfide and methane production is extremely expensive. This study aims to propose a novel physical control approach-intermittent surface sediment flushing to synchronously address sediment-induced multiple issues. The proposed approach was established investigating the suppression and recovery characteristics of sulfidogenic and methanogenic activities of sediments including the in-situ activity analysis by using the diffusive gradients in thin films (DGT). The results showed that ∼70% of total sulfide and methane production in sediments was contributed by surface sediments (0-1.5 cm), which could be easily flushed away by a low shear stress (<0.1 N/m2). Surface sediment flushing resulted in an immediate reduction in sulfidogenic and methanogenic activities, which both required about one week to recover to 50% of the maximum. These novel insights hopefully provide a feasible approach, i.e., intermittent surface sediment flushing, to effectively reduce sulfide and methane production in sewers. Compared with chemical dosing methods, the proposed approach, which has no chemical input, greatly reduces operating cost and environment impact. Moreover, intermittent surface flushing is expected to keep sediment thickness within a certain range to alleviate sewer blockage.

Changes in the Mitochondria-Related Nuclear Gene Expression Profile during Human Oocyte Maturation by the IVM Technique.

To address which mitochondria-related nuclear differentially expressed genes (DEGs) and related pathways are altered during human oocyte maturation, single-cell analysis was performed in three oocyte states: in vivo matured (M-IVO), in vitro matured (M-IVT), and failed to mature in vitro (IM-IVT). There were 691 DEGs and 16 mitochondria-related DEGs in the comparison of M-IVT vs. IM-IVT oocytes, and 2281 DEGs and 160 mitochondria-related DEGs in the comparison of M-IVT vs. M-IVO oocytes, respectively. The GO and KEGG analyses showed that most of them were involved in pathways such as oxidative phosphorylation, pyruvate metabolism, peroxisome, and amino acid metabolism, i.e., valine, leucine, isoleucine, glycine, serine, and threonine metabolism or degradation. During the progress of oocyte maturation, the metabolic pathway, which derives the main source of ATP, shifted from glucose metabolism to pyruvate and fatty acid oxidation in order to maintain a low level of damaging reactive oxygen species (ROS) production. Although the immature oocytes could be cultured to a mature stage by an in vitro technique (IVM), there were still some differences in mitochondria-related regulations, which showed that the mitochondria were regulated by nuclear genes to compensate for their developmental needs. Meanwhile, the results indicated that the current IVM culture medium should be optimized to compensate for the special need for further development according to this disclosure, as it was a latent strategy to improve the effectiveness of the IVM procedure.

Exploring the reciprocal relations between mothers' and fathers' use and attitudes of corporal punishment in China: A cross-lagged analysis.

BACKGROUND: Corporal punishment is a commonly used form of disciplinary technique. Sanctified parental attitudes of corporal punishment have been found to be a significant predictor of parental use of corporal punishment in previous research, while little is currently known about the reciprocal relations between parental use and their attitudes of corporal punishment. OBJECTIVE: This research aimed to examine the reciprocal relations between mothers' and fathers' use and attitudes of corporal punishment in China. METHODS: Data were collected on a total of 320 Chinese father-mother dyads with their children (10-11 years of age at baseline) through convenience sampling techniques at two time points, one year apart. Parents completed self-report measures of mothers' and fathers' use and attitudes of corporal punishment. Children completed self-report measures of parental corporal punishment. RESULTS: The cross-lagged analysis indicated that parental attitudes of corporal punishment in a given year predicted their use of corporal punishment in the subsequent year both for mothers (ß = 0.15, p < .01) and fathers (ß = 0.10, p < .05), while their corporal punishment in a given year did not predict their attitudes of it in the subsequent year (ßs < 0.11, ps > .05). CONCLUSIONS: Findings indicate that the reciprocal relations do not emerge, with only attitude-behavior effects being evident for both mothers and fathers, while behavior-attitude effects were not present. Findings in the present study highlight the importance of changing both mothers' and fathers' favorable attitudes toward corporal punishment when conducting appropriate prevention intervention to decrease its use.