Carbon and Nutrient Limitations of Microbial Metabolism in Xingkai Lake, China: Abiotic and Biotic Drivers.

Microbial communities are crucial for water quality and biogeochemical cycling in freshwaters. Microbes secrete extracellular enzymes to decompose organic matter for their needs of nutrients and scarce elements. Yet, there is a lack of knowledge on microbial metabolic limitations in freshwaters, especially in lake sediments. Here, we examined the carbon, nitrogen, and phosphorus-acquiring extracellular enzyme activities and the bacterial and fungal communities of 30 sediments across Xingkai Lake, the largest freshwater lake in Northeast Asia. We further analyzed the microbial metabolic limitations via extracellular enzyme stoichiometry and explored the direct and indirect effects of abiotic and biotic factors on the limitations. We found that microbial metabolisms were primarily limited by phosphorus in Xingkai Lake. For instance, microbial carbon and phosphorus limitations were closely correlated to abiotic factors like water depth, total dissolved solids, sediment total carbon, and conductivity. The metabolic limitations were also affected by biotic factors, such as showing positive relationships with the alpha and beta diversity of bacteria, and with the beta diversity of fungi. In addition, community compositions of bacteria and fungi were mainly correlated to abiotic factors such as total carbon and dissolved organic carbon, respectively. Collectively, microbial metabolic limitations were affected directly or indirectly by abiotic factors and microbial communities. Our findings indicate that microbial metabolic limitations are not only driven by bacteria and fungi but also by abiotic factors such as water depth and total nitrogen, and thus provide empirical evidence for effective management of freshwater lakes under climate warming and intensified human activities.

Timing considerations for removal of early cumulus cells in short-term insemination strategies.

Context The timing of early cumulus cell removal (ECCR) can be changed within a range. The change has an effect on the multiple pronuclei (MPN) rate and the exposure time of oocytes to sperm waste products. The timing of ECCR effects the outcomes of assisted reproductive technology, however, it is still unclear what time is best for ECCR. Aims To find the best time for ECCR based on clinical outcomes in order to increase the success rate of assisted reproductive technology. Methods A retrospective study was performed. Cycles were categorised into six groups according to the timing of ECCR. The clinical outcomes of these six groups were compared by Kruskal-Wallis test and Pearson X 2 test. Key results The timing of ECCR had a significant effect on the MPN rate, 0PN without cleavage rate and grade 1-2 embryo rate at Day3. Among our six time groups of ECCR, the cumulus cell removal ≤4h post-insemination group had the highest MPN rate and grade 1-2 embryo rate at Day3, and the 5.5h

Platelet Antibodies in Infertility Patients.

BACKGROUND: Platelet antigens can stimulate the body to produce platelet alloimmune antibodies through blood transfusion, pregnancy, and autoimmunity. In the blood of pregnant women, anti-platelet antibodies can cause embryo implantation failure, abortion, etc. if they are present. METHODS: The platelet antibody was screened in 326 infertile patients (282 primary infertility and 44 secondary infertility) and 522 healthy controls in the physical examination center of our hospital by solid phase agglutination of red blood cells. RESULTS: The positive rate of anti-platelet antibody was 9.51% in the infertility group and 2.30% in the healthy control group. There was a significant difference between them (χ2 = 4.51, p < 0.05). The positive rate of anti-platelet antibody in the infertility group was significantly higher than that in the control group. The positive rate of anti-platelet antibody in the secondary infertility patients was significantly higher than that in the primary infertility patients (χ2 = 1.62, p < 0.05), and the positive rate of serum anti-platelet antibody increased gradually with the increase of infertility years. CONCLUSIONS: The positive rate of anti-platelet antibody is closely related to infertility and gradually increases with the age of infertility.

The effect of ionomycin-induced oocyte activation on multiple morphological abnormalities of the sperm flagella.

Artificial oocyte activation (AOA) is considered an effective method to improve clinical outcomes in patients with some forms of male factor infertility and does not increase the risk of birth defects. However, the effects of AOA on patients with multiple morphological abnormalities of the sperm flagella (MMAF) caused by a DNAH1 mutation are still unknown. To explore the effects, our study analyzed a case with MMAF due to DNAH1 homozygous mutation that underwent testicular sperm extraction (TESE) combined with intracytoplasmic sperm injection (ICSI). The case had 28 MII oocytes. The 28 oocytes were divided randomly and equally into AOA and non-AOA groups. Ionomycin was used for AOA. We compared the clinical outcomes of two groups and selected three blastulation failure embryos from each group for transcriptome analysis (Data can be accessed through GSE216618). Differentially expressed genes (DEGs) were determined with an adjusted p-value <0.05 and a |log2-fold change| ≥1. The comparison of clinical outcomes showed that the two pronuclei (2PN) rate and grade 1-2 embryo rate at day 3 were not significantly different between the two groups. Transcriptome analyses of blastulation failed embryos showed that the use of AOA had potential risks of chromosome structure defects, transcriptional regulation defects, and epigenetic defects. In conclusion, when the case with MMAF due to DNAH1 mutation underwent TESE-ICSI, ionomycin-induced oocyte activation could not improve the clinical outcomes and introduced the risks of chromosome structure defect, transcriptional regulation defect, and epigenetic defect.

Female BMI has an effect on oocyte gene expression pattern.

Purpose: To explore the mechanisms by which abnormal female BMI affects oocyte quality, particularly whether it involves the alteration of gene expression patterns and how these patterns may impact clinical outcomes. Methods: In Part 1, we performed a retrospective study to compare the clinical outcomes between the female BMI ≥25 kg/m2 and female BMI ≤20 kg/m2 groups. In Part 2, we performed the transcriptome analyses based on the GSE87201 dataset. Results: In Part 1, among the clinical outcomes, only the grade 1-2 embryo rate at day 3 of ICSI cycles was significantly different between the two BMI groups; the other outcomes were not. In Part 2, compared with the BMI ≤20 kg/m2 group, the oocyte gene expression pattern of the BMI ≥25 kg/m2 group seemed to result in better oocyte tolerance to exogenous stress, such as intracytoplasmic sperm injection (ICSI). It seemed to explain the result of Part 1 that the BMI ≥25 kg/m2 group had better day-3 embryo quality after ICSI than the BMI ≤20 kg/m2 group. Conclusions: Abnormal female BMI affects oocyte quality by altering the gene expression patterns of oocytes. While a female BMI ≥25 kg/m2 is known to have certain detrimental effects on ART, our findings suggest that it can also confer some benefits to oocytes.

DDIT4 Novel Mutations in Pancreatic Cancer.

Pancreatic cancer is one of the most common malignancies worldwide. This study is aimed at searching the possible genetic mutations and the value of novel gene mutation in the DNA damage-inducible transcript 4 (DDIT4) and signaling pathway in pancreatic cancer. Polymerase chain reaction (PCR) was performed to amplify the DNA sequences of DDIT4 from patients with pancreatic ductal adenocarcinoma. In addition, we used IHC to detect the expression level of DDIT4 in patients with pancreatic cancer in different types of gene mutation. Double-labeled immunofluorescence was employed to explore the expression levels of DDIT4/LC3 and their potential correlation. Our work indicated the two novel stable gene mutations in DDIT4 mRNA 3'-untranslated region (m.990 U>A and m.1246 C>U). Thirteen samples were found to have mutation in the DDIT4 3'-untranslated regions (UTR). To further verify the influence of gene mutation on protein expression, we performed immunohistochemistry on different gene mutation types, and we found a correlation between DDIT4 expression and gene mutation, which is accompanied by nuclear staining deepening. In order to further discuss the clinical value of DDIT4 gene mutation, immunofluorescence suggested that the expression of DDIT4 colocated with LC3; thus, we speculated that DDIT4 mutation may be involved in autophagy in pancreatic cancer cell. In this study, we found mutation in the 3'-UTR region of DDIT4, which may be associated with DDIT4 expression and tumor autophagy in pancreatic cancer tissues.

DNA Methylation Pattern of CALCA and CALCB in Extremely Premature Infants with Monochorionic Triplets after Single-Embryo Transfer.

Compared with full-term peers, premature infants are more likely to suffer from neonatal diseases and death. Variations in DNA methylation may affect these pathological processes. Calcitonin gene-related peptide (CGRP) plays a complex and diversified role in reproduction and chronic inflammation, and participates in the functional maintenance of vascular adaptation and trophoblast cells during pregnancy. Here, premature live births with single-chorionic triple embryos after single-embryo transfer were used as research objects, while full-term infants with double embryos and double-chorionic twins were used as controls. DNA was extracted from umbilical cord tissues for pyrosequencing to detect the methylation level of CpG island in CGRP promoter region. The average values of CGRP methylation in the umbilical cord tissues of very premature fetuses were higher than that of normal controls obtained from the databases. Immunofluorescence results showed that the expression of αCGRP was decreased in the blood vessel wall of the umbilical cord of monozygotic triplets, especially in death cases, while the ßCGRP had a compensatory expression. In conclusion, our findings suggest that hypermethylation of CGRP might be considered as an important cause of serious neonatal morbidities.

HNF-1 binding point mutation of the AFP gene promotes cirrhosis in post-menopausal women.

OBJECTIVE: α-fetoprotein (AFP) expression is activated during the embryonic stage or hepatocellular carcinogenesis, so it is presumed that AFP is a key endogenous molecule to promote cell proliferation or differentiation. We carried out gene screening in an unknown family with hyper-alpha-fetoproteinemia and some sporadic menopausal women, and discussed the relationship between AFP expression and liver cirrhosis. METHODS: Peripheral blood samples from family members, patients with malignant liver tumors, and normal controls were collected. Full-length sequence of AFP was amplified and directly sequenced, and compared with normal controls. HNF-1α and HNF-1ß in plasma levels of family members, patients with liver cancer, newborns, pregnant women, and normal subjects were detected by ELISA, and the relationship between HNF-1 and AFP mutation or high expression was evaluated. RESULTS: There was a mutation in AFP promoter region at c.-200 C>T, which was located at the binding site of AFP hepatocyte nuclear factor 1 (HNF-1). AFP was higher than 4000 ng/L in all members carrying the mutation, but liver cancer was excluded in the family with hyper-alpha-fetoprotein. However, cirrhosis occurred in post-menopausal women. The cases reviewed showed that unknown hyper-alpha-fetoprotein was closely related to HNF-1 binding point of AFP in post-menopausal women with cirrhosis (7/11), while the plasma levels of HNF-1α and HNF-1ß were not significantly different. CONCLUSION: The mutation of the HNF-1 binding point of AFP may lead to an abnormal high expression of AFP by altering the binding of HNF transcription factors, which is closely related to cirrhosis in menopausal women.

Analysis of embryo quality with luteal phase ovarian stimulation after failed in vitro fertilization-embryo transfer with long or ultra-long protocol.

OBJECTIVE: To investigate the effect of luteal phase ovarian stimulation on embryo quality after failed in vitro fertilization-embryo transfer with long or ultra-long protocol. METHODS: In patients who underwent luteal phase ovarian stimulation after failed in vitro fertilization-embryo transfer with long or ultra-long protocol in the reproductive center between January 2015 and October 2017, self-control observations and statistical analyses were carried out for the number of oocytes retrieved, the rate of fertilization, the rate of D3 high-quality embryos, the rate of transplantable blastocyst formation, and the pregnancy rate of transfer cycle between long or ultra-long protocol and luteal phase ovarian stimulation. RESULTS: The rate of fertilization and blastocyst were significantly increased after luteal phase ovarian stimulation (P < 0.05). However, the difference of the number of oocytes retrieved and the rate of D3 high-quality embryos was not statistically significant (p > 0.05). CONCLUSIONS: The patients who failed with long protocol or ultra-long protocol due to low quality embryos, the protocol changed over to luteal phase ovarian stimulation. Can significantly improve the rate of fertilization oocytes and the transplantable blastocyst, and improve the outcome of clinical pregnancy. It provides an alternative ovarian stimulation protocol in patients with IVF-ET failure.

Erratum: Comparative Transcriptome Analysis Identifies Candidate Genes Related to Skin Color Differentiation in Red Tilapia.

This corrects the article DOI: 10.1038/srep31347.

Comparative Transcriptome Analysis Identifies Candidate Genes Related to Skin Color Differentiation in Red Tilapia.

Red tilapia is becoming more popular for aquaculture production in China in recent years. However, the pigmentation differentiation in genetic breeding is the main problem limiting its development of commercial red tilapia culture and the genetic basis of skin color variation is still unknown. In this study, we conducted Illumina sequencing of transcriptome on three color variety red tilapia. A total of 224,895,758 reads were generated, resulting in 160,762 assembled contigs that were used as reference contigs. The contigs of red tilapia transcriptome had hits in the range of 53.4% to 86.7% of the unique proteins of zebrafish, fugu, medaka, three-spined stickleback and tilapia. And 44,723 contigs containing 77,423 simple sequence repeats (SSRs) were identified, with 16,646 contigs containing more than one SSR. Three skin transcriptomes were compared pairwise and the results revealed that there were 148 common significantly differentially expressed unigenes and several key genes related to pigment synthesis, i.e. tyr, tyrp1, silv, sox10, slc24a5, cbs and slc7a11, were included. The results will facilitate understanding the molecular mechanisms of skin pigmentation differentiation in red tilapia and accelerate the molecular selection of the specific strain with consistent skin colors.