Phylogenomics and biogeographical diversification of Collabieae (Orchidaceae) and its implication in the reconstruction of the dynamic history of Asian evergreen broadleaved forests.

The tribe Collabieae (Epidendroideae, Orchidaceae) comprises approximately 500 species. Generic delimitation within Collabieae are confusing and phylogenetic interrelationships within the Collabieae have not been well resolved. Plastid genomes and nuclear internal transcribed spacer (ITS) sequences were used to estimate the phylogenetic relationships, ancestral ranges, and diversification rates of Collabieae. The results showed that Collabieae was subdivided into nine clades with high support. We proposed to combine Ancistrochilus and Pachystoma into Spathoglottis, merge Collabium and Chrysoglossum into Diglyphosa, and separate Pilophyllum and Hancockia as distinctive genera. The diversification of the nine clades of Collabieae might be associated with the uplift of the Himalayas during the Late Oligocene/Early Miocene. The enhanced East Asian summer monsoon in the Late Miocene may have promoted the rapid diversification of Collabieae at a sustained high diversification rate. The increased size of terrestrial pseudobulbs may be one of the drivers of Collabieae diversification. Our results suggest that the establishment and development of evergreen broadleaved forests facilitated the diversification of Collabieae.

Investigating the long-term response of plateau vegetation productivity to extreme climate: insights from a case study in Qinghai Province, China.

Over the past three decades, there has been a significant global climate change characterized by an increase in the intensity and frequency of extreme climate events. The vegetation status in Qinghai Province has undergone substantial changes, which are more pronounced than other regions in the Qinghai-Tibet Plateau. However, a clear understanding of the response characteristics of plateau vegetation to extreme climate events is currently lacking. In this study, we investigated the response of net primary productivity (NPP) to different forms of extreme climate events across regions characterized by varying levels of aridity and elevation gradients. Specifically, we observed a significant increase in NPP in relatively arid regions. Our findings indicate that, in relatively arid regions, single episodes of high-intensity precipitation have a pronounced positive effect (higher correlation) on NPP. Furthermore, in high-elevation regions (4000-6000 m), both the intensity and frequency of precipitation events are crucial factors for the increase in regional NPP. However, continuous precipitation can have significant negative impacts on certain areas within relatively wet regions. Regarding temperature, a reduction in the number of frost days within a year has been shown to lead to a significant increase in NPP in arid regions. This reduction allows vegetation growth rate to increase in regions where it was limited by low temperatures. Vegetation conditions in drought-poor regions are expected to continue to improve as extreme precipitation intensifies and extreme low-temperature events decrease.

Population genomic analysis provides evidence of the past success and future potential of South China tiger captive conservation.

BACKGROUND: Among six extant tiger subspecies, the South China tiger (Panthera tigris amoyensis) once was widely distributed but is now the rarest one and extinct in the wild. All living South China tigers are descendants of only two male and four female wild-caught tigers and they survive solely in zoos after 60 years of effective conservation efforts. Inbreeding depression and hybridization with other tiger subspecies were believed to have occurred within the small, captive South China tiger population. It is therefore urgently needed to examine the genomic landscape of existing genetic variation among the South China tigers. RESULTS: In this study, we assembled a high-quality chromosome-level genome using long-read sequences and re-sequenced 29 high-depth genomes of the South China tigers. By combining and comparing our data with the other 40 genomes of six tiger subspecies, we identified two significantly differentiated genomic lineages among the South China tigers, which harbored some rare genetic variants introgressed from other tiger subspecies and thus maintained a moderate genetic diversity. We noticed that the South China tiger had higher FROH values for longer runs of homozygosity (ROH > 1 Mb), an indication of recent inbreeding/founder events. We also observed that the South China tiger had the least frequent homozygous genotypes of both high- and moderate-impact deleterious mutations, and lower mutation loads than both Amur and Sumatran tigers. Altogether, our analyses indicated an effective genetic purging of deleterious mutations in homozygous states from the South China tiger, following its population contraction with a controlled increase in inbreeding based on its pedigree records. CONCLUSIONS: The identification of two unique founder/genomic lineages coupled with active genetic purging of deleterious mutations in homozygous states and the genomic resources generated in our study pave the way for a genomics-informed conservation, following the real-time monitoring and rational exchange of reproductive South China tigers among zoos.

Organelle Genomes of Epipogium roseum Provide Insight into the Evolution of Mycoheterotrophic Orchids.

Epipogium roseum, commonly known as one of the ghost orchids due to its rarity and almost transparent color, is a non-photosynthetic and fully mycoheterotrophic plant. Given its special nutritional strategies and evolutionary significance, the mitogenome was first characterized, and three plastomes sampled from Asia were assembled. The plastomes were found to be the smallest among Orchidaceae, with lengths ranging from 18,339 to 19,047 bp, and exhibited high sequence variety. For the mitogenome, a total of 414,552 bp in length, comprising 26 circular chromosomes, were identified. A total of 54 genes, including 38 protein-coding genes, 13 tRNA genes, and 3 rRNA genes, were annotated. Multiple repeat sequences spanning a length of 203,423 bp (45.47%) were discovered. Intriguingly, six plastid regions via intracellular gene transfer and four plastid regions via horizontal gene transfer to the mitogenome were observed. The phylogenomics, incorporating 90 plastomes and 56 mitogenomes, consistently revealed the sister relationship of Epipogium and Gastrodia, with a bootstrap percentage of 100%. These findings shed light on the organelle evolution of Orchidaceae and non-photosynthetic plants.

Molecular transformation of dissolved organic matter in manganese ore-mediated constructed wetlands for fresh leachate treatment.

The organic matter (OM) and nitrogen in Fresh leachate (FL) from waste compression sites pose environmental and health risks. Even though the constructed wetland (CW) can efficiently remove these pollutants, the molecular-level transformations of dissolved OM (DOM) in FL remain uncertain. This study reports the molecular dynamics of DOM and nitrogen removal during FL treatment in CWs. Two lab-scale vertical-flow CW systems were employed: one using only sand as substrates (act as a control, CW-C) and the other employing an equal mixture of manganese ore powder and sand (experimental, CW-M). Over 488 days of operation, CW-M exhibited significantly higher removal rates for chemical oxygen demand (COD), ammonia nitrogen (NH4+-N), and dissolved organic matter (represented by dissolved organic carbon, DOC) at 98.2 ± 2.5%, 99.2 ± 1.4%, and 97.9 ± 1.9%, respectively, in contrast to CW-C (92.8 ± 6.8%, 77.1 ± 28.1%, and 74.7 ± 9.5%). The three-dimensional fluorescence excitation-emission matrix (3D-EEM) and Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS) analyses unveiled that the influent DOM was predominantly composed of readily biodegradable protein-like substances with high carbon content and low unsaturation. Throughout treatment, it led to the degradation of low O/C and high H/C compounds, resulting in the formation of DOM with higher unsaturation and aromaticity, resembling humic-like substances. CW-M showcased a distinct DOM composition, characterized by lower carbon content yet higher unsaturation and aromaticity than CW-C. The study also identified the presence of Gammaproteobacteria, reported as Mn-oxidizing bacteria with significantly higher abundance in the upper and middle layers of CW-M, facilitating manganese cycling and improving DOM removal. Key pathways contributing to DOM removal encompassed adsorption, catalytic oxidation by manganese oxides, and microbial degradation. This study offers novel insights into DOM transformation and removal from FL during CW treatment, which will facilitate better design and enhanced performance.

Origin and diversification of a Himalayan orchid genus Pleione.

Pleione is an orchid endemically distributed in high mountain areas across the Hengduan Mountains (HDM), Himalayas, Southeast Asia and South of China. The unique flower shapes, rich colors and immense medicinal importance of Pleione are valuable ornamental and economic resources. However, the phylogenetic relationships and evolutionary history of the genus have not yet been comprehensively resolved. Here, the evolutionary history of Pleione was investigated using single-copy gene single nucleotide polymorphisms and chloroplast genome datasets. The data revealed that Pleione could be divided into five clades. Discordance in topology between the two phylogenetic trees and network and D-statistic analyses indicated the occurrence of reticulate evolution in the genus. The evolution could be attributed to introgression and incomplete lineage sorting. Ancestral area reconstruction suggested that Pleione was originated from the HDM. Uplifting of the HDM drove rapid diversification by creating conditions favoring rapid speciation. This coincided with two periods of consolidation of the Asian monsoon climate, which caused the first rapid diversification of Pleione from 8.87 to 7.83 Mya, and a second rapid diversification started at around 4.05 Mya to Pleistocene. The interaction between Pleione and climate changes, especially the monsoons, led to the current distribution pattern and shaped the dormancy characteristic of the different clades. In addition to revealing the evolutionary relationship of Pleione with orogeny and climate changes, the findings of this study provide insights into the speciation and diversification mechanisms of plants in the East Asian flora.

Effective protection: the embryonic development and clinical outcomes of emergency vitrification of 1246 oocytes and Day 0-Day 5 embryos in a natural disaster.

STUDY QUESTION: Can emergency vitrification protect embryos and oocytes during natural disasters or other events that prevent normal practice to achieve satisfactory embryonic development and clinical outcomes at a later time? SUMMARY ANSWER: Emergency vitrification of oocytes and Day 0-Day 5 (D0-D5) embryos during disasters is a safe and effective protective measure. WHAT IS KNOWN ALREADY: When some destructive events such as floods, earthquakes, tsunamis, and other accidents occur, emergency vitrification in embryo laboratories to protect human embryos, oocytes, and sperm is one of the important measures of an IVF emergency plan. However, there are few detailed reports on emergency vitrification in a state of disaster, especially about oocytes and D0 zygotes. Therefore, the effectiveness and safety of emergency vitrification of oocytes and D0-D5 embryos in disaster states are still unclear. STUDY DESIGN, SIZE, DURATION: A retrospective study was made in the Reproductive Medicine Center of the First Affiliated Hospital of Zhengzhou University from January 2018 to November 2022. The record rainstorms in Zhengzhou, China, caused severe flooding, traffic disruptions, and power outages. From 17:30, 20 July 2021 to 17:30, 21 July 2021, 1246 oocytes and D0-D5 embryos of 155 patients were vitrified whilst the laboratory had only an emergency power supply. PARTICIPANTS/MATERIALS, SETTING, METHODS: As of 21 December 2021, 1149 emergency vitrified oocytes and D0-D5 embryos of 124 patients underwent frozen-thawed embryo transfer (FET). They were divided into the following four groups according to the days of embryo culture in vitro: oocyte group, Day 0-Day 1 (D0-D1) group, Day 2-Day 3 (D2-D3) group, and Day 4-Day 5 (D4-D5) group. Control groups for each were selected from fresh cycle patients who underwent IVF/ICSI from January 2018 to October 2021. Control and emergency vitrification patients were matched on criteria that included age, fertilization method, days of embryonic development, and number and grade of transferred embryos. A total of 493 control patients were randomly selected from the eligible patients and matched with the emergency vitrification groups in a ratio of 4:1. The results of assisted reproduction and follow-up of pregnancy were analyzed. The embryonic development, clinical outcomes, and birth outcomes in each group were statistically analyzed. MAIN RESULTS AND THE ROLE OF CHANCE: A significant difference was observed in fertilization rate (81% versus 72%, P = 0.022) between the oocyte group and the control group. Significant differences were also observed in the monozygotic twin pregnancy rate (10% versus 0%, P = 0.038) and ectopic pregnancy rate (5% versus 0%, P = 0.039) between the D0-D1 group and the control group. No significant differences (P > 0.05) were observed between vitrified oocytes/D0-D1 embryos/D2-D3 embryos and the control group on the number of high-quality embryos (3.17 ± 3.00 versus 3.84 ± 3.01, P = 0.346; 5.04 ± 3.66 versus 4.56 ± 2.87, P = 0.346; 4.85 ± 5.36 versus 5.04 ± 4.64, P = 0.839), the number of usable blastocysts (1.22 ± 1.78 versus 1.21 ± 2.03, P = 0.981; 2.16 ± 2.26 versus 1.55 ± 2.08, P = 0.090; 2.82 ± 3.23 versus 2.58 ± 3.32, P = 0.706), clinical pregnancy rate (56% versus 57%, P = 0.915; 55% versus 55%, P = 1.000; 40% versus 50%, P = 0.488), miscarriage rate (30% versus 15%, P = 0.496; 5% versus 11%, P = 0.678; 17% versus 20%, P = 1.000), and live birth rate (39% versus 49%, P = 0.460; 53% versus 50%, P = 0.772; 33% versus 40%, P = 0.635). No significant differences (P > 0.05) were observed between the D4-D5 group and the control group on clinical pregnancy rate (40% versus 55%, P = 0.645), miscarriage rate (0% versus 18%, P = 1.000), and live birth rate (40% versus 45%, P = 1.000). LIMITATIONS, REASONS FOR CAUTION: The retrospective study design is a limitation. The timing and extent of natural disasters are unpredictable, so the sample size of vitrified oocytes, zygotes, and embryos is beyond experimental control. WIDER IMPLICATIONS OF THE FINDINGS: This study is the first study analyzing embryonic development, clinical outcomes, and birth outcomes of large samples of oocytes, D0 zygotes, and D1-D5 embryos after emergency vitrification under the disaster conditions. The results show that emergency vitrification is a safe and effective protective measure applicable to oocytes and D0-D5 embryos. The embryology laboratories need to be equipped with an emergency uninterrupted power supply capable of delivering for 6-8 h at full load. STUDY FUNDING/COMPETING INTEREST(S): This work was supported by the National Natural Science Foundation of China (grant 81871206). The authors declare that they have no conflicts of interest. All authors have completed the ICMJE Disclosure form. TRIAL REGISTRATION NUMBER: N/A.

High-Spatial-Resolution NDVI Reconstruction with GA-ANN.

The normalized differential vegetation index (NDVI) for Landsat is not continuous on the time scale due to the long revisit period and the influence of clouds and cloud shadows, such that the Landsat NDVI needs to be filled in and reconstructed. This study proposed a method based on the genetic algorithm-artificial neural network (GA-ANN) algorithm to reconstruct the Landsat NDVI when it has been affected by clouds, cloud shadows, and uncovered areas by relying on the MODIS characteristics for a wide coverage area. According to the self-validating results of the model test, the RMSE, MAE, and R were 0.0508, 0.0557, and 0.8971, respectively. Compared with the existing research, the reconstruction model based on the GA-ANN algorithm achieved a higher precision than the enhanced spatial and temporal adaptive reflectance fusion model (ESTARFM) and the flexible space-time data fusion algorithm (FSDAF) for complex land use types. The reconstructed method based on the GA-ANN algorithm had a higher root mean square error (RMSE) and mean absolute error (MAE). Then, the Sentinel NDVI data were used to verify the accuracy of the results. The validation results showed that the reconstruction method was superior to other methods in the sample plots with complex land use types. Especially on the time scale, the obtained NDVI results had a strong correlation with the Sentinel NDVI data. The correlation coefficient (R) of the GA-ANN algorithm reconstruction's NDVI and the Sentinel NDVI data was more than 0.97 for the land use types of cropland, forest, and grassland. Therefore, the reconstruction model based on the GA-ANN algorithm could effectively fill in the clouds, cloud shadows, and uncovered areas, and produce NDVI long-series data with a high spatial resolution.

Increased expression of prokineticin 2 and its receptor in endometrium of recurrent implantation failure patients decreased the expression of MMP9 important for decidualization.

BACKGROUND: Studies have shown that abnormalities in the decidualization process were closely related to recurrent implantation failure (RIF). Prokineticin 2 (PK2) is a secreted protein with angiogenic and tissue remodeling functions but its role in the endometrium is unknown. METHODS: PK2 levels and its receptor PKR1 mRNA and protein levels in mid-secretory endometrium from normal and RIF women were examined by real-time PCR and western blotting, respectively. The effects of PK2 were evaluated by overexpressed PK2 in immortalized endometrial T-HESC cells using lentivirus vector and found different expression of Matrix metalloproteinase 9(MMP9) and lncRNA LUCAT1 by RNA-sequencing. The ability of PK2 to regulate LUCAT1 and MMP9 was verified in endometrial cells by real-time PCR and western blotting. RESULTS: Using endometrial biopsies from normal and RIF patients, we found increased expression of PK2, together with its receptor PKR1 in RIF patients. We then overexpressed PK2 in immortalized endometrial T-HESC cells using lentivirus vector and found decreased expression of Matrix metalloproteinase 9(MMP9), and increased expression of lncRNA LUCAT1. We verified the ability of PK2 to stimulate LUCAT1 and decrease MMP9 in endometrial cells. We further demonstrated that increased expression of a long noncoding RNA LUCAT1 and decreased expression of MMP9 in endometrial biopsies of patients with RIF. Thus, we highlighted the important role of PK2 and its receptor PKR1 in decidualization and RIF. CONCLUSION: Prokineticin 2 and its receptor are important in endometrium decidualization. PK2 may affect endometrial decidualization through the LUCAT1- MMP9 pathway, thereby affecting embryo implantation.

Effects of seasonal variations and meteorological factors on IVF pregnancy outcomes: a cohort study from Henan Province, China.

OBJECTIVE: To investigate whether seasonal variations and meteorological factors influence pregnancy outcomes in women undergoing in vitro fertilization-embryo transfer (IVF-ET) treatment. DESIGN: Retrospective cohort study. SETTING: University-affiliated reproductive medical center. SUBJECTS: Women aged < 35 years undergoing IVF from June 1, 2015, to June 1, 2019. INTERVENTIONS: Cycles were divided into four groups according to the date of the beginning of ovulation induction: spring (659 cycles), summer (578 cycles), autumn (519 cycles), and winter (534 cycles). RESULTS: The high-quality embryo rate was higher in autumn and winter than in cycles in which ovulation induction occurred in spring and summer (58.70% vs. 58.78% vs. 62.67% vs. 63.42%; P < 0.001). The results of linear regression analysis showed that the high-quality embryo rate was significantly correlated with the daily average temperature of ovulation induction (P = 0.037). The clinical pregnancy rates of cycles starting ovulation induction in spring, summer, and autumn were significantly higher than those starting in winter (70.71% vs. 73.18% vs. 70.13% vs. 65.17%; P = 0.031), while the biochemical pregnancy rate, early abortion rate, and live birth rate were not significantly different (P > 0.050). Multivariate logistic regression analysis showed significant seasonal variation in clinical pregnancy (OR = 1.643, 95% CI = 1.203-2.243; P = 0.002), and that a higher daily average temperature at the time of ovulation induction increased the clinical pregnancy rate (OR = 1.012, 95% CI = 1.001-1.022; P = 0.031). CONCLUSIONS: In women younger than 35 years who undergo IVF treatment, the season and ambient temperature on the date of the beginning of ovulation induction may have an impact on embryo development and clinical pregnancy.

Is early-follicular long-acting GnRH agonist protocol an alternative for patients with polycystic ovary syndrome undergoing in vitro fertilization?

BACKGROUND: We aimed to compare the clinical and perinatal outcomes of patients with polycystic ovary syndrome (PCOS) undergoing in vitro fertilization/intracytoplasmic sperm injection (IVF/ICSI) treatment with either an early-follicular long-acting gonadotropin-releasing hormone agonist (GnRH-a) long protocol (EFLL) or a midluteal short-acting GnRH-a long protocol (MLSL). METHODS: This single-center, retrospective study, included patients with PCOS who underwent IVF/ICSI from January 2013 to June 2019 at the First Affiliated Hospital of Zhengzhou University, Zhengzhou, China. Patients underwent either MLSL (1179 cycles) or EFLL (2390 cycles). The primary outcomes were pregnancy outcomes, perinatal and maternal complications. RESULTS: Fresh embryo transfer (59.12% vs. 55.47%, P = 0.038), clinical pregnancy (75.23% vs. 53.82%, P = 0.001), and live birth rates (63.27% vs. 42.05%, P = 0.010) were higher in the EFLL group. However, the proportion of patients "freezing all" for high risk of ovarian hyperstimulation syndrome (OHSS) (24.27% vs. 32.06%, P = 0.001) and ectopic pregnancy (1.51% vs. 5.97%, P = 0.002) were lower in the EFLL group than in the MLSL group. The incidence of gestational diabetes was higher in the EFLL group than in the MLSL group (5.08% vs. 1.42%, RR 3.714, 95% confidence interval (CI) 1.474-9.360, P = 0.003). There were no significant differences in the incidence of hypertension, premature rupture of membranes, placenta previa, congenital heart disease, or neonatal weight between the two groups. Logistic regression results showed that age (OR 0.966, 95% CI 0.941-0.993, P = 0.013), treatments (OR 2.380, 95% CI 1.833-3.089, P = 0.001), and endometrial thickness on trigger day (OR 1.115, 95% CI 1.070-1.162, P = 0.001) were correlated with clinical pregnancy. Pre-pregnancy BMI (OR 1.098, 95% CI 1.002-1.204, P = 0.046), fasting plasma glucose (FPG) (OR 3.096, 95% CI 1.900-5.046, P = 0.001), and treatments (OR 3.458, 95% CI 1.359-8.800, P = 0.009) were correlated with gestational diabetes mellitus (GDM). Treatments (OR 0.291, 95% CI 0.148-0.575, P = 0.001) and endometrial thickness on trigger day (OR 0.834, 95% CI 0.722-0.962, P = 0.013) were correlated with ectopic pregnancy. CONCLUSION: The early-follicular long-acting GnRH agonist long protocol can be used as an ideal assisted reproductive technology (ART) pregnancy assistance program for patients with PCOS, but obese patients should be encouraged to lose weight before ART treatments to reduce the risk of GDM.

Effect of elevated progesterone levels the day before ovulation on pregnancy outcomes in natural cycles of frozen thawed embryo transfer.

OBJECTIVE: We aimed to analyze whether elevated progesterone levels on the day before ovulation affected pregnancy outcomes in natural cycles of frozen thawed embryo transfer (NC-FET). METHODS: A retrospective analysis was conducted in a public university hospital. Data on clinical pregnancy, live birth, ectopic pregnancy, and miscarriage rates were collected, along with other patient data. Patients were divided into two groups according to their progesterone levels the day before ovulation: the progesterone elevation (PE) group (progesterone level >1.0 ng/mL) and the normal progesterone (NP) group (progesterone level ≤1.0 ng/mL). We assessed the effect of elevated progesterone levels in NC-FET by performing multivariate logistic regression analysis. RESULTS: Overall 1159 women with tubal factor infertility who underwent NC-FET were enrolled, including 666 women who received cleavage-stage embryo transfers and 493 women who received blastocyst embryo transfers. When two cleavage-stage embryos were transferred, the clinical pregnancy rate was significantly higher in the PE than in the NP group following NC-FET (p < .05). After correcting for various confounders, we found that elevated progesterone levels (adjusted odds ratio [OR]: 1.672; 95% confidence interval [CI]: 1.089-2.566, p = .018) improved the clinical pregnancy rate following transfer of two cleavage-stage embryos but did not affect the pregnancy rate when blastocyst-stage embryos were transferred (adjusted OR: 0.856; 95% CI: 0.536-1.369; p = .517). CONCLUSIONS: The results showed that in patients undergoing cleavage-stage NC-FET, progesterone levels >1.0 ng/mL improved the clinical pregnancy rates. However, the level of progesterone had no effect on the clinical pregnancy rate for patients undergoing blastocyst-stage NC-FET.

Complex Uterine Cavity Abnormalities Increase the Risk of Miscarriage in In Vitro Fertilization/Intracytoplasmic Sperm Injection in Fresh Cycle-Assisted Pregnancies.

STUDY OBJECTIVE: Any abnormality of the uterine cavity can result in reduced endometrial receptivity, which negatively affects embryo implantation and leads to lower clinical pregnancy rates. The effects of improved uterine cavity environment on in vitro fertilization (IVF)/intracytoplasmic sperm injection (ICSI)-embryo transfer (ET) treatment outcome are unclear. This study aimed to investigate the impact of improved uterine cavity abnormalities on the pregnancy outcomes of infertile patients undergoing IVF/ICSI-ET. DESIGN: Retrospective study. SETTING: Single-center. PATIENTS: Women with infertility who underwent fresh cycles of IVF/ICSI-ET. INTERVENTIONS: We retrospectively analyzed the clinical data of 31 057 cycles of women with infertility undergoing IVF/ICSI-ET with hysteroscopy and treated at the First Affiliated Hospital of Zhengzhou University Reproductive Medicine Center from August 2009 to May 2018. According to the previous condition of their uterine cavity, patients were divided into the normal uterine cavity, single uterine cavity abnormality, and complex uterine cavity abnormality groups. Differences in general conditions and pregnancy-assisted outcomes were compared among the groups, which were screened according to propensity score matching. MEASUREMENTS AND MAIN RESULTS: In 3005 cycles after propensity score matching screening, there were no statistically significant differences in the implantation and clinical pregnancy rates of patients with successfully treated uterine cavity abnormalities and lesions (p > .05). The miscarriage rate was significantly higher in the complex uterine cavity abnormality group than in the normal (p = .001) and single uterine cavity abnormality groups (p = .002). Logistic regression analysis showed that the female partner's age (adjusted odds ratio, 1.12; 95% confidence interval, 1.05-1.19; p = .001) and history of intrauterine adhesions (adjusted odds ratio, 1.44; 95% confidence interval, 1.12-1.85; p = .005) were independent risk factors for miscarriage. CONCLUSION: The age of the female partner and history of intrauterine adhesions increased the miscarriage rate in patients undergoing IVF/ICSI-ET.

Hypermethylation-mediated downregulation of lncRNA PVT1 promotes granulosa cell apoptosis in premature ovarian insufficiency via interacting with Foxo3a.

Long noncoding RNA PVT1 is involved in the progression of female gynecological cancers. However, the role of PVT1 in ovarian granulosa cell apoptosis-mediated premature ovarian insufficiency (POI) remains unclear. This study aims to elucidate the role of PVT1 in ovarian granulosa cell apoptosis-mediated POI. The expression of PVT1 was compared between ovarian tissues from POI patients and normal controls. The methylation level in the PVT1 promoter region was detected by methylation-specific polymerase chain reaction. The interaction between PVT1 and forkhead box class O3A (Foxo3a) was confirmed by RNA pull-down and RNA immunoprecipitation assays. Granulosa cell apoptosis was detected using flow cytometry. The effect of PVT1 on transcription activity of Foxo3a was detected by luciferase reporter assay. The expression of PVT1 was low in the POI ovarian tissues compared with the controls, and such a low expression was related to the hypermethylation of the PVT1 promoter. PVT1 was localized in both the cytoplasm and the nucleus of granulosa cells. We determined that PVT1 could bind with Foxo3a and that downregulating PVT1 by small interfering RNAs inhibited Foxo3a phosphorylation by promoting SCP4-mediated Foxo3a dephosphorylation, resulting in an increase in Foxo3a transcription activity. Moreover, downregulating PVT1 promoted granulosa cell apoptosis by increasing the Foxo3a protein levels. An in vivo experiment showed that the injection of PVT1 overexpressing vectors restored the ovarian function in POI mice. Hypermethylation-induced downregulation of PVT1 promotes granulosa cell apoptosis in POI by inhibiting Foxo3a phosphorylation and increases the Foxo3a transcription activity.

Photosynthetic compensation of non-leaf organ stems of the invasive species Sphagneticola trilobata (L.) Pruski at low temperature.

Biological invasion is a hot topic in ecological research. Most studies on the physiological mechanisms of plants focus on leaves, but few studies focus on stems. To study the tolerance of invasive plant (Sphagneticola trilobata L.) to low temperature, relevant physiological indicators (including anthocyanin and chlorophyll) in different organs (leaves and stems) were analyzed, using a native species (Sphagneticola calendulacea L.) as the control. The results showed that, upon exposure to low temperature for 15 days, the stems of two Sphagneticola species were markedly reddened, their anthocyanin content increased, chlorophyll and chlorophyll fluorescence parameters decreased, and the accumulation of reactive oxygen species in the stem increased. The percentage increases of antioxidants and total antioxidant capacities in stems were significantly higher in S. trilobata than in S. calendulacea. This showed that S. trilobata had higher cold tolerance in stems while leaves were opposite. To further verify the higher cold tolerance of the stem of S. trilobata, a defoliation experiment was designed. We found that the defoliated stem of S. trilobata reduced anthocyanin accumulation and increased chlorophyll content, while alleviating membrane lipid damage and electrical conductivity, and the defoliated stem still showed an increase in stem diameter and biomass under low temperature. The discovery of the physiological and adaptive mechanisms of the stem of S. trilobata to low temperature will provide a theoretical basis for explaining how S. trilobata maintains its annual growth in South China. This is of great significance for predicting the future spread of cloned and propagated invasive plants.

Autotransplantation of the ovarian cortex after in-vitro activation for infertility treatment: a shortened procedure.

STUDY QUESTION: Is it possible to establish a new in-vitro activation (IVA) protocol for infertility treatment? SUMMARY ANSWER: A new IVA procedure is an efficient and easily performed approach for infertility treatment of patients with diminished ovarian reserve (DOR). WHAT IS KNOWN ALREADY: IVA of primordial follicles with or without stimulators has been developed to treat patients with primary ovarian insufficiency (POI) successfully. However, the efficiency of the procedure is still very low. There is a requirement to optimize the protocol with increased efficiency for clinical application. STUDY DESIGN, SIZE, DURATION: Newborn mouse ovaries were used to establish a new 1-h IVA protocol with the mechanistic target of rapamycin (mTOR) stimulator phosphatidic acid (PA, 200 µM) and the phosphatidylinositol-3-kinase (PI3K) stimulator 740Y-P (250 µg/ml); a prospective observational cohort study in POI patients was performed on 15 POI patients and 3 poor ovarian response (POR) patients in three different centers of reproductive medicine in China. PARTICIPANTS/MATERIALS, SETTING, METHODS: One-third of ovarian cortex was removed and processed into bigger strips (1 × 1 cm2, 1-2 mm thickness). Strips were then sutured back after treatment. The new approach only requires one laparoscopic surgery. MAIN RESULTS AND THE ROLE OF CHANCE: Follicular activation and development increased in cultured mouse and human ovarian tissues after 1 h of stimulator treatment. Compared with tiny ovarian cortex pieces (1 × 1 mm2), large ovarian strips (1 × 1 cm2) showed the lowest apoptotic signals after incubation. We applied the orthotropic transplantation procedure with large strips in the clinic, and 9 of 15 POI patients showed at least one-wave follicular growth during the monitoring period. One patient was reported with one healthy delivery after natural conception and another patient with a healthy singleton delivery after IVF. All the contacted patients (n = 13) responded with no side effects on their health 2-4 years after IVA procedure. LIMITATIONS, REASONS FOR CAUTION: Further clinical trials with a large number of well-defined patients are required to compare different IVA protocols. A long-term follow-up system should be set up to monitor patient's health in the future cohort study. WIDER IMPLICATIONS OF THE FINDINGS: By using stimulators, the findings in the study provide a more efficient IVA protocol for the treatment of patients with DOR. It requires only one laparoscopic surgery and thus minimizes patients' discomfort and costs. This strategy could be useful for patients diagnosed with POI and desire pregnancy as soon as possible after the operation. STUDY FUNDING/COMPETING INTEREST(S): This work was supported by the National Key Research and Development Program of China (2018YFC1003703 and 2018YFC1004203); the National Natural Science Foundation of China (81871221); Co-construction of Provincial Department (201601006). The authors have no conflict of interest to disclose. TRIAL REGISTRATION NUMBER: ChiCTR2000030872.

Comparison of nitrogen and VFA removal pathways in autotrophic and organotrophic anammox reactors.

Organotrophic anammox is a promising process for treating both nitrogen and organic containing wastewater than that of the traditional autotrophic anammox for sole nitrogen removal. However pathways of nitrogen removal particularly at metagenomic level in both processes are still unknown. Here we report, metabolic pathways of nitrogen removal in two lab-scale sequencing batch reactors (SBR), one autotrophic and another organotrophic (TOC/TN = 0.1) anammox bacteria incubated over 220 days. Both reactors showed satisfactory nitrogen removal with 840.31 mg N/L.d and 786.81 mg N/L.d for autotrophic and organotrophic anammox reactors respectively. Four anammox species namely Candidatus B. fulgida, B. sinica, J. caeni and Candidatus K. stuttgartiensis were identified in both reactors. The Candidatus K. stuttgartiensis (4%) was dominant in autotrophic reactor whereas Candidatus J. caeni (10%) in the organotrophic reactor. The supply of organic promoted the growth of anammox bacteria more than three times higher than that of the autotrophic anammox reactor. The functional genes related to the DNRA pathway was obtained in all anammox species except for Candidatus K. stuttgartiensis. The co-existence of other DNRA (Armatimonadetes and Thauera) and partial denitrifying bacteria (Chloroflexi) was also found in both reactors. Moreover, functional genes related to acetate metabolism by acetyl-CoA way were obtained in all anammox bacteria except Candidatus B. fulgida which showed alternative ackA/Pac-t pathways in organic anammox reactor. Overall current results suggest that the anammox, DNRA and partial denitrification were the key nitrogen transformation pathways, particularly in organotrophic anammox reactor. Our findings will improve understanding of the practical application of organotrophic anammox for wider wastewater treatment.

ARF4 regulates shoot regeneration through coordination with ARF5 and IAA12.

KEY MESSAGE: ARF4-regulated shoot regeneration through competing with ARF5 for the interaction with IAA12. Plant possess the ability to regenerate shoot meristem and subsequent the whole individual. This process is the foundation for in vitro propagation and genetic engineering and provides a system for studying fundamental biological questions, such as hormonal signaling. Auxin response factor (ARF) family transcription factors are critical components of auxin signaling pathway that regulate the transcription of target genes. To date, the mechanisms underlying the functions of class-B ARFs which act as transcription repressors remains unclear. In this study, we found that ARF4, the transcriptional repressor, was involved in regulating shoot regeneration. ARF4 interacted with auxin/Indole-3-Acetic-Acid12 (IAA12). The expression signals of ARF4 displayed a dynamic pattern similar with those of ARF5 and IAA12 during shoot meristem formation. Enhanced expression of IAA12 compromised the shoot regeneration capacity. Induced expression of ARF4 complemented the regeneration phenotype of IAA12-overexpression but did not rescued the defects in the arf5 mutant, mp-S319. Further analysis revealed that ARF4 competed with ARF5 for the interaction with IAA12. The results indicate that ARF4-regulated shoot regeneration through cooperating with ARF5 and IAA12. Our findings provided new information for deciphering the function of class-B ARFs.

Association between hyperandrogenism and adverse pregnancy outcomes in patients with different polycystic ovary syndrome phenotypes undergoing in vitro fertilization/intracytoplasmic sperm injection: a systematic review and meta-analysis.

OBJECTIVE: To study the association between hyperandrogenism (HA) and adverse pregnancy outcomes in patients with different polycystic ovary syndrome phenotypes undergoing in vitro fertilization (IVF)/intracytoplasmic sperm injection (ICSI). METHODS: We reviewed all eligible articles published up to October 2020 after searching in PubMed, Embase, Cochrane Library, Web of Science, Wanfang Data, and CNKI databases. The primary outcomes were the clinical pregnancy rate (CPR), miscarriage rate (MR), and live birth rate (LBR), whereas the secondary outcomes were the number of retrieved oocytes and endometrial thickness. Risk ratios (RRs) or mean differences with 95% confidence intervals (CIs) were calculated to estimate the HA impact on IVF/ICSI outcomes in patients with polycystic ovary syndrome (PCOS) phenotypes. RESULTS: Of the 789 trials identified, nine retrospective studies involving 3037 patients with PCOS were included. Compared to the PCOS group with normal androgen levels, the PCOS group with HA exhibited increased MR (RR: 1.56, 95% CI: 1.13, 2.16); the CPR (RR: 0.88, 95% CI: 0.77, 1.01) and LBR (RR: 0.79, 95% CI: 0.55, 1.11) were not significantly different between these groups. Subgroup analysis revealed that the CPR was lower in the polycystic ovarian (PCO)-morphology + HA + oligo-anovulation (AO) group than in the PCO + AO group (RR: 0.81, 95% CI: 0.67, 0.99). Among Asians, the PCOS/HA group had increased MR (RR: 1.56, 95% CI: 1.06, 2.31) and showed thinner endometrial thickness. However, among Caucasians, no differences were observed between the two groups. CONCLUSIONS: HA may have adverse effects on clinical pregnancy and miscarriage outcomes in different PCOS phenotypes, particularly among Asians.

Factors related to early spontaneous miscarriage during IVF/ICSI treatment: an analysis of 21,485 clinical pregnancies.

RESEARCH QUESTION: Which factors are related to early spontaneous miscarriage in IVF-conceived clinical pregnancies? DESIGN: A total of 21,485 clinical pregnancies were included in the analysis. First, early spontaneous miscarriage rates were compared among different groups according to female age, body mass index (BMI), number of previous miscarriages, infertility diagnosis and type and cycle characteristics. Then, the spontaneous miscarriage rate in patients with polycystic ovary syndrome (PCOS), uterus malformation and endometriosis was compared with that in patients with male factor infertility alone. Last, logistic regression was used to analyse factors affecting the early spontaneous miscarriage rate. RESULTS: Of the 21,485 cycles, 2703 cycles (12.58%) resulted in early spontaneous miscarriage. In patients <35 years old, those with uterus malformation or PCOS experienced significantly higher spontaneous miscarriage rates (14.44% versus 9.47%, P = 0.027; 11.43% versus 9.47%; P = 0.003) compared with controls (male factor only). In multivariate logistic regression analysis, the spontaneous miscarriage rate increased in frozen embryo transfer cycles in patients <35 years old (odds ratio [OR] 1.449, 95% confidence interval [CI] 1.303-1.611, P = 0.000), but decreased in patients ≥35 years old (OR 0.794, 95% CI 0.671-0.939, P = 0.007) compared with fresh cycles. CONCLUSIONS: Female age, number of previous miscarriages and endometrial thickness on the day of embryo transfer were independent factors associated with early spontaneous miscarriage. PCOS, uterus malformation and frozen embryo transfer significantly increased spontaneous miscarriage rate in patients <35 years old compared with male factor alone controls. However, frozen embryo transfer decreased the spontaneous miscarriage rate in patients ≥35 years old compared with fresh cycles.