Responses of Cynanchum taiwanianum and Its Bioactive Compound Biosynthesis to Levels of Nitrogen and Potassium Fertilization

Cynanchum taiwanianum is an important plant used in traditional medicine. The increasing demand and lack of information regarding its cultivation have become concerns for sustainability. This study examined the effects of nitrogen and potassium fertilization rates on the growth and biosynthesis of main bioactive compounds, including cynandione A and polyphenolic compounds, in field-cultivated C. taiwanianum. Two field experiments were conducted using three levels of nitrogen (N100, N150 and N200) and three levels of potassium (K100, K150 and K200) treatments. The experimental variables were either N or K fertilizer. The results showed that, aside from N200, N and K fertilization significantly increased C. taiwanianum shoot and tuber biomass. High N fertilization resulted in low total phenolic and total flavonoid contents in shoots and tubers, but the effects of K fertilization were minimal. Cynandione A, an important bioactive compound, was only detected in tubers; its content were enhanced with the increasing K fertilization, but reduced with excess N fertilization (N200). Although N and K fertilizers are important for C. taiwanianum tuber production, the yield of cynandione A was associated with K but not N fertilization rates. These results provide some essential information for the optimal production of C. taiwanianum tubers and functional compounds. Further studies are required to examine the mechanism(s) of cynandione A biosynthesis and its compartmentation in plant tissues.

Impact of Living in Different Altitudes on Intracytoplasmic Sperm Injection (ICSI) Outcomes for Infertile Couples in Southwestern Region of Saudi Arabia, a Retrospective Study

Objective: The aim of this study was to determine the effect of different altitudes in the Southwestern region of Saudi Arabia on ICSI outcomes, fertilization rate, embryo quality, pregnancy rate, and miscarriage rates for infertile couples. Materials and Methods: This is a retrospective study on 551 infertile couples carried out in the Assisted Reproductive Technology unit at the Maternity and Childern Hospital in Abha, Saudi Arabia between 2018 and 2019 to compare ICSI outcomes in different altitudes. Low altitude (205 patients), mild altitude (86 patients) and high altitude (260 patients). Main result measurements: fertilization rates, embryo quality, clinical pregnancy and miscarriage rates at different altitudes. Results: The data showed that there were no significant differences (P>0.67) in fertilization rates and the number of good-quality embryos between different altitudes. Importantly, clinical pregnancy rates were similar between groups and there was no significant difference in the miscarriage rates between high, mild and low altitudes. Conclusions: This study demonstrates that there is now increased risk of miscarriage or low pregnancy rates with different altitudes in Southwestern region of Saudi Arabia. This suggests that altitude changes has no obvious risk on pregnancy rate and pregnancy outcome.

Randomized allocation of oocytes to IVF or ICSI for IVF-naïve cases with unexplained infertility in an IVF-ICSI Split protocol favors ICSI to optimize live birth outcomes

In assisted reproduction treatments (ART), applying the ICSI method for fertilization of oocytes rather than traditional IVF method, is regarded as controversial for two reasons, namely utility and safety. Our study examines an IVF-ICSI Split model for couples with unexplained infertility, where male factor is meticulously excluded and ART is conducted by a strict algorithm, a commitment to blastocyst culture, along with single embryo transfers and a high commitment to cryopreservation. From 242 treatment cycles, 3346 oocytes recovered (13.8 per OPU) were randomly allocated to IVF or ICSI and the fertilization rates standardized to the number of 2PNS arising from each group applying the metaphase II oocyte number identified for the ICSI group, as the denominator for both groups. The fertilization rates were significantly higher overall for ICSI (83.2% vs 65.4%; p<0.0001), being most pronounced for women under 40 years. The resultant embryos had equivalent implantation rates in both fresh ET and frozen (FET) cycles with no significant differences in pregnancy rates, miscarriage rates or live birth outcomes indicating equivalent embryo quality. However, there were significantly higher numbers of ICSI-generated embryos cryopreserved and subsequent FET procedures showed higher live birth rates (21 births vs 6 births; p<0.005) and potential livebirths (214 births vs 104 births; p<0.0001). No congenital fetal abnormalities were detected in any of the 199 babies delivered during the study period to December 2020, neither IVF-generated nor ICSI-generated. Whilst the data strongly favors ICSI, there were 2 women (from 26 with fertilization in one arm only) who demonstrated fertilization only in the IVF arm of the study. We conclude that the IVF-ICSI Split model should be undertaken on all IVF-naïve women with unexplained infertility to determine the appropriate fertilization mode, albeit ICSI will be safely preferred for >90% of cases.

Microbial Signatures in Fertile Soils Under Long-Term N Management

Long-term reliance on inorganic N to maintain and increase crop yields in overly simplified cropping systems in the U.S. Midwest region has led to soil acidification, potentially damaging biological N2 fixation and accelerating potential nitrification activities. Building on this published work, rRNA gene-based analysis via Illumina technology with QIIME 2.0 processing was used to characterize the changes in microbial communities associated with such responses. Amplicon sequence variants (ASVs) for each archaeal, bacterial, and fungal taxa were classified using the Ribosomal Database Project (RDP). Our goal was to identify bioindicators from microbes responsive to crop rotation and N fertilization rates following 34–35 years since the initiation of experiments. Research plots were established in 1981 with treatments of rotation [continuous corn (Zea mays L.) (CCC) and both the corn (Cs) and soybean (Glycine max L. Merr.) (Sc) phases of a corn-soybean rotation], and of N fertilization rates (0, 202, and 269 kg N/ha) arranged as a split-plot in a randomized complete block design with three replications. We identified a set of three archaea, and six fungal genera responding mainly to rotation; a set of three bacteria genera whose abundances were linked to N rates; and a set with the highest number of indicator genera from both bacteria (22) and fungal (12) taxa responded to N fertilizer additions only within the CCC system. Indicators associated with the N cycle were identified from each archaeal, bacterial, and fungal taxon, with a dominance of denitrifier- over nitrifier- groups. These were represented by a nitrifier archaeon Nitrososphaera, and Woesearchaeota AR15, an anaerobic denitrifier. These archaea were identified as part of the signature for CCC environments, decreasing in abundance with rotated management. The opposite response was recorded for the fungus Plectosphaerella, a potential N2O producer, less abundant under continuous corn. N fertilization in CCC or CS systems decreased the abundance of the bacteria genera Variovorax and Steroidobacter, whereas Gp22 and Nitrosospira only showed this response under CCC. In this latter system, N fertilization resulted in increased abundances of the bacterial denitrifiers Gp1, Denitratisoma, Dokdonella, and Thermomonas, along with the fungus Hypocrea, a known N2O producer. The identified signatures could help future monitoring and comparison across cropping systems as we move toward more sustainable management practices. At the same time, this is needed primary information to understand the potential for managing the soil community composition to reduce nutrient losses to the environment.

Reproductive barriers in cassava: Factors and implications for genetic improvement

Cassava breeding is hampered by high flower abortion rates that prevent efficient recombination among promising clones. To better understand the factors causing flower abortion and propose strategies to overcome them, we 1) analyzed the reproductive barriers to intraspecific crossing, 2) evaluated pollen-pistil interactions to maximize hand pollination efficiency, and 3) identified the population structure of elite parental clones. From 2016 to 2018, the abortion and fertilization rates of 5,748 hand crossings involving 91 parents and 157 progenies were estimated. We used 16,300 single nucleotide polymorphism markers to study the parents’ population structure via discriminant analysis of principal components, and three clusters were identified. To test for male and female effects, we used a mixed model in which the environment (month and year) was fixed, while female and male (nested to female) were random effects. Regardless of the population structure, significant parental effects were identified for abortion and fertilization rates, suggesting the existence of reproductive barriers among certain cassava clones. Matching ability between cassava parents was significant for pollen grains that adhered to the stigma surface, germinated pollen grains, and the number of fertilized ovules. Non-additive genetic effects were important to the inheritance of these traits. Pollen viability and pollen-pistil interactions in cross- and self-pollination were also investigated to characterize pollen-stigma compatibility. Various events related to pollen tube growth dynamics indicated fertilization abnormalities. These abnormalities included the reticulated deposition of callose in the pollen tube, pollen tube growth cessation in a specific region of the stylet, and low pollen grain germination rate. Generally, pollen viability and stigma receptivity varied depending on the clone and flowering stage and were lost during flowering. This study provides novel insights into cassava reproduction that can assist in practical crossing and maximize the recombination of contrasting clones.

Ovarian Endometrioma Negatively Impacts Oocyte Quality and Quantity But Not Pregnancy Outcomes in Women Undergoing IVF/ICSI Treatment: A Retrospective Cohort Study

PurposeTo determine the impact of ovarian endometrioma per se on in vitro fertilization/intracytoplasmic sperm injection (IVF/ICSI) outcomes.MethodsThis retrospective study was conducted using two groups. The endometrioma group consisted of 862 women with infertility who had ovarian endometriomas and underwent their first ovarian stimulation for IVF/ICSI treatment between January 2011 to December 2019 at a public university hospital. A non-endometrioma comparison group, comprising 862 women with other infertility factors, was matched according to maternal age, body mass index (BMI), and infertility duration. Ovarian reserve and response and IVF/ICSI and pregnancy outcomes between the two groups were analyzed. Multivariate logistic regression (MLR) analysis was conducted on the basis of clinical covariates assessed for their association with live birth.ResultsThe results showed that significantly lower antral follicle count (AFC), anti-Müllerian hormone (AMH), ovarian sensitivity index (OSI), oocyte maturation and fertilization rates, blastocyst rate, number of oocytes retrieved, and available embryos were found in women with endometrioma compared with the control, respectively (P &lt; 0.05). The cumulative live birth rate per patient in women with endometrioma was lower than that of women without endometrioma (39.32% vs. 46.87%, P = 0.002). In women with endometrioma, those who underwent surgical intervention prior to IVF/ICSI treatment had higher maturation (86.03% vs. 83.42%, P = 0.003), fertilization (78.16% vs. 74.93%, P = 0.004), and top-quality embryo rates (42.94% vs. 39.93%, P = 0.097) but had fewer oocytes retrieved (8.01 ± 5.70 vs. 9.12 ± 6.69, P = 0.013) than women without surgery. However, live birth rates were comparable between women with endometrioma and women in the control group, regardless of whether they had a prior history of ovarian surgery. MLR analysis showed no correlation between endometrioma per se and live birth after being adjusted for number of top-quality embryos transferred and stage of embryo transfer.ConclusionsThe data from this study supported the conclusion that ovarian endometrioma negatively impacts oocyte quality and quantity, but not overall pregnancy outcomes, in women undergoing IVF/ICSI treatment. Endometrioma lowers the cumulative live birth rate by decreasing the number of embryos. Surgical excision of endometrioma prior to IVF/ICSI can partly improve oocyte maturation and fertilization rates but not pregnancy outcomes.

A simple and economic protocol for efficient in vitro fertilization using cryopreserved mouse sperm

The advent of genome editing tools like CRISPR/Cas has substantially increased the number of genetically engineered mouse models in recent years. In support of refinement and reduction, sperm cryopreservation is advantageous compared to embryo freezing for archiving and distribution of such mouse models. The in vitro fertilization using cryopreserved sperm from the most widely used C57BL/6 strain has become highly efficient in recent years due to several improvements of the procedure. However, purchase of the necessary media for routine application of the current protocol poses a constant burden on budgetary constraints. In-house media preparation, instead, is complex and requires quality control of each batch. Here, we describe a cost-effective and easily adaptable approach for in vitro fertilization using cryopreserved C57BL/6 sperm. This is mainly achieved by modification of an affordable commercial fertilization medium and a step-by-step description of all other necessary reagents. Large-scale comparison of fertilization rates from independent lines of genetically engineered C57BL/6 mice upon cryopreservation and in vitro fertilization with our approach demonstrated equal or significantly superior fertilization rates to current protocols. Our novel SEcuRe (Simple Economical set-up for Rederivation) method provides an affordable, easily adaptable and harmonized protocol for highly efficient rederivation using cryopreserved C57BL/6 sperm for a broad application of colony management in the sense of the 3Rs.

Lower Spermatozoal PIWI-LIKE 1 and 2 Transcript Levels Are Significantly Associated with Higher Fertilization Rates in IVF

The four human PIWI-LIKE gene family members PIWI-LIKE 1–4 play a pivotal role in stem cell maintenance and transposon repression in the human germline. Therefore, dysregulation of these genes negatively influences the genetic stability of the respective germ cell and subsequent development and maturation. Recently, we demonstrated that a lower PIWI-LIKE 2 mRNA expression in ejaculated spermatozoa is more frequent in men with oligozoospermia. In this study, we analysed how PIWI-LIKE 1–4 mRNA expression in ejaculated spermatozoa predicts ART outcome. From 160 IVF or ICSI cycles, portions of swim-up spermatozoa used for fertilization were collected, and the total RNA was isolated. PIWI-LIKE 1–4 mRNA expression was measured by qPCR using TaqMan probes with GAPDH as a reference gene. PIWI-LIKE 1 and 2 transcript levels in the spermatozoa of the swim-up fraction were positively correlated to each other (rS = 0.78; p < 0.001). Moreover, lower PIWI-LIKE 2 mRNA levels, as well as lower PIWI-LIKE 1 mRNA levels, in these spermatozoa were positively associated with a fertilization rate ≥ 50% in the respective ART cycles (p = 0.02 and p = 0.0499, Mann–Whitney U-Test). When separately analysing IVF and ICSI cycles, PIWI-LIKE 1 and 2 transcript levels were only significantly associated to increased fertilization rates in IVF, yet not in ICSI cycles. Spermatozoal PIWI-LIKE 3 and 4 transcript levels were not significantly associated to fertilization rates in ART cycles. In conclusion, lower levels of spermatozoal PIWI-LIKE 1 and 2 mRNA levels are positively associated with a higher fertilization rate in IVF cycles.

Tuning Potassium and Magnesium Fertilization of Potato in the South of West Siberia

Imbalance of nutrients limits crop yields. Although K fertilization receives sufficient attention in research and practice, Mg supply is rather neglected. The effect of Mg fertilization (0, 5 and 10 g Mg/m2), combined with two K fertilization rates (10 and 15 g K/m2), on potato production and soil exchangeable K and Mg was studied in a three-season microplot field experiment in the Novosibirsk region, Russia. Tuber yield did not respond to the increased K fertilization, but increased at 5 and decreased at 10 g Mg/m2. Total Mg concentration in tubers increased at 15 g K/m2, whereas N, P and K were not affected by fertilization. The tuber yield was maximal (3.6 kg/m2) at 10 g K/m2 and 5 g Mg/m2. Soil exchangeable Mg increased by the year, resulting in preferential development of the aboveground phytomass due to apparently increased Mg availability and K/Mg imbalance. Potato production depended on the year, strongly implicating weather conditions. Therefore, the weather and the chemical nature of K and Mg fertilizers (as pertinent to their release mode from fertilizer in soil), are important for balancing their proportions and amounts while assessing interactions among nutrients in potato production and adjusting regional fertilization strategies.