Conserved genes regulating human sex differentiation, gametogenesis and fertilization.

The study of the functional genome in mice and humans has been instrumental for describing the conserved molecular mechanisms regulating human reproductive biology, and for defining the etiologies of monogenic fertility disorders. Infertility is a reproductive disorder that includes various conditions affecting a couple's ability to achieve a healthy pregnancy. Recent advances in next-generation sequencing and CRISPR/Cas-mediated genome editing technologies have facilitated the identification and characterization of genes and mechanisms that, if affected, lead to infertility. We report established genes that regulate conserved functions in fundamental reproductive processes (e.g., sex determination, gametogenesis, and fertilization). We only cover genes the deletion of which yields comparable fertility phenotypes in both rodents and humans. In the case of newly-discovered genes, we report the studies demonstrating shared cellular and fertility phenotypes resulting from loss-of-function mutations in both species. Finally, we introduce new model systems for the study of human reproductive biology and highlight the importance of studying human consanguineous populations to discover novel monogenic causes of infertility. The rapid and continuous screening and identification of putative genetic defects coupled with an efficient functional characterization in animal models can reveal novel mechanisms of gene function in human reproductive tissues.

Implementation of a safer conception programme for HIV-affected men and women in rural Uganda.

We integrated safer conception care into a Ugandan HIV clinic. People with HIV (PWH), or partnered with a PWH, and desiring children were eligible for the Healthy Families Clinic Program. Clients completed quarterly safer conception counselling visits and questionnaires to provide information around method preferences and outcomes (partner pregnancy, partner seroconversion). We used clinic level data to evaluate longitudinal viral suppression among PWH. Between November 2016 and January 2020, 361 clients (53% men) accessed services. 75% were PWH (51% women, 96% men): 99% were on antiretroviral therapy (ART) and most reported HIV-sero-different partnerships (97%). Frequently selected safer conception methods included ART (86%), timed condomless sex (74%), and PrEP (40%) with important differences by HIV-serostatus and gender. 22.5% reported pregnancy. Most (97%) PWH were virally suppressed at enrolment and 81% of non-virally suppressed PWH were virally suppressed at 15 months. Two HIV-negative clients (2%) had HIV seroconversion. There is demand for safer conception care in a public sector HIV-clinic in Uganda. Men and women have unique safer conception care preferences. The majority of clients engaged in safer conception care had viral suppression at follow up.

Recent advances in early pregnancy loss diagnosis in dairy cows: New approaches.

Early pregnancy loss is a primary cause of low reproductive rates in dairy cows, posing severe economic losses to dairy farming. The accurate diagnosis of dairy cows with early pregnancy loss allows for oestrus synchronization, shortening day open, and increasing the overall conception rate of the herd. Several techniques are available for detecting early pregnancy loss in dairy cows, including rectal ultrasound, circulating blood progesterone, and pregnancy-associated glycoproteins (PAGs). Yet, there is a need to improve on existing techniques and develop novel strategies to identify cows with early pregnancy loss accurately. This manuscript reviews the applications of rectal ultrasound, circulating blood progesterone concentration, and PAGs in the diagnosis of pregnancy loss in dairy cows. The manuscript also discusses the recent progress of new technologies, including colour Doppler ultrasound (CDUS), interferon tau-induced genes (ISGs), and exosomal miRNA in diagnosing pregnancy loss in dairy cows. This study will provide an option for producers to re-breed cows with pregnancy loss, thereby reducing the calving interval and economic costs. Meanwhile, this manuscript might also act as a reference for exploring more economical and precise diagnostic technologies for early pregnancy loss in dairy cows.

Estimation of genetic parameters for bull conception rate and its genetic correlations with semen production traits in Japanese Black bulls.

The P of achieving pregnancy is an important trait of bull fertility in beef cattle and is defined as the bull conception rate (BCR). This study aimed to clarify and better understand the genetic architecture of the BCR calculated using artificial insemination and pregnancy diagnosis records from a progeny testing program in Japanese Black bulls. In this study, we estimated the genetic parameters of the BCR and their correlation with semen production traits. In addition, we assessed the correlated responses in BCR by considering the selection of semen production traits. Nine hundred and sixteen Japanese Black bulls were selected based on fertility, with 28 869 pregnancy diagnostic records from the progeny testing program. Our results showed that the heritability estimate was 0.04 in the BCR at the first service and 0.14 in BCR for the three services, and an increase in the inbreeding coefficient led to a significant decrease in BCR. The phenotypic trend of BCR remained almost constant over the years, whereas the genetic trend increased. In addition, the changes in the progeny testing year effect showed a similar tendency to the phenotypic trends, suggesting that the phenotypic trends could be mainly due to non-genetic effects, including progeny testing year effects. The estimated genetic correlation of BCR with sperm motility traits was favorably moderate to high (ranging from 0.49 to 0.97), and those with sperm quantity traits such as semen volume were favorably low to moderate (ranging from 0.23 to 0.51). In addition, the correlated responses in BCR at the first service by selection for sperm motility traits resulted in a higher genetic gain than direct selection. This study provides new insights into the genetic factors affecting BCR and the possibility of implementing genetic selection to improve BCR by selecting sperm motility traits in Japanese Black bulls.

Influence of Fertilization Methods and Types on Wheat Rhizosphere Microbiome Community and Functions.

To investigate the effects of fertilization methods and types on wheat rhizosphere microorganisms, macroelement (N, K) and microelement (Zn) fertilizers were applied on wheat by foliar spraying (FS) and root irrigation (RI) methods in a field experiment. The results indicated that fertilization methods and types can have significant impacts on the diversity and structure of rhizospheric microorganisms in wheat. The application method produced more significant effects than the fertilizer type. RI-N played a more important role in improving the wheat yield and quality and affected the changes in some nitrogen-fixing bacterial communities. Finally, eight strains of bacteria belonging to Pseudomonas azotoformans and P. cedrina showed positive effects on the growth of wheat seedlings. Overall, our study provides a better understanding of the dynamics of wheat rhizosphere microbial communities and their relation to fertilization, yield, and quality, showing that plant growth-promoting rhizobacteria with nitrogen fixing may be a potential approach for more sustainable agriculture production.

SMA20/PMIS2 Is a Rapidly Evolving Sperm Membrane Alloantigen with Possible Species-Divergent Function in Fertilization.

Immunodominant alloantigens in pig sperm membranes include 15 known gene products and a previously undiscovered Mr 20,000 sperm membrane-specific protein (SMA20). Here we characterize SMA20 and identify it as the unannotated pig ortholog of PMIS2. A composite SMA20 cDNA encoded a 126 amino acid polypeptide comprising two predicted transmembrane segments and an N-terminal alanine- and proline (AP)-rich region with no apparent signal peptide. The Northern blots showed that the composite SMA20 cDNA was derived from a 1.1 kb testis-specific transcript. A BLASTp search retrieved no SMA20 match from the pig genome, but it did retrieve a 99% match to the Pmis2 gene product in warthog. Sequence identity to predicted PMIS2 orthologs from other placental mammals ranged from no more than 80% overall in Cetartiodactyla to less than 60% in Primates, with the AP-rich region showing the highest divergence, including, in the extreme, its absence in most rodents, including the mouse. SMA20 immunoreactivity localized to the acrosome/apical head of methanol-fixed boar spermatozoa but not live, motile cells. Ultrastructurally, the SMA20 AP-rich domain immunolocalized to the inner leaflet of the plasma membrane, the outer acrosomal membrane, and the acrosomal contents of ejaculated spermatozoa. Gene name search failed to retrieve annotated Pmis2 from most mammalian genomes. Nevertheless, individual pairwise interrogation of loci spanning Atp4a-Haus5 identified Pmis2 in all placental mammals, but not in marsupials or monotremes. We conclude that the gene encoding sperm-specific SMA20/PMIS2 arose de novo in Eutheria after divergence from Metatheria, whereupon rapid molecular evolution likely drove the acquisition of a species-divergent function unique to fertilization in placental mammals.

Parthenogenesis, identity, and value.

Parthenogenesis is a form of asexual reproduction in which a gamete (ovum or sperm) develops without being fertilized. Tomer Jordi Chaffer uses parthenogenesis to challenge Don Marquis' future-like-ours (FLO) argument against abortion. According to Marquis, (1) what makes it morally wrong to kill us is that it would deprive us of a possible future that we might come to value-a future "like ours" (FLO) and (2) human fetuses are numerically identical to any adult human organism they may develop into, and thus have a FLO. Chaffer contends that if human ova are capable of parthenogenesis, then they would have a FLO, which contraception may deprive them of, but contends this is absurd. Bruce P. Blackshaw challenges Chaffer, contending sexually fertilized embryos are not identical to unfertilized ovum, but this would yield a more absurd implication, that fertilization deprives an ovum of a FLO! Here I show Marquis' account of identity rules out both Chaffer's and Blackshaw's accounts.

Effect of intravenous anesthetic drugs on fertilization rate in oocyte retrieval.

BACKGROUND: The purpose of this study was to investigate the effects of intravenous anesthetic drugs on fertilization rate in subjects receiving oocyte retrieval by assisted reproduction technology (ART). METHODS: A retrospective cohort study was designed. The clinical information of subjects who received oocyte retrieval procedure was collected. The subjects were divided into two groups based on the type of anesthesia used: the no-anesthesia group and the intravenous anesthesia group. Propensity score matching (PSM) was performed and multiple linear regression analyses were conducted. Fertilization rate was compared between the two groups before and after PSM. RESULTS: A total of 765 subjects were divided into two groups: the no-anesthesia group (n = 482) and the intravenous anesthesia group (n = 283). According to propensity scores, 258 pairs of subjects were well matched, and the baseline data between the two groups were not significantly different (P > 0.05). Fertilization rate was 77% in the intravenous anesthesia group, and 76% in the no-anesthesia group, without significant between-group difference (P = 0.685). Before matching, Poisson regression analysis showed no effect of intravenous anesthetic drugs on fertilization rate (RR = 0.859, 95%CI: 0.59 to 1.25, P = 0.422). After matching, no difference was found either (RR = 0.935, 95%CI: 0.67 to 1.29, P = 0.618). CONCLUSION: Intravenous anesthetic drugs may exert no effects on fertilization rate in subjects receiving ART.

Deciphering dynamic interactions between spermatozoa and the ovarian microenvironment through integrated multi-omics approaches in viviparous Sebastes schlegelii.

The ovarian microenvironment plays a crucial role in ensuring the reproductive success of viviparous teleosts. However, the molecular mechanism underlying the interaction between spermatozoa and the ovarian microenvironment has remained elusive. This study aimed to contribute to a better understanding of this process in black rockfish (Sebastes schlegelii) using integrated multi-omics approaches. The results demonstrated significant upregulation of ovarian complement-related proteins and pattern recognition receptors, along with remodeling of glycans on the surface of spermatozoa at the early spermatozoa-storage stage (1 month after mating). As spermatozoa were stored over time, ovarian complement proteins were progressively repressed by tryptophan and hippurate, indicating a remarkable adaptation of spermatozoa to the ovarian microenvironment. Before fertilization, a notable upregulation of cellular junction proteins was observed. The study revealed that spermatozoa bind to ZPB2a protein through GSTM3 and that ZPB2a promotes spermatozoa survival and movement in a GSTM3-dependent manner. These findings shed light on a key mechanism that influences the dynamics of spermatozoa in the female reproductive tract, providing valuable insights into the molecular networks regulating spermatozoa adaptation and survival in species with internal fertilization.

Strigolactones Might Regulate Ovule Development after Fertilization in Xanthoceras sorbifolium.

Strigolactones (SLs) were recently defined as a novel class of plant hormones that act as key regulators of diverse developmental processes and environmental responses. Much research has focused on SL biosynthesis and signaling in roots and shoots, but little is known about whether SLs are produced in early developing seeds and about their roles in ovule development after fertilization. This study revealed that the fertilized ovules and early developing pericarp in Xanthoceras sorbifolium produced minute amounts of two strigolactones: 5-deoxystrigol and strigol. Their content decreased in the plants with the addition of exogenous phosphate (Pi) compared to those without the Pi treatment. The exogenous application of an SL analog (GR24) and a specific inhibitor of SL biosynthesis (TIS108) affected early seed development and fruit set. In the Xanthoceras genome, we identified 69 potential homologs of genes involved in SL biological synthesis and signaling. Using RNA-seq to characterize the expression of these genes in the fertilized ovules, 37 genes were found to express differently in the fertilized ovules that were aborting compared to the normally developing ovules. A transcriptome analysis also revealed that in normally developing ovules after fertilization, 12 potential invertase genes were actively expressed. Hexoses (glucose and fructose) accumulated at high concentrations in normally developing ovules during syncytial endosperm development. In contrast, a low ratio of hexose and sucrose levels was detected in aborting ovules with a high strigolactone content. XsD14 virus-induced gene silencing (VIGS) increased the hexose content in fertilized ovules and induced the proliferation of endosperm free nuclei, thereby promoting early seed development and fruit set. We propose that the crosstalk between sugar and strigolactone signals may be an important part of a system that accurately regulates the abortion of ovules after fertilization. This study is useful for understanding the mechanisms underlying ovule abortion, which will serve as a guide for genetic or chemical approaches to promote seed yield in Xanthoceras.

Medical treatment for early pregnancy loss following in vitro fertilization compared to spontaneous pregnancies.

BACKGROUND: Misoprostol is a well-studied medical treatment for early pregnancy loss (EPL), with success rates ranging between 70 and 90%. However, treatment failure is associated with major patient discomfort, including the need for surgical intervention to evacuate the uterus. It was previously reported that medical treatment was especially successful among women who conceived after in vitro fertilization (IVF). We aimed to study if there is a difference in rates of medical treatment failures for EPL between pregnancies conceived by IVF and spontaneous pregnancies. METHODS: In this retrospective cohort study, we included all women who underwent medical treatment for EPL at our institute between 07/2015 and 12/2020. Treatment outcome was compared between IVF and spontaneous pregnancies. Treatment failure was defined as a need for surgical intervention, namely, dilation & curettage (D&C) and/or hysteroscopy, due to retained products of conception, which was defined as a gestational sac or endometrial thickness greater than 15 mm in a TVS scan. RESULTS: Overall, 775 patients were included, of which 195 (169/775 = 25.1%) ultimately required surgical intervention. There was no difference between the study groups in the rate of treatment failure. However, among IVF pregnancies, the rate of emergency D&C was lower (3.6% vs. 9.8%, p = 0.001), compared to spontaneous group. CONCLUSION: In cases of medical treatment for EPL, IVF pregnancies had no differences in rates of treatment failure compared to spontaneous pregnancies. That being said, IVF pregnancies have lower chances to undergo emergency D&C, compared to spontaneous pregnancies.

N-fertilization and disturbance exert long-lasting complex legacies on subarctic ecosystems.

Subarctic ecosystems are subjected to increasing nitrogen (N) enrichment and disturbances that induce particularly strong effects on plant communities when occurring in combination. There is little experimental evidence on the longevity of these effects. We applied N-fertilization (40 kg urea-N ha-1 year-1 for 4 years) and disturbance (removal of vegetation and organic soil layer on one occasion) in two plant communities in a subarctic forest-tundra ecotone in northern Finland. Within the first four years, N-fertilization and disturbance increased the share of deciduous dwarf shrubs and graminoids at the expense of evergreen dwarf shrubs. Individual treatments intensified the other's effect resulting in the strongest increase in graminoids under combined N-fertilization and disturbance. The re-analysis of the plant communities 15 years after cessation of N-fertilization showed an even higher share of graminoids. 18 years after disturbance, the total vascular plant abundance was still substantially lower and the share of graminoids higher. At the same point, the plant community composition was the same under disturbance as under combined N-fertilization and disturbance, indicating that multiple perturbations no longer reinforced the other's effect. Yet, complex interactions between N-fertilization and disturbance were still detected in the soil. We found higher organic N under disturbance and lower microbial N under combined N-fertilization and disturbance, which suggests a lower bioavailability of N sources for soil microorganisms. Our findings support that the effects of enhanced nutrients and disturbance on subarctic vegetation persist over decadal timescales. However, they also highlight the complexity of plant-soil interactions that drive subarctic ecosystem responses to multiple perturbations across varying timescales.

Optimization of the morphological, structural, and physicochemical properties of maize starch using straw returning and nitrogen fertilization in Northeast China.

The combination of straw returning and nitrogen (N) fertilization is a popular tillage mode and essential strategy for achieving stable yield and high quality. However, the optimal combination strategy and the influence of tillage mode on the morphological, crystalline, and molecular structures of maize starch remain unclear. We conducted a long-term field experiment over 7 years in Northeast China using two tillage modes, rotary tillage with straw returning (RTS) and plow tillage with straw returning (PTS), and four N application rates. The relative crystallinity, 1045/1022 cm-1 value, and B2 and B3 chains of maize starch were higher under RTS than under PTS, resulting in increased stability of starch and improvements in gelatinization enthalpy and temperature. The surface of the starch granules induced by N fertilizer was smoother than that under the N0 (0 kg N ha-1) treatment. The proportion of amylose content, solubility, swelling power, and light transmittance increased under N2 (262 kg N ha-1) treatment, along with improvement in starch pasting properties. These results suggest that RTS combined with N2 treatment can regulate the morphological, structural, and physicochemical characteristics of maize starch, providing an essential reference for improving the quality of maize starch from an agronomic point of view.

Microbial mechanisms in nitrogen fertilization: Modulating the re-mobilization of clay mineral-bound cadmium in agricultural soils.

Soil cadmium (Cd) can affect crop growth and food safety, and through the enrichment in the food chain, it ultimately poses a risk to human health. Reducing the re-mobilization of Cd caused by the release of protons and acids by crops and microorganisms after stabilization is one of the significant technical challenges in agricultural activities. This study aimed to investigate the re-mobilization of stabilized Cd within the clay mineral-bound fraction of soil and its subsequent accumulation in crops utilizing nitrogen ammonium nitrogen (NH4+-N) and nitrate nitrogen (NO3--N), at 60 and 120 mg kg-1. Furthermore, the study harvested root exudates at various growth stages to assess their direct influence on the re-mobilization of stabilized Cd and to evaluate the indirect effects mediated by soil microorganisms. The results revealed that, in contrast to the NO3--N treatment, the NH4+-N treatment significantly enhanced the conversion of clay mineral-bound Cd in the soil to NH4NO3-extractable Cd. It also amplified the accumulation of Cd in edible amaranth, with concentrations in roots and shoots rising from 1.7-6.0 mg kg-1 to 4.3-9.8 mg kg-1. The introduction of NH4+-N caused a decrease in the pH value of the rhizosphere soil and stimulated the production and secretion organic and amino acids, such as oxalic acid, lactic acid, stearic acid, succinic acid, and l-serine, from the crop roots. Furthermore, compared to NO3--N, the combined interaction of root exudates with NH4+-N has a more pronounced impact on the abundance of microbial genes associated with glycolysis pathway and tricarboxylic acid cycle, such as pkfA, pfkB, sucB, sucC, and sucD. The effects of NH4+-N on crops and microorganisms ultimately result in a significant increase in the re-mobilization of stabilized Cd. However, the simulated experiments showed that microorganisms only contribute to 3.8-6.6 % of the re-mobilization of clay mineral-bound Cd in soil. Therefore, the fundamental strategy to inhibit the re-mobilization of stabilized Cd in vegetable cultivation involves the regulation of proton and organic acid secretion by crops.

Foliar nitrogen uptake in broadleaf evergreen Mediterranean forests: Fertilisation experiment with labelled nitrogen.

Atmospheric nitrogen (N) deposition in Mediterranean sclerophyllous forests of Holm oak (Quercus rotundifolia, Q. ilex) in Spain often exceeds empirical critical loads established for ecosystem conservation. There are still uncertainties on the capacity of canopy retention and uptake of the atmospheric N deposited of these forests. Studying and analysing all the forest nitrogen-cycle processes is essential to understand the potential effect of N deposition in these ecosystems. This study conducted a year-long short-term fertilisation experiment with labelled ammonium (15N-NH4) and nitrate (15N-NO3) to estimate foliar N absorption rates and assess the influence of leaf phenology and meteorological seasonal variations. Fertilising solutions were prepared to simulate low and high wet N deposition concentration, based on data reported from previous studies. Additionally, ecophysiological and meteorological measurements were collected to explore potential relationships between absorption rates, plant activity, and weather conditions. The results showed that Holm oak leaves were able to absorb both oxidised and reduced N compounds, with higher rates of NH4+ absorption. N recovery of both NH4+ and NO3- was higher in the low concentration treatments, suggesting reduced effectiveness of absorption as concentration increases. Foliar absorption rates were leaf-age dependent, with the highest values observed in young developing leaves. Foliar uptake showed seasonal changes with a clear reduction during the summer, linked to drought and dry weather conditions, and showing also smaller leaf net assimilation and stomatal conductance. During the rest of the year, foliar N absorption was not clearly associated to plant physiological activity but with environmental conditions. Our findings suggest that Holm oak canopies could absorb an important part of the incoming N deposition, but this process is compound, season and leaf phenology dependent. Further research is therefore needed to better understand and model this part of the N cycle.

Carbon footprint of maize-wheat cropping system after 40-year fertilization.

Two main challenges which human society faces for sustainable development goals are the maintenance of food security and mitigation of greenhouse gas (GHG) emissions. Here, we examined the impacts of six fertilization treatments including unfertilized control (CK), mineral nitrogen (N, 90 kg N ha-1), mineral N plus 30 kg P ha-1 phosphorus (NP), NP combined with 3.75 Mg ha-1 straw (NP + Str), farmyard manure (Man, 75 Mg ha-1), and NP combined with manure (NP + Man) on crop productivity and carbon emissions (soil GHG emission; GHGI, yield-based GHG intensity; NGHGB, net GHG balance; carbon footprint, CF) in a maize-wheat cropping system during two years (April 2018-June 2020) in a semi-arid continental climate after 40 years of fertilization in the Northwest China. Manure and straw increased total GHG by 38-60 % compared to the mineral fertilizers alone, which was mainly due to the 49-80 % higher direct emissions of carbon dioxide (CO2) rather than nitrous oxide (N2O). Compared to the N fertilizer alone, organic amendments and NP increased cumulative energy yield by 134-202 % but decreased GHGI by 38-55 %, indicating that organic fertilizers increased crop productivity at the cost of higher GHG emissions. When the soil organic carbon changes (ΔSOC) were accounted for in the C emission balance, manure application acted as a net C sink due to the NGHGB recorded with -123 kg CO2-eq ha-1 year-1. When producing the same yield and economic benefits, the manure and straw addition decreased the CF by 59-85 % compared to N fertilization alone. Overall, the transition from mineral to organic fertilization in the semi-arid regions is a two-way independent solution to increase agricultural productivity along with the reduction of C emissions.

Maize functional requirements drive the selection of rhizobacteria under long-term fertilization practices.

Rhizosphere microbiomes are pivotal for crop fitness, but the principles underlying microbial assembly during root-soil interactions across soils with different nutrient statuses remain elusive. We examined the microbiomes in the rhizosphere and bulk soils of maize plants grown under six long-term (≥ 29 yr) fertilization experiments in three soil types across middle temperate to subtropical zones. The assembly of rhizosphere microbial communities was primarily driven by deterministic processes. Plant selection interacted with soil types and fertilization regimes to shape the structure and function of rhizosphere microbiomes. Predictive functional profiling showed that, to adapt to nutrient-deficient conditions, maize recruited more rhizobacteria involved in nutrient availability from bulk soil, although these functions were performed by different species. Metagenomic analyses confirmed that the number of significantly enriched Kyoto Encyclopedia of Genes and Genomes Orthology functional categories in the rhizosphere microbial community was significantly higher without fertilization than with fertilization. Notably, some key genes involved in carbon, nitrogen, and phosphorus cycling and purine metabolism were dominantly enriched in the rhizosphere soil without fertilizer input. In conclusion, our results show that maize selects microbes at the root-soil interface based on microbial functional traits beneficial to its own performance, rather than selecting particular species.

Farmland protection and fertilization intensity: Empirical evidence from preservation policy of Heilongjiang's black soil.

The preservation of cultivated land quality stands as a vital prerequisite for ensuring food security and sustainability. In the black soil area of northeast China, a large amount of fertilizer was used to stabilize grain production in its early stages, which damaged soil structure and polluted the ecological environment. Based on the panel data of fertilization intensity of 48 districts and counties in Heilongjiang Province from 2010 to 2020, this study takes the implementation of the "Three-Year Action Plan for the Protection of Black Soil Farmland in Heilongjiang Province for the (2018-2020)" (TYAP) policy as a natural experiment, and uses the difference-in-differences (DiD) method to identify the causal effect of the policy on the local fertilization intensity. The results of the empirical study showed that the implementation of the TYAP policy significantly reduced the fertilization intensity of the black soil cultivated land implemented by the policy during the implementation period, which resulted in a decrease of 11.97% on average compared with the areas without the policy implementation. Several robustness tests provided additional confirmation of the aforementioned findings. This study further revealed that the policy mitigated fertilization intensity by fostering advancements in agricultural mechanization.

Urea fertilization increased CO2 and CH4 emissions by enhancing C-cycling genes in semi-arid grasslands.

Global climate change is predicted to increase exogenous N input into terrestrial ecosystems, leading to significant changes in soil C-cycling. However, it remains largely unknown how these changes affect soil C-cycling, especially in semi-arid grasslands, which are one of the most vulnerable ecosystems. Here, based on a 3-year field study involving N additions (0, 25, 50, and 100 kg ha-1 yr-1 of urea) in a semi-arid grassland on the Loess Plateau, we investigated the impact of urea fertilization on plant characteristics, soil properties, CO2 and CH4 emissions, and microbial C cycling genes. The compositions of genes involved in C cycling, including C fixation, degradation, methanogenesis, and methane oxidation, were determined using metagenomics analysis. We found that N enrichment increased both above- and belowground biomasses and soil organic C content, but this positive effect was weakened when excessive N was input (N100). N enrichment also altered the C-cycling processes by modifying C-cycle-related genes, specifically stimulating the Calvin cycle C-fixation process, which led to an increase in the relative abundance of cbbS, prkB, and cbbL genes. However, it had no significant effect on the Reductive citrate cycle and 3-hydroxypropionate bi-cycle. N enrichment led to higher soil CO2 and CH4 emissions compared to treatments without added N. This increase showed significant correlations with C degradation genes (bglA, per, and lpo), methanogenesis genes (mch, ftr, and mcr), methane oxidation genes (pmoA, pmoB, and pmoC), and the abundance of microbial taxa harboring these genes. Microbial C-cycling genes were primarily influenced by N-induced changes in soil properties. Specifically, reduced soil pH largely explained the alterations in methane metabolism, while elevated available N levels were mainly responsible for the shift in C fixation and C degradation genes. Our results suggest that soil N enrichment enhances microbial C-cycling processes and soil CO2 and CH4 emissions in semi-arid ecosystems, which contributes to more accurate predictions of ecosystem C-cycling under future climate change.

Determining optimal range of reduction rates for nitrogen fertilization based on responses of vegetable yield and nitrogen losses to reduced nitrogen fertilizer application.

Vegetable production is commonly accompanied by high nitrogen fertilizer rates but low nitrogen use efficiency in China. Reduced fertilization has been frequently recommended in existing studies as an efficient measurement to avoid large amount of nutrient loss and subsequent nonpoint source pollution. However, the reported responses of vegetable yield and nitrogen losses to reduced fertilization rates varied in a large range, which has resulted into large uncertainties in the potential benefits of those recommended reduction rates. Thus, we constructed the relationship between responses of nitrogen losses and vegetable yield to reduced nitrogen fertilization rates to determine the optimal range of reduction rates for nitrogen fertilization in a proportional form based on data reported in literatures across China's mainland, and evaluated the roles of greenhouse, managing options, and vegetable species on the responses. The relationships were constructed separately for 4 subregions: Northern arid and semiarid, loess plateau regions (NSL), Temperate monsoon zone (TMZ), Southeast monsoon zone (SMZ), Southwest zone (SWZ). The optimal nitrogen fertilizer reduction range for the TMZ, SMZ and SWZ were 51 % to 67 %, 40 % to 66 % and 54 % to 80 %, respectively and no reduction for NSL. Vegetable yields were not be sacrificed when fertilizations were reduced within the optimal ranges. Greenhouse and managing options showed no significant effect on the responses of both vegetable yield and nitrogen losses by the optimal reduction range but vegetable species played a relatively important role on the responses of vegetable yield. This indicated that the optimal reduction rates can be effective on reducing nitrogen loss in both open-field and greenhouse conditions across China's mainland without extra managing options. Therefore, the optimal reduction rates can still serve as a good starting point for making regional plans of nitrogen reduction that help balancing the chasing of high vegetable yield and low nitrogen loss.