Plastid phylogenomics and fossil evidence provide new insights into the evolutionary complexity of the 'woody clade' in Saxifragales.

BACKGROUND: The "woody clade" in Saxifragales (WCS), encompassing four woody families (Altingiaceae, Cercidiphyllaceae, Daphniphyllaceae, and Hamamelidaceae), is a phylogenetically recalcitrant node in the angiosperm tree of life, as the interfamilial relationships of the WCS remain contentious. Based on a comprehensive sampling of WCS genera, this study aims to recover a robust maternal backbone phylogeny of the WCS by analyzing plastid genome (plastome) sequence data using Bayesian inference (BI), maximum likelihood (ML), and maximum parsimony (MP) methods, and to explore the possible causes of the phylogenetic recalcitrance with respect to deep relationships within the WCS, in combination with molecular and fossil evidence. RESULTS: Although the four WCS families were identically resolved as monophyletic, the MP analysis recovered different tree topologies for the relationships among Altingiaceae, Cercidiphyllaceae, and Daphniphyllaceae from the ML and BI phylogenies. The fossil-calibrated plastome phylogeny showed that the WCS underwent a rapid divergence of crown groups in the early Cretaceous (between 104.79 and 100.23 Ma), leading to the origin of the stem lineage ancestors of Altingiaceae, Cercidiphyllaceae, Daphniphyllaceae, and Hamamelidaceae within a very short time span (∼4.56 Ma). Compared with the tree topology recovered in a previous study based on nuclear genome data, cytonuclear discordance regarding the interfamilial relationships of the WCS was detected. CONCLUSIONS: Molecular and fossil evidence imply that the early divergence of the WCS might have experienced radiative diversification of crown groups, extensive extinctions at the genus and species levels around the Cretaceous/Paleocene boundary, and ancient hybridization. Such evolutionarily complex events may introduce biases in topological estimations within the WCS due to incomplete lineage sorting, cytonuclear discordance, and long-branch attraction, potentially impacting the accurate reconstruction of deep relationships.

Short-term test for the toxicogenomic assessment of ecotoxic modes of action in Myriophyllum spicatum.

In environmental risk assessment of substances, the 14-day growth inhibition test following OECD test guideline 239 is employed to assess toxicity in the macrophyte Myriophyllum spicatum. Currently, this test evaluates physiological parameters and does not allow the identification of the mode of action (MoA) by which adverse effects are induced. However, for an improved ecotoxicity assessment of substances, knowledge about their ecotoxic MoA in non-target organisms is required. It has previously been suggested that the identification of gene expression changes can contribute to MoA identification. Therefore, we developed a shortened three-day assay for M. spicatum including the transcriptomic assessment of global gene expression changes and applied this assay to two model substances, the herbicide and photosynthesis inhibitor bentazone and the pharmaceutical and HMG-CoA reductase inhibitor atorvastatin. Due to the lack of a reference genome for M. spicatum we performed a de novo transcriptome assembly followed by a functional annotation to use the toxicogenomic results for MoA discrimination. The gene expression changes induced by low effect concentrations of these substances were used to identify differentially expressed genes (DEGs) and impaired biological functions for the respective MoA. We observed both concentration-dependent numbers and differentiated patterns of DEGs for both substances. While bentazone impaired genes involved in the response to reactive oxygen species as well as light response, and also genes involved in developmental processes, atorvastatin exposure led to a differential regulation of genes related to brassinosteroid response as well as potential metabolic shifts between the mevalonate and methyl erythritol 4-phosphate pathway. Based on these responses, we identified biomarker candidates for the assessment of MoA in M. spicatum. Utilizing the shortened assay developed in this study, the investigation of the identified biomarker candidates may contribute to the development of future MoA-specific screening approaches in the ecotoxicological hazard prediction using aquatic non-standard model organisms.

The Complete Mitochondrial Genome of Paeonia lactiflora Pall. (Saxifragales: Paeoniaceae): Evidence of Gene Transfer from Chloroplast to Mitochondrial Genome.

Paeonia lactiflora (P. lactiflora), a perennial plant renowned for its medicinal roots, provides a unique case for studying the phylogenetic relationships of species based on organelle genomes, as well as the transference of DNA across organelle genomes. In order to investigate this matter, we sequenced and characterized the mitochondrial genome (mitogenome) of P. lactiflora. Similar to the chloroplast genome (cpgenome), the mitogenome of P. lactiflora extends across 181,688 base pairs (bp). Its unique quadripartite structure results from a pair of extensive inverted repeats, each measuring 25,680 bp in length. The annotated mitogenome includes 27 protein-coding genes, 37 tRNAs, 8 rRNAs, and two pseudogenes (rpl5, rpl16). Phylogenetic analysis was performed to identify phylogenetic trees consistent with Paeonia species phylogeny in the APG Ⅳ system. Moreover, a total of 12 MTPT events were identified and 32 RNA editing sites were detected during mitogenome analysis of P. lactiflora. Our research successfully compiled and annotated the mitogenome of P. lactiflora. The study provides valuable insights regarding the taxonomic classification and molecular evolution within the Paeoniaceae family.

The uptake and degradation of polychlorinated biphenyls in constructed wetlands planted with Myriophyllum aquaticum.

The unregulated dismantling and improper disposal of electronic waste lead to severe soil contamination by polychlorinated biphenyls (PCBs). Constructed wetlands (CWs) play an important role in PCBs removal as a result of the co-existence of anaerobic and aerobic conditions. However, the effects and mechanisms of different PCBs concentrations in soils on plant uptake and PCBs degradation within CWs are unclear. We evaluated the uptake and degradation of PCBs at different concentrations by Myriophyllum aquaticum (Vell.) Verdc. Planting significantly increased PCBs removal by 8.70% (p < 0.05) in soils with 1500 and 2500 µg/kg PCBs, whereas no significant effect was observed at 500 and 1000 µg/kg. PCBs levels did not significantly affect plant growth and PCBs accumulation. The contribution of plant uptake to PCBs removal was only 0.10-0.12%, indicating that microbial degradation was the dominant pathway for PCBs removal after planting with M. aquaticum. In the treatments with PCBs ≥ 1500 µg/kg, M. aquaticum increased the microbial population, altered the microbial community structure and enriched PCB-degrading bacteria. Functional prediction revealed that microbes in M. aquaticum rhizosphere secreted more peroxidase and glycosyltransferase than non-plant control, which were likely involved in PCBs metabolism.

Polyphenol-enriched Penthorum chinense Pursh ameliorates alcohol-related liver injury through Ras/Raf/MEK/ERK pathway: Integrating network pharmacology and experiment validation.

ETHNOPHARMACOLOGICAL RELEVANCE: Penthorum chinense Pursh (PCP) has acknowledged as an edible herbal medicinal plant for the prevention and treatment of alcoholic liver injury (ALI). However, only few of researches focus on the chemical material basis and potential mechanisms of PCP against ALI. AIM OF THE STUDY: Herein, we explored the therapeutic effects of PCP extract against ALI based on the integration of network pharmacology, molecular docking, and experiment validation. METHODS: Based on the standard quality control of PCP herbs by UPLC fingerprint and quantitative determination, 80% ethanol extract fraction of PCP containing more polyphenols, compared to aqueous extract fraction of PCP, were chosen for further experiments. After oral administration of PCP ethanol extract, serum pharmacochemistry based on UPLC-Q-Exactive-MS analysis was implemented to evaluate the potential effective compounds. These absorbed prototypes in PCP were used to construct network pharmacology and predict the potential mechanisms of PCP extract against ALI. Then, the predicted targets and biological mechanisms of PCP extract were validated using animal experiments and molecular docking analysis. RESULTS: Although totally 19 polyphenol compounds were identified in PCP ethanol extract by UPLC-MS analysis, only 18 absorbed prototypes were found in the serum collected from mice at 1 h post-administration with PCP extract. These candidate active compounds were further screened into 13 compounds to construct network pharmacology and 433 targets were identified as PCP targets. GO and KEGG pathway enrichment analyses indicated that the effects of PCP extract would involve in Ras signaling pathway. The animal experiments on chronic ALI model mice shown that the oral administration of PCP can alleviate ALI by attenuating hepatic oxidative stress, inflammation and down-regulating the target proteins in Ras/Raf/MEK/ERK pathway. Molecular docking analysis revealed the good binding ability between the three polyphenols (i.e. quercetin, apigenin, thonningianin B) in PCP with the top contribution in network pharmacology, and these target proteins (Ras, Raf, MEK1/2, and ERK1/2). CONCLUSION: Our results clarified that PCP ethanol extract could effectively alleviate ALI by down-regulating Ras/Raf/MEK/ERK signaling pathway promisingly. Quercetin, apigenin, and thonningianin B may be the active compounds of PCP, attributing to the intervention benefits of PCP against ALI.

Response strategies of stem/leaves endophyte communities to nano-plastics regulate growth performance of submerged macrophytes.

Research on the toxicity effects of nano-plastics on submerged macrophytes has been increasing over the past several years. However, how the endophytic bacteria of submerged macrophytes respond to nano-plastics remains unknown, although they have been widely shown to help terrestrial plants cope with various environmental stressors. Here, a microcosm experiment was performed to unravel the effects of high concentration of nano-plastics (20 mg/L) on three submerged macrophyte (Vallisneria natans, Potamogeton maackianus, Myriophyllum spicatum) and their endophytic bacterial communities. Results indicated that nano-plastics induced antioxidative stress in plants, but significantly reduction in relative growth rate (RGR) only occurred in V. natans (from 0.0034 to -0.0029 day-1), accompanied by change in the stem/leaves endophyte community composition. Further analysis suggested nano-plastics caused a reduction in environmental nutrient availability and the proportion of positive interactions between endophyte communities (43%), resulting in the lowest RGR of V. natans. In contrast, endophytes may help P. maackianus and M. spicatum cope with nano-plastic stress by increasing the proportion of positive correlations among communities (70% and 75%), leaving their RGR unaffected. Collectively, our study elucidates the species-specific response strategies of submerged macrophyte-endophyte to nano-plastics, which helps to reveal the different phytoremediation potential of submerged macrophytes against nano-plastic pollution.

Effects of nitrogen stress on uptake and translocation of organophosphate esters by watermifoil (Myriophyllum aquaticum L.) in an aquatic ecosystem.

Although organophosphate esters (OPEs) and nitrogen (N) are normally present in aquatic environments, the effects of the plant uptake, accumulation, and translocation of OPEs in different levels of N remain ambiguous. To better understand these processes, watermifoil (Myriophyllum aquaticum L.) as tested plant was chosen to investigate the effects of different N levels on the uptake and translocation of OPEs by plants in matched water-sediment-plant samples. After two months, we found the root-water concentration factors, root-sediment concentration factors, and translocation factors (TFs) were significantly changed with the levels of N (p < 0.05), implying that the presence of N could alter uptake, accumulation, and translocation of OPEs in M. aquaticum, particularly the process of root absorption. Low concentrations of N could remarkably promote the uptake of OPEs by M. aquaticum. However, when the concentrations of N in water were higher than 200 mg/L, the plants' growth and OPE accumulation by M. aquaticum were obviously inhibited with the elevated N contents. Moreover, the enrichment and environmental transport of OPEs in M. aquaticum seemed to be closely associated with physicochemical parameters; the octanol-water partition coefficient had significant relationships with measured organic carbon-normalized sediment-water partition coefficients and TFs in the present study. Additionally, the substituents and structures of OPEs could also affect the accumulation and translocation of OPEs in M. aquaticum, including the chlorination degree and alkyl chain length. This study could improve our understanding of uptake and translocation of OPEs in aquatic plants under different levels of N.

Effects of eutrophic water with ammonium chloride, urea, potassium dihydrogen phosphate and sodium-ß-glycerophosphate on Myriophyllum verticillatum and epiphytic bacteria.

With the rapid development of industry and agriculture, excessive nitrogen and phosphorus released into natural surface water have caused eutrophication. Applying submerged plants to manage eutrophic water has attracted widespread attention. However, there are limited studies on the effects of different nitrogen and phosphorus in the water environment on submerged plants and their epiphytic biofilm. Therefore, this paper investigated the effects of eutrophic water with ammonium chloride (IN), urea (ON), potassium dihydrogen phosphate (IP), and sodium-ß-glycerophosphate (OP) on Myriophyllum verticillatum and epiphytic biofilms. The results showed that Myriophyllum verticillatum exhibited a good purification effect on the eutrophic water with inorganic phosphorus, the removal rates of IP were 68.0%, and the plants grew best in this condition. The fresh weight of the IN group and ON group increased by 12.24% and 7.12%, and the shoot length of the IN group and the ON group increased by 17.71% and 8.33%; the fresh weight of the IP group and OP group increased by 19.19% and 10.83%, the shoot length of the IP group and the OP group increased by 21.09% and 18.23%. In addition, the enzyme activities of superoxide dismutase, catalase, nitrate reductase, and acid phosphatase in plant leaves were significantly changed in eutrophic water with different forms of nitrogen and phosphorus. Finally, the analysis of the epiphytic bacteria showed that different forms of nitrogen and phosphorus nutrients could significantly alter the abundance and structure of microorganisms and microbial metabolism also had significant changes. This study provides a new theoretical basis for evaluating the removal of different forms of nitrogen and phosphorus by Myriophyllum verticillatum, and it also provides new insights for the subsequent engineering of epiphytic microorganisms to improve the capability of submerged plants to treat eutrophic water.

Hippuris vulgaris could replace Myriophyllum aquaticum for efficiently removing water phosphorus under low temperature conditions in China.

Phytoremediation is widely used for the restoration of aquatic environments. However, the phytoremediation effects and mechanisms of special submerged species of native aquatic plants, especially under low-temperature conditions, are not yet clear. In this study, two typical submerged plants, Myriophyllum aquaticum (M. aquaticum; an exotic species) and Hippuris vulgaris (H. vulgaris; a native species), in China were investigated for their phosphorus (P) removal efficiencies (REp) and the related mechanisms of phytophysiology and microorganisms in a low-temperature incubator (10 °C during the day and 2 °C at night). At an initial P level of 0.5 mg L-1, the two plants exhibited similar REp, with the highest values (73.5%-92.1%) observed on days 3-6. After 18 days, the residual P concentration in the water was less than the Grade III limit value (0.2 mg L-1; GB 3838-2002). However, M. aquaticum had a faster REp velocity than H. vulgaris at an initial P level of 3.0 mg L-1, which was attributed to the mechanisms of plant and its interactions with microorganisms. Compared to the control group, the superoxide dismutase activity of H. vulgaris was significantly increased and its catalase activity was decreased, whereas for that of M. aquaticum was the opposite. Micro region X-ray fluorescence analysis revealed that there may be synergic absorption effects between P, S, and K, and antagonistic absorption action between P and Mn in H. vulgaris. In addition, Acinetobacter, Novosphingobium and Pseudomonas were enriched at 3.0 mg L-1 P level with these two plants, but Chlorophyta only accumulated with H. vulgaris, respectively. Overall, the native species, H. vulgaris, could replace the exotic M. aquaticum to efficiently remove P from polluted water at low temperatures. These findings provide a theoretical foundation for submerged plants P removal capabilities, and the protection of local ecosystem diversity at low temperatures.

Codon Usage Analyses Reveal the Evolutionary Patterns among Plastid Genes of Saxifragales at a Larger-Sampling Scale.

Saxifragales is a 15-family order of early-divergent Eudicots with a rich morphological diversity and an ancient rapid radiation. Codon usage bias (CUB) analyses have emerged as an essential tool for understanding the evolutionary dynamics in genes. Thus far, the codon utilization patterns had only been reported in four separate genera within Saxifragales. This study provides a comprehensive assessment of the codon manipulation based on 50 plastid genes, covering 11 constituent families at a larger sampling scale. Our results first showed a high preference for AT bases and AT-ending codons. We then used effective number of codons (ENC) to assess a range of codon bias levels in the plastid genes. We also detected high-informative intrafamilial differences of ENC in three families. Subsequently, parity rule 2 (PR2) plot analyses revealed both family-unique and order-shared bias patterns. Most importantly, the ENC plots and neutrality analyses collectively supported the dominant roles of selection in the CUB of Saxifragales plastid genes. Notably, the phylogenetic affinities inferred by both ML and BI methods were consistent with each other, and they all comprised two primary clades and four subclades. These findings significantly enhance our understanding of the evolutionary processes of the Saxifrage order, and could potentially inspire more CUB analyses at higher taxonomic levels.

Transcriptomics Insights into Phosphorus Stress Response of Myriophyllum aquaticum.

Through excellent absorption and transformation, the macrophyte Myriophyllum (M.) aquaticum can considerably remove phosphorus from wastewater. The results of changes in growth rate, chlorophyll content, and roots number and length showed that M. aquaticum could cope better with high phosphorus stress compared with low phosphorus stress. Transcriptome and differentially expressed genes (DEGs) analyses revealed that, when exposed to phosphorus stresses at various concentrations, the roots were more active than the leaves, with more DEGs regulated. M. aquaticum also showed different gene expression and pathway regulatory patterns when exposed to low phosphorus and high phosphorus stresses. M. aquaticum's capacity to cope with phosphorus stress was maybe due to its improved ability to regulate metabolic pathways such as photosynthesis, oxidative stress reduction, phosphorus metabolism, signal transduction, secondary metabolites biosynthesis, and energy metabolism. In general, M. aquaticum has a complex and interconnected regulatory network that deals efficiently with phosphorus stress to varying degrees. This is the first time that the mechanisms of M. aquaticum in sustaining phosphorus stress have been fully examined at the transcriptome level using high-throughput sequencing analysis, which may indicate the direction of follow-up research and have some guiding value for its future applications.

Complete Chloroplast Genomes and the Phylogenetic Analysis of Three Native Species of Paeoniaceae from the Sino-Himalayan Flora Subkingdom.

Paeonia delavayi var. lutea, Paeonia delavayi var. angustiloba, and Paeonia ludlowii are Chinese endemics that belong to the Paeoniaceae family and have vital medicinal and ornamental value. It is often difficult to classify Paeoniaceae plants based on their morphological characteristics, and the limited genomic information has strongly hindered molecular evolution and phylogenetic studies of Paeoniaceae. In this study, we sequenced, assembled, and annotated the chloroplast genomes of P. delavayi var. lutea, P. delavayi var. angustiloba, and P. ludlowii. The chloroplast genomes of these strains were comparatively analyzed, and their phylogenetic relationships and divergence times were inferred. These three chloroplast genomes exhibited a typical quadripartite structure and were 152,687-152,759 bp in length. Each genome contains 126-132 genes, including 81-87 protein-coding genes, 37 transfer RNAs, and 8 ribosomal RNAs. In addition, the genomes had 61-64 SSRs, with mononucleotide repeats being the most abundant. The codon bias patterns of the three species tend to use codons ending in A/U. Six regions of high variability were identified (psbK-psbL, trnG-UCC, petN-psbM, psbC, rps8-rpl14, and ycf1) that can be used as DNA molecular markers for phylogenetic and taxonomic analysis. The Ka/Ks ratio indicates positive selection for the rps18 gene associated with self-replication. The phylogenetic analysis of 99 chloroplast genomes from Saxifragales clarified the phylogenetic relationships of Paeoniaceae and revealed that P. delavayi var. lutea, P. delavayi var. angustiloba, and P. ludlowii are monophyletic groups and sisters to P. delavayi. Divergence time estimation revealed two evolutionary divergences of Paeoniaceae species in the early Oligocene and Miocene. Afterward, they underwent rapid adaptive radiation from the Pliocene to the early Pleistocene when P. delavayi var. lutea, P. delavayi var. angustiloba, and P. ludlowii formed. The results of this study enrich the chloroplast genomic information of Paeoniaceae and reveal new insights into the phylogeny of Paeoniaceae.

Functional Teas from Penthorum chinense Pursh Alleviates Ethanol-Induced Hepatic Oxidative Stress and Autophagy Impairment in Zebrafish via Modulating the AMPK/p62/Nrf2/mTOR Signaling Axis.

Penthorum chinense Pursh (PCP), a medicinal and edible plant, is widely used in many clinical liver diseases. Oxidative stress and autophagy impairment play crucial roles in the pathophysiology of alcoholic liver disease (ALD). Therefore, the aim of this study was to elucidate the mechanism of PCP in attenuating ethanol-induced liver injury. The liver-specific transgenic zebrafish larvae (lfabp: EGFP) at three days post-fertilization (3 dpf) were treated with different concentrations of PCP (100, 50 and 25 µg/mL) for 48 h, after soaked in a 350 mM ethanol for 32 h. Whole-mount oil red O, H&E staining and biochemical kits were used to detect fatty liver function and fat accumulation, western blot (WB) and immunofluorescence were used to determine proteins expression, and RT-qPCR was used to further verify the related gene expression. PCP restored zebrafish liver function. Additionally, PCP (as dose-dependent) blocked the expression of cytochrome P450 2E1 (CYP2E1), the production of intracellular reactive oxygen species (ROS) and alleviated liver fat accumulation and oxidative damage. PCP exerted its hepatoprotective function by downregulating the expression of kelch-like ECH-associated protein 1 (Keap1), up-regulating the expression of nucleus factor-E2-related factor 2 (Nrf2) (transferring to the nucleus), and attenuating systemic oxidative stress. Furthermore, PCP reduced the expression of sequestosome 1 (p62/SQSTM1, p62), Atg13, and Beclin 1, up-regulating autophagy signaling pathway. Taken together, the molecular evidence that PCP protected the ethanol-induced hepatic oxidative stress and autophagy impairment through activating AMPK/p62/Nrf2/mTOR signaling axis.

Combined toxic effects of enrofloxacin and microplastics on submerged plants and epiphytic biofilms in high nitrogen and phosphorus waters.

With the wide application of plastic products, microplastic pollution has become a major environmental issue of global concern. Microplastics in aquatic environments can interact with organic pollutants, causing a combined effect on submerged macrophytes. This study investigated the response mechanisms of the submerged plant Myriophyllum verticillatum and epiphytic biofilm to the antibiotic enrofloxacin, microplastics, and their combined exposure in a high nitrogen and phosphorus environment. The results indicated that Myriophyllum verticillatum was not sensitive to enrofloxacin of 1 mg L-1, while 10 and 50 mg L-1 enrofloxacin inhibited the uptake of nitrogen and phosphorus by the plants, as well as triggered oxidative stress in the plant leaves, causing irreversible damage to the plant cells. In addition, enrofloxacin altered the structure of the leaf epiphytic biofilm community. Interestingly, 1, 5, and 20 mg L-1 microplastics had no significant effect on the plant, while they facilitated the aggregation of microorganisms, increasing the abundance of the leaf epiphyte biofilm. The combination of enrofloxacin and microplastics induced a synergistic effect on Myriophyllum verticillatum. Specifically, the rate of nitrogen and phosphorus uptake by the plant was reduced, the content of photosynthetic pigments decreased, and antioxidant enzyme activity was further increased. In addition, the diversity of the leaf epiphytic biofilm community was similar to the single enrofloxacin exposure. These results demonstrated the differences between single and combined exposures and provided a new theoretical basis to evaluate the harmful effects of enrofloxacin and microplastics on submerged macrophytes.

Supplementary honey bee (Apis mellifera L.) pollination enhances fruit growth rate and fruit yield in Paeonia ostii (family: Paeoniaceae).

Insufficient pollination leads to low and unstable production of oil tree peony. Supplementary managed honeybees (Apis mellifera L.) in agricultural ecosystems is a common practice for addressing the problem. At this study site (N 34°38'30″ and E 112°39'43″, with an altitude of 125.5 m), we set up four pollination areas (low-density bee pollination group (LDBP), high-density bee pollination group (HDBP), blank control group (CK1) and field control group (CK2)) to examine the pollination effectiveness of different densities of honeybee supplementation on oil tree peony (Paeonia ostii). Our work demonstrated that bee-pollination increased fruit size and growth rate. On average, bee-pollinated (LDBP) plants produced 63.16% more number of seeds per plant, showed also 53.47% more weight of seeds per plant than those in CK2. Also, seeds of LDBP contained, on average, 26.15% more oil content than CK2. Kernel percent and seed oil fatty acid content, however, were unaffected (F = 1.759, p = 0.074). Compared with LDBP, weight of seeds per plant and oil content with HDBP decreased by 21.89% and 2.63%, respectively. Following the same trend, compared with LDBP, HDBP slowed fruit growth and reduced fruit size. Our results showed that insufficient pollination limits fruit set in oil tree peony, while supplementary reasonable bee density in the field for pollination is an important strategy to maximize fruit yield.

Glutamine synthetase plays an important role in ammonium tolerance of Myriophyllum aquaticum.

High-strength ammonium (NH4+), the main characteristic of swine wastewater, poses a significant threat to the rural ecological environment. As a novel phytoremediation technology, Myriophyllum aquaticum wetlands have high tolerance and removal rate of NH4+. Glutamine synthetase (GS), a pivotal enzyme in nitrogen (N) metabolism, is hypothesized to play an important role in the tolerance of M. aquaticum to high NH4+. Herein, the responses of M. aquaticum to GS inhibition by 0.1 mM methionine sulfoximine (MSX) under 15 mM NH4+ were investigated. After 5 days, visible NH4+ toxicity symptoms were observed in MSX-treated plants. Compared with the control, the NH4+ accumulation in the leaves increased by 20.99 times, while that of stems and roots increased by 3.27 times and 47.76 %, suggesting that GS inhibition had a greater impact on the leaves. GS inhibition decreased pigments in the leaves by 8.64 %-41.06 %, triggered oxidative stress, and affected ions concentrations in M. aquaticum. The concentrations of glutamine (Gln) and asparagine decreased by 63.46 %-97.43 % and 12.37 %-76.41 %, respectively, while the concentrations of most other amino acids increased after 5 days of MSX treatment, showing that GS inhibition reprogrammed the amino acids synthesis. A decrease in Gln explains the regulations of N-related genes, including increased expression of AMT in roots and decreased expression of GS, GOGAT, GDH, and AS, which would cause further NH4+ accumulation via promoting NH4+ uptake and decreasing NH4+ assimilation in M. aquaticum. This study revealed for the first time that GS inhibition under high NH4+ condition can lead to phytotoxicity in M. aquaticum due to NH4+ accumulation. The physiological and molecular responses of the leaves, stems, and roots confirmed the importance of GS in the high NH4+ tolerance of M. aquaticum. These findings provide new insights into NH4+ tolerance mechanisms in M. aquaticum and a theoretical foundation for the phytoremediation of high NH4+-loaded swine wastewater.

Partial nitrification in free nitrous acid-treated sediment planting Myriophyllum aquaticum constructed wetland strengthens the treatment of black-odor water.

Black-odor water pollution in rural areas, especially swine wastewater, can lead to the deterioration of water quality and thus seriously affect the daily life of people in the area. However, there is a lack of effective treatment measures with simultaneous attention to carbon, nitrogen and sulfur pollution in rural black-odor water bodies. This study evaluated the feasibility of an in-situ pilot-scale constructed wetland (CW) for the synchronous removal of COD, ammonium, and sulfur compounds in the swine wastewater. In this study, the operation strategy of CW sediment pretreated with free nitrous acid (FNA) and Myriophyllum aquaticum plantation was established. Throughout the 114-day operation, the total removal efficiencies of COD and ammonium nitrogen in experimental groups were 81.2 ± 4.2 % and 72.8 ± 1.8 %, respectively, which were significantly higher than CW without any treatment. Removal efficiencies of Sulfur compounds, i.e. sulfide, sulfate, thiosulfate, and sulfite, were 92.3 ± 1.9 % (61.2 % higher than the no-treatment group), 42.1 ± 3.8 %, 97.9 ± 1.7 %, and 42.7 ± 4.5 % respectively. High-throughput sequencing and qPCR revealed that experimental group significantly increased denitrification genes (nirK, nosZ) and sulfur oxidation genes (soxB, fccAB) and enriched the corresponding microbial taxa (Bacillus, Conexibacter and Clostridium sensu stricto). Moreover, metabolic pathways related to nitrogen and sulfur and the degradation of organic matter were up-regulated. These results indicated that partial nitrification in CW planted with M. aquaticum promoted sulfur oxidation denitrification and heterotrophic denitrification. Overall, the in-situ pilot-scale study revealed that the cultivation of M. aquaticum in FNA-treated CW can be a sustainable approach to treat black-odor water bodies.

Performance of various fillers in ecological floating beds planted with Myriophyllum aquaticum treating municipal wastewater.

The performance of different suspended fillers (zeolite, drinking water treatment residual, biochar, woodchip and stereo-elastic packing) and their combinations in treating municipal wastewater in ecological floating beds (Eco-FBs) planted with Myriophyllum aquaticum was assessed. Six sets of enhanced Eco-FBs were developed to assess the individual and synergistic effects of combinations of the various fillers and microorganisms on nutrient elimination. The results demonstrated mean TN, NH4-N, TP and COD purification efficiencies of 99.2 ±â€¯11.2 %, 99.82 ±â€¯16.4 %, 98.3 ±â€¯14.3 %, and 96.1 ±â€¯12.3 %, respectively in the Eco-FBs strengthened with all five fillers. The corresponding purification rates were 0.89 ±â€¯0.14, 0.75 ±â€¯0.12, 0.08 ±â€¯0.016, and 7.05 ±â€¯1.09 g m-2 d-1, which were 2-3 times higher than those of the conventional Eco-FB system. High-throughput sequencing showed that some genera related to nutrient transformation, including Proteobacteria (24.13-51.95 %), followed by Chloroflexi (5.64-25.01 %), Planctomycetes (8.48-14.43 %) and Acidobacteria (2.29-11.65 %), were abundantly enriched in the strengthened Eco-FBs. Enhancement of the Eco-FBs with various fillers significantly increased microbial species richness and diversity as demonstrated by Chao1, Shannon and Simpson's indexes, particularly when all the five fillers were combined. Therefore, introducing suspended fillers into Eco-FBs is an appropriate approach for improving nutrient elimination from municipal wastewater.

Myriophyllum spicatum Leaves: Aerophily for Gas Collection and Transportation in Water.

The unique living environment of aquatic plants makes them produce many fantastic properties different from land ones. For instance, the leaves of Myriophyllum spicatum show excellent hydrophobicity and aerophily characteristics. In this paper, the abundant morphological structure, composition, and aerophily properties of Myriophyllum spicatum leaves are revealed. The contact angle of the leaf surface can reach 122° in air, exhibiting wonderful gas collection ability under water. The results showed that the aerophily of the leaves is attributed to the multistage micro-nanostructure and waxy layer on the surface. The gas transportation toward the tips of leaves is based on the void gradient formed by the nanoscale morphology at different growth stages and the buoyancy as well. These features provide bionic experience for gas collection, bubble transportation, and liquid resistance reduction in water environments.

Physiological response and tolerance of Myriophyllum aquaticum to a wide range of ammonium concentrations.

Myriophyllum aquaticum (M. aquaticum) can be used in constructed wetlands (CWs) to effectively purify swine wastewater with high-ammonia nitrogen (NH3-N and NH4+-N) concentrations. However, the understanding of its tolerance mechanism to ammonia nitrogen is limited. The physiological response and tolerance mechanism of M. aquaticum to a wide range of NH4+ concentrations (0-35 mM) were investigated in the present study. The results indicated that M. aquaticum can tolerate NH4+ concentrations of up to 30 mM for 21 days and grow well with high nutrient (N, P) uptake. A suitable concentration of NH4+ for a better growth of M. aquaticum was 0.5-20 mM. The free NH4+ content was no obviously increase at NH4+ concentration below 15 mM, indicated there was no obviously ammonium accumulation. Exogenous NH4+ inhibited K+ absorption and improved Ca2+ absorption, indicating mineral cation could mediate NH4+ homeostasis under NH4+ stress. Moreover, comparison with those in the control group, the activities of glutamine synthetase (GS), glutamate synthetase (GOGAT) in M. aquaticum increased by 52.7%-115% at 1-20 mM NH4+, and superoxide dismutase (SOD) increased by 29.2-143% at 1-35 mM NH4+. This indicated that the high NH4+ tolerance of M. aquaticum was mainly due to the balance of free NH4+ content in tissues, as well as improved nitrogen metabolism and antioxidant system. This could be attributed to the role of the GS-GOGAT cycle and SOD. In conclusion, M. aquaticum, which tolerates high NH4+ concentration and has a high N uptake ability, can be used as a good candidate specie to help develop more efficient management strategies for treating high-NH4+ wastewater in CW systems.