Nitrogen fertilization modulates rice phyllosphere functional genes and pathogens through fungal communities.

The phyllosphere is a vital yet often neglected habitat hosting diverse microorganisms with various functions. However, studies regarding how the composition and functions of the phyllosphere microbiome respond to agricultural practices, like nitrogen fertilization, are limited. This study investigated the effects of long-term nitrogen fertilization with different levels (CK, N90, N210, N330) on the functional genes and pathogens of the rice phyllosphere microbiome. Results showed that the relative abundance of many microbial functional genes in the rice phyllosphere was significantly affected by nitrogen fertilization, especially those involved in C fixation and denitrification genes. Different nitrogen fertilization levels have greater effects on fungal communities than bacteria communities in the rice phyllosphere, and network analysis and structural equation models further elucidate that fungal communities not only changed bacterial-fungal inter-kingdom interactions in the phyllosphere but also contributed to the variation of biogeochemical cycle potential. Besides, the moderate nitrogen fertilization level (N210) was associated with an enrichment of beneficial microbes in the phyllosphere, while also resulting in the lowest abundance of pathogenic fungi (1.14 %). In contrast, the highest abundance of pathogenic fungi (1.64 %) was observed in the highest nitrogen fertilization level (N330). This enrichment of pathogen due to high nitrogen level was also regulated by the fungal communities, as revealed through SEM analysis. Together, we demonstrated that the phyllosphere fungal communities were more sensitive to the nitrogen fertilization levels and played a crucial role in influencing phyllosphere functional profiles including element cycling potential and pathogen abundance. This study expands our knowledge regarding the role of phyllosphere fungal communities in modulating the element cycling and plant health in sustainable agriculture.

Cadmium inhibits powdery mildew colonization and reconstructs microbial community in leaves of the hyperaccumulator plant Sedum alfredii.

Understanding the influence of the heavy metal cadmium (Cd) on the phyllosphere microbiome of hyperaccumulator plants is crucial for enhancing phytoremediation. The characteristics of the phyllosphere of Sedum alfredii Hance, a hyperaccumulator plant, were investigated using 16S rRNA and internal transcribed spacer amplicon sequencing of powdery mildew-infected leaves treated or untreated with Cd. The results showed that the colonization of powdery mildew caused severe chlorosis and necrosis in S. alfredii leaves, and the relative abundance of Leotiomycetes in infected leaves increased dramatically and significantly decreased phyllosphere microbiome diversity. However, S. alfredii preferentially accumulated higher concentrations of Cd in the leaves of infected plants than in uninfected plants by powdery mildew, which in turn significantly inhibited powdery mildew colonization in leaves; the relative abundance of the fungal class Leotiomycetes in infected leaves decreased, and alpha and beta diversities of the phyllosphere microbiome significantly increased with Cd treatment in the infected plants. In addition, the inter-kingdom networks in the microbiota of the infected leaves treated with Cd presented many nodes and edges, and the highest inter-kingdom modularity compared to the untreated infected leaves, indicating a highly connected microbial community. These results suggest that Cd significantly inhibits powdery mildew colonization by altering the composition of the phyllosphere microbiome in S. alfredii leaves, paving the way for efficient heavy metal phytoremediation and providing a new perspective on defense strategies against heavy metals.

Effect of Current Density on Preparation and Properties of TF/ß-PbO2 in MSA Media.

The lead-based alloy and DSA anodes have drawbacks, such as poor corrosion resistance, easy peeling of coating, low electrocatalytic activity, and environmental pollution in electrode preparation processes. In this study, titanium foam/ß-PbO2 (TF/ß-PbO2) was prepared by electrodeposition in methanesulfonic acid (MSA) media. The current efficiency and the deposition rate were 89.7% and 5.36 v/(µm·min-1) at the best current density of 80 mA·cm-2, respectively. The TF/ß-PbO2 was characterized by electrochemical tests, X-ray diffraction (XRD), and scanning electron microscopy (SEM). The cyclic voltammetry (CV) test shows that the anodic peak potential of the optimum TF/ß-PbO2 was as low as 2.135 V and anodic voltammetry charge was up to the maximum value of 3.564 × 10-2 C. The linear sweep voltammetry (LSV) test indicates that exchange current density of the optimum TF/ß-PbO2 reached the maximum value of 8.284 × 10-6 A·cm-2. CV and LSV tests indicate that the optimum TF/ß-PbO2 had a high electrocatalytic activity. Electrochemical impedance spectroscopy test Tafel polarization curves show that the optimum TF/ß-PbO2 had better corrosion resistance. The XRD test shows that the crystal was mainly ß-PbO2 of the optimum TF/ß-PbO2 surface and the current density affected the preferential growth of the crystal surface of PbO2. SEM tests show that grains of the optimum TF/ß-PbO2 coating prepared were tightly bound and uniform in size.

Multimode fiber surface plasmon resonance sensor based on a down-up taper.

A multimode fiber surface plasmon resonance sensor based on a down-up taper for refractive index measurement is proposed and demonstrated. The device is fabricated by splicing two multimode fibers in a heating and pushing process to form an up taper, followed by heating and pulling the fiber adjacent to the up-taper area to form a down taper, and then using a magnetron sputtering technique to deposit a Cr + Ag layer of 50 nm on the surface of the down-up taper. Such a structure effectively increases the taper ratio and hence improves the measurement sensitivity. The experimental results obtained show that in the refractive index range of 1.345-1.375, the refractive index sensitivity achieved is ∼3264.01 nm/RIU. The device has a compact size and its entire length is ∼2.75 mm. Moreover, the robustness of the device is better than that of the previously reported fiber surface plasmon resonance sensors due to its relatively large waist diameter of 40 µm for the down taper. The device is expected to have potential applications in biological and chemical sensing.

Responses of earthworm Metaphire vulgaris gut microbiota to arsenic and nanoplastics contamination.

The growing contamination of arsenic and plastics has severely effects on the soil fauna health, including shifts of gut microbiota community. A few studies have focused on effects of microplastics and metal(loid) in soil and fauna gut microbiome. However, the environmental effect of nanoplastics and arsenic on the earthworm gut microbiota, especially on arsenic biotransformation in the gut, remain largely unknown. Here, a microcosm study was performed to explore the effects of nanoplastics and arsenic on the microbiota characteristics in earthworm Metaphire vulgaris gut using Illumina high throughput sequencing, and to investigate changes in the gut microbiota-mediated arsenic biotransformation genes (ABGs) by using high-throughput quantitative PCR. Our results demonstrated that the concentration of arsenic in the earthworm body tissues after exposure to arsenic and nanoplastics was significantly lower from that with arsenic alone exposure. Moreover, the clearly different bacterial community was observed in the soil compared with the earthworm gut, which was dominated by Proteobacteria, Actinobacteria, and Firmicutes at phylum level. Arsenic exposure significantly disturbed bacterial community structure in the earthworm gut, but exposure to nanoplastics did not induce gut microbiota changes. More interestingly, nanoplastics can relieve adverse effect of arsenic on the gut microbiota possibly by adsorbing arsenic. In addition, a total of 16 ABGs were detected, and predominant genes involved in arsenic reduction and transport process were observed in the earthworm guts. In short, this study provides a new picture of the effects of nanoplastics and arsenic on the gut microbiota and arsenic biotransformation in soil fauna gut.

Variations of earthworm gut bacterial community composition and metabolic functions in coastal upland soil along a 700-year reclamation chronosequence.

Most ecosystem functions attributed to earthworms are mediated by their internal microbiomes, and these are sensitive to disturbances in the external environment. However, few studies have focused on the response of the earthworm gut microbiome to soil chronosequence. Here, we used 16S rRNA high-throughput sequencing and high-throughput quantitative PCR to investigate the variations in bacterial communities and functional gene abundance in earthworm (Lumbricina sp.) guts and upland soils under 700 years of cultivation. Our results indicated that 700 years of upland cultivation significantly shaped bacterial communities and increased functional traits of microbes in earthworm guts, which were more sensitive to cultivation age compared to the surrounding soils. The earthworm gut bacterial community changed rapidly over the first 300 years of cultivation and then changed slowly in the following centuries. Along with the cultivation age, we also observed that the earthworm gut microbiota was successive towards a copiotrophic strategy (e.g., Xanthobacteraceae, Nocardioidaceae, Hyphomicrobiaceae, and Bacillaceae) and higher potential functions (e.g., ureC, nirS, nosZ, phoD, and pqqC). Furthermore, canonical correspondence analysis further revealed that soil pH, C:N ratio, soil organic carbon, and total nitrogen were key abiotic drivers shaping earthworm gut bacterial communities. Taken together, this study reveals the succession of bacterial communities and potential functions in earthworm guts within 700 years of upland cultivation, which may provide a broader space for us to rationally exploit and utilize the interactions between soil and earthworm gut microbiotas to benefit the soil nutrient cycling process.

How can fertilization regimes and durations shape earthworm gut microbiota in a long-term field experiment?

The positive roles of earthworms on soil functionality has been extensively documented. The capacity of the earthworm gut microbiota on decomposition and nutrient cycling under long-term fertilization in field conditions has rarely been studied. Here, we report the structural, taxonomic, and functional responses of Eisenia foetida and Pheretima guillelmi gut microbiota to different fertilization regimes and durations using 16S rRNA gene-based Illumina sequencing and high-throughput quantitative PCR techniques. Our results revealed that the core gut microbiota, especially the fermentative bacteria were mainly sourced from the soil, but strongly stimulated with species-specificity, potential benefits for the host and soil health. The functional compositions of gut microbiota were altered by fertilization with fertilization duration being more influential than fertilization regimes. Moreover, the combination of organic and inorganic fertilization with the longer duration resulted in a higher richness and connectivity in the gut microbiota, and also their functional potential related to carbon (C), nitrogen, and phosphorus cycling, particularly the labile C decomposition, denitrification, and phosphate mobilization. We also found that long-term inorganic fertilization increased the abundance of pathogenic bacteria in the P. guillelmi gut. This study demonstrates that understanding earthworm gut microbiota can provide insights into how agricultural practices can potentially alter soil ecosystem functions through the interactions between soil and earthworm gut microbiotas.

Inhibition of C3a/C3aR Axis in Diverse Stages of Ulcerative Colitis Affected the Prognosis of UC by Modulating the Pyroptosis and Expression of Caspase-11.

Ulcerative colitis (UC) is a serious digestive system disease. Furthermore, the activation of C3a/C3aR axis promoted the expression of caspase-11. And higher levels of caspase-11 could induce the pyroptosis and inflammation of cells. However, some studies suggested that caspase-11 could promote and suppress the inflammation during the development of UC. In addition, whether C3a/C3aR axis could affect the development of UC by modulating the expression of caspase-11 is unclear. We established the UC rat model in this study. Next, the C3aR inhibitor was used to treat these rats at diverse stages of UC. Next, the HE staining was performed to detect the intestinal damage. ELISA was performed to reveal the expression of IL-6 and TNF-α in different stages of UC. Western blotting was used to detect the expression of caspase-11 and C3aR in different stages of UC. Stimulation of C3aR inhibitor in early stage of UC promoted the expression of C3aR and caspase-11 in later stage of UC. Treatment of C3aR inhibitor in later stage of UC inhibited the expression of C3aR and caspase-11 in later stage of UC. Furthermore, application of C3aR inhibitor in early stage of UC aggravates the damage of colon tissue and enhanced the secretion of TNF-α and IL-6 in the later stage of UC. Treatment of C3aR inhibitor in later stage of UC relieved the symptoms of UC and suppressed the production of TNF-α and IL-6 in the later stage of UC. Application of C3aR inhibitor in early stage of UC induced the poor prognosis of UC by upregulating the expression of caspase-11. Treatment of C3aR inhibitor in later stage of UC relieved the symptoms of UC and lead to the favorable prognosis of UC by inhibiting the expression of caspase-11.

An effective separation process of arsenic, lead, and zinc from high arsenic-containing copper smelting ashes by alkali leaching followed by sulfide precipitation.

Separation of arsenic and valuable metals (Pb, Zn, Cu, Bi, Sn, In, Ag, Sb, etc.) is a core problem for effective utilization of high arsenic-containing copper smelting ashes (HACSA). This study developed an effective separation process of arsenic, lead, and zinc from HACSA via alkali leaching followed by sulfide precipitation. The separation behaviors and optimum conditions for alkali leaching of arsenic and sulfide precipitation of lead and zinc were established respectively as follows: NaOH concentration 3.81 M; temperature 80°C; time 90 minutes; liquid-to-solid ratio 4:1; agitation speed 450 revolutions/minute (r/min) and 2.0 times of theoretical quantity of sodium sulfide (Na2S); temperature 70°C; and time 60 minutes. The results indicated that the leaching rates of As, Pb, and Zn were 92.4%, 36.9% and 13.4%, respectively. More than 99% of lead and zinc were precipitated from the alkali leachate. The scanning electron microscopy/energy dispersive X-ray spectroscopy study confirmed that arsenic was dissolved from HACSA into the alkali leachate. Furthermore, lead and zinc were precipitated as sulfides from the alkali leachate. The proposed process was a good technique for separation of arsenic and enrichment of valuable metals for further centralized treatment separately. It provided high separation efficiency of arsenic and valuable metals, as well as low environmental pollution.

Partial replacement of inorganic phosphorus (P) by organic manure reshapes phosphate mobilizing bacterial community and promotes P bioavailability in a paddy soil.

The optimization of more sustainable fertilization practice to relieve phosphorus (P) resource scarcity and increase P fertilizer utilization, a better understanding of the regulatory roles of microbes in P mobilization is urgently required to reduce P input. The genes phoD and pqqC are responsible for regulating organic and inorganic P mobilization, respectively. Using high-throughput sequencing, the corresponding bacterial communities harbored by these genes were determined. We conducted a 4-year rice-rice-crop rotation to investigate the responses of phoD- and pqqC-harboring bacterial communities to the partial replacement of inorganic P fertilizer by organic manure with reduced P input. The results showed that a combination of organic and inorganic fertilization maintained high rice yield, and also produced a more complex and stable phosphate mobilizing bacterial community, which contributed to phosphatase activities more than their gene abundances in the model analysis. Compared with the conventional mineral fertilization, organic-inorganic fertilization with the reduced P input slightly increased pqqC gene abundance while significantly enhanced the abundance of phoD-harboring bacteria, especially the genera Bradyrhizobium and Methylobacterium known as potential organic P mineralizers which can maintain high rice production. Moreover, the increased pH was the most impactful factor for the phoD- and pqqC-harboring bacterial communities, by promoting microbial P turnover and greatly increasing bioavailable P pools (H2O-Pi and NaHCO3-Pi, NaOH-Pi) in this P-deficient paddy soil. Hence, our study demonstrated that the partial replacement of mineral P with organic manure could reshape the inorganic phosphate solubilizing and alkaline-phosphomonoesterase encoding bacterial communities towards more resilient and effective to the high P utilization and productivity over intense cultivation, providing insights into the potential of soil microbes in the efficient management of agricultural P fertilization.

MicroRNA-186-5p Inhibits Proliferation And Metastasis Of Esophageal Cancer By Mediating HOXA9.

OBJECTIVE: MicroRNA (miRNA) is an endogenous, non-coding small RNA that plays a key role in regulating organism biology and pathology. The aim of this study was to investigate the expression characteristics of microRNA-186-5p in esophageal cancer (ECa) and its correlation with clinical progression and prognosis, and to further explore its underlying mechanisms. METHODS: Real-time quantitative PCR (qRT-PCR) was used to detect microRNA-186-5p level in 45 pairs of ECa tissue samples and adjacent ones, and to analyze the expression of microRNA-186-5p and clinical progression of ECa and prognosis. The relationship between microRNA-186-5p level in ECa cell lines was further verified by qRT-PCR. Finally, the potential mechanism was explored using luciferase reporter gene assay and cell recovery experiment. RESULTS: QRT-PCR results revealed that the expression of microRNA-186-5p in ECa tissues was remarkably lower than that in adjacent tissues, and the difference was statistically significant. Compared with patients with high expression of microRNA-186-5p, patients with low expression of microRNA-186-5p had higher incidence of pathological stage and lower overall survival rate. Besides, compared with the miR-NC group, the microRNA-186-5p mimics group had a significant decrease in proliferation and metastasis ability of ECa cells. Subsequent qRT-PCR validation in ECa cell lines and tissues indicated a significant increase in HOXA9 expression and a negative correlation with microRNA-186-5p. CONCLUSION: The expression of microRNA-186-5p was remarkably decreased in ECa, which was remarkably correlated with pathological stage, distant metastasis and poor prognosis of ECa. The results suggested that microRNA-186-5p may inhibit cell proliferation of ECa by regulating HOXA9.

Reduced nitrogen supply enhances the cellular antioxidant potential of phenolic extracts through alteration of the phenolic composition in lettuce (Lactuca sativa L.).

BACKGROUND: Nitrogen availability is an important environmental factor that determines the production of phenolic compounds in vegetables, but the relationship between low nitrogen-induced alterations of phenolic compounds in vegetable crops and the cellular antioxidant activities of these compounds remains unclear. This study investigated the effect of reduced nitrogen supply (0.05 mmol L-1 nitrate) on phenolic metabolism in lettuce and the protective role of phenolic extracts against H2 O2 -induced oxidative stress in Caco-2 cells by determining cell damage, reactive oxygen species (ROS) content and antioxidant enzyme activities. RESULTS: Reduced nitrogen supply significantly improved the accumulation of phenolic compounds in lettuce, which was partially correlated with the upregulation of genes related to the phenolic synthesis pathway. Phenolic extracts from lettuce cultivated in low-nitrogen medium exhibited a better protective effect against H2 O2 -induced oxidative damage in Caco-2 cells than those from lettuce cultivated with adequate nitrogen. These extracts act by increasing the activities of antioxidant enzymes and, subsequently, by inhibiting ROS overproduction, which leads to a decrease in mitochondrial membrane and DNA damage. The results of HPLC and correlation analyses implied that the improvement in the protective capacity of lettuce extracts after low-nitrogen treatment may be related, not only to the increased content of phenolic compounds, but also to the increased percentage contribution of chlorogenic acid and quercetin derivatives to the total phenolic content. CONCLUSION: Reduction in nitrogen supply can be a powerful strategy to modify phenolic metabolism and composition in lettuce and, consequently, to improve their antioxidant capacity. © 2019 Society of Chemical Industry.

Deep sequencing analysis of phage libraries using Illumina platform.

This paper presents an analysis of phage-displayed libraries of peptides using Illumina. We describe steps for the preparation of short DNA fragments for deep sequencing and MatLab software for the analysis of the results. Screening of peptide libraries displayed on the surface of bacteriophage (phage display) can be used to discover peptides that bind to any target. The key step in this discovery is the analysis of peptide sequences present in the library. This analysis is usually performed by Sanger sequencing, which is labor intensive and limited to examination of a few hundred phage clones. On the other hand, Illumina deep-sequencing technology can characterize over 10(7) reads in a single run. We applied Illumina sequencing to analyze phage libraries. Using PCR, we isolated the variable regions from M13KE phage vectors from a phage display library. The PCR primers contained (i) sequences flanking the variable region, (ii) barcodes, and (iii) variable 5'-terminal region. We used this approach to examine how diversity of peptides in phage display libraries changes as a result of amplification of libraries in bacteria. Using HiSeq single-end Illumina sequencing of these fragments, we acquired over 2×10(7) reads, 57 base pairs (bp) in length. Each read contained information about the barcode (6bp), one complimentary region (12bp) and a variable region (36bp). We applied this sequencing to a model library of 10(6) unique clones and observed that amplification enriches ∼150 clones, which dominate ∼20% of the library. Deep sequencing, for the first time, characterized the collapse of diversity in phage libraries. The results suggest that screens based on repeated amplification and small-scale sequencing identify a few binding clones and miss thousands of useful clones. The deep sequencing approach described here could identify under-represented clones in phage screens. It could also be instrumental in developing new screening strategies, which can preserve diversity of phage clones and identify ligands previously lost in phage display screens.