Proteomics analysis of PK-15 cells infected with porcine parvovirus and the effect of PCBP1 on PPV replication.

Porcine parvovirus (PPV) is one of the most important pathogens that cause reproductive failure in pigs. However, the pathogenesis of PPV infection remains unclear. Proteomics is a powerful tool to understand the interaction between virus and host cells. In the present study, we analyzed the proteomics of PPV-infected PK-15 cells. A total of 32 and 345 proteins were differentially expressed at the early and replication stages, respectively. Subsequent gene ontology annotation and Kyoto Encyclopedia of Genes and Genomes enrichment analysis showed these differentially expressed proteins were significantly enriched in pathways including toll-like receptor signaling pathway, tumor necrosis factor signaling pathway, and viral carcinogenesis. The expression of poly (rC) binding protein 1 (PCBP1) was observed to decrease after PPV infection. Overexpressed or silenced PCBP1 expression inhibited or promoted PPV infection. Our studies established a foundation for further exploration of the multiplication mechanism of PPV. IMPORTANCE: Porcine parvovirus (PPV) is a cause of reproductive failure in the swine industry. Our knowledge of PPV remains limited, and there is no effective treatment for PPV infection. Proteomics of PPV-infected PK-15 cells was conducted to identify differentially expressed proteins at 6 hours post-infection (hpi) and 36 hpi. Gene ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analysis showed that various pathways participate in PPV infection. Poly (rC) binding protein 1 was confirmed to inhibit PPV replication, which provided potential targets for anti-PPV infection. Our findings improve the understanding of PPV infection and pave the way for future research in this area.

Engineering lentivirus envelope VSV-G for liver targeted delivery of IDOL-shRNA to ameliorate hypercholesterolemia and atherosclerosis.

Lentiviral vectors (LVs) have been widely used as a tool for gene therapies. However, tissue-selective transduction after systemic delivery remains a challenge. Inducible degrader of low-density lipoprotein receptor is an attractive target for treating hypercholesterolemia. Here, a liver-targeted LV, CS8-LV-shIDOL, is developed by incorporating a hepatocyte-targeted peptide derived from circumsporozoite protein (CSP) into the lentivirus envelope for liver-targeted delivery of IDOL-shRNA (short hairpin RNA) to alleviate hypercholesterolemia. Tail-vein injection of CS8-LV-shIDOL results in extremely high accumulation in liver and nearly undetectable levels in other organs in mice. In addition, it shows superior therapeutic efficacy in lowering serum low-density lipoprotein cholesterol (LDL-C) and reducing atherosclerotic lesions over unmodified LV-shIDOL in hyperlipidemic mice. Mechanically, the envelope-engineered CS8-LV-shIDOL can enter liver cells via low-density lipoprotein receptor-related protein (LRP). Thus, this study provides a novel approach for liver-targeted delivery of IDOL-shRNA to treat hypercholesterolemia by using an envelope-engineered LV, and this delivery system has great potential for liver-targeted transgene therapy.

A novel fully human anti-NT-ANGPTL3 antibody from phage display library exhibits potent ApoB, TG, and LDL-C lowering activities in hyperlipidemia mice.

Dyslipidemia is characterized by elevated plasma levels of low-density lipoprotein cholesterol (LDL-C), triglycerides (TG), and TG-rich lipoprotein (TGRLs) in circulation, and is closely associated with the incidence and development of cardiovascular disease. Angiopoietin-like protein 3 (ANGPTL3) deficiency has been identified as a cause of familial combined hypolipidemia in humans, which allows it to be an important therapeutic target for reducing plasma lipids. Here, we report the discovery and characterization of a novel fully human antibody F1519-D95aA against N-terminal ANGPTL3 (NT-ANGPTL3), which potently inhibits NT-ANGPTL3 with a KD as low as 9.21 nM. In hyperlipidemic mice, F1519-D95aA shows higher apolipoprotein B (ApoB) and TG-lowering, and similar LDL-C reducing activity as compared to positive control Evinacumab (56.50% vs 26.01% decrease in serum ApoB levels, 30.84% vs 25.28% decrease in serum TG levels, 23.32% vs 22.52% decrease in serum LDLC levels, relative to vehicle group). Molecular docking and binding energy calculations reveal that the F1519-D95aA-ANGPTL3 complex (10 hydrogen bonds, -65.51 kcal/mol) is more stable than the Evinacumab-ANGPTL3 complex (4 hydrogen bonds, -63.76 kcal/mol). Importantly, F1519-D95aA binds to ANGPTL3 with different residues in ANGPTL3 from Evinacumab, suggesting that F1519-D95aA may be useful for the treatment of patients resistant to Evinacumab. In conclusion, F1519-D95aA is a novel fully human anti-NT-ANGPTL3 antibody with potent plasma ApoB, TG, and LDL-C lowering activities, which can potentially serve as a therapeutic agent for hyperlipidemia and relevant cardiovascular diseases.

Molecular and functional characterization of porcine poly C binding protein 1 (PCBP1).

BACKGROUND: Poly C Binding Protein 1 (PCBP1) belongs to the heterogeneous nuclear ribonucleoprotein family. It is a multifunctional protein that participates in several functional circuits and plays a variety of roles in cellular processes. Although PCBP1 has been identified in several mammals, its function in porcine was unclear. RESULTS: In this study, we cloned the gene of porcine PCBP1 and analyzed its evolutionary relationships among different species. We found porcine PCBP1 protein sequence was similar to that of other animals. The subcellular localization of PCBP1 in porcine kidney cells 15 (PK-15) cells was analyzed by immunofluorescence assay (IFA) and revealed that PCBP1 was mainly localized to the nucleus. Reverse transcription-quantitative PCR (RT-qPCR) was used to compare PCBP1 mRNA levels in different tissues of 30-day-old pigs. Results indicated that PCBP1 was expressed in various tissues and was most abundant in the liver. Finally, the effects of PCBP1 on cell cycle and apoptosis were investigated following its overexpression or knockdown in PK-15 cells. The findings demonstrated that PCBP1 knockdown arrested cell cycle in G0/G1 phase, and enhanced cell apoptosis. CONCLUSIONS: Porcine PCBP1 is a highly conserved protein, plays an important role in determining cell fate, and its functions need further study.

A bifunctional anti-PCSK9 scFv/Exendin-4 fusion protein exhibits enhanced lipid-lowering effects via targeting multiple signaling pathways in HFD-fed mice.

Fusion protein which encompasses more than one functional component, has become one of the most important representatives of macromolecular drugs for disease treatment since that monotherapy itself might not be effective enough to eradicate the disease. In this study, we sought to construct a bifunctional antibody fusion protein by fusing anti-PCSK9 scFv with Exendin-4 for simultaneously lowering both LDL-C and TG. Firstly, three Ex4-anti-PCSK9 scFv fusion proteins were constructed by genetically connecting the C-terminal of Exendin-4 to the N-terminal of anti-PCSK9 scFv through various flexible linker peptides (G4S)n (n = 2, 3, 4). After soluble expression in E. coli, the most potent Ex4-(G4S)4-anti-PCSK9 scFv fusion protein was selected based on in vitro activity assays. Then, we investigated the in vivo therapeutic effects of Ex4-(G4S)4-anti-PCSK9 scFv on the serum lipid profile and bodyweight changes as well as underlying molecular mechanism in HFD-fed C57BL/6 mice. The data showed that Ex4-(G4S)4-anti-PCSK9 scFv exhibits enhanced effects of lowering both LDL-C and TG in serum, reducing food intake and body weight via blocking PCSK9/LDLR, activating AMPK/SREBP-1 pathways, and up-regulating sirt6. Conclusively, Ex4-(G4S)4-anti-PCSK9 has the potential to serve as a promising therapeutic agent for effectively treating dyslipidemia with high levels of both LDL-C and TG.

TLR2-mediated NF-κB signaling pathway is involved in PPV1-induced apoptosis in PK-15 cells.

Porcine parvovirus 1 (PPV1) mainly induces severe reproductive failure in pregnant swine, and causes huge economic losses to the swine industry. Cell apoptosis induced by PPV1 infection has been identified the major cause of reproductive failure. However, the molecular mechanism was not fully elucidated. In this study, the potential mechanism of PPV1 induced apoptosis in PK-15 cells was investigated. Our results showed that PPV1 induced apoptosis in PK-15 cells. Further studies revealed toll-like receptor 2 (TLR2) was involved in the PPV1-mediated apoptosis. TLR2 siRNA significantly decreased the apoptosis. Finally, our study showed NF-κB was activated by TLR2 during PPV1-induced apoptosis. The activation of NF-κB signaling was demonstrated by the phosphorylation of p65, p65 nuclear translocation and degradation of inhibitor of kappa B α (IκBα). Together, these results provided evidence that the recognition between PPV1 and PK-15 cells was mainly through TLR2, and then induction of the NF-κB signaling pathway activation, which further induces apoptosis. Our study could provide information to understand the molecular mechanisms of PPV1 infection.

Purification and water resource circulation utilization of Cd-containing wastewater during microbial remediation of Cd-polluted soil.

The purification and water resource circulation utilization of cadmium-containing leachate is a key link in the field application of microbial remediation in Cd-polluted soil. In this study, through a simulation experiment of microbial remediation of Cd-polluted paddy soil, the feasibility of the purification and recycling process of wastewater derived from microbial remediation of Cd-polluted soil was explored. The results of the microbial mobilization and removal experiment showed that the concentrations of Cd, N, P, and K in the leachate were 88.51 µg/L, 38.06, 0.53, and 98.87 mg/L, respectively. The leachate also contained a large number of microbial resources, indicating that it had high recovery values. To recycle this wastewater, activated carbon (C), humic acid (H), and self-assembled monolayers on mesoporous supports (SAMMS; S) were used as adsorbents. The results showed that the co-existing cations in the leachate had a major influence on the adsorption of Cd. In the ternary system of Fe, Al, and Cd, the removal efficiency of Cd increased to 91.2% when the S dosage was increased to 5‰, and the sorption of Cd occurred after Fe and Al. However, C and H exhibited poor adsorption performances. The isotherm models further showed that the maximum adsorption capacities of S, H, and C were 13.96, 6.41 and 2.94 mg/g, respectively. The adsorption kinetics of S showed that adsorption was a rapid process, and the C-H and O-Si-O of S were the key functional groups. The pH of the leachate significantly affected the adsorption efficiency of Cd. Finally, the purified leachate was successfully applied to microbial cultivation and soil remediation. Overall, the reclamation of Cd-containing wastewater can not only dampen the impacts of water shortages, but also achieve the purposes of Cd removal and resource recovery to lower costs by approximately 1166-3499 yuan per mu.

Computational Optimizing the Electromagnetic Wave Reflectivity of Double-Layered Polymer Nanocomposites.

Double-layered absorption-dominated electromagnetic interference (EMI) shielding composites are highly desirable to prevent secondary electromagnetic wave pollution. However, it is a tremendous challenge to optimize the shielding performance via the trial-and-error method due to the low efficiency. Herein, a novel approach of computation-aided experimental design is proposed to efficiently optimize the reflectivity of the double-layered composites. A normalized input impedance (NII) method is presented to calculate the electromagnetic wave reflectivity of multilayered EMI shielding composites. The calculated results are a good match with the experimental results. Then, the NII method is utilized to design polyvinylidene difluoride/MXene/carbon nanotube (PVDF/MXene/CNT) composites. According to the optimization of the NII method, the prepared PVDF/MXene/CNT composite has an ultralow reflectivity of 0.000057, which outperforms that reported in current work and satisfies the requirement of electromagnetic wave absorbing material. Additionally, its average EMI shielding effectiveness is 30 dB, demonstrating that PVDF/MXene/CNT composite simultaneously achieves shielding and absorption. The ultralow reflection mechanism can be ascribed to the ideal impedance match. Both the PVDF/MXene and the PVDF/CNT layers can attenuate electromagnetic energy, which subverts the traditional cognition of double-layered absorption-dominated EMI shielding composites. The NII method opens a way for the practical fabrication of double-layered absorption-dominated EMI shielding composites.

Co-infection of porcine epidemic diarrhoea virus and porcine deltacoronavirus enhances the disease severity in piglets.

Porcine epidemic diarrhoea virus (PEDV) and porcine deltacoronavirus (PDCoV) are the main enteric coronaviruses that cause acute diarrhoea and dehydration in pigs. The co-infection of PDCoV and PEDV is common in natural swine infections, but the clinical outcomes of the interaction between the co-circulating PDCoV and PEDV are unknown. In current study, we established a co-infection model by inoculating the cell culture-adapted PDCoV HNZK-02 strain and PEDV CV777 simultaneously or sequentially using 4-day-old piglets. The weight loss, clinical scores, viral load and titre, histopathological changes and serum cytokines expression were compared with piglets challenged by either virus. Our results indicated the piglets co-inoculated with PDCoV and PEDV showed more serious diarrhoeal symptoms, mainly characterized by longer diarrhoeal period when compared to those of the mono-infection piglets. Furthermore, we observed that PEDV could promote PDCoV replication in the co-inoculated piglets with evidence of prolonged faecal viral shedding, high viral titres in faeces and intestine tissues. Histological analysis indicated the co-infected piglets showed more extensive and serious pathological lesions in small intestine tissues than the mono-infection piglets. Our data also suggested that the co-infection of PDCoV and PEDV caused the excessive expression of pro-inflammatory cytokines (IL-6, IL-8 and TNF-α) in serum. These results proved there existed obvious synergistic pathogenic effects between PDCoV and PEDV co-infection, which provided new insights into the synergistic pathogenic mechanism caused by these two porcine coronaviruses.

Generation of a Novel High-Affinity Antibody Binding to PCSK9 Catalytic Domain with Slow Dissociation Rate by CDR-Grafting, Alanine Scanning and Saturated Site-Directed Mutagenesis for Favorably Treating Hypercholesterolemia.

Inhibition of proprotein convertase subtilisin/kexin type 9 (PCSK9) has become an attractive therapeutic strategy for lowering low-density lipoprotein cholesterol (LDL-C). In this study, a novel high affinity humanized IgG1 mAb (named h5E12-L230G) targeting the catalytic domain of human PCSK9 (hPCSK9) was generated by using CDR-grafting, alanine-scanning mutagenesis, and saturated site-directed mutagenesis. The heavy-chain constant region of h5E12-L230G was modified to eliminate the cytotoxic effector functions and mitigate the heterogeneity. The biolayer interferometry (BLI) binding assay and molecular docking study revealed that h5E12-L230G binds to the catalytic domain of hPCSK9 with nanomolar affinity (KD = 1.72 nM) and an extremely slow dissociation rate (koff, 4.84 × 10-5 s-1), which interprets its quite low binding energy (-54.97 kcal/mol) with hPCSK9. Additionally, h5E12-L230G elevated the levels of LDLR and enhanced the LDL-C uptake in HepG2 cells, as well as reducing the serum LDL-C and total cholesterol (TC) levels in hyperlipidemic mouse model with high potency comparable to the positive control alirocumab. Our data indicate that h5E12-L230G is a high-affinity anti-PCSK9 antibody candidate with an extremely slow dissociation rate for favorably treating hypercholesterolemia and relevant cardiovascular diseases.

Development of a novel, fully human, anti-PCSK9 antibody with potent hypolipidemic activity by utilizing phage display-based strategy.

BACKGROUND: Proprotein convertase subtilisin/kexin type 9 (PCSK9) regulates serum LDL cholesterol (LDL-C) levels by facilitating the degradation of the LDL receptor (LDLR) and is an attractive therapeutic target for hypercholesterolemia intervention. Herein, we generated a novel fully human antibody with favourable druggability by utilizing phage display-based strategy. METHODS: A potent single-chain variable fragment (scFv) named AP2M21 was obtained by screening a fully human scFv phage display library with hPCSK9, and performing two in vitro affinity maturation processes including CDR-targeted tailored mutagenesis and cross-cloning. Thereafter, it was transformed to a full-length Fc-silenced anti-PCSK9 antibody FAP2M21 by fusing to a modified human IgG1 Fc fragment with L234A/L235A/N297G mutations and C-terminal lysine deletion, thus eliminating its immune effector functions and mitigating mAb heterogeneity. FINDINGS: Our data showed that the generated full-length anti-PCSK9 antibody FAP2M21 binds to hPCSK9 with a KD as low as 1.42 nM, and a dramatically slow dissociation rate (koff, 4.68 × 10-6 s-1), which could be attributed to its lower binding energy (-47.51 kcal/mol) than its parent counterpart FAP2 (-30.39 kcal/mol). We verified that FAP2M21 potently inhibited PCSK9-induced reduction of LDL-C uptake in HepG2 cells, with an EC50 of 43.56 nM. Further, in hPCSK9 overexpressed C57BL/6 mice, a single tail i.v. injection of FAP2M21 at 1, 3 and 10 mg/kg, dose-dependently up-regulated hepatic LDLR levels, and concomitantly reduced serum LDL-C by 3.3% (P = 0.658, unpaired Student's t-test), 30.2% (P = 0.002, Mann-Whitney U-test) and 37.2% (P = 0.002, Mann-Whitney U-test), respectively. INTERPRETATION: FAP2M21 with potent inhibitory effect on PCSK9 may serve as a promising therapeutic agent for treating hypercholesterolemia and associated cardiovascular diseases.

Exploring and exploiting plant cyclic peptides for drug discovery and development.

Ever since the discovery of insulin, natural peptides have become an important resource for therapeutic development. Decades of research has led to the discovery of a long list of peptide drugs with broad applications in clinics, from antibiotics to hypertension treatment to pain management. Many of these US FDA-approved peptide drugs are derived from microorganisms and animals. By contrast, the great potential of plant cyclic peptides as therapeutics remains largely unexplored. These macrocyclic peptides typically have rigid structures, good bioavailability and membrane permeability, making them appealing candidates for drug development and engineering. In this review, we introduce the three major classes of plant cyclic peptides and summarize their potential medical applications. We discuss how we can leverage the genome information of many different plants to quickly search for new cyclic peptides and how we can take advantage of the insights gained from their biosynthetic pathways to transform the process of production and drug development. These recent developments have provided a new angle for exploring and exploiting plant cyclic peptides, and we believe that many more peptide drugs derived from plants are about to come.

Spatial Distribution of Toxic Metal(loid)s and Microbial Community Analysis in Soil Vertical Profile at an Abandoned Nonferrous Metal Smelting Site.

In this study soils at different depths were collected in a Zn smelting site located in Zhuzhou City, China, in order to understand toxic metal(loid)s distribution and microbial community in vertical soil profile at a smelting site. Except Soil properties and metal(loid)s content, the richness and diversity of microbial communities in soil samples were analyzed via high-throughput Illumina sequencing of 16s rRNA gene amplicons. The results showed that the content of As, Pb, Cu, Cd, Zn, and Mn was relatively high in top soil in comparison to subsoil, while the concentration of Cr in subsoil was comparable with that in top soil due to its relative high background value in this soil layer. The bioavailability of Cd, Mn, Zn, and Pb was relative higher than that of As, Cr, and Cu. The diversity of soil microbial communities decreased with increasing depth, which might be ascribed to the decrease in evenness with increase in depth duo to the influence by environmental conditions, such as pH, TK (total potassium), CEC (cation exchange capacity), ORP (oxidation reduction potential), and Bio-Cu (bioavailable copper). The results also found Acidobacteria, Proteobacteria, Firmicutes, and Chloroflexi were dominant phyla in soil samples. At the genus level, Acinetobacter, Pseudomonas, and Gp7 were dominant soil microorganism. Besides, Environmental factors, such as SOM (soil organic matter), pH, Bio-Cu, Bio-Cd (bioavailable cadmium), and Bio-Pb (bioavailable lead), greatly impacted microbial community in surface soil (1-3 m), while ORP, TK, and AN concentration influenced microbial community in the subsoil (4-10 m).

Assessments of different inactivating reagents in formulating transmissible gastroenteritis virus vaccine.

BACKGROUND: Transmissible gastroenteritis virus (TGEV) causes enteric infection in piglets, characterized by vomiting, severe diarrhea and dehydration, and the mortality in suckling piglets is often high up to 100%. Vaccination is an effective measure to control the disease caused by TGEV. METHODS: In this study, cell-cultured TGEV HN-2012 strain was inactivated by formaldehyde (FA), ß-propiolactone (BPL) or binaryethylenimine (BEI), respectively. Then the inactivated TGEV vaccine was prepared with freund's adjuvant, and the immunization effects were evaluated in mice. The TGEV-specific IgG level was detected by ELISA. The positive rates of CD4+, CD8+, CD4+IFN-γ+, CD4+IL-4+ T lymphocytes were detected by flow cytometry assay. Lymphocyte proliferation assay and gross pathology and histopathology examination were also performed to assess the three different inactivating reagents in formulating TGEV vaccine. RESULTS: The results showed that the TGEV-specific IgG level in FA group (n = 17) was earlier and stronger, while the BEI group produced much longer-term IgG level. The lymphocyte proliferation test demonstrated that the BEI group had a stronger ability to induce spleen lymphocyte proliferation. The positive rates of CD4+ and CD8+ T lymphocyte subsets of peripheral blood lymphocyte in BEI group was higher than that in FA group and BPL groups by flow cytometry assay. The positive rate of CD4+IFN-γ+ T lymphocyte subset was the highest in the BPL group, and the positive rate of CD4+IL-4+ T lymphocyte subset was the highest in the FA group. There were no obvious pathological changes in the vaccinated mice and the control group after the macroscopic and histopathological examination. CONCLUSIONS: These results indicated that all the three experimental groups could induce cellular and humoral immunity, and the FA group had the best humoral immunity effect, while the BEI group showed its excellent cellular immunity effect.

A natural product solution to aging and aging-associated diseases.

Aging is a natural biological progress accompanied by the gradual decline in physiological functions, manifested by its close association with an increased incidence of human diseases and higher vulnerability to death. Those diseases include neurological disorders, cardiovascular diseases, diabetes, and cancer, many of which are currently without effective cures. Even though aging is inevitable, there are still interventions that can be developed to prevent/delay the onset and progression of those aging-associated diseases and extend healthspan and/or lifespan. Here, we review decades of research that reveals the molecular pathways underlying aging and forms the biochemical basis for anti-aging drug development. Importantly, due to the vast chemical space of natural products and the rich history of herb medicines in treating human diseases documented in different cultures, natural products have played essential roles in aging research. Using several of the most promising natural products and their derivatives as examples, we discuss how natural products serve as an inspiration resource that helped the identification of key components/pathways underlying aging, their mechanisms of action inside the cell, and the functional scaffolds or targeting mechanisms that can be learned from natural products for drug engineering and optimization. We argue that natural products might eventually provide a solution to aging and aging-associated diseases.

Reduction mechanism of Cd accumulation in rice grain by Chinese milk vetch residue: Insight into microbial community.

Chinese milk vetch is an efficient approach to reduce Cd accumulation in rice, nevertheless, its reduction mechanism is not well understood. In this study, we investigated the rice grain Cd, soil properties and microbial community in a Cd-polluted paddy field amended with milk vetch residue (MV) or without (CK) during rice growth period. We found that milk vetch residue averagely decreased the Cd content in rice grain by 45%. Decrease of Cd in rice mainly attributed to the inhibition of Cd activation by milk vetch residue at heading stage probably by the formation of HA-Cd (Humic Acid) and CdS. Increased pH and organic matter (OM) promoted the reduction of available Cd. In addition, nonmetric multidimensional scaling (NMDS) analysis revealed that microbial community structure was significantly different between MV and CK treatment (r = 0.187, p = 0.002), and the core functions of differentially abundant genera were mainly associated with N-cycling, organic matter degradation and sulfate-reducing. The application of milk vetch residue increased the abundance of sulfate-reducing bacteria (SRB) by 8-112% during the rice growth period, which may involve in promoting the transformation of Cd to a more stably residual Cd (CdS). Canonical correspondence analysis (CCA) and mantel test analysis indicated that available K (p = 0.004) and available N (p = 0.005) were the key environmental factors of shaping the SRB. Altogether, changes in soil properties affected microbial structure and functional characteristics, especially the response of SRB in MV treatment would provide valuable insights into reducing the bioavailability of Cd in soil.

Precision Delivery of Multiscale Payloads to Tissue-Specific Targets in Plants.

The precise deployment of functional payloads to plant tissues is a new approach to help advance the fundamental understanding of plant biology and accelerate plant engineering. Here, the design of a silk-based biomaterial is reported to fabricate a microneedle-like device, dubbed "phytoinjector," capable of delivering a variety of payloads ranging from small molecules to large proteins into specific loci of various plant tissues. It is shown that phytoinjector can be used to deliver payloads into plant vasculature to study material transport in xylem and phloem and to perform complex biochemical reactions in situ. In another application, it is demonstrated Agrobacterium-mediated gene transfer to shoot apical meristem (SAM) and leaves at various stages of growth. Tuning of the material composition enables the fabrication of another device, dubbed "phytosampler," which is used to precisely sample plant sap. The design of plant-specific biomaterials to fabricate devices for drug delivery in planta opens new avenues to enhance plant resistance to biotic and abiotic stresses, provides new tools for diagnostics, and enables new opportunities in plant engineering.

Assessing Chromium Contamination in Red Soil: Monitoring the Migration of Fractions and the Change of Related Microorganisms.

The improper stacking of chromium (Cr) slag poses a great threat to the environment and human health. The toxicity of Cr in soil is not only related to its total amount, but also to its fractions. A simulated experiment was conducted in laboratory to assess the environmental risk of Cr fractions migration and distribution in red soil. The results showed the content of acid-soluble and reducible Cr significantly decreased (P < 0.05) in top layer but increased in middle and substratum layers over time. This indicated that acid-soluble and reducible Cr migrated downward with time and the relative mobility of acid-soluble Cr (0.038 mg/kg·d·m) was higher than that of reducible Cr (0.028 mg/kg·d·m). Furthermore, correlation analysis between microbial community and chromium fraction showed the relative abundance of Lysobacter, Flavihumibacter, Flavisolbacter, and Altererythrobacter was significantly (P < 0.05) correlated with acid-soluble and reducible fractions. Thus, these microorganisms might be evaluators to assess the migration of acid-soluble and reducible fractions in red soil. In summary, this study provided a new comprehension on remediation of Cr-contaminated soil by monitoring the migration of acid-soluble and reducible fractions and the changes of related microbial groups.

Bioremediation of cadmium-contaminated paddy soil using an autotrophic and heterotrophic mixture.

Cadmium (Cd) pollution poses a serious risk to human health and ecological security. Bioremediation can be a promising and effective remediation technology for treating Cd contaminated soils. In this study, seven heterotrophic strains were isolated from Cd contaminated soil and 7 autotrophic strains were isolated from acid mine drainage. Cd removal efficiencies were compared after leaching with autotrophic bacteria (Att-sys), heterotrophic isolates (Htt-sys) and cooperative leaching systems (Co-sys) in laboratory agitating reactors. The results indicated that Cd removal efficiency of Co-sys (32.09%) was significantly higher than that of Att-sys (23.24%) and Htt-sys (0.74%). By analyzing the soil microbial community in different bioleaching systems, we found that the addition of heterotrophic isolates significantly promoted the growth of some heavy metal resistant inhabitants (Massilia, Alicyclobacillus, Micromonospora, etc.), and Co-sys had a minor effect on the growth of soil indigenous microbes. In Co-sys, the content of the four Cd fractions all decreased compared with other leaching systems. The analysis of soil physicochemical parameters during the leaching process showed that pH and ORP (oxidation reduction potential) were not the only determinants for Cd removal efficiency in Co-sys, synergistic metabolic activities of autotrophic and heterotrophic strains may be other determinants. This study demonstrated that cooperative bioremediation may prove to be a safe and efficient technique for field application in heavy metal soil pollution.

Repeated evolution of cytochrome P450-mediated spiroketal steroid biosynthesis in plants.

Diosgenin is a spiroketal steroidal natural product extracted from plants and used as the single most important precursor for the world steroid hormone industry. The sporadic occurrences of diosgenin in distantly related plants imply possible independent biosynthetic origins. The characteristic 5,6-spiroketal moiety in diosgenin is reminiscent of the spiroketal moiety present in anthelmintic avermectins isolated from actinomycete bacteria. How plants gained the ability to biosynthesize spiroketal natural products is unknown. Here, we report the diosgenin-biosynthetic pathways in himalayan paris (Paris polyphylla), a monocot medicinal plant with hemostatic and antibacterial properties, and fenugreek (Trigonella foenum-graecum), an eudicot culinary herb plant commonly used as a galactagogue. Both plants have independently recruited pairs of cytochromes P450 that catalyze oxidative 5,6-spiroketalization of cholesterol to produce diosgenin, with evolutionary progenitors traced to conserved phytohormone metabolism. This study paves the way for engineering the production of diosgenin and derived analogs in heterologous hosts.