Iron/ROS/Itga3 mediated accelerated depletion of hippocampal neural stem cell pool contributes to cognitive impairment after hemorrhagic stroke.

Hemorrhagic stroke, specifically intracerebral hemorrhage (ICH), has been implicated in the development of persistent cognitive impairment, significantly compromising the quality of life for affected individuals. Nevertheless, the precise underlying mechanism remains elusive. Here, we report for the first time that the accumulation of iron within the hippocampus, distal to the site of ICH in the striatum, is causally linked to the observed cognitive impairment with both clinical patient data and animal model. Both susceptibility-weighted imaging (SWI) and quantitative susceptibility mapping (QSM) demonstrated significant iron accumulation in the hippocampus of ICH patients, which is far from the actual hematoma. Logistical regression analysis and multiple linear regression analysis identified iron level as an independent risk factor with a negative correlation with post-ICH cognitive impairment. Using a mouse model of ICH, we demonstrated that iron accumulation triggers an excessive activation of neural stem cells (NSCs). This overactivation subsequently leads to the depletion of the NSC pool, diminished neurogenesis, and the onset of progressive cognitive dysfunction. Mechanistically, iron accumulation elevated the levels of reactive oxygen species (ROS), which downregulated the expression of Itga3. Notably, pharmacological chelation of iron accumulation or scavenger of aberrant ROS levels, as well as conditionally overexpressed Itga3 in NSCs, remarkably attenuated the exhaustion of NSC pool, abnormal neurogenesis and cognitive decline in the mouse model of ICH. Together, these results provide molecular insights into ICH-induced cognitive impairment, shedding light on the value of maintaining NSC pool in preventing cognitive dysfunction in patients with hemorrhagic stroke or related conditions.

Endophytic fungi of Lycium barbarum: isolation, determination, bioactivity and separation of compounds.

Lycium barbarum is widely distributed in China and used as a traditional Chinese medicine herb to treat dizziness, abdominal pain, dry cough, headache and fatigue. Several studies have examined the endophytes of L. barbarum from northwest China; however, few have focused on that from eastern China. The objective of this study was to isolate and determine the endophytic fungi of L. barbarum from Shandong province, as well as to obtain and identify active secondary metabolites from the endophytes. In this study, 17 endophytic fungi were isolated from L. barbarum and denoted as GQ-1 to GQ-17, respectively. These fungi were further classified into ten genera based on the morphological and ITS identification. The crude extracts of these fungi were obtained by using liquid fermentation and EtOAc extraction, and their antibacterial, cytotoxic, and antioxidant activities were evaluated. The results showed that GQ-6 and GQ-16 exhibited high inhibitory activity; GQ-6 and GQ-9 showed high cytotoxic activity and GQ-5 exhibited high scavenging capability for DPPH free radicals. Additionally, Cladosporium sp. GQ-6 was used to investigate the secondary metabolites. The crude extracts were purified by using column chromatography, reverse column, and liquid chromatography, and four monomeric compounds were identified, including two known compounds (α-acetylorcinol (1) and cladosporester B (2)) and two new compounds (cladosporacid F (3) and cladosporester D (4)). The anti-fungal and antibacterial activities of these compounds were confirmed, but no cytotoxic activity was observed. In conclusion, endophytic fungi of L. barbarum from eastern China can serve as a potential source of active natural products with antibacterial and antioxidant properties.

Study on secondary metabolites of endophytic fungus Diaporthe sp. AC1 induced by tryptophan analogs.

Small molecule-induced fermentation of the endophytic fungus Diaporthe sp. AC1 originated from Artemisia argyi was executed to investigate its secondary metabolites. It was fermented in a culture medium containing 5-hydroxytryptophan (5-HTP), 1-methyl-L-tryptophan (1-MT), and tryptamine (TA), respectively. The antibacterial activities of crude extracts against pathogenic bacteria and pathogenic fungi were determined by using the Oxford cup method, while the cytotoxicity of crude extracts against cancer cells was determined by using the MTT method. The results showed that the secondary metabolites of Diaporthe sp. AC1 induced by 1-MT exhibited optimal antibacterial activity and tumor cytotoxicity. The induction conditions of 1-MT were optimized, and the antibacterial activities and tumor cytotoxicity of crude extracts under different induction conditions were investigated. As indicated, the optimal moment for 1-MT addition was before inoculation and its optimal concentration was 0.25 mM. Under these conditions, Diaporthe sp. AC1 was fermented and approximately 12 g of crude extracts was obtained. The crude extracts were then separated and purified to acquire nine monomer compounds, including three new compounds (1-3) and six known compounds (4-9). The antibacterial activities of the compounds against pathogenic bacteria and pathogenic fungi were investigated by using the microdilution method, while their cytotoxicity against cancer cells was analyzed by using the MTT method. The results demonstrated that Compound 1 exhibited moderate antibacterial activities against Verticillium dahlia, Fusarium graminearum, and Botrytis cinerea, as well as a low inhibitory activity against Listeria monocytogenes. Nevertheless, Compound 1 showed significant cytotoxicity against five cancer cells, with IC50 ranging from 12.26 to 52.52 µM. Compounds 2 and 3 exhibited negligible biological activity, while other compounds showed detectable inhibitory activities against pathogenic bacteria and cancer cells.

Exosomes from young healthy human plasma promote functional recovery from intracerebral hemorrhage via counteracting ferroptotic injury.

Intracerebral hemorrhage (ICH), as a type of life-threatening and highly disabled disease, has limited therapeutic approaches. Here, we show that exosomes derived from young healthy human plasma exhibiting typical exosomes features could facilitate functional recovery of ICH mice. When these exosomes are intraventricularly delivered into the brain after ICH, they mainly distribute around the hematoma and could be internalized by neuronal cells. Strikingly, exosomes administration markedly enhanced the behavioral recovery of ICH mice through reducing brain injury and cell ferroptosis. MiRNA sequencing revealed that microRNA-25-3p (miR-25-3p) was differentially expressed miRNA in the exosomes from young healthy human plasma, compared with exosomes from the old control. Importantly, miR-25-3p mimicked the treatment effect of exosomes on behavioral improvement, and mediated the neuroprotective effect of exosomes against ferroptosis in ICH. Furthermore, luciferase assay and western blotting data illustrated that P53 as assumed the role of a downstream effector of miR-25-3p, thereby regulating SLC7A11/GPX4 pathway to counteract ferroptosis. Taken together, these findings firstly reveal that exosomes from young healthy human plasma improve functional recovery through counteracting ferroptotic injury by regulating P53/SLC7A11/GPX4 axis after ICH. Given the easy availability of plasma exosomes, our study provides a potent therapeutic strategy for ICH patients with quick clinical translation in the near future.

Antimicrobial and Cytotoxic Activity of Endophytic Fungi from Lagopsis supina.

In this study, five endophytic fungi belonging to the Aspergillus and Alternaria genera were isolated from Lagopsis supina. The antimicrobial activity of all fungal fermented extracts against Staphylococcus and Fusarium graminearum was tested using the cup-plate method. Among them, Aspergillus ochraceus XZC-1 showed the best activity and was subsequently selected for large-scale fermentation and bioactivity-directed separation of the secondary metabolites. Four compounds, including 2-methoxy-6-methyl-1,4-benzoquinone (1), 3,5-dihydroxytoluene (2), oleic acid (3), and penicillic acid (4) were discovered. Here, compounds 1 and 4 displayed anti-fungal activity against F. graminearum, F. oxysporum, F. moniliforme, F. stratum, Botrytis cinerea, Magnaporthe oryzae, and Verticillium dahliae with diverse MIC values (128-512 µg/ml), which were close to that of the positive control antifungal, actidione (64-128 µg/ml). Additionally, compounds 1 and 4 also exhibited moderate antibacterial activity against S. aureus, Listeria monocytogenes, Escherichia coli, and Salmonella enterica, with low MIC values (8-64 µg/ml). Moreover, compounds 1 and 4 displayed selective cytotoxicity against cancer cell lines as compared with the normal fibroblast cells. Therefore, this study proposes that the endophytic fungi from L. supina can potentially produce bioactive molecules to be used as lead compounds in drugs or agricultural antibiotics.

Development of a sensitive phage-mimotope and horseradish peroxidase based electrochemical immunosensor for detection of O,O-dimethyl organophosphorus pesticides.

In this work, a green, harmless and signal-amplified electrochemical immunosensor based on phage-mimotope M31 (C-P-D-G-N-H-V-P-F-C) and horseradish peroxidase (HRP) was constructed for detecting O,O-dimethyl organophosphorus pesticides (OPs). The glassy carbon electrode (GCE) was modified by nitrogen and boron doped carbon quantum dots and graphene oxide (NBCQDs@GO) which can provide sufficient surface area and enhance the conductivity of the electrode. The O,O-dimethyl OPs class specific antibody mAb3C9 was assembled onto the NBCQDs@GO and the phage-mimotope M31 competitively bound to mAb3C9 with OPs. Furthermore, large amounts of anti-M13 mAb-HRP were introduced to the electrode through thousands of binding sites on the capsid of phage. HRP can catalyze 4-chloro-1-naphthol (4-CN) to produce insoluble precipitates (Benzo-4-chlorhexanedione, 4-CD). Hence, the concentration of OPs can be quantified by measuring impedance signal with electrochemical impedance spectrum (EIS). Under the optimal detection conditions, the 50% inhibitory concentration (IC50) and limits of detection (LODs) values of 9 O,O-dimethyl OPs were in range of 0.989-4.017 ng/mL and 0.003-0.014 ng/mL, respectively. The recovery rates of spiked OPs in cucumber, cabbage and lettuce were 88.20-112.50% with coefficient of variation from 2.97 to 15.64%. Therefore, the immunosensor showed very good sensitivity and demonstrating potential application for the detection of O,O-dimethyl OPs in food samples.

Antimicrobial Potential of Endophytic Fungi From Artemisia argyi and Bioactive Metabolites From Diaporthe sp. AC1.

Endophytic fungi of medicinal plants are important sources of active natural products. In this study, 26 fungi were isolated from Artemisia argyi, which were belonging to eight genera, namely, Alternaria, Fusarium, Chaetomium, Phoma, Diaporthe, Trichoderma, Gibberella, and Colletotrichum. The antimicrobial activities of all fungal extracts were tested by using the cup-plate method against Staphylococcus aureus, Salmonella enteritidis, and Fusarium graminearum. The results demonstrated that 25 extracts (96%) exhibited inhibitory activity against at least one of the tested pathogenic microorganisms. The strain Diaporthe sp. AC1, which showed good antimicrobial activity and high yield of crude extract from fermentation, was selected for the study of secondary metabolites. The crude extract of strain AC1 was purified by silica gel column chromatography, Sephadex LH-20 gel column chromatography, and HPLC, and finally, a new compound phomopsolide G (1), together with three known phomopsolides (2-4) and four other known compounds (5-8), was obtained. The structures of the compounds were elucidated by NMR and/or HR-MS spectroscopy. Microdilution method and MTT colorimetry were used to determine the bioactivity of the compounds. The study demonstrated that the new compound 1 had moderate antifungal activity against F. graminearum, Fusarium moniliforme, and Botrytis cinerea and weak antibacterial activity against Staphylococcus aureus. Compound 1 also showed weak cytotoxicity against HepG2, A549, and MDA-MB-231, with IC50 values of 89.91, 107.65, and 53.97 µM. Additionally, other compounds also exhibited antimicrobial and/or cytotoxic activities. The findings provided the basis for searching drug and agricultural lead compounds from A. argyi-associated fungi resources.

A three-dimensional matrix system containing melatonin and neural stem cells repairs damage from traumatic brain injury in rats.

Brain lesions can cause neural stem cells to activate, proliferate, differentiate, and migrate to the injured area. However, after traumatic brain injury, brain tissue defects and microenvironment changes greatly affect the survival and growth of neural stem cells; the resulting reduction in the number of neural stem cells impedes effective repair of the injured area. Melatonin can promote the survival, proliferation, and differentiation of neural stem cells under adverse conditions such as oxidative stress or hypoxia that can occur after traumatic brain injury. Therefore, we investigated the therapeutic effects of melatonin combined with neural stem cells on traumatic brain injury in rats. First, in vitro studies confirmed that melatonin promoted the survival of neural stem cells deprived of oxygen and glucose. Then, we established a three-dimensional Matrigel-based transplantation system containing melatonin and neural stem cells and then used it to treat traumatic brain injury in rats. We found that treatment with the Matrigel system containing melatonin and neural stem cells decreased brain lesion volume, increased the number of surviving neurons, and improved recovery of neurological function compared with treatment with Matrigel alone, neural stem cells alone, Matrigel and neural stem cells combined, and Matrigel and melatonin combined. Our findings suggest that the three-dimensional Matrigel-based transplantation system containing melatonin and neural stem cells is a potential treatment for traumatic brain injury.

Rapid and sensitive noncompetitive immunoassay for detection of aflatoxin B1 based on anti-immune complex peptide.

Noncompetitive immunoassays for small molecules are generally considered to be more sensitive than competitive ones. In this study, a phage-peptide against immune complex of aflatoxin B1 (AFB1) and nanobody Nb28 was obtained by phage-display technology. The phage-peptide was labeled with peroxidase and used to develop a direct noncompetitive magnetic-chemiluminescent enzyme-linked immunoassay (Nc-MCLEIA) for AFB1. The 50% signal saturation concentration (SC50) and limit of detection (LOD) of Nc-MCLEIA for AFB1 were 0.089 and 0.006 ng/mL, respectively. Compared with competitive immunoassays developed by the Nb28, the sensitivity and efficiency of Nc-MCLEIA were greatly improved. The recoveries of AFB1 from spiked corn, rice, flour, peanut, peanut oil and corn oil samples ranged from 83.8% to 119.2% with coefficient of variable under 8.9%. Furthermore, parallel analysis of natural corn, rice and flour samples by Nc-MCLEIA and HPLC (high performance liquid chromatography) proved that the Nc-MCLEIA was reliable.

Development of a noncompetitive magnetic-phage anti-immunocomplex assay for detecting of organophosphorus pesticides with a thiophosphate group.

Organophosphorus pesticides (OPs) are widely used in agriculture and the monitoring of their residues is very important to protect human health. Immunoassays are important tools for the analysis of small molecules. Generally, noncompetitive mode of immunoassay is considered to be more sensitive than competitive mode. In this study, peptides that can identify immunocomplex of OPs were screened from a phage display library. Subsequently, a second-generation peptide library was constructed and peptides with better performance were isolated. Then, a rapid and sensitive noncompetitive magnetic-phage anti-immunocomplex assay (MPHAIA) for OPs was developed based on the best phage-peptide and single chain antibody immunomagnetic beads. The MPHAIA showed broad specificity for OPs with a thiophosphate group. The half-saturated concentration (SC50) values and limits of detection (LODs) of MPHAIA to 12 OPs were ranged from 15.04 to 105.48 ng/mL and 4.07-14.19 ng/mL, respectively. The accuracy and reliability of MPHAIA were verified by gas chromatography-tandem mass spectrometry (GC-MS/MS) parallel analysis of six kinds of OPs in spiked cucumber samples. The recovery rates were in range of 81.2-116.3% with coefficient of variation from 4.1% to 14.1%, which were consistent with the results of GC-MS/MS.

Tetrahydrofolate Alleviates the Inhibitory Effect of Oxidative Stress on Neural Stem Cell Proliferation through PTEN/Akt/mTOR Pathway.

Neural stem cell (NSC) proliferation is the initial step for NSC participating in neurorehabilitation after central nervous system (CNS) injury. During this process, oxidative stress is always involved in restricting the regenerative ability of NSC. Tetrahydrofolate (THF) is susceptible to oxidative stress and exhibits a high antioxidant activity. While its effect on NSC proliferation under oxidative stress condition remains obscure. Here, NSC were isolated from embryonic mice and identified using immunofluorescent staining. Meanwhile, the results showed that THF (5 µM and 10 µM) attenuated oxidative stress induced by 50 µM hydrogen peroxide (H2O2) in NSC using mitochondrial hydroxyl radical detection and Western blotting assays. Afterward, administration of THF markedly alleviated the inhibitory effect of oxidative stress on NSC proliferation, which was evidenced by Cell Counting Kit-8 (CCK8), neurosphere formation, and immunofluorescence of Ki67 assays. Thereafter, the results revealed that PTEN/Akt/mTOR signaling pathway played a pivotal role in counteracting oxidative stress to rescue the inhibitory effect of oxidative stress on NSC proliferation using Western blotting assays and gene knockdown techniques. Collectively, these results demonstrate that THF mitigates the inhibitory effect of oxidative stress on NSC proliferation via PTEN/Akt/mTOR signaling pathway, which provides evidence for administrating THF to potentiate the neuro-reparative capacity of NSC in the treatment of CNS diseases with the presence of oxidative stress.

Association between postoperative hypoalbuminemia and postoperative pulmonary imaging abnormalities patients undergoing craniotomy for brain tumors: a retrospective cohort study.

Hypoalbuminemia is associated with poor outcome in patients undergoing surgery intervention. The main aim for this study was to investigate the incidence and the risk factors of postoperative hypoalbuminemia and assessed the impact of postoperative hypoalbuminemia on complications in patients undergoing brain tumor surgery. This retrospective study included 372 consecutive patients who underwent brain tumors surgery from January 2017 to December 2019. The patients were divided into hypoalbuminemia (< 35 g/L) and non-hypoalbuminemia group (≥ 35 g/L) based on postoperative albumin levels. Logistic regression analyses were used to determine risk factors. Of the total 372 patients, 333 (89.5%) developed hypoalbuminemia after surgery. Hypoalbuminemia was associated with operation time (OR 1.011, P < 0.001), preoperative albumin (OR 0.864, P = 0.015) and peroperative globulin (OR 1.192, P = 0.004). Postoperative pulmonary imaging abnormalities had a higher incidence in patients with than without hypoalbuminemia (41.1% vs 23.1%, P = 0.029). The independent predictors of postoperative pulmonary imaging abnormalities were age (OR 1.053, P < 0.001), operation time (OR 1.003, P = 0.013) and lower postoperative albumin (OR 0.946, P = 0.018). Pulmonary imaging abnormalities [OR 19.862 (95% CI 2.546-154.936, P = 0.004)] was a novel independent predictors of postoperative pneumonia. Postoperative hypoalbuminemia has a higher incidence with the increase of operation time, and may be associated with postoperative complications in patients undergoing brain tumor surgery.

Combination of the Distance From Tumor Edge to Subventricular Zone and IDH Mutation Predicts Prognosis of Patients With Glioma.

Both subventricular zone (SVZ) contact and isocitrate dehydrogenase 1 (IDH1) mutation have been reported to be related to the outcome of glioma, respectively. However, far too little attention has been paid to the role of tumor edge-SVZ distance in the outcome of glioma. We aim to assess the value of tumor-SVZ distance, as well as combined tumor-SVZ distance and IDH status, in predicting the outcome of gliomas (WHO grade II-IV). Here, the MR images and clinical data from 146 patients were included in the current study. The relationship between survival and the tumor-SVZ distance as well as survival and combination of tumor-SVZ distance and IDH status were determined via univariate and multivariate analyses. In univariate analysis of tumor-SVZ distance, the patients were divided into three types (SVZ involvement, tumor-SVZ distance from 0 to 10 mm, and tumor-SVZ distance >10 mm). The results showed that the OS (p = 0.02) and PFS (p = 0.002) for the patients had a positive correlation with the tumor-SVZ distance. In addition, simple linear correlation found a significant relationship between the two parameters (OS and PFS) and tumor-SVZ distance in patients with non-SVZ-contacting glioma. Combination analysis of the tumor-SVZ distance and IDH status showed that IDH1 mutation and SVZ non-involvement enable favorable outcomes, whereas IDH1 wild type with SVZ involvement indicates a significantly worse prognosis in all patients. Moreover, in patients with non-SVZ-contacting glioma, IDH1 mutation concurrent with tumor-SVZ distance >10 mm has better OS and PFS. IDH1 wild type and tumor-SVZ distance from 0 to 10 mm suggest poorer OS and PFS. Multivariate analysis showed WHO grade IV, SVZ involvement, tumor-SVZ distance from 0 to 10 mm, IDH1 mutation, gross total resection, and chemotherapy serve as independent predictors of OS. WHO grade IV, SVZ involvement, tumor-SVZ distance from 0 to 10 mm, IDH1 mutation, and chemotherapy serve as independent predictors of PFS of patients with glioma. In conclusion, tumor-SVZ distance and IDH1 mutation status are the determinants affecting patient outcome.

Application of phage-display developed antibody and antigen substitutes in immunoassays for small molecule contaminants analysis: A mini-review.

Toxic small molecule contaminants (SMCs) residues in food threaten human health. Immunoassays are popular and simple techniques for SMCs analysis. However, immunoassays based on polyclonal antibodies, monoclonal antibodies and chemosynthetic antigens have some defects, such as complicated preparation of antibodies, risk of toxic haptens using for antigen chemosynthesis and so on. Phage-display technique has been proven to be an attractive alternative approach to producing antibody and antigen substitutes of SMCs, and opened up new realms for developing immunoassays of SMCs. These substitutes contain five types, including anti-idiotypic recombinant antibody (AIdA), anti-immune complex peptide (AIcP), anti-immune complex recombinant antibody (AIcA) and anti-SMC recombinant antibody (anti-SMC RAb). In this review, the principle of immunoassays based on the five types of substitutes, as well as their application and advantages are summarized and discussed.

Lactobacillus fermentum JX306 Restrain D-galactose-induced Oxidative Stress of Mice through its Antioxidant Activity.

Oxidative stress-induced series of related degenerative diseases have received widespread attention. To screen new lactic acid bacteria (LAB) strains to resist oxidative stress, traditional Chinese fermented vegetables were used as a resource library to screen of LAB. The Lactobacillus fermentum JX306 strain, which showed high scavenging activity of DPPH free radical and hydrogen radical, and a strong lipid peroxidation inhibition rate in vitro was selected. L. fermentum JX306 was also examined for its antioxidant capacity in D-galactose-induced aging mice. The results showed that L. fermentum JX306 could significantly decrease malondialdehyde (MDA) levels and improve the activity of glutathione peroxidase (GSH-Px), and total antioxygenic capacity (TOC) in the serum, kidney, and liver. Meanwhile, the strain could remarkably upregulate the transcriptional level of the antioxidant-related enzyme genes, such as peroxiredoxin1 (Prdx1), glutathione reductase (Gsr), glutathione peroxidase (Gpx1), and thioredoxin reductase (TR3) encoding genes in the liver. Besides, histopathological observation proves that this probiotic strain could effectively inhibit oxidative damage to the liver and kidney in aging mice. Therefore, this unique antioxidant strain may have a high application value in the functional food industry and medicine industry.Oxidative stress-induced series of related degenerative diseases have received widespread attention. To screen new lactic acid bacteria (LAB) strains to resist oxidative stress, traditional Chinese fermented vegetables were used as a resource library to screen of LAB. The Lactobacillus fermentum JX306 strain, which showed high scavenging activity of DPPH free radical and hydrogen radical, and a strong lipid peroxidation inhibition rate in vitro was selected. L. fermentum JX306 was also examined for its antioxidant capacity in D-galactose-induced aging mice. The results showed that L. fermentum JX306 could significantly decrease malondialdehyde (MDA) levels and improve the activity of glutathione peroxidase (GSH-Px), and total antioxygenic capacity (TOC) in the serum, kidney, and liver. Meanwhile, the strain could remarkably upregulate the transcriptional level of the antioxidant-related enzyme genes, such as peroxiredoxin1 (Prdx1), glutathione reductase (Gsr), glutathione peroxidase (Gpx1), and thioredoxin reductase (TR3) encoding genes in the liver. Besides, histopathological observation proves that this probiotic strain could effectively inhibit oxidative damage to the liver and kidney in aging mice. Therefore, this unique antioxidant strain may have a high application value in the functional food industry and medicine industry.

Highly sensitive phage-magnetic-chemiluminescent enzyme immunoassay for determination of zearalenone.

Here we demonstrate a novel phage-magnetic-chemiluminescent enzyme immunoassay (P-MCLEIA) for detection of zearalenone (ZEN). The P-MCLEIA was more efficient than conventional ELISA through several improvements. In the P-MCLEIA, magnetic nanoparticles were replaced of microplates as solid phases to reduce the whole incubation time within 40 min. Phage-mimotope was replaced of chemosynthetic antigen to improve the sensitivity of immunoassay. Chemiluminescence substrate was replaced of chromogenic substrate to further improve the sensitivity. The IC50 value of P-MCLEIA was 31.4 pg/mL, which was about 11 times lower than that of phage-magnetic-enzyme linked immunosorbent assay (P-MELISA) and 72 times lower than that of conventional ELISA. The LOD of P-MCLEIA was 4.3 pg/mL. Recovery study of P-MCLEIA was performed by analyzing ZEN levels in spiked corn samples, intra- and inter-assay recoveries were 80.0-119.8% and 82.7-112.7%, respectively. Furthermore, parallel analysis of natural corn samples showed a good correlation between the P-MCLEIA and high performance liquid chromatography.

Antimicrobial potential of endophytic fungi from Astragalus chinensis.

The purpose of the present study was to discover antimicrobial endophytic fungi from Astragalus chinensis. Three fungal endophytes with antibacterial activity were isolated and determined as Chaetomium sp. HQ-1, Fusarium sp. HQ-7 and Fusarium sp. HQ-9 based on the neighbor-joining phylogenetic tree. Chaetomium sp. HQ-1 showed the best antibiotic potential and was thus selected for large-scale fermentation. Bioactivity-directed separation of ME fermentation of strain HQ-1 led to the discovery of three compounds, which were identified as differanisole A (1), 2,6-dichloro-4-propylphenol (2) and 4,5-dimethylresorcinol (3), from the HR-ESI-MS and NMR data analysis. All three compounds exhibited moderate antibacterial activity against Listeria monocytogenes, Staphylococcus aureus, and methicillin-resistant S. aureus, with MIC values ranging from 16 to 128 µg/mL. Compounds 1 and 3 also displayed promising antifungal activity against Selerotium rolfsii with IC50 values of less than 16 and 32 µg/mL, respectively, which were comparable to that of actidione (8 µg/mL). The findings of the present study suggest that the endophytic fungi from A. chinensis have the potential to be used as bactericides and fungicides.

Engineered Pichia pastoris production of fusaruside, a selective immunomodulator.

BACKGROUD: Fusaruside is an immunomodulatory fungal sphingolipid which has medical potentials for treating colitis and liver injury, but its poor natural abundance limits its further study. RESULTS: In this study, we described a synthetic biology approach for fusaruside production by engineered Pichia pastoris that was based on polycistronic expression. Two fusaruside biosynthesis genes (Δ3(E)-sd and Δ10(E)-sd), were introduced into P. pastoris to obtain fusaruside producing strain FUS2. To further enhance the yield of fusaruside, three relevant biosynthetic genes (Δ3(E)-sd, Δ10(E)-sd and gcs) were subsequently introduced into P. pastoris to obtain FUS3. All of the biosynthetic genes were successfully co-expressed in FUS2 and FUS3. Compared to that produced by FUS2, fusaruside achieved from FUS3 were slightly increased. In addition, the culture conditions including pH, temperature and methanol concentration were optimized to improve the fusaruside production level. CONCLUSIONS: Here a novel P. pastoris fusaruside production system was developed by introducing the biosynthetic genes linked by 2A peptide gene sequences into a polycistronic expression construct, laying a foundation for further development and application of fusaruside.

A novel nanobody and mimotope based immunoassay for rapid analysis of aflatoxin B1.

Mimotopes could replace mycotoxins and their conjugates to develop immunoassay methods. The mimotopes obtained by phage display technology were mainly using monoclonal antibodies or polyclonal antibodies as targets. However, the mimotope of recombinant antibody has not been selected and applied to immunoassay for mycotoxin. The purpose of this study was to prove that an immunoassay for mycotoxin could be developed based on both recombinant antibody and its mimotope. Using aflatoxin B1 (AFB1) as a model system, mimotopes of an aflatoxin nanobody Nb28 were screened by phage display. A rapid magnetic beads-based directed competitive ELISA (MB-dcELISA) was developed utilizing Nb28 and its mimotope ME17. The 50% inhibitory concentration and the detection limit of the MB-dcELISA were 0.75 and 0.13 ng/mL, respectively, with a linear range of 0.24-2.21 ng/mL. Further validation study indicated good recovery (84.2-116.2%) with low coefficient of variable (2.2%-15.9%) in spiked corn, rice, peanut, feedstuff, corn germ oil and peanut oil samples. The developed immunoassay based on nanobody and mimotope provides a new strategy for the monitoring of AFB1 and other toxic small molecular weight compounds.

Secondary Metabolites from Polar Organisms.

Polar organisms have been found to develop unique defences against the extreme environment environment, leading to the biosynthesis of novel molecules with diverse bioactivities. This review covers the 219 novel natural products described since 2001, from the Arctic and the Antarctic microoganisms, lichen, moss and marine faunas. The structures of the new compounds and details of the source organism, along with any relevant biological activities are presented. Where reported, synthetic and biosynthetic studies on the polar metabolites have also been included.