Cool, wet conditions in the southern US during the maturing stages of rice in 1998 contributed to outbreaks of false smut caused by Ustilaginoidea virens. Water extracts of false smut galls in Asia have been reported ...Cool, wet conditions in the southern US during the maturing stages of rice in 1998 contributed to outbreaks of false smut caused by Ustilaginoidea virens. Water extracts of false smut galls in Asia have been reported to contain ustiloxin toxins, cyclic peptide antibiotics that interfered with microtubule function and caused “lupinosis”-like lesions in mice. Cell-free extracts from false smut galls on rice grown in Arkansas were fractionated by a published procedure for the purification of ustiloxins. The ustiloxin fraction was phytotoxic to Lemnapausicostata (duckweed) at ≥19 μg/ml, but the host plant, rice, was much less susceptible, exhibiting phytotoxic effects in germinating seeds at ≥1000 μg/ml. The aqueous extract of rice false smut galls showed no cytotoxicity to mammalian cell cultures at 200 μg/ml, but the ustiloxin fractionwas cytotoxic at 10 - 100 μg/ml. However, rice false smut galls were not toxic when fed to mice at 10% of chow, but caused feed refusal at higher concentrations. We conclude that for 1) the U. virens which causes false smut in southern USA differs from Asian isolates in that does not produce detectable ustiloxins;and 2) false smut affects the appearance, but not the food safety of rice in the United States.展开更多
Ustiloxins are vital cyclopeptide mycotoxins originally isolated from rice false smut balls that form in rice spikelets infected by the fungal pathogen Ustilaginoidea virens.The toxicity of the water extract of rice f...Ustiloxins are vital cyclopeptide mycotoxins originally isolated from rice false smut balls that form in rice spikelets infected by the fungal pathogen Ustilaginoidea virens.The toxicity of the water extract of rice false smut balls(RBWE) remains to be investigated.Studies have shown that RBWE may be toxic to animals,but toxicological evidence is still lacking.In this study,we found that the IC50 values of RBWE to BNL CL.2 cells at 24 and 48 h were 40.02 and 30.11 μg/m L,respectively,with positive correlations with dose toxicity and time toxicity.After treatment with RBWE,the number of BNL CL.2 cells decreased significantly,and the morphology of BNL CL.2 cells showed atrophy and wall detachment.RBWE induced DNA presynthesis phase arrest of BNL CL.2 cells,increased the proportion of apoptotic cells and inhibited cell proliferation.RBWE up-regulated reactive oxygen species(ROS) levels and lowered mitochondrial membrane potentials.Additionally,Western blot and q RT-PCR results suggested that RBWE exerted the above effects by promoting the Nrf2/HO-1 and caspase-induced apoptosis pathways in vitro and in vivo.The contents of alanine aminotransferase,aspartate aminotransferase,alkaline phosphatase,and total bile acids in the serum of mice from Institute of Cancer were significantly up-regulated by RBWE.At the same time,RBWE can lead to increases in ROS and malondialdehyde contents,decreases in contents of oxidized glutathione,glutathione and reduced glutathione,as well as decrease in catalase and superoxide dismutase activities in mouse liver tissues,demonstrating that oxidative stress occurred in mice.Moreover,liver damage was further detected by haematoxylin-eosin staining and electron microscopy to verify the damage to the mice caused by RBWE.In general,RBWE may cause hepatotoxicity in vivo and in vitro via the apoptosis pathway,which provides a reference for hepatotoxicity and its mechanism of action.展开更多
稻曲病菌在PD 液体培养基中生长良好,并能产生对植物细胞具有高度生物抑制活性的毒素。生物学活性测定袁明,用100%的甲醇能提取稻曲病菌液体培养物中的粗毒素。粗毒素对小麦胚根胚芽的生长有强烈的抑制作用。把毒素主要成分Ustiloxin A ...稻曲病菌在PD 液体培养基中生长良好,并能产生对植物细胞具有高度生物抑制活性的毒素。生物学活性测定袁明,用100%的甲醇能提取稻曲病菌液体培养物中的粗毒素。粗毒素对小麦胚根胚芽的生长有强烈的抑制作用。把毒素主要成分Ustiloxin A 和BSA 偶联后,制备了抗血清,ELISA 检测表明用两种偶联剂偶联所制备的抗体效价分别为1∶20000和1∶6000。进一步的免疫胶体金标记分析表明,所制备的抗体能与茼丝中分泌的毒素特异性结合,说明所获得的抗体是特异性的。展开更多
目的明确黄曲霉中Zn(II)2Cys6转录因子稻曲菌素B调控蛋白R(ustiloxin B biosynthesis protein R,UstR)功能,解析UstR对黄曲霉生长、次级代谢和黄曲霉毒素(aflatoxin,AF)合成的调控。方法本研究利用聚乙二醇介导的同源重组构建ustR基因...目的明确黄曲霉中Zn(II)2Cys6转录因子稻曲菌素B调控蛋白R(ustiloxin B biosynthesis protein R,UstR)功能,解析UstR对黄曲霉生长、次级代谢和黄曲霉毒素(aflatoxin,AF)合成的调控。方法本研究利用聚乙二醇介导的同源重组构建ustR基因缺失突变体;比较敲除突变体和野生型菌株在黄曲霉生长发育、胁迫响应、侵染能力、AF合成及次级代谢产物合成等方面的差异,并通过酶联免疫吸附测定(enzyme-linked immunosorbent assay,ELISA)试剂盒测定AF合成底物变化,利用实时聚合酶链式反应(real-time polymerase chain reaction,RT-qPCR)验证AF合成相关基因的转录水平改变,明确转录因子UstR对AF合成的调控作用。结果基因敲除突变体的分生孢子形成受抑制,孢子产量降低,但菌核形成能力显著提高;ustR缺失不影响黄曲霉细胞壁完整性、胁迫响应能力以及侵染能力;UstR能够影响黄曲霉多种次级代谢产物产量,其中UstR通过正调控AF合成基因簇表达影响AF生物合成。结论UstR是黄曲霉重要的Zn(II)2Cys6转录因子,调控黄曲霉生长发育及AF产生,研究进一步完善了AF合成调控网络,为AF污染防控提供了新的防控潜在靶点。展开更多
Rice false smut is an emerging plant disease worldwide.Ustiloxin A(UstA)is the major mycotoxin found in rice false smut balls,which are fungal colonies in rice florets.In this study,a new method consisting of macropor...Rice false smut is an emerging plant disease worldwide.Ustiloxin A(UstA)is the major mycotoxin found in rice false smut balls,which are fungal colonies in rice florets.In this study,a new method consisting of macroporous resin column chromatography and high-speed countercurrent chromatography(HSCCC)was developed for UstA separation.UstA was extracted by a 3.81%HCOOH solution and adsorbed by XAD-4 resin.UstA was then eluted by a 40%methanol solution supplemented with 0.1%trifluoroacetic acid(TFA).Further purification was achieved by HSCCC using a two-phase solvent system consisting of n-butanol/TFA/H_(2)O(1/0.05/1,v/v/v).Under the optimized conditions,225 mg of UstA was obtained with a purity of 97.39%in a single run,with a final recovery of 65.2%.An inhibitory effect on seed germination of wheat and maize caused by UstA was observed in a preliminary phytotoxicity assay.展开更多
文摘Cool, wet conditions in the southern US during the maturing stages of rice in 1998 contributed to outbreaks of false smut caused by Ustilaginoidea virens. Water extracts of false smut galls in Asia have been reported to contain ustiloxin toxins, cyclic peptide antibiotics that interfered with microtubule function and caused “lupinosis”-like lesions in mice. Cell-free extracts from false smut galls on rice grown in Arkansas were fractionated by a published procedure for the purification of ustiloxins. The ustiloxin fraction was phytotoxic to Lemnapausicostata (duckweed) at ≥19 μg/ml, but the host plant, rice, was much less susceptible, exhibiting phytotoxic effects in germinating seeds at ≥1000 μg/ml. The aqueous extract of rice false smut galls showed no cytotoxicity to mammalian cell cultures at 200 μg/ml, but the ustiloxin fractionwas cytotoxic at 10 - 100 μg/ml. However, rice false smut galls were not toxic when fed to mice at 10% of chow, but caused feed refusal at higher concentrations. We conclude that for 1) the U. virens which causes false smut in southern USA differs from Asian isolates in that does not produce detectable ustiloxins;and 2) false smut affects the appearance, but not the food safety of rice in the United States.
基金funded by the Education Department of Zhejiang Province Foundation of China(Grant No.Y202249221)。
文摘Ustiloxins are vital cyclopeptide mycotoxins originally isolated from rice false smut balls that form in rice spikelets infected by the fungal pathogen Ustilaginoidea virens.The toxicity of the water extract of rice false smut balls(RBWE) remains to be investigated.Studies have shown that RBWE may be toxic to animals,but toxicological evidence is still lacking.In this study,we found that the IC50 values of RBWE to BNL CL.2 cells at 24 and 48 h were 40.02 and 30.11 μg/m L,respectively,with positive correlations with dose toxicity and time toxicity.After treatment with RBWE,the number of BNL CL.2 cells decreased significantly,and the morphology of BNL CL.2 cells showed atrophy and wall detachment.RBWE induced DNA presynthesis phase arrest of BNL CL.2 cells,increased the proportion of apoptotic cells and inhibited cell proliferation.RBWE up-regulated reactive oxygen species(ROS) levels and lowered mitochondrial membrane potentials.Additionally,Western blot and q RT-PCR results suggested that RBWE exerted the above effects by promoting the Nrf2/HO-1 and caspase-induced apoptosis pathways in vitro and in vivo.The contents of alanine aminotransferase,aspartate aminotransferase,alkaline phosphatase,and total bile acids in the serum of mice from Institute of Cancer were significantly up-regulated by RBWE.At the same time,RBWE can lead to increases in ROS and malondialdehyde contents,decreases in contents of oxidized glutathione,glutathione and reduced glutathione,as well as decrease in catalase and superoxide dismutase activities in mouse liver tissues,demonstrating that oxidative stress occurred in mice.Moreover,liver damage was further detected by haematoxylin-eosin staining and electron microscopy to verify the damage to the mice caused by RBWE.In general,RBWE may cause hepatotoxicity in vivo and in vitro via the apoptosis pathway,which provides a reference for hepatotoxicity and its mechanism of action.
文摘目的明确黄曲霉中Zn(II)2Cys6转录因子稻曲菌素B调控蛋白R(ustiloxin B biosynthesis protein R,UstR)功能,解析UstR对黄曲霉生长、次级代谢和黄曲霉毒素(aflatoxin,AF)合成的调控。方法本研究利用聚乙二醇介导的同源重组构建ustR基因缺失突变体;比较敲除突变体和野生型菌株在黄曲霉生长发育、胁迫响应、侵染能力、AF合成及次级代谢产物合成等方面的差异,并通过酶联免疫吸附测定(enzyme-linked immunosorbent assay,ELISA)试剂盒测定AF合成底物变化,利用实时聚合酶链式反应(real-time polymerase chain reaction,RT-qPCR)验证AF合成相关基因的转录水平改变,明确转录因子UstR对AF合成的调控作用。结果基因敲除突变体的分生孢子形成受抑制,孢子产量降低,但菌核形成能力显著提高;ustR缺失不影响黄曲霉细胞壁完整性、胁迫响应能力以及侵染能力;UstR能够影响黄曲霉多种次级代谢产物产量,其中UstR通过正调控AF合成基因簇表达影响AF生物合成。结论UstR是黄曲霉重要的Zn(II)2Cys6转录因子,调控黄曲霉生长发育及AF产生,研究进一步完善了AF合成调控网络,为AF污染防控提供了新的防控潜在靶点。
基金This work was financially supported by the National Key Research and Development Project(2018YFE0206000)National Natural Science Foundation of China(31901805,U1604234,31872914)Jiangsu Agriculture Science and Technology Innovation Fund(CX(19)3004).
文摘Rice false smut is an emerging plant disease worldwide.Ustiloxin A(UstA)is the major mycotoxin found in rice false smut balls,which are fungal colonies in rice florets.In this study,a new method consisting of macroporous resin column chromatography and high-speed countercurrent chromatography(HSCCC)was developed for UstA separation.UstA was extracted by a 3.81%HCOOH solution and adsorbed by XAD-4 resin.UstA was then eluted by a 40%methanol solution supplemented with 0.1%trifluoroacetic acid(TFA).Further purification was achieved by HSCCC using a two-phase solvent system consisting of n-butanol/TFA/H_(2)O(1/0.05/1,v/v/v).Under the optimized conditions,225 mg of UstA was obtained with a purity of 97.39%in a single run,with a final recovery of 65.2%.An inhibitory effect on seed germination of wheat and maize caused by UstA was observed in a preliminary phytotoxicity assay.
