光频微波对黄曲霉的杀灭试验

黄曲霉是污染粮食、食品、中药材和烟丝的常见霉菌。由于黄曲霉的产物黄曲霉毒素具有很强的致癌作用。因此 ,从这些食品或物品中消除黄曲霉可减少甚至消除黄曲霉毒素的危害。报告了微波辐射杀灭烟丝中黄曲霉的效果。光频微波对烟丝上的黄曲霉菌具有良好的杀灭作用 ,微波辐射 30s后菌落数明显减少 ,经 4 5s作用后可完全杀灭黄曲霉的细胞 (包括菌丝和孢子 )。应用光频微波辐射杀灭黄曲霉具有经济、方便和高效的优点 ,既可以用于大规模工业生产 ,亦可在家庭生活中使用。     

黄曲霉和米曲霉的多相鉴定方法

【背景】黄曲霉(Aspergillus flavus)和米曲霉(Aspergillus oryzae)形态特征相近,基因组高度相似,较难区分。【目的】旨在总结一套准确鉴别二者的分类方法。【方法】利用22株标准菌株对传统形态学、产毒培养基、酶联免疫毒素检测、系统发育分析、产毒基因检测等5种鉴别方法分别进行验证。【结果】各鉴定方法的结果存在异同,单一的鉴定方法容易出现假阴性或假阳性结果。【结论】利用单一方法区分黄曲霉和米曲霉具有潜在风险,多相鉴定方法可以准确鉴别二者。     

黄曲霉菌原生质体的制备和再生技术优化

黄曲霉菌的遗传转化是研究黄曲霉菌致病相关功能基因的前提和基础,而原生质体是研究和建立真菌遗传转化系统的重要工具。本文分别以黄曲霉孢子和菌丝为材料,研究不同条件下黄曲霉原生质体的形成和再生,结果表明,黄曲霉孢子在酶液浓度为纤维素酶∶蜗牛酶∶溶壁酶=1.5%∶1.5%∶1.5%,30℃酶解3 h,原生质体制备率高达97.3%,再生率达89.2%;黄曲霉菌丝在菌龄为42 h,酶液浓度为纤维素酶∶蜗牛酶∶溶壁酶=1.5%∶1.5%∶1.5%,30℃酶解1 h,可获得最高原生质体产量为2.0×10^6个/m L,再生培养基中以1 mol/L蔗糖作为渗透压稳定剂时,原生质体再生率达5.5%。故本实验条件下,黄曲霉孢子原生质体的形成和再生优于菌丝。     

Molecular characterisation and biological control of Aspergillus flavus isolates from Saudi Arabia

Abstract

Aspergillus flavus is a phytopathogenic fungus that produces toxic compounds, aflatoxins, in infected plant tissues which harm human and animal health. In this study, 57 food/feed products were collected from 9 locations in Aseer, KSA. A total of 93 isolates were recovered from the samples and were identified as Aspergillus spp. based on their cultural and microscopic characteristics. Six isolates (Af3, Af23, Af24, Af26, Af45 and Af48) were selected and confirmed as A. flavus using polymerase chain reaction (PCR), sequencing of ITS1-5.8s-ITS2 rDNA region and phylogenetic analyses. The six sequences were deposited in the GenBank under the accession numbers of KU561932, KU561934, KU561935, KU561936, KU561937 and KU561938, respectively. Random amplification of polymorphic DNA (RAPD-PCR) of the six isolates using five primers (OPA-2, OPA-3, OPA-9, OPA-11 and OPA-15), produced polymorphic DNA bands of 12, 36, 25, 1 and 1, respectively. The band sizes ranged from 130 to 1600?bp, whereas no monomorphic bands were observed. The bio-control of the six selected A. flavus isolates using three locally isolated yeasts (Candida davisiana, Rhodotorula graminis and Exophiala dermatitidis) was assessed. On solid media, the three yeast strains inhibited all tested A. flavus isolates. The most effective yeast strain was R. graminis. In liquid media, both yeast strains C. davisiana and R. graminis inhibited the dry weights of the six A. flavus isolates. Bio-control approaches of A. flavus could help controlling the pathogen, ultimately, reduce the risk of aflatoxins in human and animal supplies and reduce the use of chemicals that affect the environment and health.     

Characterization of Nonaflatoxigenic Aspergillus flavus/oryzae Strains Isolated from Korean Traditional Soybean Meju

Filamentous fungi that could be classified into Aspergillus flavus/oryzae were isolated from traditionally fermented meju commercially available in Korea. The samples were analyzed for aflatoxin B1 and ochratoxin A contamination by HPLC; however, no toxin was detected. In addition, fungal and bacterial metagenomic sequencing were performed to analyze the microbial distribution in the samples. The results revealed that the distribution and abundance of fungi and bacteria differed considerably depending on the production regions and fermentation conditions of the meju samples. Through morphological analysis, ITS region sequencing, and assessment of the aflatoxin-producing ability, a total of 32 A. flavus/oryzae strains were identified. PCR analysis of six regions with a high mutation frequency in the aflatoxin gene cluster (AGC) revealed a total of six types of AGC breaking point patterns. The A. flavus/oryzae strains did not exhibit the high amylase activity detected in the commercial yellow koji strain (starter mold). However, their peptidase and lipase activities were generally higher than that of the koji isolates. We verified the safety of the traditionally fermented meju samples by analyzing the AGC breaking point pattern and the enzyme activities of A. flavus/oryzae strains isolated from the samples. The isolated strains could possibly be used as starter molds for soybean fermentation.     

柠檬醛胁迫环境下黄曲霉线粒体的畸变

通过对黄曲霉细胞受柠檬醛损伤后线粒体形态畸变的透射电镜观察,发现柠檬醛所产生胁迫环境影响线粒体DNA复制系统,产生增生变异的巨型线粒体而与之应答。丙二醛法测定黄曲霉细胞内自由基,结果表明药物进入细胞后还通过诱发自由基使线粒体损伤,致使氧化还原系统及细胞能量代谢途径受到影响。     

Heat- and alkali-induced deamination of 5-methylcytosine and cytosine residues in DNA

5-Methylcytosine residues in DNA underwent deamination at high temperatures. Furthemore, their rate of deamination at neutral or alkaline pH was greater than that of cytosine residues in DNA. As sources of [¹⁴C]5-methylcytosine-containing DNA, we used bacteriophage XP-12 DNA, in which 5-methylcytosine residues completely replace C residues, and calf thymus DNA experimentally substituted with [¹⁴C]5-methylcytosine residues. Upon incubation at 95°C in a physiological buffer or at 60°C in 1 M NaOH, the respective rates of deamination of 5-methylcytosine residues were about 3- and 1.5-times those of cytosine residues. Under the same conditions, the free 5-methyldeoxycytidine was converted to thymidine more rapidly than deoxycytidine was converted to deoxyuridine. The reactions at physiological pH and elevated temperature suggest that deamination of 5-methylcytosine residues may yield a significant portion of spontaneous mutations in vivo, especially in view of the lack of thymine-specific mismatch repair systems with specificity and efficiency comparable to that of uracil excision repair systems.     

Total DNA 5-methylcytosine level of rice calli derived from protoplasts is independent of regeneration ability

To study the roles of m⁵C in the differentiation of rice calli derived from protoplasts (protoclones), the m⁵C level of the total DNA was analyzed using the³²P post-labeling method. The level of m⁵C in regenerable and nonregenerable protoclones was similar, as was in calli and leaves of plants regenerated from the same protoclones. Treatment with 0.5 mM 5-azacytidine caused significant reduction of the level of m⁵C and of the regeneration frequency of callus. Significantly increased m⁵C levels were observed during prolonged culture.     

杜仲抗真菌肽对黄曲霉的抑制作用(英文)

用纸片琼脂扩散法,研究了杜仲抗真菌肽EAFP3对黄曲霉(Aspergillus flavus)的抗真菌活性.研究结果表明,EAFP3对黄曲霉有明显抑制作用,最小抑菌浓度为125 μg/mL.     

Substrate-induced lipase gene expression and aflatoxin production in Aspergillus parasiticus and Aspergillus flavus

AIMS: To establish a relationship between lipase gene expression and aflatoxin production by cloning the lipA gene and studying its expression pattern in several aflatoxigenic and nontoxigenic isolates of Aspergillus flavus and A. parasiticus. METHODS AND RESULTS: We have cloned a gene, lipA, that encodes a lipase involved in the breakdown of lipids from aflatoxin-producing A. flavus, A. parasiticus and two nonaflatoxigenic A. flavus isolates, wool-1 and wool-2. The lipA gene was transcribed under diverse media conditions, however, no mature mRNA was detected unless the growth medium was supplemented with 0.5% soya bean or peanut oil or the fungus was grown in lipid-rich medium such as coconut medium. The expression of the lipase gene (mature mRNA) under substrate-induced conditions correlated well with aflatoxin production in aflatoxigenic species A. flavus (SRRC 1007) and A. parasiticus (SRRC 143). CONCLUSIONS: Substrate-induced lipase gene expression might be indirectly related to aflatoxin formation by providing the basic building block 'acetate' for aflatoxin synthesis. No direct relationship between lipid metabolism and aflatoxin production can be ascertained, however, lipase gene expression correlates well with aflatoxin formation. SIGNIFICANCE AND IMPACT OF THE STUDY: Lipid substrate induces and promotes aflatoxin formation. It gives insight into genetic and biochemical aspects of aflatoxin formation.     

黑曲霉对黄曲霉生长、产毒及黄曲霉毒素B1的影响

目的研究黑曲霉对黄曲霉生长、产毒的抑制作用及对AFB1的降解作用。方法将黑曲霉分别与黄曲霉、AFB1共同培养,定期测定培养液pH、菌丝体干重、黄曲霉孢子数、AFB1含量。结果黑曲霉与黄曲霉混合培养时,黄曲霉孢子数、AFB1含量均比单独培养的低,2组之间差异有统计学意义(P<0.05),抑制率达到68.06%~91.52%;加入黑曲霉后,AFB1含量降低,实验组与对照组之间差异有统计学意义(P<0.05),降解率为46.19%。结论黑曲霉既能抑制黄曲霉生长、产毒,又能降解AFB1。     

5-methylcytosine content in the vertebrate deoxyribonucleic acids: Species specificity

Summary RNA-free native DNA samples, isolated by four methods, from different vertebrate tissues and species, were hydrolyzed chemically and enzymatically and analyzed by paper chromatography to estimate the base composition. It was noted that (i) all the DNA preparations analyzed contained 5-methylcytosine, (ii) on the basis of mole percent of 5-methylcytosine, the composition of DNA varied in different species, but not so much in different tissues of the same species, (iii) the method of DNA hydrolysis, but not the method of deproteinization, affected the mole percent of 5-methylcytosine, and (iv) no 5-hydroxymethylcytosine (5-HMC) was detected in any of the DNA preparations analyzed.     

黄曲霉毒素B1降解菌的筛选及在玉米贮藏中的应用

【目的】本研究分离筛选出一株对黄曲霉既有抑制作用又能降解其毒素的拮抗细菌菌株,并将其应用于玉米中的黄曲霉污染防治研究。【方法】试验通过平板筛选法结合玉米活体筛选法对黄曲霉毒素B1 (AFB1)的拮抗细菌进行初筛,以AFB1的降解率和抑制率为指标进行复筛。【结果】分离到的菌株对黄曲霉菌的抑制率为79.20%,对1μg/mL的黄曲霉毒素B1的降解率为68.39%。贮藏期玉米含水量在15%–30%时,该菌株对黄曲霉污染的抑制率与玉米含水量成反比,即玉米含水量在15%时其抑制率达92.46%,玉米含水量在30%时其抑制率为19.41%。玉米含水量在28%时,菌株对黄曲霉污染玉米的防治效果达到36.39%。【结论】筛选出的拮抗菌株为枯草芽孢杆菌,该菌株不仅对黄曲霉菌有抑制作用,而且能减少AFB1对玉米的污染。     

Biochemical properties of a serine protease from Aspergillus flavus and application in dehairing

The proteases are enzymes produced by several filamentous fungi with important biotechnological applications. In this work, a protease from Aspergillus flavus was characterized. The culture filtrate of A. flavus was purified to homogeneity by Sephacryl S-200 column chromatography followed by CM–cellulose. The molecular weight of the purified enzyme was estimated to be approximately 32?kDa by SDS–PAGE. The enzyme hydrolysed BTpNA (N-α-benzoyl-dl-tyrosyl-p-nitroanilide), azo-casein and casein as substrates. Optimal temperature and pH were 55?°C and 6.5, respectively. The enzyme was stimulated by Mg²⁺, Ca²⁺, Zn²⁺ and inhibited by Hg²⁺ and Ag²⁺ and Cu²⁺. The protease showed increased activity with detergents, such as Tween 80 and Triton X, and was stable to the reducing agents, such as β-mercaptoethanol. The protease activity was strongly inhibited in the presence of phenylmethylsulfonyl fluoride, indicating it is a serine protease. The enzyme entrapped in calcium alginate beads retained its activity for longer time and could be reused up to 10 times. The thermostability was increased after the immobilization and the enzyme retained 100% of activity at 45?°C after 60?min of incubation, and 90% of residual activity at 50?°C after 30?min. In contrast, the free enzyme only retained 10% of its residual activity after 60?min at 50?°C. The enzymatic preparation was demonstrated to be efficient in the capability of dehairing without destruction of the hide. The remarkable properties such as temperature, pH and immobilization stability found with this enzyme assure that it could be a potential candidate for industrial applications.     

对多烯类药物耐药的1株黄曲霉临床株耐药性的初步研究

目的对1株分离自疑似侵袭性肺曲霉病患者肺泡灌洗液的黄曲霉进行常用抗真菌药物敏感性的测定,判断其药物敏感性。方法以形态学方法对该菌株进行菌种鉴定;然后按照美国临床和实验室标准研究所(CLSI)的丝状真菌抗真菌药物敏感性试验方案M38-A,测定常用抗真菌药物对该菌株的最低抑菌浓度、最低杀菌浓度;同时以E-test法测定该菌对两性霉素B和伊曲康唑的敏感性。结果微量液基稀释法显示,两性霉素B或制霉菌素对该菌的MIC值均为4μg/ml,MFC值均为16μg/ml;伊曲康唑的MIC值为0.5μg/ml,MFC值为2μg/ml;特比萘芬的MIC为0.03μg/ml,MFC为0.03μg/ml。E-test法结果显示,该菌对伊曲康唑敏感,对两性霉素B耐药。结论临床上可以分离到对多烯类抗真菌药物耐药的黄曲霉株,应该引起重视。     

Decolorization of Synthetic Dyes by Aspergillus flavus

A problem of paramount importance that has attracted the attention of environmental biologists is the discharge of highly colored effluents into the environment by various industries, which use a wide range of synthetic dyes. The existing chemical methods for dye degradation are not only expensive but also contributes to secondary pollution due to high dose of the chemicals used. Hence an alternative is to exploit the potential of microorganisms to alleviate this problem. The current paper deals with the isolation, characterization, and sugar utilization for better growth of Aspergillus flavus, a marine fungus from the Bay of Bengal. The goal is to assess the bioremediation potential of a variety of synthetic, paper mill, and color photography dyes. A correlation between the amount of sugar used, biomass, and quality of protein produced was observed. This fungus is capable of reducing between 80% and 90% of synthetic dyes and 100% color photography effluents within 3 to 7 days, and 8 days, respectively. Significant effect of carbon sources was observed in the decolorization of the synthetic dye crystal violet, up to 90% in 3 to 7 days, by Aspergillus flavus. The organism showed better growth with fructose as the sole carbon source for the least sugar consumption. Therefore, this fungus can be used as an economical and eco-friendly tool to minimize the pollution by industries to a significant extent.