辣椒籽抗菌肽对黄曲霉的抑制作用

研究辣椒籽抗菌肽对玉米中分离的致腐菌(黄曲霉)的作用方式,包括最低抑菌浓度(minimum inhibitory concentration,MIC)、以及对黄曲霉生长、孢子萌发、养分吸收及细胞膜透性的影响。辣椒籽抗菌肽能够抑制黄曲霉生长,在1MIC浓度下萌发率降至18. 51%,浓度达到4MIC时能完全抑制黄曲霉孢子的萌发。在黄曲霉菌快速生长但还未产生孢子的时期(24 h)加入抗菌肽,能有效抑制孢子的产生。在黄曲霉生长的不同阶段(0、24、48 h)加入辣椒籽抗菌肽,其利用培养基中营养物质能力显著降低;通过核酸染料Sytox Green(SG)结合荧光显微镜来研究辣椒籽抗菌肽对黄曲霉的细胞通透性,结果显示,抗菌肽处理后黄曲霉菌丝体细胞内出现荧光,说明细胞膜受到损伤导致通透性发生了改变。     

薄荷精油对黄曲霉生长的抑制作用研究

目的 研究薄荷精油对黄曲霉生长及其毒素B1合成的抑制作用。方法 在培养基中加入不同体积浓度的薄荷精油, 以不加薄荷精油作为对照, 通过测定其对产毒黄曲霉的最小抑菌浓度(minimum inhibitory concentration, MIC)、孢子萌发、菌丝干重、菌落直径、微观结构及黄曲霉毒素B1合成量的影响, 探究薄荷精油对黄曲霉的抑制效果。结果 薄荷精油对产毒黄曲霉的MIC为1.6 μL/mL; 2MIC薄荷精油处理对黄曲霉孢子萌发抑制率、黄曲霉菌丝生物量抑制率及黄曲霉毒素B1合成抑制率达到了99%以上，且扫描电镜结果表明，薄荷精油会影响黄曲霉菌丝形态; 同时, 通过熏蒸处理, 薄荷精油能够有效抑制花生中的霉菌增长, 防止霉菌侵染花生。结论 薄荷精油对黄曲霉具有良好的抑制作用, 在粮食储藏、食品防霉等方面具有进一步开发成安全防腐抑菌剂的潜力。     

石香薷提取物抑制黄曲霉机制的初步研究

目的探讨石香薷提取物抑制黄曲霉的作用机制.方法采用杯碟法测抑菌圈,参考Victor方法测最低抑菌浓度(MIC)和最低杀菌浓度(MBC),平板法计数黄曲霉孢子萌发,显微镜观察石香薷提取物作用后的黄曲霉的形态.结果石香薷提取物抑制黄曲霉MIC为0.15 mg/mL,MBC为0.20 mg/mL;石香薷提取物对黄曲霉孢子萌发有显著的抑制作用,可使黄曲霉菌丝、足细胞、孢子梗的形态发生明显改变.结论石香薷提取物对黄曲霉有明显的抑制作用,其抑菌作用与浓度呈正相关,其作用机制是药物首先破坏细胞壁、膜结构,影响细胞正常新陈代谢,从而影响孢子萌发、菌体形态的改变,进而抑制菌体生长.     

山苍子精油抑制黄曲霉菌的生长和产毒作用研究

山苍子精油是一种纯天然植物精油,本文研究了其对黄曲霉生长、代谢和毒素产生的抑制作用,探讨了山苍子精油对黄曲霉菌的抑菌能力和作用机理。本研究将花生放置于自然环境染菌并分离纯化目标菌,采用形态学并结合ITS序列法进行菌株分类鉴定;结合抑菌圈、抑菌率和最低抑菌浓度(MIC)的测定探讨山苍子精油对黄曲霉菌的抑制能力;进行了山苍子精油影响黄曲霉孢子萌发率、生长曲线和黄曲霉毒素B1产生的实验研究;从细胞膜渗透性、细胞酶活性的变化探讨了山苍子精油抑制黄曲霉的作用机理。实验结果表明:从腐败花生中分离筛选出菌株HB₂,经ITS序列法鉴定为黄曲霉(Aspergillus flavus);黄曲霉素测定结果显示其含有黄曲霉素B1(AFB1),质量浓度为3.4×10³μg·kg^-1(纯湿菌体);抑菌圈随精油浓度的增大明显变大,对黄曲霉的最低抑菌体积分数(MIC)为0.800μL·mL^-1;孢子萌发率、牙管长度、黄曲霉菌体的生长量和AFB1的浓度随培养液中精油浓度的增大呈显著下降趋势,当山苍子精油浓度为0.100μL·mL     

肉桂醛、柠檬醛抗黄曲霉作用的研究

目的:研究肉桂醛、柠檬醛的抗黄曲霉作用。方法:在含不同浓度药物的蔡氏培养液中接种一定量的黄曲霉孢子,每隔 4h分别取定量培养液涂布于蔡氏培养皿上进行霉菌计数,直到48h。 26.5℃培养 72h后计数萌发的霉菌孢子;用扫描电镜观察药物作用后黄曲霉的形态、表面结构的变化。结果:与对照组比较,肉桂醛、柠檬醛的浓度分别在≥0.05μg/ml、0.28μg/ml作用4h后,萌发的黄曲霉孢子数明显减少;肉桂醛、柠檬醛作用后黄曲霉孢子菌丝的形态、表面结构明显改变。结论:肉桂醛、柠檬醛对黄曲霉孢子萌发有显著的抑制作用,柠檬醛抑制黄曲霉孢子的萌发更迅速,两者可能通过破坏真菌细胞结构起抗菌作用。     

吲哚对黄曲霉生长及产毒的抑制作用

本文研究了吲哚对黄曲霉的生长以及产毒的抑制作用。通过96孔板微量法发现吲哚能够抑制黄曲霉的生长,其最小抑制浓度(MIC)为100μg/m L;进而采用差量法测定了吲哚对黄曲霉菌丝生长量的抑制作用,结果表明200μg/m L的吲哚处理可以完全抑制黄曲霉的菌丝生长。此外,通过高效液相色谱分析发现吲哚浓度达到50μg/m L时,尽管对菌丝生长量没有明显影响,但是可以有效地抑制黄曲霉毒素B1的产生。这些结果说明吲哚抑制毒素产生并不是通过抑制生长来实现的。为了进一步了解吲哚对毒素产生的抑制机制,本研究通过RT-PCR分析了吲哚对黄曲霉产毒相关基因表达的影响。结果显示吲哚对产毒调控基因afl R的影响与其对毒素的作用趋势相同,同时吲哚还能够下调其他一些产毒相关基因afl K和alf D的表达。我们的研究结果表明吲哚具有很高的开发价值应用于粮食和饲料中黄曲霉毒素污染的控制。     

茶多酚对黄曲霉生长及产毒能力的影响

基于氧化应激与黄曲霉毒素生物合成的持续相关性,研究了天然抗氧化剂茶多酚对黄曲霉生长及产毒的影响,并通过扫描电镜观察茶多酚对黄曲霉菌丝体形态的影响,旨在寻找一种安全、高效、环境友好型的抑菌剂来控制食品中黄曲霉毒素的污染。结果表明:茶多酚对黄曲霉菌落生长有一定抑制作用,在10 mg/m L浓度下,抑制率达到48.41%,使黄曲霉菌落生长速率为(5.57±0.16)mm/d,延滞期为(1.32±0.13)d;而茶多酚对黄曲霉产毒抑制效果显著,在1,5和10 mg/m L浓度下,茶多酚对AFB1生物合成的抑制率分别达到了60.15%,87.12%和97.48%;且茶多酚可严重破坏黄曲霉菌丝体的超微结构。因此,茶多酚可作为天然抑菌剂应用于食品与饲料中,控制黄曲霉毒素的污染。     

花生黄曲霉毒素污染拮抗菌B25-5的筛选、鉴定及控制效果

黄曲霉毒素是花生中重要的污染物,对人畜的健康具有极强的危害性。本研究筛选对黄曲霉毒素污染具有较强抑制作用的有益微生物,为花生中黄曲霉毒素污染治理提供支持。本研究从河北省滦南县、滦县、玉田、丰南等主产区县的花生中分离有益微生物,通过室内外实验,获得对产毒黄曲霉较强拮抗菌株B25-5。经分子生物学及理化性鉴定,该菌株为解淀粉芽孢杆菌。进一步开展了B25-5对黄曲霉孢子萌发的抑制作用、黄曲霉毒素的消减作用及黄曲霉毒素的分解作用等几个方面的研究,结果表明,拮抗菌可以显著的抑制产毒黄曲霉孢子的萌发、生长、菌丝的生长,降低黄曲霉毒素的产生及消除黄曲霉毒素。     

使用气态臭氧体外处理控制储存埃及带壳花生中黄曲霉和黄曲霉毒素B1的潜力（网络首发、推荐阅读）

研究考察在不同水分活度（aw, 0.89~0.97=13.1%~24%水分含量）和体外25 ℃条件下,气态臭氧（O3）对黄曲霉菌株（a）孢子萌发,（b）菌丝生长,（c）产黄曲霉毒素B1（AFB1）的影响。此外,以埃及花生为研究对象,在0.93 aw下,带壳花生接种两种浓度的黄曲霉分生孢子（103、105分生孢子/g）置于气态O3中然后储存。在所有情况下,臭氧处理设置为30 min,流速为6 L/min。通常,大于100 ppm浓度的O3 48 h内,可显著抑制黄曲霉菌株（EGP-B07;SRRC-G 1907）在特定酵母蔗糖培养基上的分生孢子萌发。然而,浓度高达300 ppm的O3,对黄曲霉的生长和菌落的扩展没有影响。相同浓度在0.89 aw时能显著影响AFB1的产生。0.93 aw条件下,储存带壳花生中的两种黄曲霉菌株在100~400 ppm O3的数量显著减少。然而,两接种量水平下,AFB1产量仅在 400 ppm O3处理中显著降低。对使用O3气体控制采后带壳花生中的黄曲霉种群和AFB1的结果进行了讨论。     

不产毒黄曲霉菌对产毒黄曲霉菌产毒抑制效果分析

本实验6株菌分离自广东、山东、辽宁和湖北四省的花生土壤中,通过形态学和分子生物学鉴定均为黄曲霉菌,HPLC测定其产毒能力,其中GZ-6为产毒菌,GZ-15、WF-5、WF-20、JZ-2和YC-8为不产毒菌。分别以花生和玉米为培养基,将不产毒黄曲霉菌和产毒菌(孢子浓度:104:105或105:105)进行混合培养,测定不产毒菌对产毒黄曲霉产毒的抑制效果。结果显示:不产毒菌对产毒菌产毒的抑制率随着其孢子浓度的增加而明显加强,当孢子浓度比为105:105(不产毒菌:产毒菌)时,5株不产毒菌在玉米培养基上对产毒菌产毒的抑制率为34.55%~75.94%,在花生培养基上对产毒菌产毒的抑制率为38.03%~83.03%,其中WF-5、WF-20和GZ-15这三株不产毒菌对产毒黄曲霉产毒的抑制效果均达到75.00%以上,可以作为田间防治黄曲霉毒素污染的候选菌株。     

二氢杨梅素抑制黄曲霉菌生长的机制

本研究探讨了二氢杨梅素对黄曲霉的抗真菌活性和潜在的抗真菌机制。首先,通过抑菌实验证明,二氢杨梅素对黄曲霉孢子和菌丝的最小抑菌质量浓度（minimum inhibitory concentration,MIC）均为4 mg/mL。通过荧光增白剂（calcofluor white,CW）和碘化丙锭（propidium iodide,PI）染色实验证明,二氢杨梅素处理后黄曲霉细胞壁和细胞膜受损。与对照组相比,1/2 MIC和MIC组黄曲霉细胞内容物释放量（OD260 nm）分别增加了3.14 倍和5.93 倍,细胞外相对电导率和pH值均升高,MIC的二氢杨梅素对黄曲霉的呼吸抑制率达25.82%。这些结果表明,二氢杨梅素通过破坏细胞壁和细胞膜的完整性以及干扰呼吸代谢发挥抗菌活性。此外,二氢杨梅素在MIC时能够完全抑制黄曲霉在花生和玉米籽粒上的萌发,因此,二氢杨梅素可作为一种有效的抗真菌天然化合物应用于粮食及农产品储藏中。     

Aqueous extracts of Tulbaghia violacea inhibit germination of Aspergillus flavus and Aspergillus parasiticus conidia

Aspergillus flavus and Aspergillus parasiticus are important plant pathogens and causal agents of pre- and postharvest rots of corn, peanuts, and tree nuts. These fungal pathogens cause significant crop losses and produce aflatoxins, which contaminate many food products and contribute to liver cancer worldwide. Aqueous preparations of Tulbaghia violacea (wild garlic) were antifungal and at 10 mg/ml resulted in sustained growth inhibition of greater than 50% for both A. flavus and A. parasiticus. Light microscopy revealed that the plant extract inhibited conidial germination in a dose-dependent manner. When exposed to T. violacea extract concentrations of 10 mg/ml and above, A. parasiticus conidia began germinating earlier and germination was completed before that of A. flavus, indicating that A. parasiticus conidia were more resistant to the antifungal effects of T. violacea than were A. flavus conidia. At a subinhibitory extract dose of 15 mg/ml, hyphae of both fungal species exhibited increased granulation and vesicle formation, possibly due to increased reactivity between hyphal cellular components and T. violacea extract. These hyphal changes were not seen when hyphae were formed in the absence of the extract. Transmission electron microscopy revealed thickening of conidial cell walls in both fungal species when grown in the presence of the plant extract. Cell walls of A. flavus also became considerably thicker than those of A. parasiticus, indicating differential response to the extract. Aqueous preparations of T. violacea can be used as antifungal treatments for the control of A. flavus and A. parasiticus. Because the extract exhibited a more pronounced effect on A. flavus than on A. parasiticus, higher doses may be needed for control of A. parasiticus infections.     

Aspergillus flavus growth in the presence of chemical preservatives and naturally occurring antimicrobial compounds

The combined effects of water activity ([a(w)] 0.99 or 0.95), pH (4.5 or 3.5) and antimicrobial agent (potassium sorbate, sodium benzoate, sodium bisulfite, carvacrol, citral, eugenol, thymol, or vanillin) concentration (0, 100, 200 up to 1800 ppm) on the growth of Aspergillus flavus were evaluated in potato dextrose agar (PDA). Mold spore germination time and radial growth rates (RGR) were significantly (p<0.05) affected by the variables. For equal antimicrobial concentration, reduction in pH or a(w) had important effects, lowering RGR and delaying germination time. Depending on a(w) and pH, increase in antimicrobial concentration slightly reduced RGR until a critical concentration where RGR was drastically reduced or mold growth was inhibited. Germination time increased as antimicrobial agent concentration increased and when a(w) and pH decreased. Important antimicrobial differences were observed, being, in general, the natural antimicrobials less pH-dependent than chemical preservatives. A. flavus exhibited higher sensitivity to thymol, eugenol, carvacrol, potassium sorbate, sodium bisulfite, and sodium benzoate (at pH 3.5) than to vanillin or citral.     

Evaluation of five essential oils from aromatic plants of Cameroon for controlling food spoilage and mycotoxin producing fungi

Five essential oils (EO) extracted from Cymbopogon citratus, Monodora myristica, Ocimum gratissimum, Thymus vulgaris and Zingiber officinale were investigated for their inhibitory effect against three food spoilage and mycotoxin producing fungi, Fusarium moniliforme, Aspergillus flavus and Aspergillus fumigatus. Five strains of each fungus were tested. The agar dilution technique was used to determine the inhibitory effect of each EO on the radial growth of the fungus, and a dose response was recorded. The EO from O. gratissimum, T. vulgaris and C. citratus were the most effective and prevented conidial germination and the growth of all three fungi on corn meal agar at 800, 1000 and 1200 ppm, respectively. Moderate activity was observed for the EO from Z. officinale between 800 and 2500 ppm, while the EO from M. myristica was less inhibitory. These effects against food spoilage and mycotoxin producing fungi indicated the possible ability of each essential oil as a food preservative. A comparative test on the preservative ability of the EO from O. gratissimum and potassium sorbate against A. flavus at pH 3.0 and 4.5 showed that the EO remained stable at both pH, whereas the efficacy of potassium sorbate was reduced at higher pH. We concluded that the EO from O. gratissimum is a potential food preservative with a pH dependent superiority against potassium sorbate, and these are novel scientific information.     

一株防治花生采后黄曲霉污染的拮抗菌的筛选及其防控效果

为预防花生储藏过程中的黄曲霉污染,从农田土中分离出2株拮抗芽孢杆菌,通过平板对峙法优选出1株作为研究对象,研究其对黄曲霉的抑制效果及对花生采后黄曲霉污染的防控效果。采用平板对峙法检测拮抗菌的抑菌广谱性并绘制生长曲线,并通过生理生化实验、形态特征观察及16S rDNA序列分析对该菌株进行鉴定。结果表明：优选出拮抗菌B419作为研究对象,其可有效抑制黄曲霉的生长,在拮抗菌浓度为1.0×10⁶ CFU/mL,黄曲霉孢子浓度为1.0×10³ CFU/mL时,黄曲霉孢子萌发抑制率达到98%;相比拮抗菌无菌发酵滤液和拮抗菌细胞悬浮液,拮抗菌发酵液防治花生黄曲霉污染效果最好;该菌株对互隔交链孢霉、枝孢霉、青霉、拟盘多毛孢等多种霉菌都有抑制作用,对大肠杆菌、酵母菌也有抑制作用;该菌株在培养4 h后进入对数期,14 h达到最大,26 h后进入衰亡期;该菌株被鉴定为贝莱斯芽孢杆菌(Bacillus velezensis)。综上,分离的拮抗菌B419对多种微生物都有抑制作用,不仅可以用来预防花生储藏过程中的真菌污染,在防治植物微生物病害方面也具有良好的应用潜力。     

复配精油对粮油中霉菌防治的增效作用及机理研究

开发新型天然防霉剂控制粮食霉变是保障粮食质量安全的重要途径之一。为研究复配植物精油对粮油储藏过程中常见霉菌赭曲霉(Aspergillus ochrator)、黄曲霉(Aspergillus flavus)和黑曲霉(Aspergillus niger)的防霉效果,挑选活性较强的植物精油进行复配并对联合防霉效果进行评价。通过复配植物精油对霉菌孢子萌发、菌丝干重和细胞完整性的影响,对其防霉机理进行初步探究。结果显示,牛至精油对赭曲霉和黑曲霉的防霉效果最好,抑菌圈直径分别为（27.83±0.58 ）、（15.33±0.29）mm,肉桂醛对黄曲霉的防霉效果最好,抑菌圈直径为（18.50±0.87 ）mm;山苍子精油与牛至精油、肉桂醛与牛至精油复配体积比为2：8时对3种霉菌的防霉效果较优;通过对部分抑菌浓度指数判读,两组植物精油复配对黑曲霉和赭曲霉的防治效果为协同作用,对黄曲霉的防治效果为相加作用;山苍子精油与牛至精油按体积比2：8复配可抑制霉菌孢子萌发和菌丝生长、破坏细胞的完整性、改变孢子和菌丝结构;山苍子精油和牛至精油按体积比2：8复配施用于含黄曲霉的玉米上,可有效降低玉米中黄曲霉毒素B1和赭曲霉毒素的含量。本研究为山苍子精油与牛至精油复配作为防霉剂提供理论支持。     

Distribution of aflatoxin-producing Aspergillus flavus and Aspergillus parasiticus in sugarcane fields in the southernmost islands of Japan

The distribution of Aspergillus flavus and Aspergillus parasiticus in sugarcane field soils and on harvested sugarcane stems was studied on seven islands of Okinawa and Kagoshima Prefectures, the southernmost prefectures in Japan. With the use of a combination of dilution plate and plant debris plate techniques, the fungi were detected on all seven islands studied and in 74% of 53 soil samples. The fungi were also found on the cut surfaces of sugarcane stems from one of the islands. A. parasiticus was the predominant fungus, although many atypical A. parasiticus isolates that produced metulated conidial heads were also obtained. The proportions of isolates testing positive for aflatoxin production were ca. 89% (146 of 164) of all isolates and ca. 69% of A. flavus isolates. More than 40% of A. flavus isolates also produced G aflatoxins. Scanning electron microscopic observation of conidial wall texture was useful in distinguishing A. parasiticus from A. flavus. Cyclopiazonic acid, an indole mycotoxin, was never synthesized by any of the A. parasiticus or G aflatoxin-producing A. flavus isolates tested.     

12种真菌钾离子通道蛋白生物信息学分析

利用生物信息学对真菌钾离子通道蛋白进行序列分析,推断并预测其结构和功能。本文筛选了酿酒酵母、黄曲霉、烟曲霉、禾谷镰刀菌、玉米黑粉菌、布拉克须霉等12种真菌的钾离子通道蛋白,综合采用Blast、ProtParam、SignalP和Swiss-Model等生物信息学网站和软件对这些蛋白的理化性质、系统进化、蛋白结构和功能域等进行了分析。结果表明:所选择的钾离子通道蛋白氨基酸序列长度在659~928 aa之间,理论等电点在5.45~9.54之间,主要定位在质膜、液泡上。二级结构分析表明,真菌钾离子通道蛋白均由α-螺旋、β-折叠、无规则卷曲和延伸链等结构元件组成,其空间结构高度相似,且12种钾离子通道蛋白都拥有保守的TxGYGD钾离子选择器特征片段。     

云南大叶种晒青毛茶提取物对产毒黄曲霉生长及产毒的影响

普洱茶在发酵过程中,微生物组成十分复杂,尤其是黑曲霉等霉菌起到主要作用,因此一些消费者担心普洱茶在发酵过程中受到黄曲霉毒素(AFS)的污染。本文通过测定菌落直径、孢子萌发及菌丝体干重等方法研究云南大叶种茶提取物对产毒黄曲霉AS3.4408生长的影响;采用紫外荧光法和HPLC法研究云南大叶种茶提取物对产毒黄曲霉AFS生物合成的影响;并将产毒黄曲霉AS3.4408接种到云南大叶种茶叶中,检测茶叶中的AFS含量,以求对普洱茶的安全性进行评估。研究表明,云南大叶种茶提取物对产毒黄曲霉AS3.4408菌落的生长及产毒均具有显著的抑制作用,且存在明显的剂量依赖关系;将产毒黄曲霉AS3.4408接种到云南大叶种茶叶中,菌株生长良好,但茶叶基质经HPLC检测,未检测到黄曲霉毒素B1、B2、G1和G2,表明云南大叶种茶中的某种(些)成分对黄曲霉毒素的生物合成具有抑制作用。     

In vitro application of selected essential oils and their major components in controlling fungal pathogens of crown rot in Embul banana (Musa acuminata– AAB)

The necessity for pesticide‐free fresh produce have prompted investigating the effect of selected essential oils and their major components on inhibition of conidial germination, appressoria formation and membrane permeability changes of the pathogens responsible for crown rot of banana. Eugenol, which is the major component of the essential oil of Ocimum basilicum, was the most effective chemical component in inhibiting conidial germination of Colletotrichum musae and Fusarium proliferatumin vitro while Cymbopogon citratus oil was the least effective. Both O. basilicum and C. citratus oils and their major components (Eugenol, citral a + b) inhibited appressoria formation by C. musae and changed the selective permeability of conidial membranes. Ocimum basilicum oil, eugenol and citral a + b could be satisfactorily used for inhibition of conidial germination and disruption of conidial activity of banana pathogens.