灭蚊真菌贵阳腐霉孢子囊和游动孢子形成的观察

贵阳腐霉是很有应用前景的灭蚊真菌.其无性繁殖的游动孢子是其侵犯蚊虫体壁的生理阶段.观察了孢子囊形成的时间及影响因素,并观察了游动孢子形成的动态过程.研究结果为其在灭蚊的实际应用提供了有价值的参考资料.     

灭蚊真菌贵阳腐霉Pythium guiyangense对大鼠的长期安全性测试

为了解灭蚊真菌贵阳腐霉Pythium guiyangense对大鼠是否有长期毒性作用。将大鼠120只随机分4组,经饮水口服贵阳腐霉菌丝体。菌丝剂量分别为200mg/kg、100mg/kg、50mg/kg,对照组饮自来水。分别于试验期的90d和180d各组取鼠总数的1/3处死进行检查,剩余1/3大鼠待停用菌丝体悬液2周后处死。观察大鼠一般情况、血常规、肝肾功能等各项生化和组织学指标。结果表明各组大鼠均发育正常,精神食欲好,体重增加;血液学、生化指标变化无明显的剂量-效应关系;内脏系数无异常;对重要脏器的解剖学及组织学检查未见明显异常。本研究结果证明灭蚊真菌贵阳腐霉对大鼠是安全的,从而为其在防治蚊虫的推广应用中的安全性提供了重要的毒理学依据。     

灭蚊真菌贵阳腐霉Pythium guiyangense对大鼠的长期安全性测试

为了解灭蚊真菌贵阳腐霉Pythium guiyangense对大鼠是否有长期毒性作用。将大鼠120只随机分4组,经饮水口服贵阳腐霉菌丝体。菌丝剂量分别为200mg/kg、100mg/kg、50mg/kg,对照组饮自来水。分别于试验期的90d和180d各组取鼠总数的1/3处死进行检查,剩余1/3大鼠待停用菌丝体悬液2周后处死。观察大鼠一般情况、血常规、肝肾功能等各项生化和组织学指标。结果表明各组大鼠均发育正常,精神食欲好,体重增加;血液学、生化指标变化无明显的剂量-效应关系;内脏系数无异常;对重要脏器的解剖学及组织学检查未见明显异常。本研究结果证明灭蚊真菌贵阳腐霉对大鼠是安全的,从而为其在防治蚊虫的推广应用中的安全性提供了重要的毒理学依据。     

灭蚊真菌贵阳腐霉Pythium guiyangense对大鼠的长期安全性测试

为了解灭蚊真菌贵阳腐霉Pythium guiyangense对大鼠是否有长期毒性作用。将大鼠120只随机分4组,经饮水口服贵阳腐霉菌丝体。菌丝剂量分别为200mg/kg、100mg/kg、50mg/kg,对照组饮自来水。分别于试验期的90d和180d各组取鼠总数的1/3处死进行检查,剩余1/3大鼠待停用菌丝体悬液2周后处死。观察大鼠一般情况、血常规、肝肾功能等各项生化和组织学指标。结果表明各组大鼠均发育正常,精神食欲好,体重增加;血液学、生化指标变化无明显的剂量-效应关系;内脏系数无异常;对重要脏器的解剖学及组织学检查未见明显异常。本研究结果证明灭蚊真菌贵阳腐霉对大鼠是安全的,从而为其在防治蚊虫的推广应用中的安全性提供了重要的毒理学依据。     

水霉拮抗菌的筛选及其拮抗作用的初步研究

为了获得水产动物水霉病病原菌的拮抗微生物,以水霉为靶标,对其拮抗菌进行了筛选,并研究了拮抗作用最强菌株的拮抗活性.实验结果表明:从发生过水霉病害的水体中分离得到了130株细菌,能抑制水霉菌落生长的有三株:LD038、LD057和LD106,其中以LD038的拮抗作用最强,经梅里埃ATB微生物自动鉴定系统鉴定为黏质沙雷氏菌(Serratia marcescens).培养基对LD038的拮抗活性影响显著,LD038在PDA平板上表现出的拮抗活性最强,大小依次为PDA＞BHI＞GY＞CA＞Sabouraud.进一步对菌株LD038体外拮抗活性进行了研究.平板抑制实验表明,LD038能抑制水霉菌丝生长和孢子萌发.琼脂扩散法中,LD038菌落周围产生的抑菌圈直径为20mm,而无菌水对照组中水霉菌丝蔓过滤纸纸片并长至平板边缘;D38菌线间孢子的萌发与孢子距菌线的距离有关,距LD038菌线22mm范围内,孢子的萌发完全被抑制.无菌发酵上清液实验表明,与对照组相比,LD038无菌发酵上清液对孢子的萌发和菌丝生长都表现出显著的抑制作用.其中孢子较菌丝对上清液更为敏感,5倍稀释上清液下的孢子萌发率仅为10%,且萌发后的菌丝生长也同样受到抑制,整个实验过程中菌丝均未形成网状,对照组中萌发形成的菌丝在16h后呈网状.无菌上清原液和2倍稀释上清液中孢子均未萌发.对菌丝而言,仅无菌上清原液能彻底抑制菌丝生长,与对照组相比,2倍和5倍稀释上清液显著延缓了菌丝的生长.当对照组中的菌丝铺满整孔时,2倍和5倍稀释上清液中的菌丝长度分别为0和2.6mm.显微观察表明LD038的无菌发酵上清液导致水霉菌丝形态发生变化,较正常菌丝短而粗大,且出现了黑色原生质聚集.本研究为水霉病害的生物防治提供了一定的理论基础和依据.     

工厂化生产海鲜菇菌包污染霉菌的鉴定及防治

对工厂化生产中海鲜菇菌包污染霉菌进行分离,根据霉菌的形态特征、培养性状及ITS序列分析,鉴定其为哈茨木霉、拟康氏木霉、脉孢霉、长枝木霉、黑曲霉、产红青霉和产黄青霉;在此基础上探讨了常用抑菌剂对霉菌的防治效果及对海鲜菇菌丝生长的影响。结果表明,质量浓度100 mg/L克霉灵对哈茨木霉、黑曲霉、产红青霉、产黄青霉、拟康氏木霉有强抑制作用,质量浓度100 mg/L多菌灵对长枝木霉、产红青霉、产黄青霉、拟康氏木霉有强抑制作用,二者对海鲜菇菌丝生长的抑制都比较弱。可为海鲜菇工厂化生产中污染霉菌的综合防治提供参考。     

蚊幼虫病原真菌贵阳腐霉的生物学研究

为了开发灭蚊真菌贵阳腐霉Pythiumguiyangense用于蚊虫防治,对贵阳腐霉生物学特性进行了研究。采用菌丝生长率、真菌产孢情况以及对致倦库蚊Culexquinquefaciatus1龄幼虫的毒力作为评价的指标。测试了7种人工培养基、7种单糖、7种氮源、真菌生长所需的pH范围和温度范围、以及4种光-暗比的光照程序对真菌的影响。结果证明该真菌生长的适合温度为5℃～35℃,最佳温度为25℃～30℃;适合pH范围是5～12;在pH9～11范围内菌丝和游动孢子生长最好;测试的人工培养基中,按照菌丝生长速度从高到低排列,依次为Czapek’SFE、PYG、KPYG2、SDAY、CMA和PDA。其中从真菌的游动孢子形成量和对蚊幼虫的毒力来看,最佳培养基为SFE;葡萄糖、果糖、乳糖、蔗糖、麦芽糖、甘露糖和可溶性淀粉都是本真菌的合适碳氢营养源;含有机氮的培养基比含无机氮的培养基好;不同的光照程序没有表现明显的影响,但是观察到紫外光对本真菌有明显的抑制作用。     

根癌农杆菌介导的Pr1基因在贵阳腐霉的组成性表达

【目的】构建组成性表达贵阳腐霉Pr1基因的载体pCambia-hph-Pr1,转化贵阳腐霉,以获得高毒力的基因工程转化菌株。【方法】以双元载体pCambia-Pnos-PNN-hph为基本骨架,将Pr1基因置于组成型启动子PgpdA和终止子TtrpC之间,获得含Pr1基因的表达元件。以贵阳腐霉菌丝体为受体材料,利用根癌农杆菌介导的遗传转化法转化贵阳腐霉,观察转化株的生物学特征和灭蚊毒力。【结果】转化株的菌丝生长速度和游动孢子产量与野生株无显著性差异。生物测定表明转化株对蚊幼虫平均致死率达到32.9%,比野生株提高了约一倍,并且使受试蚊幼虫生长速度明显减缓。【结论】利用根癌农杆菌介导的遗传转化获得了遗传稳定的高毒力菌株。     

水霉拮抗菌的筛选与拮抗物特点初步分析

水霉病是淡水和咸淡水鱼类被水霉菌感染引起的常见疾病。该研究从多个养殖水体中分离得到6株水霉拮抗菌,SZK15、XL03、TG08、DJ12、HD05和DS08,以SZK15的拮抗作用最强。对SZK15研究发现,在PDA平板上,其对水霉菌丝抑制距离为11.5 mm,对水霉孢子抑制圈直径为29.7 mm,其10倍浓缩的无细胞发酵液的抑水霉菌圈直径达30.2 mm;在液体培养条件下,SZK15无细胞发酵原液能完全抑制水霉菌丝和孢子的萌发生长,其5倍稀释液中,水霉菌丝生长受抑制而长度稳定在2.0 mm,有30%的孢子萌发,且萌发后菌丝受抑制而未形成菌丝网,其10倍稀释液中水霉菌丝生长慢于对照组。SZK15分泌的拮抗活性物质对121℃高温敏感,对80℃、100℃高温、蛋白酶K和胰蛋白酶均部分敏感。本研究为水霉病害的生物防治提供了潜在新途径。     

铁皮石斛软腐病菌终极腐霉生物学特性及其抑菌植物筛选CSCD

开展铁皮石斛软腐病菌终极腐霉(Pythium ultimum)生物学特性及其抑菌植物筛选研究,为铁皮石斛软腐病防治奠定理论基础。采用菌丝生长速率法测定不同中药材水提物对铁皮石斛软腐病菌终极腐霉的抑制作用及其生物学特性。终极腐霉在PDA培养基上菌丝平均生长速度为0.22 cm/h,最适宜的温度为30℃,最适宜的pH在6~7,最适碳源为葡萄糖、可溶性淀粉,最适氮源为酵母浸膏,光照对菌丝生长影响不显著。当水提物浓度为40 mg干样/mL时,丁香和薄荷的抑制率均分别为100%和96.48%。丁香和薄荷水提物具有防治铁皮石斛软腐病的良好应用潜力。     

Effects of temperature, pH and salinity on the infection of Leptolegnia chapmanii Seymour (Peronosporomycetes) in mosquito larvae

The effects of temperature, pH, and NaCl concentrations on the infectivity of zoospores of Leptolegnia chapmanii (Argentine isolate) were determined for Aedes aegypti and Culex pipiens under laboratory conditions. Zoospores of L. chapmanii were infectious at temperatures between 10 and 35 degrees C but not at 5 or 40 degrees C. At the permissive temperatures, mortality rates in young instars were much higher than in older instars and larvae of Ae. aegypti were more susceptible to L. chapmanii than larvae of Cx. pipiens. At 25 degrees C, Ae. aegypti larvae challenged with L. chapmanii zoospores resulted in 100% infection at pH levels ranging from 4 to 10. Larvae of Cx. pipiens exposed to similar pH and zoospore concentrations resulted in increasing mortality rates from 62% to 99% at pH 4 to 7, respectively, and then decreased to 71% at pH 10. Aedes aegypti larvae exposed to L. chapmanii zoospores in NaCl concentrations ranging from 0 to 7 parts per thousand (ppt) at 25 degrees C resulted in 100% mortality while mortality rates for Cx. pipiens decreases from 96% in distilled water to 31.5% in water with 6 ppt NaCl. Control Cx. pipiens larvae died when exposed at a NaCl concentration of 7 ppt. Vegetative growth of L. chapmanii was negatively affected by NaCl concentrations. These results have demonstrated that the Argentinean isolate of L. chapmanii tolerated a wide range of temperatures, pH, and salinity, suggesting that it has the potential to adapt to a wide variety of mosquito habitats.     

Use of a Novel Sporulation Monitor to Quantify the EVects of Formulation and Storage on Conidiation by Dried Mycelia of the Entomopathogenic Fungus Zoophthora radicans

A novel piece of equipment, the sporulation monitor, is described for the comparison of conidia production from mycelia receiving diVerent formulation and storage treatments. This equipment was used to compare the viability of Zoophthora radicans mycelial samples treated either with 10% maltose solution or with distilled water before drying and storage for 0-12 weeks at 4oC. Freshly dried maltose-treated mycelial mat samples produced significantly more conidia for a significantly longer time than distilled water-treated mat samples of the same age. Very few conidia were produced from mats in either treatment after storage for 4 weeks or longer. There were great diVerences in conidia production from mycelial mat samples produced in diVerent fermenter runs. These results are discussed in relation to the potential for the use of dried mycelia in biological control programmes.     

The effects of fungicides on the growth rates of thermophilous fungi

The effects of the fungicides Captan, Dicloran, Thiram, Verdasan and mercuric chloride (HgCl₂) on the radial and mycelial growth rates of thermophilous fungi isolated from living green leaves were studied. On controlplates Thermoascus aurantiacus andTorula thermophila had the highest radial growth rates whileMalbranchea pulchella var.sulfurea andTalaromyces duponti had the lowest growth rates. Good sporulation was seen on all control plates.With increasing concentrations of fungicides the growth rates were correspondingly retarded and sporulation was reduced. At het higher fungicide concentrations the hyphal tips and young hyphae were thicker and warty or granulated.Of the fungi studies,Aspergillus andMucor pusillus were the most tolerant species; they were capable of growth in more concentrations of fungicides compared with the other species.Talaromyces duponti andThermomyces lanuginosus emerged as the most sensitive species.Most marked reductions in the growth rates were produced by Thiram and Verdasan than by the other fungicides. All the ten thermophilous species studied were capable of growth with more concentrations of HgCl₂ than with the organo mercurial Verdasan.Six of the ten thermophilous species studied showed higher radial growth rates compared with the two fastest growing mesophilic species studied;Trlchoderma viride andZygorhynchus moelleri. While M. pusillus was capable of growth in dry weight at 0.05 ppm of all fine fungicides,Sporotrichum thermophile did not show mycelial growth with Verdasan at 0.05 ppm.Thermomyces lanuginosus showed reduced growth rates with Thiram and no mycelial growth with both HgCl₂ and Verdasan.     

Influence of Water Potential on Growth, Enzyme Secretion and In Vitro Enzyme Activities of Trichoderma harzianum at Different Temperatures

The influence of water potential on linear mycelial growth, secretion, and the in vitro activities of enzymes β-glucosidase, cellobiohydrolase, β-xylosidase, exochitinase, and chymotrypsin of Trichoderma harzianum strain T66 was studied at different temperatures. Nearly linear correlation was found between water potential and colony growth rate at both 25°C and 10°C, with higher growth rates at the higher temperature and higher water potentials. The amounts of enzyme secretion depended on the water potential and not on the quality of salt (NaCl or KCl) used as osmoticum. Enzyme activities were significantly affected by water potential. Significant enzyme activities were measured for most of the enzymes even at −14.800 megapascal (MPa), which is below the water potential where mycelial growth ceased. These results suggest the possibility of using mutants with improved xerotolerance for biocontrol purposes in soils with lower water potentials. Received: 5 October 1999 / Accepted: 22 November 1999     

Toxicity of zinc to fungi, bacteria, and coliphages: influence of chloride ions.

A 10 mM concentration of Zn2+ decreased the survival of Escherichia coli; enhanced the survival of Bacillus cereus; did not significantly affect the survival of Pseudomonas aeruginosa, Norcardia corallina, and T1, T7, P1, and phi80 coliphages; completely inhibited mycelial growth of Rhizoctonia solani; and reduced mycelial growth of Fusarium solani, Cunninghamella echinulata, Aspergillus niger, and Trichoderma viride. The toxicity of zinc to the fungi, bacteria, and coliphages was unaffected, lessened, or increased by the addition of high concentrations of NaCl. The increased toxicity of zinc in the presence of high concentrations of NaCl was not a result of a synergistic interaction between Zn2+ and elevated osmotic pressures but of the formation of complex anionic ZnCl species that exerted greater toxicities than did cationic Zn2+. Conversely, the decrease in zinc toxicity with increasing concentrations of NaCl probably reflected the decrease in the levels of Zn2+ due to the formation of Zn-Cl species, which was less inhibitory to these microbes than was Zn2+. A. niger tolerated higher concentrations of zinc in the presence of NaCl at 37 than at 25 degrees C.     

A procedure for preservation of the mosquito pathogen Culicinomyces clavisporus

Summary An effective storage procedure has been demonstrated for the mosquito pathogen Culicinomyces clavisporus. A mycelial preparation was harvested by filtration, sprayed with a sucrose solution and air dried at 20°C in a laminar flow cabinet until the mycelial mat became crisp. This material was then ground in a hammer mill and particles of less than 355 m were sieved out. Viability of the particles was assessed by studying mycelial growth and conidial formation when particles were added to water agar plates or to water. The particles retained 100% viability after 9 weeks storage at-20°C or 6 days at 4°C. Preparations lost activity rapidly if stored at 20°C. Conidia produced by this method were pathogenic to mosquito larvae.     

Sensitivity of Various Bacteria, Including Actinomycetes, and Fungi to Cadmium and the Influence of pH on Sensitivity

A variety of microorganisms, including gram-negative and gram-positive eubacteria, actinomycetes, yeasts, and filamentous fungi, were tested for their sensitivity to cadmium (Cd). In general, the actinomycetes were more tolerant to Cd than were the eubacteria; gram-negative eubacteria were more tolerant to Cd than were gram-positive eubacteria. The period of exponential growth of the eubacteria and actinomycetes was extended in the presence of Cd. Wide extremes in sensitivity to Cd were noted among the fungi; there was no correlation between the class of fungus and tolerance to Cd. Fungal sporulation was more sensitive to Cd than was mycelial growth, as spore formation was inhibited at Cd concentrations that were noninhibitory to mycelial proliferation. The toxicity of Cd to the eubacteria, actinomycetes, and fungi appeared to be pH dependent, as toxicity was generally potentiated at pH 8 or 9.     

Effect of chromium compounds on sporulation of Eimeria piriformis oocysts.

Freshly defecated unsporulated oocysts of Eimeria piriformis from rabbit were treated with various concentrations (1%, 2.5%, 5%, and 10%) of chromium compounds, potassium dichromate, potassium chromate, chromium oxide and chromium nitrate, to examine their effect on sporulation. The sporulation time of oocysts treated with 1 to 10% K(2)Cr(2)O(7) was 28 h. However, much longer sporulation times of about 60 h were required for oocysts treated with 2.5% CrO(3) and Cr(NO(3))(3). Moreover, for oocysts treated with distilled water, 1% K(2)CrO(4) and 10% K(2)CrO(4), the sporulation times required were 216, 156 and 96 h, respectively. Thus, potassium dichromate was found to have higher catalytic activity for the sporulation of E. piriformis oocysts than other chromium compounds.