真空干燥球孢白僵菌纯孢粉的活孢率、毒力与贮存期

采用真空冷冻干燥法 (冻干 )和真空室温干燥法 (抽干 )对球孢白僵菌 (Beauveriabassiana)原菌粉进行了干燥工艺的比较。在固体产孢基质上生产的新鲜纯孢粉 (原菌粉 )含水量达 5 8 65 % ,经 36h冻干或抽干 ,孢子粉的含水量分别降至 3 97%和 4 2 6% ,含孢量分别为 1 2 9× 1 0 1 1 孢子 /g和 1 2 5× 1 0 1 1 孢子 /g ,活孢率分别为 99 0 %和 97 9%。冻干粉和抽干粉之间的活孢率无显著差异 ,但萌发速度不同。原菌粉、抽干粉及冻干粉对桃蚜(Myzuspersicae)在接种后第 7d的LC50 分别为 1 1 5 ,5 89和 2 95× 1 0 4 个孢子 /mL ;在 1 0 6 个孢子 /mL的剂量下对桃蚜的LT50 分别为 3 6d、 4 4d和 3 9d。从生产成本低得多 ,抽干法比冻干法在白僵菌纯孢粉的生产中更有应用前景 ,尽管冻干粉的质量略优于抽干粉。在4℃和 2 0℃下贮存冻干粉并进行为期一年的孢子萌发率和毒力检测。结果显示 ,冻干粉在2 0℃下贮存至 2 5 5d时完全失活而丧失染虫能力 ,而在 4℃下贮存一年后的活孢率仍达90 2 % ,LT50 仅从初始的 3 9d延长至 4 7d。这说明仅通过降低含水量以延长白僵菌孢子粉在常温下的贮存期仍是不可能的。     

不同含水量和温度下贮存中球孢白僵菌分生孢子活力与内贮营养的衰变

将球孢白僵菌(Beauveriabassiana)BBSG8702的未干燥孢子粉(含水量58.9±1.6%)和真空冷冻干燥孢子粉(含水量7.4±0.9%)置于4℃和20℃下贮存1个月,每隔5d取样测定活孢率和孢子内贮总糖和蛋白含量,发现含水量和贮存温度交互影响孢子的活力以及内贮总糖和蛋白质的代谢水平,各组合中的活孢率一般与内贮总糖和蛋白质代谢水平均存在显著或极显著相关性.在1个月的贮存期间,4℃下冻干粉总糖含量下降13.4%,蛋白质含量下降39.2%,清水中的萌发率下降32.0%,营养液中的萌发率仅下降6.7%,而未干燥孢子粉的相同指标分别下降42.4%、66.3%、96.4%和99%;在20℃下,冻干粉的上述指标分别下降了14.1%、38.2%、55.8%和 10.4%,而未干燥孢子粉则分别下降了 43.2%、65.4%、99.4%和98.4%. 显然,含水量影响活孢率和内贮营养衰变的幅度,而温度影响衰变的速度,但内贮营养的耗尽并不立即引起孢子失活,在供给外源营养之后孢子仍能萌发.将含水量降至4.0±0.9%的冻干粉贮存1年,4℃下活孢率由初始的99.0%下降至90.2%,而20℃下贮存的前165d活孢率下降较为缓慢,但此后急剧下降,至第240d时几乎全部失活.模拟分析表明,低含水量冻干粉在4℃和20℃下贮存的半衰期(即活孢率减少一半所需的时间)分别为1006d和197d.这些结果说明,白僵菌纯孢粉的含水量     

害虫防治用玫烟色拟青霉分生孢子粉的干燥工艺优化

用液－固两相法生产的玫烟色拟青霉Paecilomyces fumosoroseus Pfr116菌株的分生孢子粉,在高真空冷冻干燥,高真空室温抽干,35℃下烘干和低真空低热干燥条件下进行不同程序的干燥处理,以筛选适合该菌孢子粉生产的干燥工艺条件。结果表明,低真空（0.1MPa),低热（30℃）抽干20－24h的干燥方法最适合用于该菌孢子粉的干燥,既能保证含水量在9.0%以下,又能保证87％以上的活孢率和1130亿－1310亿／g的含孢量,而且操作简便,成本较低,可作为高纯度孢子粉生产的首选干燥工艺,高真空（15.86Pa)条件下无论冻干还是室温抽干,虽然孢子粉的含水量（2．2％－8．7％）和含孢量（1270亿－1360亿/g)指标符合生产要求,但活孢率仅62％,说明该菌孢子不适合在高真空条件干燥,在35℃下烘干24h所获孢子粉含水量,24h萌发率和含孢量分别为9．6％,82．8％和1200亿／g。该方法也可在生产中应用,但其活孢率显著低于（P＜0．05）,低真空低热抽干≤24h的孢子粉。     

害虫防治用玫烟色拟青霉分生孢子粉的干燥工艺优化*

用液—固两相法生产的玫烟色拟青霉Paecilomyces fumosoroseus Pfr116菌株的分生孢子粉,在高真空冷冻干燥、高真空室温抽干、35℃下烘干和低真空低热干燥条件下进行不同程序的干燥处理,以筛选适合该菌孢子粉生产的干燥工艺条件。结果表明,低真空(0.1 MPa)低热(30℃)抽干20~24 h的干燥方法最适合用于该菌孢子粉的干燥,既能保证含水量在9.0%以下,又能保证87%以上的活孢率和1130亿~1310亿/g的含孢量,而且操作简便,成本较低,可作为高纯度孢子粉生产的首选干燥工艺。高真空(15.86 Pa)条件下无论冻干还是室温抽干,虽然孢子粉的含水量(2.2%~8.7%)和含孢量(1270亿~1360亿/g)指标符合生产要求,但活孢率仅62%,说明该菌孢子不适合在高真空条件下干燥。在35℃下烘干24 h所获孢子粉含水量、24 h萌发率和含孢量分别为9.6%、82.8%和1200亿/g,该方法也可在生产中应用,但其活孢率显著低于(P<0.05)低真空低热抽干24 h的孢子粉。     

球孢白僵菌对桃蚜接种后特定时间内的侵染率

用球孢白僵菌 (Beauveriabassiana)BBSG870 2菌株的分生孢子悬液 (5× 10 6个·ml-1孢子 )对桃蚜 (Myzuspersicae)始产若蚜的成蚜进行表面接种 ,在接种后 5 6h内 ,每隔 8h取样用 0 .2 %百菌清处理蚜虫 ,使其体表残存的孢子全部失活 ,分别置于 10℃和 2 0℃下逐日观察感染引起的死亡 .结果表明 ,10℃下接种后 5 6h、2 0℃下接种后 40h内各时间段有效侵染引起的死亡率相互间存在显著差异 ,并与不用杀菌剂的对照处理差异显著 (P <0 .0 5 ) .与对照相比 ,接种后 8、16、2 4、32、40和 5 6h内 ,10℃下的有效侵染率分别为 2 2 .9、48.8、6 4.9、80 .4、72 .7和 98.3% ,2 0℃下分别为 31.6、48.8、5 8.6、86 .9、97.2和 98.7% .由此表明 ,在 10～ 2 0℃范围内 ,接种后 2 4h内是该菌有效侵染桃蚜的关键时段 ,有效侵染率达 5 9～ 6 5 % .     

白僵菌孢子悬乳剂田间控蚜效果与常温货架寿命测定

将高纯度球孢白僵菌分生孢子粉兑入一种含悬浮稳定剂及紫外保护剂的矿物油中,配成含孢量为100亿/ml的孢子悬乳剂(纯菌剂),再添加3%的吡虫啉10%可湿剂而得菌药混配剂,于2003年5月在浙江余姚进行了防治蔬菜蚜虫的田间药效试验和农户示范。在两块夏熟白菜地中,纯菌剂和菌药混配剂均用1000倍稀释液(107个孢子/ml)常规喷雾2次,并设清水对照和低用量吡虫啉对照。结果表明,混配剂在两块菜地的相对防效分别达94.5%和86.2%,纯菌剂分别达86.4%和73.1%,而低用量吡虫啉处理的最高防效仅为50%左右。在入夏豇豆、甘蓝、白菜、萝卜及梨树上用菌药混配剂1000倍稀释液对多种蚜虫进行的农户示范试验中,一次性喷雾后5~10d内对菜蚜防效为48%~68%,尤对梨二叉蚜的防效高达99%。在对含有和不含添加剂的两配方孢子悬乳剂进行的常温避光18个月贮存中,活孢率在头7个月均稳定在95%左右且未受配方影响,第10个月活孢率仍有75%左右。而后,含添加剂配方的活孢率下降加速,至16个月时基本失活;而不含添加剂配方的活孢率在第16、17及18个月时还有57%、51%和42%。     

变温干燥固体发酵产物对球孢白僵菌分生孢子性能的影响

[目的]评价球孢白僵菌固体发酵产物的干燥温度对产后分生孢子性能的影响.[方法]采用28℃2和35℃组合的7种恒温或变温处理干燥发酵产物,分析收获的分生孢子质量.[结果]变温干燥可显著降低产后孢子粉的杂菌污染.干燥温度对活孢率和孢子萌发速度影响不一致.35℃恒温干燥5 h后活孢率与新鲜孢子无明显差异,但萌发中时缩短了9.3%.干燥处理提高了孢子对高温和紫外辐射的耐受性.适当的变温干燥比恒温干燥有利于增强孢子抗逆性.干燥温度影响分生孢子胞内海藻糖积累,但其含量与抗逆性无直接相关性.优化干燥温度可提高产后分生孢子毒力.在370～450孢子/mm2剂量下,经28℃ 24 h后升至35℃干燥2 h或35℃恒温干燥5 h的分生孢子对桃蚜的致死中时分别比新鲜孢子缩短了10.6 h和7.5 h.[结论]球孢白僵菌固体发酵产物的干燥温度是影响产后孢子粉杂菌污染、孢子活力、抗逆性和毒力的重要因素.     

顽拗性竹柏种子的贮藏特性?

文章对竹柏( Podocarpus nagi)种子的脱水耐性和贮藏特性进行了研究,结果表明：竹柏种子成熟时初始含水量约为(35±0?7)％,种子对脱水敏感,其最低安全含水量约为(16?86±0?73)％,具有顽拗性种子的典型特征;湿藏和半干藏都可以作为短期保存竹柏种子的方法,且以4℃保存优于15℃保存,但不管种子含水量如何,零下低温保存对竹柏种子都是致命的;半干藏法保存实验中,未进行脱水处理的种子(对照)在4℃贮藏6个月,种子萌发率没有发生明显下降,但贮藏期延长到9个月时,临界含水量的种子萌发力保存最高;不管贮藏介质的含水量高低,也无论贮藏在4℃还是15℃,湿藏种子在9个月的贮藏期内萌发率均没有明显的降低,但当贮藏到12个月时,15℃湿藏种子的萌发率显著高于4℃贮藏的种子,但15℃湿藏的种子在贮藏到3个月时即发现种子在贮藏期间萌发,且随着贮藏介质含水量的升高和贮藏期的延长,萌发的种子增多;竹柏的离体胚经过2h硅胶快速脱水至含水量7％后再冷冻即可获得90％以上的融后存活率,且超低温保存1年的离体胚解冻后,与只保存1周的存活率没有明显差异,表明超低温长期保存竹柏种子是可行的。本研究可以为进一步探究顽拗性种子的短期贮藏和长期保存提供理论基础和基础资料。     

家兔供体卵裂球细胞周期对核移植胚胎发育的影响

通过化学药物处理使家兔桑椹胚卵裂球的细胞周期分别同期化于G1、S或G2期 ,然后分别移入去核的MⅡ期卵母细胞中 ,以研究供体核细胞周期对家兔胚胎细胞核移植效率的影响。试验结果表明供体核细胞周期对家兔核移植胚胎的发育潜力有明显影响 ,以G1期卵裂球为供体的核移植重组胚的激活率、卵裂率、桑椹胚、囊胚和孵化囊胚率及囊胚平均细胞数分别为 87 3% (32 2 / 36 9)、 84 9% (2 99/ 35 2 )、 71 5 % (193/ 2 70 )、5 8 0 % (76 / 131)、 35 1% (4 6 / 131)和 12 6± 4 8,显著高于S期 [79 6 % (10 9/ 137)、 74 4% (93/ 12 5 )、30 3% (33/ 10 9)、19 3% (17/ 88)、 6 8% (6 / 88)和 118 8± 3 5 ]、G2期 [6 3 6 % (70 / 110 )、6 0 % (6 0 / 10 0 )、16 9% (11/ 6 5 )、 16 3% (7/ 43)、 4 7% (2 / 43)和 110 6± 5 8]和未经同期化处理的卵裂球 [78 1% (185 /2 37)、 73 2 % (15 8/ 2 16 )、 49 4% (4 3/ 87)、 32 2 % (2 8/ 87)、 12 6 % (11/ 87)和 12 0 5± 4 4](P <0 0 5 )。来源于G1期卵裂球的 144枚克隆胚胎移植到 12只受体中 ,6只妊娠并产下 19只活仔 ,产仔率为 13 2 % ,显著高于来源于S期 (5 8% ,7/ 12 0 )、G2期 (0 ,0 / 12 6 )或未同期化卵裂球的克隆胚胎 (5 3% ,8/ 15 0 ) (P <0     

顽拗性竹柏种子的贮藏特性

文章对竹柏(Podocarpus nagi)种子的脱水耐性和贮藏特性进行了研究,结果表明:竹柏种子成熟时初始含水量约为(35±0.7)%,种子对脱水敏感,其最低安全含水量约为(16.86±0.73)%,具有顽拗性种子的典型特征;湿藏和半干藏都可以作为短期保存竹柏种子的方法,且以4℃保存优于15℃保存,但不管种子含水量如何,零下低温保存对竹柏种子都是致命的;半干藏法保存实验中,未进行脱水处理的种子(对照)在4℃贮藏6个月,种子萌发率没有发生明显下降,但贮藏期延长到9个月时,临界含水量的种子萌发力保存最高;不管贮藏介质的含水量高低,也无论贮藏在4℃还是15℃,湿藏种子在9个月的贮藏期内萌发率均没有明显的降低,但当贮藏到12个月时,15℃湿藏种子的萌发率显著高于4℃贮藏的种子,但15℃湿藏的种子在贮藏到3个月时即发现种子在贮藏期间萌发,且随着贮藏介质含水量的升高和贮藏期的延长,萌发的种子增多;竹柏的离体胚经过2 h硅胶快速脱水至含水量7%后再冷冻即可获得90%以上的融后存活率,且超低温保存1年的离体胚解冻后,与只保存1周的存活率没有明显差异,表明超低温长期保存竹柏种子是可行的。本研究可以为进一步探究顽拗性种子的短期贮藏和长期保存提供理论基础和基础资料。     

常用化学杀螨剂对两种生防真菌孢子的相容性测定

用营养液萌发法和平板萌发法测定了10种常用化学杀螨剂与球孢白僵菌（Beauveria bassiana）和玫烟色拟青霉（Paecilomyces fumosoroseus）分生孢子的生物相容性.结果表明,营养液萌发法比平板法灵敏.以24 h内的孢子萌发率作为指标,供试杀螨剂的常用浓度及其5和10倍稀释液对活孢率的影响在不同杀螨剂间差异极显著,并在两种真菌间也存在差异.三唑锡、浏阳霉素、三氯杀螨醇和阿维菌素对生防真菌孢子的抑制力很强,不宜混用.速螨酮、克螨特、乐斯本、尼索朗及双甲脒等杀螨剂与真菌孢子的相容性优良,可与菌剂田间混用.将速螨酮、克螨特和乐斯本分别按3种浓度与白僵菌的孢子悬乳剂混配后在4 ℃恒温和室内变温下长期共贮,结果显示,3种杀螨剂对孢子活力的影响迥异,混配制剂在室内变温下的贮存期均未达到12个月,离商品化的要求差距较大.乐斯本的长期相容性相对较优,与菌剂低浓度混配可在常温下贮存6.5个月,中浓度和低浓度混配在4 ℃下可贮存12个月.     

Effects of stabilizers on shelf-life of Epicoccum nigrum formulations and their relationship with biocontrol of postharvest brown rot by Monilinia of peaches

AIM: To find a formulation of Epicoccum nigrum conidia that maintains a high viability over time and which proves efficient to biocontrol peach rot caused by Monilinia spp. METHODS AND RESULTS: We tested the effect of stabilizers and desiccants on the shelf-life of Epicoccum nigrum conidia. Conidial samples were dried for 40 min at 40 degrees C in a fluidized bed-dryer to obtain moisture contents <15%. The toxicity of additives was tested by assaying production of conidia in fermentations and germinability of the produced conidia: 50% PEG300, 10%-5% KCl (stabilizers) and 95.24% Cl(2)Ca (desiccant) significantly (P = 0.05) reduced conidial germination. To enhance shelf-life of dried conidia, nontoxic stabilizers were added at the following different stages of the production-drying process: (i) to substrate contained in bags before production, (ii) to conidial centrifuge pellets obtained after production, before filtering and drying, (iii) to conidial centrifuge pellets obtained after production, before adding talc and drying, and (iv) to conidial centrifuge pellets obtained after production, before adding silica powder and drying. Conidial germinability was tested at 0, 180 and 365 days after storage at room temperature. Shelf-life of formulations retaining the highest viability were conidia produced with 1% KCl or 50% PEG 8000, conidia dried with 2.5% methylcellulose, and conidia dried with 1% KCl + silica powder. All these formulations improved the shelf-life of E. nigrum conidia and significantly reduced brown rot on peaches. CONCLUSIONS: Our results show that additives improve the shelf-life of E. nigrum and assist controlling brown rot on peaches. SIGNIFICANCE AND IMPACT OF THE STUDY: New improved formulations of a biocontrol agent have been obtained which will improve the control of Monilinia on peach.     

Characterization of Beauveria bassiana and Metarhizium anisopliae isolates for management of tarnished plant bug,Lygus lineolaris (Hemiptera: Miridae)

Selected morphological and physiological characteristics of four Beauveria bassiana (Balsamo) Vuillemin isolates and one Metarhizium anisopliae (Metschnikoff) Sorokin isolate, which are highly pathogenic to Lygus lineolaris (Palisot de Beauvois) (Hemiptera: Miridae), were determined. There were significant differences in conidial size, viability, spore production, speed of germination, relative hyphal growth, and temperature sensitivity. Spore viability after incubation for 24h at 20 degrees C ranged from 91.4 to 98.6% for the five isolates tested. Spore production on quarter-strength Sabouraud dextrose agar plus 0.25% (w/v) yeast extract after 10 days incubation at 20 degrees C ranged from 1.6x10(6) to 15.5x10(6)conidia/cm(2). One B. bassiana isolate (ARSEF 1394) produced significantly more conidia than the others. Spore germination was temperature-dependant for both B. bassiana and M. anisopliae. The time required for 50% germination (TG(50)) ranged from 25.0 to 30.9, 14.0 to 16.6, and 14.8 to 18.0h at 15, 22, and 28 degrees C, respectively. Only the M. anisopliae isolate (ARSEF 3540) had significant spore germination at 35 degrees C with a TG(50) of 11.8h. A destructive sampling method was used to measure the relative hyphal growth rate among isolates. Exposure to high temperature (40-50 degrees C) for 10min had a negative effect on conidial viability. The importance of these characteristics in selecting fungal isolates for management of L. lineolaris is discussed.     

基于分生孢子热胁迫反应的球孢白僵菌耐热菌株筛选

将寄主和原产地不同的16株球孢白僵菌Beauveriabassiana的分生孢子分别悬浮于1mL萌发液中,接受48℃恒温水浴45min的热胁迫后,再在25℃下振荡培养24h,以残存活孢率作为各菌株对热胁迫反应的耐热力指标。结果显示,供试菌株的残存活孢率在4.6%~87.1%之间变化,相互间差异极显著。热胁迫后有三株菌的残活孢率均在40%以上,耐热力强;四株菌的残存活孢率低于10%,耐热力弱;其余菌株的残存活孢率为11.0%~23.6%,耐热力居中。从上述菌株中挑选耐热力强、居中和弱的菌株各一株(残存活孢率分别为40.8%、15.8%和4.6%),分别置于25℃、30℃及35℃下平板培养,其菌落生长表现明显与耐热力的强弱相关。虽然三菌株在25℃下的菌落生长无显著差异,但只有耐热力强的菌株能在35℃下正常生长并产孢,是罕见的球孢白僵菌耐热菌株。     

Glucose improves the in vitro viability of Microsporum canis and Trichophyton mentagrophytes var. mentagrophytes

We describe simple and cost-effective methods using carbohydrates to improve the in vitro viability of dermatophytes. Glucose and sucrose in different concentrations (3, 6, 9 and 12%) were used to maintain fifteen strains of M. canis and T. mentagrophytes var. mentagrophytes at 4 and -20 degrees C. The strains were phenotypically analyzed before storage and reevaluated at 1, 3, 6 and 9 months. At 1 and 3 months, any alterations in the viability or phenotype pattern of the stored strains were noted. At 6 months, both dermatophytes were 100% viable, when preserved in glucose (3, 6, 9 and 12%) at -20 degrees C. All T. mentagrophytes strains were also viable in sucrose (12%), at 4 degrees C and -20 degrees C. However, sucrose failed to improve the viability of M. canis at both temperatures. At 9 months, the higher viabilities without pleomorphism were seen for both dermatophytes preserved in glucose (9 and 12%) at -20 degrees C.     

Effect of temperature and poultry litter in Beauveria bassiana (bals.) Vuill. and Metarhizium anisopliae (Metsch) virulence against the lesser mealworm Alphitobius diaperinus (Panzer) (Coleoptera: Tenebrionidae)

This study was carried out to evaluate the influence of temperature and poultry litter on germination vegetative growth virulence and conidial production of Beauveria bassiana (Bals.) Vuill. and Metarhizium anisopliae (Metsch.) isolates on larvae and adults of the lesser mealworm (Alphitobius diaperinus) (Panzer). The vegetative growth and conidial production were evaluated on culture media. Virulence was studied submerging larvae and adults in a conidial suspension (1 x 10(8) conidia/ml). All the experiments were carried out in growth chamber (26 degrees C and 32 degrees C and 14h photophase). Fungus-killed insects were daily collected and used for microscopic conidial counts. The poultry litter effect was evaluated by submerging the insects in a fungal suspension (10(8) conidia/ml) and then transferring them to cups containing poultry litter (new and used). B. bassiana isolates were more sensitive than M. anisopliae to high temperature because conidia viability, vegetative growth and virulence were negatively affected (P < 0.05). The conidial production was higher to B. bassiana in 26 degrees C (7 to 11 x 10(8) conidia/larval cadaver and 8 x 10(8) conidia/adult cadaver) (P < 0,05). Larval stage was about 10 times more sensitive to M. anisopliae at 26 degrees C than adults stage. Regarding B. bassiana, differences on sensitivity between larval stages and adults were not observed at this temperature. However, at 32 degrees C, larval stage was more sensitive for CB116 and UEL50 isolates. Mortality was higher when larvae and adults (15.7 and 66.7% respectively) were treated by B. bassiana and maintained on new poultry litter at 26 degrees C) (P < 0.05).     

Immersion of 'Coscia' pear fruit in water at 55 degrees C for 60 sec controls Penicillium expansum decay and delays ripening during short storage

'Coscia' is an early ripening pear with a short postharvest Life (1 month) and chemical treatments to prevent decay are generally not undertaken. This, along with the fast deterioration under shelf-life conditions, makes it difficult to contain postharvest moulds. 1-methylcyclopropene (1-MCP) has been employed with success to delay ripening and, as a co-effect, the development of decay was contained, but this treatment is not allowed for organically grown crops. Here we report the results of an alternative approach employing immersion treatments in water at 45, 50, 55 or 60 degrees C for 0 (control), 15, 30, 45 or 60 sec. Fruit was harvested after the climacteric peak, immediately subjected to the heat treatment and stored for 2 weeks at 1 degrees C followed by a 3 days of a simulated shelf-life at 17 degrees C and 75% RH. Half of the fruit was wounded (3 x 3mm) and inoculated with Penicillium expansum (20 microL of 10(4) conidia mL(-1). Decay inhibition and fruit appearance, rated from 0 to 3 (0 = excellent; 1 = good; 2 = scarce; 3 = not marketable), were monitored and compared to the control after storage and shelf-life. All heat treatments affected the mould development when performed for 45 or 60 sec. The best result in terms of decay control and appearance after storage and shelf-life occurred when fruit was immersed at 55 degrees C for 60 sec with a decay reduction of about 85% compared to control (75% decay) and with a good appearance.