摘要
球孢白僵菌(Beauveria bassiana(Balsamo)Vuillemin，以下简称白僵菌)是一种最常见的昆虫病原真菌，一直以来在微生物防治中起着重要作用，国外已被开发成多种商品制剂，但国内目前尚无关于白僵菌杀虫剂商品制剂登记。主要原因之一是由于制剂和保存方面上的技术问题一直未得到很好解决，从而成为阻止其进一步开发和应用的重要因素。本实验是在白僵菌油剂前期工作基础之上，对白僵菌孢子粉制备、贮存及其相应制剂的改进和林间应用进行了较为系统的研究，旨在探索适合林间应用的白僵菌制剂开发的技术，揭示孢子在贮存过程中各助剂与孢子活性之间的关系，并通过测定其林间应用效果确定其实践中的可行性。同时，本实验还初步研究并筛选了适合枯叶蛾虫瘴霉(Furia gastropachae)的液体培养基和固体培养基。对马尾松毛虫进行的初步感染实验表明，枯叶蛾虫瘴霉具有应用于马尾松毛虫防治试验的潜力。
1．国内产品调查
收集目前国内白僵菌主要生产厂家生产的白僵菌分生孢子粉，检测产品含孢量、含水量以及室内对马尾松毛虫的毒力。结果表明，不同厂家产品在各项指标之间都存在一定差异。含孢量测定结果表明不同产品含孢量相差较大，其只进行粗加工产品含孢量均小于0.4×10~(11)孢子／g。而经过深加工产品含孢量基本都大于0.6×10~(11)孢子／g，最高能达到0.94×10~(11)孢子／g。各产品测定的含水量普遍偏高，都在12％以上，低于企业标准。将幼虫在孢子浓度1×10~8孢子／ml的孢悬液中浸10秒钟进行室内毒力测定，结果表明，不同产品对马尾松毛虫致死率基本可达到80％以上，多数感染率达60％以上，其中最高为84％。LT_(50)多在5.81天以内，其中最短的为4.12天。因此从各项指标结果可以看出，目前我国各厂家生产的白僵菌粉大多具有较强毒力，但要实现白僵菌制剂商品化其菌粉的其它指标仍需要改进和提高。
2．白僵菌油剂的改进
对本实验室研制的951白僵菌油剂的孢子粉以及溶剂油和其它助剂进行了改进和优化。采用冻干、抽干、和38℃烘干对孢子粉进行干燥及测定其含水量、含孢量和活孢率，结果表明抽干或冻干方法均可在14h后使孢子粉含水量降到5％以下、含孢量达到1.34±0.03×10~(11)孢子／g、活孢率96％以上，而烘干显然不适合孢子粉干燥，所得孢子粉含水量仍然过高并导致萌发率下降。溶剂油的筛选表明，色拉油和液体石蜡油对孢子存活的影响较小。在加入5％V_E+0.1％V_c后可显著提高油剂的贮存期，自二月份起室温贮存5个月后孢子萌发率仍可达50％以上。溴氰菊酯与球孢白僵菌分生孢子的生物学相容性很好。在溴氰菊酯有效含量为240×10~(-6)mg／l的白僵菌油剂中，常温贮存3个月后对孢子萌发率无显著影响。室内测定表明在油剂中溴氰菊酯有效浓度为2.0-4.0mg／l、油剂的虫体用量为0.5μl时，2天后致死率可达66％，7天死亡率为
8()%。
3.油剂的林间应用效果
用Bb!3菌株的高纯度分生袍子粉，配制成含饱量为1了袍子/mL的袍子油剂，
在此基础_匕按不同比例加入澳氰菊酷得至lJ含不同浓度澳氰菊酷的分生袍子油剂。于
2002年5月和20仍年3月两次在安徽桐城市郊林场进行了自然条件下的林间试验。
试验共6个处理，1个对照，每处理2个重复。每亩剂量分别为30Oml油剂十300川
澳氰菊酷、30Oml油剂+1 50林l澳氰菊酷、20Oml油剂+200件l澳氰菊醋、20oml油剂+100协l
澳氰菊酷、100ml油剂+100州澳氰菊酷、30Oml油剂和空白对照。在喷后2天的林间
抽样调查中表明，要使快速击倒率达到70%以_匕每亩澳氰菊酷用量约为470闪。若
要在实践中防治3龄松毛虫，澳氰菊酷的用量应适量减少。
4.枯叶蛾虫瘴霉发酵条件和应用的研究初步
自美国引入的枯叶娥病原真菌，枯叶娥虫瘴霉(Furiag’otropachae comb.nov.)，通
过对其不同的碳、氮源、脂肪酸、起始pH、接种量以及正交试验，以菌丝生物量和
产饱量为主要指标筛选确定了适合该虫霉液体培养的配方:黄豆粉10%、白砂糖40q/0、
进口酵母浸出粉15%、色拉油0.5%，pH为6.5，初始接种量为10%。固体培养以黍
米(Panicum milzaeeum L.，俗称黄米)和慧仁米(coix lacryma一bi)为最佳。将生长良好
的菌丝液按10%接入经灭菌并适度熟化的，含水量在50%以上的米粒中，25’C条件
下培养。黍米培养物培养7天的累计产袍量达19.3士1 .5 xl口抱子/粒，慧米仁培养物
培养7天累计产袍量为20.6士1 .5 xl少袍子/粒。因此通过谷物直接培养枯叶蛾虫瘴霉
可避免制作SEMA培养基的烦琐过程。用对马尾松毛虫幼虫进行了感染试验，马尾
松毛虫感染率为23.3士0.2%。
Beauveria bassiana(Balsamo) Vuillemin is a common entomopathogenic fungus and a classical fungal agent which plays an very important role in microbiai control of insect pests. It has been developed into a variety of commercial formulations as mycoinsecticides in some countries. However, there was still no formulation of B. bassiana registered in China although its great contribution has been made in the past three decades. One of the main reasons that prevent it from further research and application is technical problems involving formulation and long-term storage. In this thesis its preparation, storage, formulation and feasible application in forest were studied in order to provide appropriate techniques for preparation of oil formulation of B. bassiana conidia powder. The studies also tried to clarify the relationship between additives added to the oil formulation and viability of the conidial powder during storage, and feasibility in practice by estimating its effects in forest. Meanwhile, pilot studies
on another entomopathogenic fungus, Furia gastropachae. were also conducted to select the optimum liquid and solid medium for production. Artificial infection of F. gastropachae was tried and the result showed that it has potential to protect pine trees from Dindrolimus pinctatus.
1. Investigation of products of B. bassiana
The gemination, conidial content, water content, and virulence on Dindrolimus pinctatus of some product, collected from different factories, were evaluated respectively. The results showed that there were some differences in various quality estimates. Conidial content of the crude conidia powder were fewer thanconidia g"1, much lower than those of other conidia powder , ranging from conidia g to conidia g Water contents of the products were all above 12%, below the standard requirements for enterprises. The indoor virulence of all the products on Dindrolimus
pinctatus was evaluated by dipping in the conidial suspension at concentration of 1 X 10 conidia ml. Mortalities were above 80% generally. Infection rates were mostly above 60%, with the most highest at 84%. LT's were mostly shorter than 5.81d with the shortest at 4.12d. The quality of conidial powder was therefore to be improved before making them into various kinds of formulation although all products caused a high mortality of the insect pests.
2. Optimization of components for oil formulations.
Main components of the B. bassiana conidia oil formulation prepared previously were further optimized. The conidial powder used in this study was freezing-dried.
vacuum-dried, and heat-dried at 38 癈. The effects of the three methods on water content, conidial content, and viability were evaluated. The water content was reduced to 5% by freezing-drying or vacuum-drying after 14h. Conidial content reached 1. 34?. 03X 10" conidia g"1 and viabilities were above 96%. However, water contents were still high even after a long-term drying at 38癈, which undoubtedly resulted in a low viability. Both salad oil and liquid paraffin had comparatively low effect on the viability of conidia after a long-term storage at ambient temperature. By adding additives of 5%VE+0.1%VC. the viability of conidia in oil formulation was still kept above 50% after stored for about 5 months since February. The chemical insecticide. Deltamefhrin, had little effect on the viability of B. bassiana at the concentration of 240 X 10~6mg/l after stored for about 3 months. The results showed that the best concentration of Deltamethrin was between 2-4mg/l when oil dosage sprayed on every insect was kept at O.Sul, and the general mortality was up to 70% in two days, and 80% in seven days.
3. Field efficacy of oil formulation of B. bassiana conidia for Dindrolimus punctatus control
Conidia powder of B. bassiana Bbl3 was diluted into a concentration of 108 conidia g"' by oil. which was added with different concentrations of Deltamethrin. The field efficacy was conducted at Forestry Extention Center of Tongcheng city, Anhui province, in May of 2002 and iMarch of 2003. resp
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