| Entomopathogenic fungi have great potential for integrated pest management programs due to their specificity, mode of action and ease of application. Nomuraea rileyi (Farlow) Samson is an entomopathogenic fungus that is considered to be a promising candidate for development into a commercial microbial insect control agent. It has been shown that many insect species belonging to Lepidoptera including Spodoptera litura and some belonging to Coleoptera are susceptible to N.rileyi. Natural epizootics have been reported from larvae attacking many important agricultural crops in North and South America, Europe, Asia and it is possible artificially to induce epizootics of N. rileyi. It is reported safely to human beings and other non-target organisms including insect parasites and predators. However, technical problems involved in mass production, virulence, and survival of N. rileyi conidia remain obstacles to intensive development and practical use of N. rileyi-based mycoinsecticides for pest control.We isolated a N. rileyi strain from infected silkworm, in previous studies, we studied on the effects of environmental factors on the growth and conidial production, the production and properties of chitinase and cloning and expression of the chitinase gene. In this study we search for a modified technology to improving production of aerial conidia, analysis virulence of aerial conidia by solid fermentation on rice medium, investigate dynamics of survival and virulence of aerial conidia in soil. The results are following:1. A modified technology for production of conidial powder on rice medium. Low-quality rice was used as solid substrate to produce aerial conidia of N. rileyi in 15-cm dishes. The rice layers piled in dishes, the source and concentration of supplemented nitrogen were round affect in yield of conidial powder, number of conidia and water content. The result indicated that, the sporulation of N. rileyi needed a certain amount of nitrogen. Mycelium of N. rileyi growthed very well on rice medium without any of nitrogen, but aerial conidia can't be seen in 26 days; adding a very small amount of yeast, peptone or potassium nitrate to the rice substrate significantly increased yield of conidial powder. Although there was big difference among the peptone, potassium nitrate and yeast, little difference between the peptone and potassium nitrate, so we chose potassium nitrate with supplement nitrogen source; in 15-cm dishes, when rice layer was 90g/dish, the conidia number produced by per gram rice reached to the biggest with 0.91×109 conidia/g; experimental results of supplementary potassium nitrate showed that a supplement of 0.8% potassium nitrate (w/w) was found most favorable to conidial production, yield of conidial powder was 29.41 mg/g, the conidial number was 1.74×109 produced by per gram rice and water content was 18.39%; a supplement of 15mL water in 100g rice was found most favorable to conidial production.2. Virulence of aerial conidia by solid formation on rice mediums was determinated through fourth instar silkworm. The results showed that aerial conidia by solid formation on rice mediums maintained a good virulence, the LC50 of the fourth instar silkworm was 1.43×105 conidia/mL, a little bigger than airial conidia by slant culture on PDA medium, but there was no significant difference.3. To investigate winter-survival of this fungus in soil, N.rileyi conidia produced on low-quality rice substrate for 12 days at 26℃were inoculated into soil, stored both outdoors and indoors under protected conditions from October to April. The changes in the density of N.rileyi were assessed by using CFUs (Colony Forming Units) counting method on a selective medium every sampling period. The virulence of N.rileyi survived in soil during each sampling period was quantified by evaluating the ability to infect fourth instar silkworm. The results indicated that the fungus can be re-isolated from soil samples even after 180 days storage. The number of CFUs/g dry soil reduced from 17667±655×104 on the day of inoculation to 3×104 (indoors) and 2×104 (outdoors) on the 180th postinoculation day, which demonstrated that the surviving rate of N.rileyi in soil is inverse correlation with the time of storage, and it is unrelated to the storage methods in this study. The virulence of N.rileyi survived in soil decreased very slowly on the first sampling period that it caused high lethality in fourth instar silkworm after being inoculated in soil for 60 days, and the average mortality was more than 90%; after 90 days of inoculation, the virulence of N.rileyi which stored indoors was still high enough to kill about 90% of silkworm larvae, but the virulence of the sample stored outdoors dropped sharply, and the average mortality of silkworm were only 58.33%. N. rileyi lost about 80% of the original virulence after 120 days. However, activated by medium, conidia of N. rileyi recovered almost all virulence. This indicated that N. rileyi could overwinter in the soil, and maintained sufficient quantity and virulence to cause a large outbreak of fungal epidemic in the coming year.
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