Study On Beauveria Bassiana Cultivated In Submerged Culture

In order to find the culture environment and culture conditions for promoting Beauveria bassiana sporulation, and provide the scientific evidences for fermented in submerged culture, we made researches on cultivation in submerged culture, and indoor the virulence test of submerged conidia to Dendrolimus Punctatus, and making a comparison with aerial conidia.Which making a study on six Beauveria bassiana strain submerged culture with the conditions of 25℃, surging frequency of 130r/min and cultivated 168h, we found that the submerged conidia number of Bb¹⁸⁷ , Bb¹³ and Bb⁰³ are higher than the other three strains, and the peak period is between 108h and 144h with the peak 120h.; the blastospores number of Bb¹⁸⁷,Bb¹³,Bb⁰³ are also the same with submerged conidia, but higher than the other three strains, and with the peak period between 48h and 120h. Among the biomass of six strains, Bb¹⁸⁷ is the highest, and then Bb¹³ and Bb⁰⁵ , the biomass of the other three has no significantly difference. Therefore, we chose Bb¹⁸⁷ and Bb¹³ as the strain cultivated in submerged culture in the following researches, and the .cultivated time is 120h.The results showed that the submerged sporulation depended on a delicate equilibrium in the composition of the medium. Various nitrogen and carbon sources greatly affected the production of submerged conidia. Peanut cake, peanut power, soybean power and (NH₄) ₂SO₄ were suitable nitrogen sources for the conidia formation. Sucrose and dextrose were effective for sporulation of the submerged conidia. For the blastospores formation, peanut cake, soybean power and (NH₄) ₂SO₄ were nitrogen sources good for sporulation, but carbon sources were different from various strains. Various nitrogen also made greatly difference on biomass, peanut cake had upper biomass, soybean power had the second. Among various carbon sources, potato fecula had the upper biomass, but Sucrose and dextrose had no difference from it.The content of nitrogen and carbon sources had important influence on sporulation of Beauveria bassiana in submerged culture. When various carbon sources with the same nitrogen sources, 20g/L-35g/L carbon sources were more effective for producing submerged conidia, 15g/L-30g/L carbon sources were had higher production of blastospores, 35g/L-45g/L carbon sources had upper biomass. When various nitrogen sources with the same carbon sources, 10g/L-25g/L nitrogen sources had more production of submerged conidia and blastospores than the other content of nitrogen sources. 20g/L-50g/L nitrogen sources made higher biomass.Adding proper accessory factor of growth into the cultivated medium, had an advanced effect on sporulation of submerged conidia and blastospores, and on growth of biomass. VB6+VB1+VB2 and compound vitamin B were more effective for production of submerged conidia. Complicated vitamin were more benefit for sporulation of submerged conidia than single vitamin. VB6+VB1+VB2 had the highest number of blastospores, compound vitamin B following it. VB6+VB1+VC had the greatest biomass.The influence of trace elements on submerged sporulation was most spectacular. When all six elements (B, Cu, Fe, Mn, Mo, Zn) were added together, it yielded highest production of submerged conidia. When adding single trace elements, Molybdenum was the most important element to formation of submerged conidia. In formation of blastospores, Mn had important function. The influence of trace elements on biomass also was great. Molybdenum had the highest biomass, adding all six elements had the following biomass.The cultural temperature affected the sporulation of conidia and the growth of biomass significantly. The preference temperature for submerged culture was between 25℃-28℃.At this temperature range, it had greatest production of submerged conidia, blastospores and biomass. Various illumination had effect on submerged culture. 12h illumination was the best way to sporulation of submerged conidia, and Oh illumination was the better one. The sporulation of blastospores was partial to 0h and 12h illumination. On the growth of biomass,12h illumination was the highest ,then 0h and 24h illumination. It is obvious that 24h illumination went against the sporulation of conidia.The pH of the cultural medium had significant effect on growth and sporulation of submerged conidia. When the pH of the cultural medium was between 6．5-7．0, it was the furthest profit for sporulation of submerged conidia. When the pH of the cultural medium was between 6．0-7．0,it was the grestest advanced for the formation of blastospores. When the pH of the cultural medium was between 6．0-7．0,it had the upper biomass.Various culture medium volume in the same triangle-flask supplied different oxygen for the sporulation of submerged conidia. So various culture medium volume had significantly effect on growth and sporulation of submerged conidia. When the culture medium volume was 20ml-50ml, it had not bad production, and the volume of 40ml was the peak. When the culture medium volume was between 30ml-50ml, the sporulation of blastospores was the better, and the volume of 50ml was the best way to sporulation of blastospores. When the culture medium volume was between 40ml-60ml, it was the best benefit for growth of biomass.And the volume of 50ml had the highest biomass.Different concentrations tween-80 was most effective for sporulation of submerged conidia. The submerged conidia production was highest in liquid media containing 0．4%-0．8% tween-80．And lower 0．4% tween-80 had the lower production. The blastospores production was upper in liquid media containing 0．2%-1．0% tween-80, beside of the range had unsatisfactory production. When the liquid media containing 0．1%-1．0% tween-80, it had a great growth of biomass.Supplied the 4 strains of Beauveria bassiana (Bb¹⁸⁷,Bb¹³,Bb⁰³,Bb¹⁰ ),the pathogenicity of the conidia was compared with that of aerial conidia in laboratory bioassay.The data indicated that there was no significant difference in the mortality to Dendrolimus punctatus between the submerged conidia and aerial conidia. There was little different on LC₅₀ between the submerged conidia and aerial conidia.And the LC₅₀ of submerged conidia was a bit lower than that of aerial conidia.The LT₅₀ of submerged conidia was no gteatly difference with that of aerial conidia.The submerged conidia had the correspond toxicity to the aerial conidia.