Investigation On Bioactive Metabolites Produced By Three Insect Gut Fungi

Microorganisms lived in insect gut are important mediators in ecological interactions. The huge amounts of insect species and multifarious of living environment lead to the particularity of insect gut microorganisms. Insect gut microorganisms is a significant place to dicover new microorganisms and bioactive metabolites.In this study, three fungi were isolated from the gut of insect:MF06, MF08, QTYC51. Three fungi were identified by morphological observation and molecular biology identification. From the culture filtrates of three strains,8 metabolites were isolated by bioactivity guided fractionation and the activity were tested. The specific research content as follows:1. The study of wasp gut fungi MF06 and MF08:Eleven strains were isolated from wasp gut. Among them the fermentaiton broth of strain MF06 showed potent herbicidal activity against Amaranthus retroflexus. By the morphological observation and molecular biology identification, MF06 was identified as Fusarium oxysporum. The ethyl acetate extract of MF06 had strong activity against A. retroflexus with inhibition rate of more than 68% under the concentration of 100 μg/mL. Fermentation product was isolated and purified by silica column chromatography, TLC and Sephadex LH-20 column chromatography. Metabolite 1 was obtained from fermentation product, and the structure was determined as a mixture of fusaric acid and 9,10-dehydrofusaric acid. Inhibitory effect of metabolite 1 against the radical growth of A. retroflexus was studied, the IC50 value is (0.51±0.18) μg/mL, comparable to that of 2,4-D ((0.30±0.14) μg/mL) used as a positive control.By the morphological observation and ITS sequence analysis, MF08 was identified as Curvularia lunata. The ethyl acetate extract of MF08 had strong activity against A. retroflexus and Echinochloa crusgalli with inhibition rate of 77% and 88%, comparable to that of 2,4-D (inhibition rate are 90% and 95%) used as a positive control. Fermentation products were isolated and purified by silica column chromatography, TLC and Sephadex LH-20 column chromatography. Metabolite 2 and metabolite 3 was obtained from fermentation product, and the structure was determined as ethyl 2-acetyl-3,5-dihydroxyphenylacetate (2) and methyl 2-acetyl-3,5-dihydroxyphenyl acetate (3). Study herbicidal activity of two metabolites. Unfortunately, they had no herbicidal activity.2. The study of Pantala flavescens gut fungal isolate QTYC51:According to the herbicidal activity, fermentation broth of strain QTYC51 showed potent herbicidal activity against A. retroflexus and E. crusgalli. By the morphological boservation and ITS sequence analysis, QTYC51 was identified as Paraconiothyrium pinophilum. The ethyl acetate extract of QTYC51 had good activity against A. Retroflexus and E. crusgalli with inhibition rate of 15% and 49%. Fermentation products were isolated and purified. Metabolites 4-8 were botained from fermentation product. The structures were determined by nuclear magnetic resonance analyses, namely 1,8-dihydroxy anthraquinone (4), 1-hydroxy-10-methoxy-dibenz[b,e]oxepin-6,11-dione (5), hydrox-yvertixanthone (6), microdiplodianone (7) and 1,3,6,8-tetrahydroxyanthraquinone (8). Tested the herbicidal activity, antibacterial activity and antifungal activity, the results are shown below. (1) Herbicidal activity:Five metabolites have varying degrees of herbicidal activity, and metabolite 7 inhibited radical growth of A. retroflexus and E. crusgalli with IC50 value of (10.23±2.70) μg/mL and (3.21±1.40) μg/mL(2,4-D is positive control, ICso value of (0.30±0.14) μg/mL and (0.0052±0.0036) μg/mL). (2) Antibacterial activity:Metabolite 4 and 7 possess potent antibacterial activity against S. aureus and B. subtilis, metabolite 4 with the MIC value of 25 μg/mL against B. subtilis and metabolite 7 with the MIC value of 12.5 μg/mL against S. aureus and B. subtilis(Gentamicin sulfate is positive control, MIC values are 6.25 μg/mL against S. aureus and B. subtilis). (3) Antifungal activity:Metabolites 5-7 had weakly activity against Magnaporthe oryzae and Dothiorella gregaria.According to the research results and study of insect gut fungal, we can predicting as a kind of pooly studied microbes, insect gut fungi may inspire the discovery of chemical agents with biological activity. These fungi is worth extensive attention and futher study.