The Selection And Evaluation Of Pathogenic Fungi Using For Microbial Herbicide

The purpose of this study is to provide more strains for the research and development of microbialherbicide. The main contents and the results are as follows:The collection and separation research of pathogenic fungi on weeds. More than300natural-infected samples were collected from different ecological conditions of paddy field fromZhejiang, Shangha, Jiangsu, Jiangxi, Malaysia.Three hundredand thirty-six strains of pathogenic fungiwere isolated from the samples.By the colony and spores morphology identification, the pathogenicfungi mainly distributed in genus as follows: Alternaria,Curvularia,Bipolaris,Exserohilum,Fusarium,Phoma,Nigroporus. Among them, Alternaria,Curvularia,Bipolaris,Exserohilum areadvantage genuses, which are accounting for about80%of the separation strains.The selection and identification research of pathogenic fungi with weeding potential. Through thepetri dish bioassay method, we screen34strains from336pathogenic fungies as the secondaryscreening strains. Through pot bioassay method, we find six strains with stronger pathogenicity tobarnyardgrass's (Echinochloa crus-galli (L.) Beauv．) leaf, and have higher inhibition rate tobarnyardgrass's height and fresh weight as well. We extract the DNA of six strains,and sequence theirrDNA ITS region, the results show that: the rDNA ITS sequences of six strains using ITS1and ITS4primer to amplificate are single clear channel,segment sizes are about600bp. According to genesequencing result, we utilize Neibour-Joining method to construct Phylogenetic Tree. Through thePhylogenetic Tree, the genetic relationships among six strains, Bipolaris oryzae (Breda DE Hann) andCochliobolus miyabeanus (Ito et Kuribayaski Drechsler ex Dastur) are closest, which gather into acluster and constitute a branch for98%bootstrap value. Based on morphological characteristics ofcolonies, spores and sequence alignments of rDNA ITS region, we can confirm that six potential strainsare Bipolaris oryzae.The research about herbicide controlling spectrum and safety test to major crop of potential strains.The7-day mycelia suspension made different degrees of pathogenicity to E. crusgalli (L.) Beauv.,Leptochloa chinensis (L.) Nees, Leersia hexandra Swartz, Monochoria vaginalis (Burm.f.) Presl.exKunth, Sagittaria pygmaea Miq. and Eleocharis plantaginerformis Tang et Wang, but made nopathogenicity to Paspalum distichum L., Cyperusdifformis L. and Alopecurus aequalis Sobol..Furthermore, which make strong pathogenicity to E. crusgalli (L.) Beauv., Leptochloa chinensis (L.)Nees, Monochoria vaginalis, Sagittaria pygmaea and Eleocharis plantaginerformis, make weakpathogenicity to Leersia hexandra Swartz and Eleocharis plantaginerformis. Especially thepathogenicity of L5to E. crusgalli and the pathogenicity of L4,L5,L8to Monochoria vaginalis,whose disease indexes achieve more than60%, therefore they demonstrate the higher herbicidal activity.Meanwhile, these six strains were safe or weak pathogenic infection to main crops, and showed thevaluable to study and develop into potential biological herbicides.The biological characteristics research of potential strains. We explore the effect of temperature,light, pH, oxygen, carbon and nitrogen source to colony growth of six strains, and ensure the optimum culture conditions for mycelial growth, which lay a solid foundation for solid, liquid fermentationmedium amplification culture research of six potential strains. According to HPLC analysis to culturefiltrate toxin during different culture time,we can make sure that ophiobolin A is the main component ofculture filtrate,and we determine the optimum culture time of toxin production in PDB medium at thesame time, which lay the foundation for the herbicide mechanism and toxin production of liquidfermentation research.