Diversity Of Ceratocystis Fimbriata And Bioactivity Of Its Volatile Organic Compounds

Ceratocystis fimbriata is a soil-borne plant pathogen with worldwide distribution. Its quick proliferation has threatened various crops and economically important trees in South China. As a plant pathogen, its host specialization, genetic variability, production of volatile organic compounds (VOCs), and diseases epidemiology have been extensively studied. However, there are few reports on the bioactivity of the VOCs, and genetic diversity of C. fimbriata isolates in China. To further understand this fungus, genetic diversity of C. fimbriata isolates, diversity and bioactivity of VOCs produced by this organism were examined with biological assay, molecular techniques, instrumental analysis and efficacy evaluation. This is the first systemic report in the regard in China. The major results are as follows.1) Diversity of biological characteristics of C. fimbriata. Among the examined22C. fimbriata isolates, obtained from9hosts in9countries, there is significant diversity in colony morphology and mycelial growth rate. Isolates Cal, El, Ml, Pll and Po1grew faster than the other17isolates. Compared with nutrition, pH value and illumination conditions, all of the isolates were much more sensitive to temperature.2) Genetic diversity of C. fimbriata. Out of the110isolates isolated from5hosts in10provinces in China,91were real C. fimbriata belong to the Latin American Clade. Twelve isolates and isolates from other countries isolated from sweet potato shared the identical genetic background. Seventy-nine isolates from pomegranate, taro, loquat and eucalyptus in China had the identical genetic background with the isolates from eucalyptus in Brazil. Twenty-one new genotypes were found based on14microsatellite loci among these91isolates from China. Ceratocystis funbriata populations are the result of an introduction, but not native to China. The other19isolates were in the Austral-Asian Clade. Seven new genotypes were found based on11microsatellite loci among these19isolates from China. According to the phylogenetic analyses; microsatellite diversity, interfertility tests and morphology,6isolates from eucalyptus in Kunming City of Yunnan Province were identified as a new species, C. oceanica Q. Li, D. McNew&TC Harrim, sp. nov., while the isolates from eucalyptus in Zhanjiang City of Guangdong Province and Lingao City of Hainan Province may be hybrids species.3) Diversity of VOCs from C. fumbriata. The VOCs produced by C. fimbriata were qualitatively and quantitatively determined by headspace gas chromatography-mass spectrometer. Total volatile compounds, kinds of compounds and major components of each isolates in variety of culture conditions showed the diversity. The total volatile compounds and the number of components produced at25℃were higher than that at4℃with different major components. In the same culture condition, the total VOCs from all of the isolates increased after the inoculation, and decreased11days post inoculation. The VOCs were still detected50days post inoculation. Sixteen new components were firstly reported, such as oxalic acid, dimethyl ether, carbon disulfide, dimethyl sulfide, furan,2-methylfuran,2-methyl-butanal, trichloromethane,2-fluoropropene, propanoic acid propyl ester, dimethyl disulfide, 2-methylpropyl ester propanoic acid, pentyl ester propanoic acid,2-pentyl-furan, styrene and acetoin. The numbers of components from the isolates of different genetic background were different. The total VOCs produced by the isolates of lineage one of Latin American Clade were significantly higher than that of other lineage or clade.4) Bioactivity of the VOCs produced by C.fimbriata. The VOCs from C. fimbriata significantly inhibited plant pathogenic fungi and oomycetes, including Botrytis cinerea, Monilinia fructicola, Fusarium verticillioides, F. oxysporum, Valsa mali, Curvularia sp., Penicillium italicum, P. digitatum, Rhizoctonia solani), bacteria (Pectobacterium carotovorum, Acidovorax citrulli, Clavibacter michiganensis subsp. michiganensis, Bacillus subtilis, Rhizobium japonicum, Phytophthora sojae, and P. capsici. The faster growing strain Ca1, E1, M1, Pl1and Po1had stronger bioactivity. The VOC-treated test fungi and oomycetes grew slowly with significantly decreased spore production, spore germination and pigment production. After exposure to C. fimbriata volatiles, none of the test bacteria were able to form regularly shaped colonies that appeared watery, not round, and discrete. Artificially inoculated peach brown rot and citrus green mold were reduced by91.5%and97.4%, respectively. VOC-treated pathogens were observed by scanning electronic microscope. The severely degenerating hyphae were collapsed, curling, distorted or wrinkled, with swollen apexes with extensive branches, and the conidia were enlarged and distorted. All of there misshapen structures were irreversible. The VOCs from strain El promoted the seed germination and seedling growth of Arabidopsis thaliana, including germination energy, germination rate, seedling length, root length, fresh weight and total chlorophyll concentration, which are safe and nontoxic to plants. Since strain E1isolated from eucaluptus in Brazil had the same genetic background with the isolates from pomegranate, taro, loquat and eucalyptus in China, it should avoid to contact the hosts and soil in the application of diseases control by means of biofumigation of the VOCs from C. fimbriata.