Study On The Characteristics And Functions Of Melanin In Exserohilum Turcicum

In order to examine the role of melanin in Exserohilum turcicum, physical and chemical characteristics and functions of melanin were investigated in the paper. The melanin biosynthetic pathway in E. turcicum was identified and a 1,3,8-trihydroxynaph-thalene reductase (3HNR) gene, associated with the melanin of biosynthetic pathway in E. turcicum, was characterized. Some mutant isolates, which are absent of melanin, were obtained by combined mutation. The relationship between melanin and fungus pathogenesis was eluciated by comparation of pathogenesis between wild isolates and mutant ones.The characteristics of melanin, extracted and purified from cell wall and zymolytic filtrate of E. turcicum, were compared with standard melanin (Sigma), by diagnostic tests, UV-Vis spectrum and infrared spectra. The results indicated that the physical and chemical characteristics of E. turcicum melanin were similar to the standard melanin, but some differences remained in infrared spectra. The fungal melanin did not display the characteristic bands of GHB melanin in 1100 cm~-1 and DOPA melanin inl600 cm~-1. The results suggested that the melanin extracted from E. turcicum was neither a kind of DOPA nor a GHB melanin.Mutant isolates, which were absent of melanin (wild type isolate 01-23 ), were obtained by combined mutation. The tests on the mycelial growth, the number of conidiophores, the activity of HT-toxin and pathogenesis were conducted, comparing the mutant isolates with wild type ones. The results indicated that the mycelial growth, the number of conidiophores and pathogenesis reduced greatly in mutant isolates, but the activity of HT-toxin didn't change obviously. Mutant isolates resumed a little pathogenesis after growing on PDA containing melanin. These findings had shown that the melanin of E. turcicum played very important roles in pathogenesis.A pair of primers were designed according to the conserved regions of 3HNR genes. A fragment of 864bp was acquired by PCR-amplified of isolates 01-23 (race 1) and 01-24 (race 0) of E. turcicum genomic DNA. The 3HNR gene of E. turcicum possesses 89-100% percentage of identity with other known 3HNRs, especially it demonstrated completehomology in the two races of E. turcicum. The results in this study provided powerful evidences that the pathway of melanin of E. turcicum might belong to DHN.Tricyclazole, a specific inhibitor of DHN melanin synthesis, can prevent the accumulation of the melanin in E. turcicum. The colour of the wild type colonies cultivating in PDA medium containing tricyclazole, turned from red to brown. In low concentration (5ug/mL), tricyclazole had neither a strong inhibitory effect on hypha growth nor production of conidiophores, but it reduced greatly the infection ratio and lesion area. Therefore, the results suggested that the melanin extracted from E. turcicum was a kind of DHN melanin and the absence of melanin in E. turcicum decreased the fungal pathogenesis.In order to find influence factors on melanin biosynthetic pathways, this paper designed different conditions to cultivate E. turcicum wild type, which established foundation for making new fungicides.