Preliminary Study On Endophytic Fungi Of Aquilaria Sinensis

Endophytic fungi is a large category fungi which has not been thoroughly studied.In recent years,many studies have shown that endophytic fungi distribute in subcutaneous tissue of root,stem,leaf and other organs of almost all plants.It generally refers to the fungus which live in healthy plants and do not cause any symptoms of the diseases in plants.Endophytic fungi dwell stably in the plant's micro-ecological system in long-term,and establish half symbiotic or symbiotic relationship between the host.Researches have shown that the majority of isolated endophytic fungus can produce same or similar active ingredients which exist in plants host.Aquilaria sinensis tree can be infected by fungi after artificially slashed,moth-eaten or totted,then the material stored in the parenchyma cells of the xylem of the trunk take a series of chemical reactions and finally generate resin,which is known as agalwood.In order to find out the timber containing resin,people have to cut down many Aquilaria sinensis trees,so that the individual with agalwood resin can be identified,which cause a large number of Aquilaria sinensis trees felled and seriously affected the survival of ethnic groups of Aquilaria sinensis.Meanwhile the high price of agalwood has led to illegal log and trade agalwood in a number of countries of origin.So the separating endophytic fungi from Chen Xiang block and study of artificial edgeworthia is of great significance.In this paper,during optimization of conditions of endophytic fungi isolation,9 agalwood endophytic fungi have been separated and purified from Chen Xiang block.Then through benzylacetone tolerance,artificial edgeworthia materials analysis et al,4 agalwood endophytic fungi with the promotion of the role of artificial edgeworthia have been screened and the species identified by molecular biology techniques.At the same time the artificial edgeworthia conditions and Hypocrea jecorina fermentation experiments have been carried out.The results can be summarized as follows:1,The experiments of optimization of conditions of agalwood endophytic fungi separation adopt NaClO(5%available chlorine)as a disinfectant on the surface and sterilize for 4 min,and choose 26℃as the isolation and culture temperature.At last 9 agalwood endophytic fungi have been separated and purified.According to the preliminary identification,they are belong to 2 Subdivisions,5 Classes,5 Orders,5 Families,5 geni.2,Through researches of extraction methods of agalwood volatile oil,we select Soxhlet anhydrous diethyl ether reflux and GC-MS as the method to analysis components of agalwood volatile oil.3,In the agalwood endophytic fungi screening experiments,Aq-2,Aq-3,Aq-5b,Aq-7 et al,4 endophytic fungi have been screened after testing the benzylacetone tolerance,the width of artificial edgeworthia samples which change color and the burning smell.4,The results of the species identification of agalwood endophytic fungi by molecular biology techniques is:Aq-2(Botryosphaeria rhodina),Aq-3(Hypocrea jecorina),Aq-5b(Trichoderma reesei),Aq-7(Trichoderma koningii).5,The studies of artificial edgeworthia conditions of 4 Aquilaria endophytic fungi:Botryosphaeria rhodina,Hypocrea jecorina,Trichoderma reesei,Trichoderma koningii,have carried out following the programs of sealing methods,srilling locations,fungi injection modes,edgeworthia time.The best conditions are:tampon and muse as sealing methods;the area 50～100cm from the boot of the trunk;a single fungus of Botryosphaeria rhodina and Hypocrea jecorina,Trichoderma reesei,Trichoderma koningii 3 endophytic fungi of Trichoderma mingling together,Trichoderma reesei or Trichoderma koningii 2 endophytic fungi mingling together;extending edgeworthia time,can be a good quality of agalwood.6,GC-MS was used to analyze the contents of broth and mycelium of Hypocrea jecorina,the results are as follows:the benzyl group and furyl group have not been found in the broth,but the benzopyrans which belong to the complex components of agalwood has been found in the mycelium of Hypocrea jecorina,but benzopyrans is very little and single,so we can conclude that the feasibility of using microbial fermentation to yield agalwood components is limited.