Preliminary Study On Biological Activities And Isolation Of Endophytes From Arundina Graminifolia

Arundina graminifolia(D.Don)Hochr.is one species of plants belonging to Orchidaceae,which can be used as medicine.A.graminifolia is mainly distributed in the tropical and subtropical regions and used to treat jaundice,bruises and snake bites and so on.Phytochemical studies have shown that A.graminifolia biologically synthesizes bibenzyl,phenanthrene and other compounds with good medicinal activities.A.graminifolia is a rare medicinal plant in Dai and its distribution is also limited,but currently the artificial cultivation technology of A.graminifolia has not yet been designed.So it is an efficient and convenient method to develop new drugs from endophyte instead of plant itself.A total of 30 cultivated endophytes including 18 endophytic fungi and 12 endophytic bacteria were isolated from these plants.It indicated that more endophytic fungi were isolated from the leaves of A.graminifolia in Baise city,Guangxi province based on comparative analysis.More endophytes were isolated from leaves of A.graminifolia than stems and roots.The experimental results showed that the generation of endophytes in A.graminifolia may be related to the growth environment(including geographical location,climatic conditions,etc.)of the host plants.Ten strains of plant pathogenic fungi(Mycosphaerella arachidicola,Penicillium italicum,Alternaria longipes,Fusarium graminearum,Penicillium digitatum,Botrytis cinerea,Colletotrichum higginsianum,Phomopsis punctatus,Botryosphaeria dothidea,Geotrichum citriaurantii)and 5 strains of pathogenic bacteria(Escherichia coli,Salmonella typhimurium,Pseudomonas aeruginosa,Staphylococcus aureus,Bacillus subtilis)were selected to test the antibacterial activity of 30 endophytic strains isolated in the above experiment.The findings showed that the fermentation broth of the endophytic fungus YNLF-1 had a strong broad-spectrum antibacterial activity with a 50%inhibition effect against 10 plant pathogenic fungi,especially for Colletotrichum higginsianum and Botryosphaeria dothidea,with an inhibition rate of 77.1%and 73.9%respectively.The strain YNLF-1 was identified as a new species of Phanerochaete sp.using ITS sequence coupled with its morphological traits.It indicated that endophytic fungus of A.graminifolia had a good broad-spectrum antagonism against phytopathogenic fungi,and plant endophytes were the main ways to find new species as well.However,the endophytic fungi of A.graminifolia had no significant effects on pathogenic bacteria.Only an endophytic bacterium,Burkholderia gladioli FCSB-1,showed a certain inhibitory effect on E.coli and B.subtilis.The antioxidant activity of 5 endophytic fungi(including YNLF-1,BSRF-1,BSRF-2,BSRF-3 and BSSF-2)from A.graminifolia was tested here.The crude extract was used to determine by DPPH free radical scavenging activity,Fe3+ determination and H2O2 ability to obtain its antioxidant activity.The results showed that the crude extracts of 5 endophytic fungi fermentation broth expressed different degrees of antioxidant activity,with an order of BSSF-2>YNLF-1>BSRF-1>BSRF-3>BSRF-2.The dominant strain BSSF-2 of antioxidant was identified as Colletotrichum karstii.The 96-well v-plate clotting evaluation method was used to study the activity of lectins from fermentation broths of 18 endophytic fungi,and the predominant lectin-producing strain YNLF-3 was obtained here.The single factor test and response surface methods were used to optimize the fermentation medium composition to gain lectin of high yield.The results showed that the optimal culture medium included 29.98 g/L mannose,13.05 g/L yeast,0.12 mol/L KCl,0.04 g/L VB1 and pH 6.8.The total activity of lectin reached 4.41×105 U and 52-fold greater,with the optimized media than with the nonoptimized basal media.Moreover,it was found that the optimized medium of strain YNLF-3 can significantly increase the antibacterial activity.