Studies On Relationships Between Swainsonine And Fungal Endophytes From Qxytropis Glabra In Hangjin Banner

Oxytropis glabra is a kind of plant that widely distributes ingrassland and desert regions of Inner Mongolia. Many plants of thisspecies contain alkaloids swainsonine which causes locoism by inhibitingthe activity of mannosidase in cells. Livestock is poisoned and thensuffered locoism when feeded certain amount of the plant. Thereforesignificant losses for animal husbandry on grassland were caused. It iswith great theoretical and applying values to study the poisoningmechanism of swainsonine in O.glabra. In addition, O.glabra hasimportant medicinal and ecological values. It would be an excellentforage species if swainsonine is removed in the plant. The swainsonine isalso a kind of potential anticancer medicament as well.In this research, fungal endophytes were cultured with the stemexplant of O.glabra, and the morphologic characters of the fungi wereinvestigated. The5.8SrDNA/ITS sequences of the fungal endophyteswere amplified and then sequenced. Then the sequences were submittedto GenBank and compared with the published sequences of fungalendophytes from O.glabra and other locoweeds by using blastn program.The taxonomy of the fungal endophytes was analyzed according to theresults of molecular biological and microbiological studies. The fungalspecific sequences of5.8SrDNA/ITS in individual plants from which thefungal endophytes were not screened were also amplified from palnt totalDNA and then sequenced. The swainsonine was extracted both fromindividual plant of O.glabra and their fungal endophytes. Theswainsonine contents in samples were determined by gas chromatography.Samples with two fungl bands in plant total DNA were cloned to the Tvectors and then sequenced in order to explore the relationship between swainsonine from O.glabra and the fungal endophytes in O.glabra. Theresults showed as follows:1. Seven fungal endophytes from O.glabra were isolated. Six ofthem were the same to the ten fungal endophytes from O.glabra and theEmbellisia sp. L12in published papers in morphologic characteristics ofthe colonies. The sequences of the5.8SrDNA/ITS of the fungalendophytes in O.glabra were the same to that of published fungalendophytes in O.glabra. Compared to the sequence of Embellisia sp.L12,there was only one variation site in the5.8SrDNA region. The fungalendophytes from O.glabra were therefore considered belonging toEmbellisia. Fungous HJ1.9was slightly different from the fungalendophytes mentioned above in morphologic characters. However its5.8SrDNA/ITS sequence were98.5%of identity compared to thesequences mentioned above from O.glabra, and98.3%of identitycompare to the sequence of Embellisia sp.L12. The Fungous HJ1.9wasalso considered to belonging to Embellisia.2. The fungal specific bands were amplified from all45plants ofO.glabra, and all of them contained swainsonine. The plant whichswainsonine was determined also had fungal endophyte in it. Thecontents of swainsonine in fungal endophytes cultured in vitro werehigher than that of swainsonine in their host plants. Swainsonine contentschanged among different populations and different single plant samples.The average content of swainsonine in HJ1, HJ2and HJ3populations was19.84ug/g,102.19ug/g and73.32ug/g sepratedly. The concentration ofswainsonine ranged from0.25μg/g to222.20μg/g,0.28μg/g to172.70μg/g and1.50μg/g to305.30μg/g in HJ1, HJ2and HJ3populations.3. The results of T-A cloning indicated that one sequence of fungalendophyte was cloned in three samples, which belonged to Embellisia.Two sequences of fungal endophytes were cloned in other ten samples, inwhich one of the two belonged to Embellisia. The other band belonged toErysiphe in two samples and Alternaria in other five samples and three samples cannot be identified.