Screening Of Immunoenhancement Mangrove Fungus And Identification Of Strain PH0016

ObjectiveTo isolate and screen fungi possessed immunoenhancement from mangrove soil collected from the sea area in Hainan.Evaluated its activities with immunological assays, and identified strains by morphological and molecular biological methods, and explore the application value of mangrove fungi in immunoloregulation.MethodsIsolation and culture fungi from mangrove soil samples on the Martin medium, screen fungi possessed immunoenhancement by using MTT method to measure the fermental supernatant. The extracts were separated from positive strains with Sephadex G-25. Animal experiments were used to evaluate the immunologic functions of fungi possessed immunoenhancement extracts. KM mice allocated randomly into the intervention group and control group, fermental supernatant extract of fungi and normal sodium were given by intragastric administration respectively for 10 days before test, test including detection methods like immune organ index, serum hemolysin HC50, hemolytic pique-forming, delayed-type hypersensitivity (DTH), macrophage phagocytosis. Identified and explore the taxonomic status of strains by means of observation on morphology and cultural characteristics,combined with rDNA ITS sequences of squence and phylogenetic analysis.ResultsⅠ.The results of isolate and screen fungi possessed immunoenhancement from mangrove soil.1.Isolation of strains:325 strains were attained from mangrove soil samples. 2.Screen fungi possessed immunoenhancement:By using MTT method to measure the fermental supernatant,12 strains were screened.The strain PH0016 can significantly stimulate lymphocyte proliferation(P=0.000).Ⅱ. Evaluation of the immunologic functions.The extracts were separated from strain PH0016 with Sephadex G-25.By using methods including immune organ index, serum hemolysin HC50, hemolytic pique-forming, delayed-type hypersensitivity (DTH) and macrophage phagocytosis, the effects of extracts from strain PH0016 were evaluated in vivo. The results showed that the extracts from strain PH0016 had no significant effects on weight (F=1.037,P=0.324) and spleen index(t=1.878,P=0.078). Analysis of Independent-Samples t Test showed that the edema of foot-plate increased significant in intervention group (t=3.858,P=0.001);the level of HC50 has significant difference between intervention group and control group (t=5.632,P=0.000);the level of PFC in intervention group significant increased than that in control group(t=2.855,P=0.008);and the phagocytic index in intervention group increased significant(t=3.268,P=0.003).III. Identification of strain PH0016The growth of strain PH0016 on PDA medium,Czapek's medium and Sobaurand sagar culture medium is generic, forming colonies about 15mm-18mm across during 7 days of culture at 28℃。The color of the colonies was lavender and white in the backside. The strain was identified as Paecilomyces on the basis of detailed morpholodical observations and analysis of the rDNA-ITS sequence.Conclusionsl.The fungi resources in mangrove are abundant and highly valuable in its development.We isolated 325 strains from mangrove soil samples and attained 12 strains possessed immunoenhancement.2. Evaluation of the immunologic functions showed that the extract of HP0016 promoted the activities of T,B lymphocyte and macrophage.3. The strain PH0016 was identified as Paecilomyces sp.4.The Paecilomyces has many species.There is no report of Paecilomyces isolated from mangrove.