Isolation And Identification Of An Endophytic Bacterium From Cymbopogon Citrates (DC.) Stapf And Its Optimal Flavor-producing Conditions

Cymbopogon citrates(DC.) Stapf is a perennial herbaceous plant, which belongs to Cymbopogon of ramineae.As an important tropical spice crop and medicinal plant, the volatile products of Cymbopogon citrates(DC.) Stapf are mainly terpene compounds which are kinds of spice source. However, the planting scale was restricted by the farming area, the climate of environment and the disease. Moreover, it is hard to meet the market demand by using the conventional extraction method of terpene compound content because of the low productivity. Therefore, the method has gradually become the focus especially in the microbial synthesis. Endophytes have the potential for secondary metabolite production which similar to the host, when live in the medicinal plants that synthesize a large number of natural metabolites. Endophytes isolated from medicinal plants have the higher possibility than the other and have specific physiological and biochemical functio under the unique living environment. Development and utilization of plant endophytes resource will be a new solution for the synthesis of natural medicine resources.Through using four kinds of separation medium(WA, TWYE, MS, TSA) after plant surface disinfection, a number of endophytic bacterias were isolated from mature Cymbopogon citrates(DC.) Stapf.Then 36 endophytic bacterias which can produce fragrance were screened according to sensory evaluation. One endophytic bacterium which have obvious and pleasant fragrance was chosen and named CcSh-1. Furthermore, according to the morphological, physiological and biochemical and 16 s rRNA sequence analysis, CcSh-1 was identified as Pantoea ananatis.Using headspace gas chromatography mass spectrometry, we identified volatile components from Cymbopogon citrates(DC.) Stapf and Pantoea ananatis CcSh-1. Volatile components from Cymbopogon citrates(DC.) Stapf were α-pinene, 1s-α-pinene, 3-carene, mugwort-ether, 2, 2-dimethyl-3, 4-symplectic diene, neral and geranial and the volatile components in CcSh-1 were camphene, α-pinene, sandalwood triene, neral, enol geranial and sandalwood oil. The citral(neral and geranial) content was the highest, which reached 2.1 μg/m L. They had the same volatile components which were: α-pinene, neral and geranial. And citral(neral and geranial) content was relatively higher.We preliminarily explored the best condition of CcSh-1 for flavor producing. The factors involved in the flavor productivity and the growth status of CcSh-1 were investigated, and the optimal liquid culture conditions for CsSh-1 were confirmed: beef extract as carbon source, peptone as nitrogen source, 30℃ and pH 7. In addition, we discovered that adding 1%(w/v) abrasive of host plant as substrate could promote flavor production of CcSh-1(particularly citral content), and flavor species had no change.In conclusion, we successfully screened an endophytic bacteria strain CcSh-1 that produces obvious fragrance. After identification analysis, we found that the volatile constituents predominantly include citral of terpenoids. We preliminarily determine the optimal flavor production conditions of CcSh-1; In addition, we researched the flavor production of CcSh-1 with the affection by Cymbopogon citrates(DC.) Stapf and this influence reflected the interaction between endophytes and their host.