Molecular identification of endophytic fungi from Aquilaria sinensis and artificial agarwood induced by pinholes-infusion technique

Abstract

Agarwood, the resinous portions of Aquilaria plants, have been used as medicines and incenses. Aquilaria sinensis is the major producer of agarwood in China. Agarwood are generally viewed as pathological products formed as defense symptom against fungal infection. In this study, microbial communities inhabiting the leaves of non-resinous and agarwood-producing wounded A. sinensis tree were investigated by cultivation-independent approaches, such as PCR, restriction fragment length polymorphism (RFLP) analysis and sequencing of rDNA internal transcribed spacer (ITS) library. Molecular phylogenetic analysis demonstrated that Botryosphaeria, Colletotrichum gloeosporioides, Phomopsis and Cylindrocladium species are members of the agarwood-producing wounded tree, while Phoma, Mycosphaerella, Sagenomella, Alternaria and Ramichloridium species is able to colonize the non-resinous tree internally. C. gloeosporioides was the only fungus shared by the two rDNA ITS libraries. C. gloeosporoides, Botryosphaeria , and Cylindrocladium were considered to be related to agarwood formation. A pinholes-infusion method to induce the generation of agarwood by chemically stimulated and/or inoculate combined method was established. One to two years after the artificial inoculation, resinous wood were collected and the inoculating effects were detected by ethanol extraction content, thin layer chromatography (TLC) and gas chromatography-mass spectrometry (GCMS) techniques. The results reveal that chemically stimulated with formic acid and infected by Botryosphaeria dothidea produced high yield and high quality artificial agarwood in a relatively short time. Keywords : Agarwood, endophytic fungi, Aquilaria sinensis , molecular identification, artificial induce of agarwood African Journal of Biotechnology Vol. 12(21), pp. 3115-3131