Molecular phylogenetic identification of endophytic fungi isolated from resinous and healthy wood of Aquilaria malaccensis, a red listed and highly exploited medicinal tree

Aquilaria malaccensis is a fast-growing, tropical tree belonging to the family Thymelaeaceae and is locally known as Agar. Agarwood formation takes place in the stem or main branches of the tree where an injury has occurred. It is believed that the tree is first attacked by a pathogenic fungus, which causes it to weaken. Most fungal diversity studies have previously been based on morphological examination and cultivation methods. In this study, we used both culture-dependent and culture-independent approaches (metagenomic) to study the endophytic fungi on wood chips of A. malaccensis. The culture-based approach revealed Alternaria, Cladosporium, Curvularia, Fusarium, Phaeoacremonium and Trichoderma as members of the agarwood community. Also analysis of ITS sequencing of these culture isolates provided further verification of the identity of the cultured groups. Analysis of community DNA (metagenome) extracted from both infected and healthy wood samples revealed that the majority of fungi present had highest sequence similarity to members of Dothideomycetes, followed by Sordariomycetes and Saccharomycetes. Thus, morphological and genetic characteristics showed that most isolates from agarwood belong to phylum Ascomycota. A neighbour-joining tree showed the relationships between the isolates sequence data and the closest identified relatives from GenBank.