During a screening of putative diterpene synthase genes found in public databases using the Aspergillus oryzae expression system, it was found that a single transformant with the ACLA_76850 gene from A. clavatus produced a sesterterpene alcohol, ophiobolin F, and three minor sesterterpene hydrocarbons. The sesterterpene synthase has two catalytically independent domains (prenyltransferase/terpene cyclase) which are homologous to those of diterpene synthase, fusicoccadiene synthase. Coevolution of both domains and reaction mechanisms of these terpene synthases are discussed.

Heterologous expression of four candidate genes found in ophiobolin gene clusters from three fungal strains was employed to elucidate the late-stage biosynthetic pathway of phytotoxin ophiobolin. Expression of oblBAc (cytochrome P450) from the cryptic gene cluster gave unexpected products, and that of oblBBm/oblBEv from the gene cluster of ophiobolin producers, with oblDBm as the transporter, yielded intermediate ophiobolin C through an unusual four-step oxidation process. The observation made in this study may provide a useful guideline for the elucidation of genuine biosynthetic pathways of natural products.

Background: Trichosporon asahii is the major causative agent of disseminated and deep-seated trichosporonosis. It is capable of forming biofilms on surfaces, leading to medical device-related infection. Trichosporon asahii may be present as yeast form, hyphae and/or arthroconidia; however, the relationship between its biofilm-forming ability and its morphological transition is unclear. Objectives: We investigated whether the T. asahii morphological transition contributes to its biofilm formation. We also determined the conditions required to induce each of the morphologies. Methods: Three high-and three low-biofilm-producing strains (HBS and LBS, respectively) were selected using a biofilm formation assay, and the cell surface hydrophobicity of these six strains was measured. For each strain, the morphology was observed and the number of each morphological form (yeast form, hypha and arthroconidium) was counted to calculate the ratio. Finally, the ability of cells each morphological type to adhere to the polystyrene substrate was evaluated. Results: The HBS exhibited abundant arthroconidia and hyphae; in contrast, the LBS produced mainly hyphae with few or no arthroconidia. The production of hyphae was increased by nitrogen-containing medium, and the production of arthroconidia was increased by nitrogen-deficient medium. Cells incubated under nitrogen-deficient conditions showed higher adherence to a polystyrene surface than those incubated in the presence of nitrogen. Conclusion: Arthroconidia of T. asahii play a key role in biofilm formation by promoting cellular adhesion.

Objective: Trichosporon asahii is the major causative fungus of disseminated or deep-seated trichosporonosis and forms a biofilm on medical devices. Biofilm formation leads to antifungal drug resistance, so biofilm-related infections are relatively difficult to treat and infected devices often require surgical removal. Therefore, prevention of biofilm formation is important in clinical settings. In this study, to identify metal cations that affect biofilm formation, we evaluated the effects of cation chelators on biofilm formation in T. asahii. Results: We evaluated the effect of cation chelators on biofilm formation, since microorganisms must assimilate essential nutrients from their hosts to form and maintain biofilms. The inhibition by N,N,N′,N′-tetrakis(2-pyridylmethyl) ethylenediamine (TPEN) was greater than those by other cation chelators, such as deferoxamine, triethylenetetramine, and ethylenediaminetetraacetic acid. The inhibitory effect of TPEN was suppressed by the addition of zinc. TPEN also inhibited T. asahii hyphal formation, which is related to biofilm formation, and the inhibition was suppressed by the addition of zinc. These results suggest that zinc is essential for biofilm formation and hyphal formation. Thus, zinc chelators have the potential to be developed into a new treatment for biofilm-related infection caused by T. asahii.