Deletion of the Aspergillus flavus orthologue of A. nidulans fluG reduces conidiation and promotes production of sclerotia but does not abolish aflatoxin biosynthesis

Abstract

The fluG gene is a member of a family of genes required for conidiation and sterigmatocystin production in Aspergillus nidulans. We examined the role of the Aspergillus flavus fluG orthologue in asexual development and aflatoxin biosynthesis. Deletion of fluG in A. flavus yielded strains with an approximately 3-fold reduction in conidiation but a 30-fold increase in sclerotial formation when grown on potato dextrose agar in the dark. The concurrent developmental changes suggest that A. flavus FluG exerts opposite effects on a mutual signaling pathway for both processes. The altered conidial development was in part attributable to delayed expression of brlA, a gene controlling conidiophore formation. Unlike the loss of sterigmatocystin production by A. nidulans fluG deletion strains, aflatoxin biosynthesis was not affected by the fluG deletion in A. flavus. In A. nidulans, FluG was recently found to be involved in the formation of dehydroaustinol, a component of a diffusible signal of conidiation. Coculturing experiments did not show a similar diffusible meroterpenoid secondary metabolite produced by A. flavus. These results suggest that the function of fluG and the signaling pathways related to conidiation are different in the two related aspergilli.

Fig 1

Morphology and production of conidia and sclerotia by A. flavus ΔfluG strains on PDA. (A) Cultures were grown at 30°C under white light and in the dark for the specified numbers of days. Washed, conidia of the 7-day-old cultures were washed off to expose sclerotia (black aggregates). (B) Quantification of conidia and sclerotia. Gray bars, wild-type CA14; crosshatched bars, ΔfluG strains; open bars, fluG complemented strains. Conidiation and sclerotial production under light by the fluG complemented strains were not determined. The complemented strains shown are those putatively having the native fluG promoter (see Fig. S1 in the supplemental material). The numbers of conidia and sclerotia are the total count per plate. The error bars indicate standard deviations.

Fig 2

Morphologies of A. flavus ΔfluG and fluG complemented strains on Wickerham medium. (Top) Cultures after 7 days of growth at 30°C in the dark. (Bottom) Conidia of the colonies were washed off to expose sclerotia (black aggregates). The control strain is A. flavus KuPG no. 1.

Fig 3

Time course semiquantitative TLC analyses of aflatoxin B₁ production by A. flavus ΔfluG and fluG complemented strains. Wt, wild-type A. flavus CA14; Pt, control A. flavus KuPG no. 1; D, ΔfluG strains; C, fluG complemented strains with fluG under the gpdA (C1) and the fluG (C2) promoter, respectively. The numbers indicate independent isolates. Sd, aflatoxin B₁ standard.

Fig 4

Morphologies of A. flavus ΔlaeA and ΔlaeA ΔfluG strains on Wickerham medium grown at 30°C in the dark for the specified numbers of days.

Fig 5

Coculture of fluG-positive and ΔfluG strains. Cultures were grown on PDA at 30°C for 5 days in the dark. (A) Morphology before (left) and after (right) conidia were washed off the PDA plates. (B) Close-up of the boundary of the wild-type and ΔfluG strains.