Redefinition of Aureobasidium pullulans and its varieties

Abstract

Using media with low water activity, a large numbers of aureobasidium-like black yeasts were isolated from glacial and subglacial ice of three polythermal glaciers from the coastal Arctic environment of Kongsfjorden (Svalbard, Spitsbergen), as well as from adjacent sea water, sea ice and glacial meltwaters. To characterise the genetic variability of Aureobasidium pullulans strains originating from the Arctic and strains originating pan-globally, a multilocus molecular analysis was performed, through rDNA (internal transcribed spacers, partial 28 S rDNA), and partial introns and exons of genes encoding beta-tubulin (TUB), translation elongation factor (EF1alpha) and elongase (ELO). Two globally ubiquitous varieties were distinguished: var. pullulans, occurring particularly in slightly osmotic substrates and in the phyllosphere; and var. melanogenum, mainly isolated from watery habitats. Both varieties were commonly isolated from the sampled Arctic habitats. However, some aureobasidium-like strains from subglacial ice from three different glaciers in Kongsfjorden (Svalbard, Spitsbergen), appeared to represent a new variety of A. pullulans. A strain from dolomitic marble in Namibia was found to belong to yet another variety. No molecular support has as yet been found for the previously described var. aubasidani. A partial elongase-encoding gene was successfully used as a phylogenetic marker at the (infra-)specific level.

Keywords: Arctic; Aureobasidium; ITS; LSU; black yeasts; elongase; glacier; phylogeny; polar environment; rDNA; sea ice; seawater; taxonomy; translation elongation factor; β-tubulin.

Fig. 1.

Detailed map of the sampling area in Svalbard, with sites of retrieved aureobasidium-like isolates marked.

Fig. 2.

Consensus phylogram (50 % majority rule) of 24000 trees resulting from a Bayesian analysis of the LSU sequence alignments using MrBayes v. 3.1.2. Bayesian posterior probabilities are indicated at the nodes, branches with posterior probabilities >95 in bold. The tree was rooted to the sequence of Elsinoe veneta (DQ678060). Ex-type and ex-neotype strains are underlined; when known origin two digit country codes are listed after strain numbers. The colour marks stand for:

1. - plant associated;

2. - originating from Arctic ice;

3. - originating from hyperosmotic environment;

4. - clinical strain.

5. - water.

Fig. 3.

Consensus phylograms (50 % majority rule) resulting from a Bayesian analysis of: A. ITS rDNA; B. elongase gene; C. translation elongation factor EF-1α gene; D. β-tubulin gene. Bayesian posterior probabilities are indicated at the nodes. The trees are not rooted. Ex-type and ex-neotype strains are underlined.

Fig. 4.

Macromorphology of different Aureobasidium pullulans varieties incubated for 7 d at 25 °C in the dark on MEA (left 2 columns) and on PDA (right two columns). a–h. A. pullulans var. pullulans: a, b. CBS 584.75 (MEA); c, d. CBS 584.75 (PDA); e. CBS 109810 (MEA); f, g. CBS 701.76 (MEA, PDA); h. MZKI B-700 (PDA). i–p. A. pullulans var. melanogenum: i, j. CBS 105.22 (MEA); k, l. CBS 105.22 (PDA); m. EXF-3382 (MEA); n. CBS 621.80 (MEA); o. EXF-924 (PDA); p. CBS 100225 (PDA). q–u. A. pullulans var. subglaciale: q, r. EXF-2481 (MEA); s. EXF-2481 after 14 d incubation (MEA); t. EXF-2481 (PDA); u. EXF-2479 (PDA). v–y. A. pullulans var. namibiae. v, w. CBS 147.97 (MEA); x, y. CBS 147.97 (PDA).

Fig. 5.

Aureobasidium pullulans var. pullulans. a. Liberated conidia transforming to budding cells. b. Synchronous production of conidia on a transformed conidium – yeast cell. c. Short hypha synchronously producing conidia. d. Dark brown conidia. e–i, m. Poorly differentiated, terminal and intercalar conidiophors performing synchronous conidiation. k. Immersed hypha with lateral accumulation of conidia. l. Hypha with lateral scars – conidiogenous loci. j. Endoconidia. a–c, e–g, k–m. CBS 584.75 (ex-neotype strain); d. CBS 100524; h–i, j, m. EXF-1702B. Scale bars: a–j, l–m= 10 μm; k= 20 μm.

Fig. 6.

Aureobasidium pullulans var. melanogenum. a–c. Liberated conidia transforming to budding cells. d. Dark brown conidia. e–h. Poorly differentiated, terminal and intercalar conidiophors performing synchronous conidiation. i, l. Hypha with long lateral conidiogenous cells. j. Hypha with prolonged lateral pegs. k. Vegetative hyphae. m. Immersed hyphae with multiple lateral pegs. n. Melanized hyphae with intercalar synchronous conidiogenesis. o. Melanized hyphae / chlamydospores. a–c, e–n. CBS 105.22 (ex-type strain); d, o. EXF-926. Scale bar: as marked on k (a–o) = 10 μm.

Fig. 7.

Aureobasidium pullulans var. subglaciale. a. Conidia. b–d. Budding conidia. e. Dark brown conidia. f, g. Hyphae with multiple lateral pegs, which develop into synchronous conidiation aparatus. h, m. Hyphae with lateral pegs. i, k. Conidiophore-like structure synchronously producing conidia. l. Endoconidia. n. Hyaline vegetative hyphae. o. Melanized hyphae. a–o. EXF–2481 (ex-type strain). Scale bar: as marked on a (a–o) = 10 μm.

Fig. 8.

Aureobasidium pullulans var. namibiae. a. Conidia. b–f. Liberated conidia transforming to budding cells. g–h. Hypha with long lateral conidiogenous cells. i. Immersed hyphae with multiple lateral pegs. j. Melanized hypha surrounded by EPS. k. Hyaline and dark brown conidia. a–k. CBS 147.97 (ex-type strain). Scale bar: as marked on a (a–k) = 10 μm.