Appressorium: The Breakthrough in Dikarya

Abstract

Phytopathogenic and mycorrhizal fungi often penetrate living hosts by using appressoria and related structures. The differentiation of similar structures in saprotrophic fungi to penetrate dead plant biomass has seldom been investigated and has been reported only in the model fungus Podospora anserina. Here, we report on the ability of many saprotrophs from a large range of taxa to produce appressoria on cellophane. Most Ascomycota and Basidiomycota were able to form appressoria. In contrast, none of the three investigated Mucoromycotina was able to differentiate such structures. The ability of filamentous fungi to differentiate appressoria no longer belongs solely to pathogenic or mutualistic fungi, and this raises the question of the evolutionary origin of the appressorium in Eumycetes.

Keywords: Eumycetes; appressorium; biomass degradation; cellophane; infection cushion; penetration; saprotrophic fungi.

Figure 1

Fusarium graminearum and Rhizoctonia solani penetrate cellophane through appressoria. For both F. graminearum (upper panel) and R. solani (bottom panel), the main steps of appressorium-based cellophane penetration have been represented. From left to right, the four first columns represent successive z-planes of the same field of view. The distances from the initial z-plane are indicated. Arrows indicate contact points and the penetration peg they emit to penetrate cellophane. (*) Haustorium-like growth. Last column (right) shows in-depth intra-cellophane mycelial development. Scale bar = 10 µm. Magnification in the bottom insets is 1.7×.

Figure 2

Appressoria in Dikarya. For a selection of the representative species of the main groups of fungi endowed with an appressorium to penetrate cellophane, the main steps of appressorium-based cellophane penetration have been represented. From left to right, the first four columns represent successive z-planes of the same field of view. The distances from the initial z-plane are indicated. For Xylaria polymorpha, the mycelium was too dense to allow imaging of the first steps of appressorial development. To image the “penetration peg” and the “haustorium-like growth” stages in X. polymorpha, pictures were taken after on-top-growing mycelium removal. For each appressorium imaged, arrows indicate the contact points and the penetration peg corresponding to the haustorium-like growth (*) in the third column; for X. polymorpha, arrows point to every observable penetration peg. Last column (right) shows in-depth intra-cellophane mycelial development. For X. polymorpha, this step is already represented in the previous “haustorium-like growth” column. Scale bar = 10µm. Magnification in the upper insets is 1.7×.

Figure 3

Phylogenetic tree of the 38 Eumycetes strains analyzed. “YES”, presence of appressoria; “R”, reorientation of hyphal growth towards cellophane; “NO”, no hyphal growth reorientation and no appressorium developed. This tree was built according to the “MycoCosm Fungal Tree of Life” of the JGI [22].

Figure 4

Schematic of appressorial development. (A) The fungi, showing clear tropism towards cellophane, reorient hyphal growth towards cellophane and establish contact points at the cellophane surface. (B,C) The fungi, able to penetrate cellophane, develop appressoria in five steps. (i) They reorient hyphal growth, (ii) establish contact points, (iii) emit a penetration peg in order to breach cellophane, and (iv) starthaustorium-like growth inside the cellophane layer. The haustorium-like enlarges with time and it becomes the base for the emission of more penetration pegs. This process allows invasion (v) and eventual breaching of the cellophane layer. In some species, concurrently to their intra-cellophane development, the penetration peg enlarges and becomes hardly distinguishable from other fungal structures inside the cellophane (B).

Figure 5

Reorientation and contact points without cellophane penetration. The Pezizomycetes Pyronema omphalodes, as well as the Agaricomycetes Schizophyllum commune and Phanerochaete chrysosporium exert clear tropism towards cellophane. Both columns represent different z-planes of the same field of view. The distances from the initial z-plane are indicated. Left column: mycelium growing on top of the cellophane layer. Right column: contact points (arrows) observed following hyphal growth reorientation. Scale bar = 10µm.

Figure 6

Botrytis Cinerea and Orbiliomycetes Arthrobotrys oligospora develop infection cushions. From left to right, the four columns represent successive z-planes of the same field of view. The distances from the initial z-plane are indicated. Upper row: when grown on cellophane, B. cinerea develops penetration cushions composed of a bundle of hyphae growing together transversally to the cellophane layer. Second row: comparably to B. cinerea, in A. oligospora, bundles of hyphae (from 2 to 20) can grow transversally to the cellophane layer, creating an infection cushion (upper and bottom frames). The last row corresponds to the 3× magnification of the infection cushion in the upper frame. Penetration pegs (arrow heads) at the tip of the hyphae compose the infection cushions. (*) Haustorium-like growth. In contrast to B. cinerea, hyphae of A. oligospora can be observed inside the cellophane layer. Scale bar = 10 µm.