Revision of Archaeosporomycetes with two old and two new fungal orders: Archaeosporales, Geosiphonales, Polonosporales, and Ambisporales

Currently, the fungal class Archaeosporomycetes consists of one order, Archaeosporales with four families: Archaeosporaceae, Ambisporaceae, Geosiphonaceae, and Polonosporaceae. In the present study, the objective was to re-analyze the phylogeny and morphology of the Archaeosporomycetes from order to genus level. The different ecological strategies and, consequently, distinct evolutionary patterns of these taxa, as well as their morphological characters and other data updated here, suggest the need to divide Archaeosporales into four orders: (i) the type order Archaeosporales, (ii) Ambisporales ord. nov., both with four genera, (iii) Geosiphonales and (iv) Polonosporales ord. nov., both with single families and genera. Remarkably, the order Geosiphonales was described in the past, but was not considered in the Archaeosporomycetes until now. Phylogenetically, the four main clades (orders here proposed) of Archaeosporomycetes are well supported, with bootstrap values higher than 95% in all analyses, except Ambisporales/Ambisporaceae for RAxML-NG FBP analysis in the SSU tree (75%). Ecologically, this class includes three orders of arbuscular mycorrhizal fungi (AMF) forming symbiotic associations with plants, while Geosiphonales form an endocytobiosis with the cyanobacterium Nostoc. Morphologically, there are at least two AMF orders with spore bimorphism, which has not (yet) been described for Polonosporales. The only known species of Polonosporales, Polonospora polonica, forms spores directly on the neck of sporiferous saccules and the spores can morphologically be differentiated from all other taxa in Archaeosporomycetes by the formation of three permanent, rather thick spore walls, of which two form de novo during spore formation. The outer spore wall of Archaeosporales and Ambisporales are semi-permanent, evanescent or even short-lived, or show multiple fissures during aging, when it is more resistant. Ambisporales can easily be differentiated from Archaeosporales for instance by larger spores of the acaulosporoid morph and thicker spore walls. Our phylogenetic analyses suggested that Archaeosporales can be divided into two families: Antiquisporaceae that was described to form intraradical hyphae, vesicles and spores, staining darkly in Trypan blue, and Archaeosporaceae whose hyphae generally do not or only faintly stain in this reagent, and vesicles and intraradical spores have been rarely, if ever reported.