Lizel Mostert

The ascomycota tree of life: A phylum-wide phylogeny clarifies the origin and evolution of fundamental reproductive and ecological traits

We present a 6-gene, 420-species maximum-likelihood phylogeny of Ascomycota, the largest phylum of Fungi. This analysis is the most taxonomically complete to date with species sampled from all 15 currently circumscribed classes. A number of superclass-level nodes that have previously evaded resolution and were unnamed in classifications of the Fungi are resolved for the first time. Based on the 6-gene phylogeny we conducted a phylogenetic informativeness analysis of all 6 genes and a series of ancestral character state reconstructions that focused on morphology of sporocarps, ascus dehiscence, and evolution of nutritional modes and ecologies. A gene-by-gene assessment of phylogenetic informativeness yielded higher levels of informativeness for protein genes (RPB1, RPB2, and TEF1) as compared with the ribosomal genes, which have been the standard bearer in fungal systematics. Our reconstruction of sporocarp characters is consistent with 2 origins for multicellular sexual reproductive structures in Ascomycota, once in the common ancestor of Pezizomycotina and once in the common ancestor of Neolectomycetes. This first report of dual origins of ascomycete sporocarps highlights the complicated nature of assessing homology of morphological traits across Fungi. Furthermore, ancestral reconstruction supports an open sporocarp with an exposed hymenium (apothecium) as the primitive morphology for Pezizomycotina with multiple derivations of the partially (perithecia) or completely enclosed (cleistothecia) sporocarps. Ascus dehiscence is most informative at the class level within Pezizomycotina with most superclass nodes reconstructed equivocally. Character-state reconstructions support a terrestrial, saprobic ecology as ancestral. In contrast to previous studies, these analyses support multiple origins of lichenization events with the loss of lichenization as less frequent and limited to terminal, closely related species.

Species of Phaeoacremonium Associated with Infections in Humans and Environmental Reservoirs in Infected Woody Plants

ABSTRACT To date, three species of Phaeoacremonium have been associated with phaeohyphomycosis. These are P. parasiticum (formerly Phialophora parasitica), P. inflatipes, and P. rubrigenum. Numerous unknown isolates resembling Phaeoacremonium spp. have in recent years been isolated from human patients as well as from woody plants that appear to be the main environmental source of these fungi. Nine new Phaeoacremonium species, of which six were obtained as etiologic agents of human opportunistic infection, are reported. They can be identified based on their cultural and morphological characters, and the identifications are strongly supported in phylogenetic analyses of partial sequences of the actin, β-tubulin, and calmodulin genes. A multiple-entry electronic key based on morphological, cultural, and β-tubulin sequence data was developed to facilitate routine species identification. Reexamination of all isolates of P. inflatipes associated with human disease showed them to be misidentified and to belong to the new taxa described here.

Togninia (Calosphaeriales) is confirmed as teleomorph of Phaeoacremonium by means of morphology, sexual compatibility and DNA phylogeny.

Petri disease, or black goo, is a serious disease of vines in most areas where grapevines are cultivated. The predominant associated fungus is Phaeomoniella chlamydospora (Chaetothyriales). Several species of Phaeoacremonium (Pm.) also are associated, of which Pm. aleophilum is the most common. Although no teleomorph is known for Phaeoacremonium, the genus Togninia previously has been linked to phaeoacremonium-like anamorphs. To investigate the possible anamorph-teleomorph connection of Phaeoacremonium to Togninia, anamorphs of Togninia minima, T. fraxinopennsylvanica and T. novae-zealandiae morphologically were compared with Pm. aleophilum and some representative cultures were mated in all combinations. Although no interspecies mating proved fertile, matings between isolates of Pm. aleophilum produced a Togninia teleomorph within 3-4 weeks. Certain field isolates of Pm. aleophilum commonly produced the teleomorph, demonstrating that both mating types can occur in the same vine and thus also explaining the genetic diversity observed for this fungus in some vineyards. To elucidate the phylogenetic relationships among these taxa, isolates were subjected to sequence analysis of the nuclear ribosomal internal transcribed spacers (ITS1, ITS2) and the 5.8S rRNA gene, as well as portions of the translation elongation factor 1 alpha (EF-1α) gene. The generic placement of teleomorphs within Togninia (Calosphaeriales) further was confirmed via phylogenetic analyses of 18S small subunit (SSU) DNA. From these sequences, morphological and mating data, we conclude that T. minima is the teleomorph of Pm. aleophilum, and that it has a biallelic heterothallic mating system. An epitype and mating type tester strains also are designated for T. minima.

Fungal trunk pathogens associated with wood decay of almond trees on Mallorca (Spain)

Severe decline of almond trees has recently been observed in several orchards on the island of Mallorca (Balearic Islands, western Mediterranean Sea). However, the identity of the causal agents has not yet been investigated. Between August 2008 and June 2010, wood samples from branches of almond trees showing internal necroses and brown to black vascular streaking were collected in the Llevant region on the island of Mallorca. Several fungal species were subsequently isolated from the margin between healthy and symptomatic tissue. Five species of Botryosphaeriaceae (namely Botryosphaeria dothidea, Diplodia olivarum, D. seriata, Neofusicoccum australe and N. parvum), Eutypa lata, Phaeoacremonium iranianum and Phomopsis amygdali were identified based on morphology, culture characteristics and DNA sequence comparisons. Neofusicoccum parvum was the dominant species, followed by E. lata, D. olivarum and N. australe. First reports from almond include D. olivarum and Pm. iranianum. Two species are newly described, namely Collophora hispanica sp. nov. and Phaeoacremonium amygdalinum sp. nov.

Pathogenicity testing of lesser-known vascular fungi of grapevines

Several hyphomycetes were recently isolated from asymptomatic or symptomatic vascular tissues of grapevines showing Petri disease symptoms in South Africa. In most cases, their status as pathogens was unknown and pathogenicity studies were, therefore, conducted to determine their potential as decline pathogens. The fungi included Acremonium cf. charticola, Acremonium cf. ochraceum, Cadophora luteo-olivacea, Phialemonium cf. curvatum, Pleurostomophora richardsiae, Phaeoacremonium (Pm.) krajdenii, Pm. parasiticum, Pm. subulatum, Pm. venezuelense and Pm. viticola. Of these, Pm. parasiticum and Pm. viticola have been associated with Petri disease symptoms, although their pathogenicity has not been tested. Phaeomoniella (Pa.) chlamydospora and Pm. aleophilum, known to be involved in Petri disease and esca, were included as positive controls. Pathogenicity studies were conducted in glasshouse experiments where grapevine rootstocks were artificially inoculated, as well as in the field. Data obtained after 3 months from a glasshouse trial were difficult to interpret, due to the small lesions and similarity in disease expression among different species. However, Pa. chlamydospora produced the largest lesions and was by far the most aggressive pathogen. To supplement the glasshouse trial, grapevine trunks and pruning wounds of Vitis vinifera cv. Periquita were artificially inoculated in the field. Field trials, rated after 14 months, confirmed Pa. chlamydospora to be the most aggressive pathogen, since it produced the largest trunk and pruning wound lesions. Furthermore, it was re-isolated more frequently than any of the other fungi, especially from the pruning wounds. All the fungi were able to infect, colonise and produce lesions statistically different to those caused by the water control and the non-pathogen in the field trial.

A review of Phaeoacremonium species involved in Petri disease and esca of grapevines

Summary. The current status of Phaeoacremonium species involved in Petri disease and esca is reviewed. The taxonomical position and classification of Phaeoacremonium as well as its teleomorph, Togninia, are discussed. The review also provides the currently known distribution and host range of Phaeoacremonium species. The epidemiology of Phaeoacremonium species together with the more commonly isolated Phaeomoniella chlamydospora, is also treated. An overview is given of the molecular methods that have been used thus far to identify and detect the fungi involved in Petri disease. The role that Phaeoacremonium species, and the morphologically closely related pathogen Pa. chlamydospora, play in disease development and the results obtained with pathogenicity trials are also discussed. Lastly, an overview is given of the effect of management strategies on the Phaeoacremonium species associated with Petri disease and esca.

Cylindrocarpon species associated with apple tree roots in South Africa and their quantification using real-time PCR

Cylindrocarpon species are known to be a component of the pathogen/pest complex that incites apple replant disease. In South Africa, no information is available on apple associated Cylindrocarpon species and their pathogenicity. Therefore, these aspects were investigated. Among the isolates recovered from apple roots in South Africa, four species (C. destructans, C. liriodendri, C. macrodidymum and C. pauciseptatum) were identified using β-tubulin gene sequencing and phylogenetic analysis. This is the first report of C. liriodendri, C. macrodidymum and C. pauciseptatum on apple trees. Cylindrocarpon macrodidymum was the most prevalent. Isolates within each of the four species were pathogenic towards apple seedlings, but varied in their virulence. With a single exception, all isolates were able to induce lesion development on seedling roots. Only 57% of the isolates, which represented all four species, were able to cause a significant reduction in seedling weight and/or height. The greatest seedling growth reductions were caused by two isolates of C. destructans, and one isolate each of C. liriodendri and C. macrodidymum. A quantitative real-time polymerase chain reaction (qPCR) method was developed for simultaneous detection of all four Cylindrocarpon species. qPCR analyses of Cylindrocarpon from the roots of inoculated seedlings showed that the amount of Cylindrocarpon DNA in roots was not correlated to seedling growth reductions (weight and height) or root rot. The qPCR method is, however, very useful for the rapid identification of apple associated Cylindrocarpon species in roots. The technique may also hold potential for being indicative of Cylindrocarpon disease potential if rhizosphere soil rather than roots are used.

Morphological and phylogenetic analyses of Pythium species in South Africa

The genus Pythium is important in agriculture, since it contains many plant pathogenic species, as well as species that can promote plant growth and some that have biocontrol potential. In South Africa, very little is known about the diversity of Pythium species within agricultural soil, irrigation and hydroponic systems. Therefore, the aim of the study was to characterise a selection of 85 Pythium isolates collected in South Africa from 1991 through to 2007. The isolates were characterised morphologically as well as through sequence and phylogenetic analyses of the internal transcribed spacer regions (ITS) and the 5.8S gene of the nuclear ribosomal DNA. Phylogenetic analyses showed that the isolates represented ten of the 11 published Pythium clades [Lévesque & De Cock, 2004. Molecular phylogeny and taxonomy of the genus Pythium. Mycological Research 108: 1363-1383]. Characterisation of isolates in clade D and J suggested that the phylogenetic concept of Pythium acanthicum and Pythium perplexum respectively, needs further investigation in order to enable reliable species identification within these clades. Our phylogenetic analyses of Pythium species in clade B also showed that species with globose sporangia group basal within this clade, and are not dispersed within the clade as previously reported. The 85 South African isolates represented 34 known species, of which 20 species have not been reported previously in South Africa. Additionally, three isolates (PPRI 8428, 8300 and 8418) were identified that may each represent putative new species, Pythium sp. WJB-1 to WJB-3.

Phaeoacremonium: from esca disease to phaeohyphomycosis.

Phaeoacremonium spp. are commonly isolated from stems and branches of diseased woody hosts, and humans with phaeohyphomycosis. The genus Phaeoacremonium (Togniniaceae, Togniniales) has recently been monographed, and presently contains 46 species, while its sexual morph, Togninia, contains 26 epithets, of which 13 are insufficiently known. In this review we summarise information pertaining to the global distribution, pathology, ecology, and detection of these species, and present a case for retaining the genus Phaeoacremonium over that of Togninia. Furthermore, to obtain a single nomenclature, the following new combinations are also proposed: Phaeoacremonium africanum, P. aquaticum, P. fraxinopennsylvanicum, P. griseo-olivaceum, P. inconspicuum, P. leptorrhynchum, P. minimum, and P. vibratile.

Phytophthora taxa associated with cultivated Agathosma, with emphasis on the P. citricola complex and P. capensis sp. nov.

Agathosma species, which are indigenous to South Africa, are also cultivated for commercial use. Recently growers experienced severe plant loss, and symptoms shown by affected plants suggested that a soilborne disease could be the cause of death. A number of Phytophthora taxa were isolated from diseased plants, and this paper reports their identity, mating type, and pathogenicity to young Agathosma plants. Using morphological and sequence data seven Phytophthora taxa were identified: the A1 mating type of P. cinnamomi var. cinnamomi, P. cinnamomi var. parvispora and P. cryptogea, the A2 mating type of P. drechsleri and P. nicotianae, and two homothallic taxa from the P. citricola complex. The identity of isolates in the P. citricola complex was resolved using reference isolates of P. citricola CIT groups 1 to 5 sensu Oudemans et al. (1994) along with multi-locus phylogenies (three nuclear and two mitochondrial regions), isozyme analyses, morphological characteristics and temperature-growth studies. These analyses revealed the isolates from Agathosma to include P. multivora and a putative novel species, P. taxon emzansi. Furthermore, among the P. citricola reference isolates the presence of a new species was revealed, described here as P. capensis. Findings of our study, along with some recent other studies, have contributed to resolving some of the species complexity within the P. citricola complex, resulting in the identification of a number of phylogenetically distinct taxa. The pathogenicity of representative isolates of the taxa from Agathosma was tested on A. betulina seedlings. The putative novel species, P. taxon emzansi, and P. cinnamomi var. parvispora were non-pathogenic, whereas the other species were pathogenic to this host.