Taxonomy and phylogeny of Ophiostoma spp. with Sporothrix anamorphs and their generic relationships in the Ophiostomatales

The ophiostomatoid fungi included more than 450 species of ascomycetes specifically adapted for insect dispersal. Many of these species have a significant economic impact as sapstaining fungi or tree pathogens harmful to forestry industries, but some are also as opportunistic human pathogens. DNA based studies in recent years have shown that the majority of these fungi belonged in either the Ophiostomatales or Microascales (Sordariomycetes), with a few Sporothrix spp. grouping in the Microstromatales (Ustilaginomycetes). However, most phylogenetic studies have focussed on restricted numbers of taxa sharing similar morphology. The aim of the studies in this thesis was to reconsider the taxonomy of all the ophiostomatoid fungi at the order and family levels, and the status of genera and species with sporothrix-like anamorphs in the Ophiostomatales and Microstromatales. All available published sequence data were screened for reliable sequences representing as many species as possible, and new data were generated where necessary for ex-type or other isolates. The resulting phylogenies enabled the formal redefinition of the Ophiostomatales and Ophiostomataceae, and the description of two new families, the Graphiaceae (Microascales) and Quambalariaceae (Microstromatales). Problems relating to the delineation of Ophiostoma s.l., Leptographium s.l., and Raffaelea s.l. were exposed and discussed, 18 species complexes were defined in the Ophiostomatales, and four genera were formally redefined: Sporothrix, Graphium, Graphilbum and Knoxdaviesia. Forty six new combinations were made, primarily in Sporothrix, Ophiostoma, Graphilbum and Knoxdaviesia. One nomen novum was erected in Ceratocystis and one new Quambalaria species was described. A comprehensive nomenclator for 596 ophiostomatoid species including references to all descriptions, synonymies and phylogenetic data was also compiled. This study represents the first comprehensive, all-inclusive assessment of the taxonomy and nomenclature of the ophisotomatoid fungi based on phylogenetic relationships and the one fungus one name principles. Finally, the immediate and indiscriminate application of the one fungus one name principles in Ophiostoma s.l. and Leptographium s.l. might result in many unnecessary name changes. Thus, several recommendations have been made to ensure nomenclatural stability in these genera in the immediate future and until more robust phylogenies become available that can refine the delineation of these genera.PhDMicrobiology and Plant PathologyUnrestricte

DNA sequence comparisons of Ophiostoma spp., including Ophiostoma aurorae sp. nov., associated with pine bark beetles in South Africa

Bark beetles (Coleoptera: Scolytinae) are well-recognized vectors of Ophiostoma species. Three non-native bark beetle species infest various Pinus species in South Africa, and they are known to carry at least 12 different species of ophiostomatoid fungi. Some of these fungi have not been identified to species level. The aim of this study was to determine or confirm the identities of Ophiostoma species associated with bark beetles in South Africa using comparisons of DNA sequence data. Identities of Ophiostoma ips, O. floccosum, O. pluriannulatum, O. quercus and O. stenoceras were confirmed. Ophiostoma abietinum, O. piliferum and Pesotum fragrans are recognised for the first time and the new species, O. aurorae sp. nov., is described from pine-infesting bark beetles in South Africa

DNA-based identification of Quambalaria pitereka causing severe leaf blight of Corymbia citriodora in China

Quambalaria spp. include serious plant pathogens, causing leaf and shoot blight of Corymbia and Eucalyptus spp. In this study, a disease resembling Quambalaria leaf blight was observed on young Corymbia citriodora trees in a plantation in the Guangdong Province of China. Comparisons of rDNA sequence data showed that the causal agent of the disease is Q. pitereka. This study provides the first report of Quambalaria leaf blight from China, and it is also the first time that this pathogen has been found on trees outside the native range of Eucalypts

Multi-gene phylogenies define Ceratocystiopsis and Grosmannia distinct from Ophiostoma

Ophiostoma species have diverse morphological features and are found in a large variety of ecological niches. Many different classification schemes have been applied to these fungi in the past based on teleomorph and anamorph features. More recently, studies based on DNA sequence comparisions have shown that Ophiostoma consists of different phylogenetic groups, but the data have not been sufficient to define clear monophyletic lineages represented by practical taxonomic units. We used DNA sequence data from combined partial nuclear LSU and β-tubulin genes to consider the phylogenetic relationships of 50 Ophiostoma species, representing all the major morphological groups in the genus. Our data showed three well-supported, monophyletic lineages in Ophiostoma. Species with Leptographium anamorphs grouped together and to accommodate these species the teleomorph-genus Grosmannia (type species G. penicillata), including 27 species and 24 new combinations, is re-instated. Another well-defined lineage includes species that are cycloheximide-sensitive with short perithecial necks, falcate ascospores and Hyalorhinocladiella anamorphs. For these species, the teleomorph-genus Ceratocystiopsis (type species O. minuta), including 11 species and three new combinations, is re-instated. A third group of species with either Sporothrix or Pesotum anamorphs includes species from various ecological niches such as Protea infructescences in South Africa. This group also includes O. piliferum, the type species of Ophiostoma, and these species are retained in that genus. Ophiostoma is redefined to reflect the changes resulting from new combinations in Grosmannia and Ceratocystiopsis. Our data have revealed additional lineages in Ophiostoma linked to morphological characters. However, these species are retained in Ophiostoma until further data for a larger number of species can be obtained to confirm monophyly of the apparent lineages

The divorce of Sporothrix and Ophiostoma : solution to a problematic relationship

One of the causal agents of human sporotrichosis, Sporothrix schenckii, is the type species of the genus Sporothrix. During the course of the last century the asexual morphs of many Ophiostoma spp. have also been treated in Sporothrix. More recently several DNA-based studies have suggested that species of Sporothrix and Ophiostoma converge in what has become known as Ophiostoma s. lat. Were the one fungus one name principles adopted in the Melbourne Code to be applied to Ophiostoma s. lat., Sporothrix would have priority over Ophiostoma, resulting in more than 100 new combinations. The consequence would be name changes for several economically important tree pathogens including O. novo-ulmi. Alternatively, Ophiostoma could be conserved against Sporothrix, but this would necessitate changing the names of the important human pathogens in the group. In this study, we sought to resolve the phylogenetic relationship between Ophiostoma and Sporothrix. DNA sequences were determined for the ribosomal large subunit and internal transcribed spacer regions, as well as the beta-tubulin and calmodulin genes in 65 isolates. The results revealed Sporothrix as a well-supported monophyletic lineage including 51 taxa, distinct from Ophiostoma s. str. To facilitate future studies exploring species level resolution within Sporothrix, we defined six species complexes in the genus. These include the Pathogenic Clade containing the four human pathogens, together with the S. pallida-, S. candida-, S. inflata-, S. gossypina- and S. stenoceras complexes, which include environmental species mostly from soil, hardwoods and Protea infructescences. The description of Sporothrix is emended to include sexual morphs, and 26 new combinations. Two new names are also provided for species previously treated as Ophiostoma.The National Research Foundation, members of the Tree Protection Cooperative Programme (TPCP), and the DST/NRF Centre of Excellence in Tree Health Biotechnology (CTHB), South Africa.http://www.studiesinmycology.org/am2016Forestry and Agricultural Biotechnology Institute (FABI)GeneticsMicrobiology and Plant Patholog

Phylogenetic analyses of diverse <i>Podaxis</i> specimens from Southern Africa reveal hidden diversity and new insights into associations with termites

<p>Although frequently found on mounds of the grass-cutting termite genus . Trinervitermes, virtually nothing is known about the natural history of the fungal genus . Podaxis (Agaricaceae) nor why it associates with termite mounds. More than 40 species of this secotioid genus have been described since Linnaeus characterised the first species in 1771. However, taxonomic confusion arose when most of these species were reduced to synonymy with . Podaxis pistillaris in 1933. Although a few more species have since been described, the vast majority of specimens worldwide are still treated as . P. pistillaris. Using 45 fresh and herbarium specimens from Southern Africa, four from North America and one each from Ethiopia, and Kenya, we constructed the first comprehensive phylogeny of the genus. Four of the genotyped specimens were more than 100 y old. With the exception of the type specimen of . Podaxis rugospora, all herbarium specimens were labelled as . P. pistillaris or . Podaxis sp. However, our data shows that the genus contains at least five well-supported clades with significant inter-clade differences in spore length, width and wall thickness, and fruiting body length, supporting that clades likely represent distinct . Podaxis species. Certain clades consistently associate with termites while others appear entirely free-living.</p

Actinomadura rubteroloni sp. nov. and Actinomadura macrotermitis sp. nov., isolated from the gut of the fungus growing-termite Macrotermes natalensis

The taxonomic positions of two novel aerobic, Gram-positive actinobacteria, designated strains RB29$^{T}$ and RB68T, were determined using a polyphasic approach. Based on 16S rRNA gene sequence analysis, the closest phylogenetic neighbours of RB29$^{T}$ were identified as Actinomadura rayongensis DSM 102126$^{T}$ (99.2 % similarity) and Actinomadura atramentaria DSM 43919$^{T}$ (98.7 %), and for strain RB68$^{T}$ was Actinomadura hibisca DSM 44148$^{T}$ (98.3 %). Digital DNA–DNA hybridization (dDDH) between RB29$^{T}$ and its closest phylogenetic neighbours, A. rayongensis DSM 102126$^{T}$ and A. atramentaria DSM 43919$^{T}$, resulted in similarity values of 53.2 % (50.6–55.9 %) and 26.4 % (24.1–28.9 %), respectively. Additionally, the average nucleotide identity (ANI) was 93.2 % (94.0 %) for A. rayongensis DSM 102126$^{T}$ and 82.3 % (78.9 %) for A. atramentaria DSM 43919$^{T}$. dDDH analysis between strain RB68$^{T}$ and A. hibisca DSM 44148$^{T}$ gave a similarity value of 24.5 % (22.2–27.0 %). Both strains, RB29$^{T}$ and RB68$^{T}$, revealed morphological characteristics and chemotaxonomic features typical for the genus Actinomadura, such as the presence of meso-diaminopimelic acid in the cell wall, galactose and glucose as major sugar components within whole-cell hydrolysates and the absence of mycolic acids. The major phospholipids were diphosphatidylglycerol, phosphatidylglycerol, phosphatidylinositol and phosphatidylinositol mannoside. Predominant menaquinones were MK-9(H$_{6}$) and MK-9(H$_{8}$) for RB29$^{T}$ and MK-9(H$_{4}$) and MK-9(H$_{6}$) for RB68T. The main fatty acids were identified as 10-methyloctadecanoic acid (10-methyl C$_{18:0}$), 14-methylpentadecanoic acid (iso-C$_{16:0}$), hexadecanoic acid (C$_{16:0}$) and cis-9-octadecanoic acid (C$_{18:1}$ ω9c). Here, we propose two novel species of the genus Actinomadura: Actinomadura rubteroloni sp. nov. with the type strain RB29$^{T}$ (=CCUG 72668$^{T}$=NRRL B-65537$^{T}$) and Actinomadura macrotermitis sp. nov. with the type strain RB68$^{T}$ (=CCUG 72669$^{T}$=NRRL B-65538$^{T}$)

Lessons from a major pest invasion : the polyphagous shot hole borer in South Africa

CITATION: Paap, T. et al. 2020. Lessons from a major pest invasion : the polyphagous shot hole borer in South Africa. South African Journal of Science, 116(11/12):8757, doi:10.17159/sajs.2020/8757.The original publication is available at https://sajs.co.zaThe arrival and establishment of invasive forest pests can cause devastating environmental damage and great economic impact. For example, the cost over the past decade of dealing with the arrival of a single invasive beetle in the USA, the emerald ash borer (Agrilus planipennis), is estimated at more than USD10 billion.1 Originating from Asia, this beetle has killed hundreds of millions of native ash trees since it became established in the USA. However, this beetle is but one of hundreds of invasive insect pests that impact forests in the USA, and that contribute to a global tree health crisis caused by invasive insects and pathogenic microorganisms.2-4 South Africa is no different from other countries and is experiencing an increasing rate of introductions of damaging forest pests.5,6 These invasions are largely unintentional and are a by-product of globalisation and increasing global trade.7,8 The movement of living plants and plant products, including untreated wood packaging materials (i.e. pallets, dunnage and crating), is known to be a major pathway for these pests.9,10 For clarification, in this commentary we use the terms ‘insect’ and ‘pathogen’ to distinguish between the two types of organisms, although we also use the general term ‘pest’ to refer to both groups. The term ‘invasive pest’ is used for introduced species that, in addition to maintaining a self-sustaining population, show evidence of spread and impact.https://sajs.co.za/article/view/8757Publisher's versio

Comparative Genomics Reveals Prophylactic and Catabolic Capabilities of Actinobacteria within the Fungus-Farming Termite Symbiosis

Actinobacteria, one of the largest bacterial phyla, are ubiquitous in many of Earth’s ecosystems and often act as defensive symbionts with animal hosts. Members of the phylum have repeatedly been isolated from basidiomycete-cultivating fungus-farming termites that maintain a monoculture fungus crop on macerated dead plant substrate. The proclivity for antimicrobial and enzyme production of Actinobacteria make them likely contributors to plant decomposition and defense in the symbiosis. To test this, we analyzed the prophylactic (biosynthetic gene cluster [BGC]) and metabolic (carbohydrate-active enzyme [CAZy]) potential in 16 (10 existing and six new genomes) termite-associated Actinobacteria and compared these to the soil-dwelling close relatives. Using antiSMASH, we identified 435 BGCs, of which 329 (65 unique) were similar to known compound gene clusters, while 106 were putatively novel, suggesting ample prospects for novel compound discovery. BGCs were identified among all major compound categories, including 26 encoding the production of known antimicrobial compounds, which ranged in activity (antibacterial being most prevalent) and modes of action that might suggest broad defensive potential. Peptide pattern recognition analysis revealed 823 (43 unique) CAZymes coding for enzymes that target key plant and fungal cell wall components (predominantly chitin, cellulose, and hemicellulose), confirming a substantial degradative potential of these bacteria. Comparison of termite-associated and soil-dwelling bacteria indicated no significant difference in either BGC or CAZy potential, suggesting that the farming termite hosts may have coopted these soil-dwelling bacteria due to their metabolic potential but that they have not been subject to genome change associated with symbiosis. IMPORTANCE Actinobacteria have repeatedly been isolated in fungus-farming termites, and our genome analyses provide insights into the potential roles they may serve in defense and for plant biomass breakdown. These insights, combined with their relatively higher abundances in fungus combs than in termite gut, suggest that they are more likely to play roles in fungus combs than in termite guts. Up to 25% of the BGCs we identify have no similarity to known clusters, indicating a large potential for novel chemistry to be discovered. Similarities in metabolic potential of soil-dwelling and termite-associated bacteria suggest that they have environmental origins, but their consistent presence with the termite system suggests their importance for the symbiosis

Phylogeny of the Quambalariaceae fam. nov., including important Eucalyptus pathogens in South Africa and Australia

The genus Quambalaria consists of plant-pathogenic fungi causing disease on leaves and shoots of species of Eucalyptus and its close relative, Corymbia. The phylogenetic relationship of Quambalaria spp., previously classified in genera such as Sporothrix and Ramularia, has never been addressed. It has, however, been suggested that they belong to the basidiomycete orders Exobasidiales or Ustilaginales. The aim of this study was thus to consider the ordinal relationships of Q. eucalypti and Q. pitereka using ribosomal LSU sequences. Sequence data from the ITS nrDNA were used to determine the phylogenetic relationship of the two Quambalaria species together with Fugomyces (= Cerinosterus) cyanescens. In addition to sequence data, the ultrastructure of the septal pores of the species in question was compared. From the LSU sequence data it was concluded that Quambalaria spp. and F. cyanescens form a monophyletic clade in the Microstromatales, an order of the Ustilaginomycetes. Sequences from the ITS region confirmed that Q. pitereka and Q. eucalypti are distinct species. The ex-type isolate of F. cyanescens, together with another isolate from Eucalyptus in Australia, constitute a third species of Quambalaria, Q. cyanescens (de Hoog & G.A. de Vries) Z.W. de Beer, Begerow & R. Bauer comb. nov. Transmission electron-microscopic studies of the septal pores confirm that all three Quambalaria spp. have dolipores with swollen lips, which differ from other members of the Microstromatales (i.e. the Microstromataceae and Volvocisporiaceae) that have simple pores with more or less rounded pore lips. Based on their unique ultrastructural features and the monophyly of the three Quambalaria spp. in the Microstromatales, a new family, Quambalariaceae Z.W. de Beer, Begerow & R. Bauer fam. nov., is described