Ecology and systematics of South African Protea-associated Ophiostoma species

Roets, Francois (2006-12)

Thesis (PhD (Botany and Zology))--University of Stellenbosch, 2006.


The well-known, and often phytopathogenic, ophiostomatoid fungi are represented in South Africa by the two phylogenetically distantly related genera Ophiostoma (Ophiostomatales) and Gondwanamyces (Microascales). They are commonly associated with the fruiting structures (infructescences) of serotinous members of the African endemic plant genus Protea. The species O. splendens, O. africanum, O. protearum, G. proteae and G. capensis have been collected from various Protea spp. in South Africa where, like other ophiostomatoid fungi, they are thought to be transported by arthropod vectors. The present study set out to identify the vector organisms of Protea-associated members of mainly Ophiostoma species, using both molecular and direct isolation methods. A polymerase chain reaction (PCR) and taxon specific primers for the two Protea-associated ophiostomatoid genera were developed. Implementation of these newly developed methods revealed the presence of Ophiostoma and Gondwanamyces DNA on three insect species. They included a beetle (Genuchus hottentottus), a bug (Oxycarenus maculates) and a psocopteran species. It was, however, curious that the frequency of these insects that tested positive for ophiostomatoid DNA was very low, despite the fact that ophiostomatoid fungi are known to colonise more than 50% of Protea infructescences. Subsequent direct isolation methods revealed the presence of reproductive propagules of Ophiostoma spp. on four Protea-associated mite species (Oodinychus sp., two Tarsonemus spp. and Proctolaelaps vandenbergi). These mites are numerous within Protea infructescences and Ophiostoma spp. were isolated from a high frequency of these individuals. The Oodinychus sp. mite was found to vector most of the Protea-associated Ophiostoma species. It was thus postulated that the mites (in particular the Oodinychus sp.) act as primary vectors of the Protea-associated Ophiostoma species. The association between Oodinychus mites collected from P. repens and O. splendens proved to be mutualistic. Mites feeding on this fungus showed significantly higher population growth than mites feeding on any of the other fungal species tested. The short- and long-distance dispersal methods of these mites were also investigated. Firstly the ability of mites to move from drying infructescences to moist and sheltered areas such as provided by intact infructescences on the same plant was investigated experimentally. Significantly more mites were found to actively disperse from drying infructescences to artificially manufactured infructescences containing moistened filter paper shreds than to artificially manufactured infructescences containing dry filter paper shreds. The frequent fires associated with the habitat of these mites would, however, require movement over larger areas than what would be possible through self-dispersal. Dispersal of mites via air currents was thus investigated using sticky traps, but no Ophiostoma-vectoring mites were captured in this way. Self-dispersal aided by air currents could thus be ruled out, and our investigations shifted to vectored dispersal. Numerous insects emerging from Ophiostoma-containing P. repens and P. neriifolia infructescences were collected using specially designed emergence cages. Scanning electron microscopy and stereo-microscopy revealed that all three Ophiostoma-vectoring mite genera were phoretic on the beetle G. hottentottus. Tarsonemus spp. and P. vandenbergi were also phoretic on the beetles Trichostetha fascicularis and T. capensis associated with P. repens and P. neriifolia flowers. Mites collected from the surface of these beetles were found to vector reproductive propagules of various Ophiostoma spp. This thus seems to be the only method of long-distance dispersal of these mites and subsequently also the Protea-associated Ophiostoma species. Molecular phylogenetic reconstruction based on large subunit, ITS and beta-tubulin DNA sequence data suggests a polyphyletic origin for the Protea-associated members of Ophiostoma, which proposes multiple invasions of this unusual niche by these fungi. These studies also revealed the presence of four new species of Ophiostoma associated with Protea spp. The new species O. palmiculminatum, O. phasma, O. gemellus and Sporothrix variecibatus were thus described. Ophiostoma palmiculminatum is associated with P. repens infructescences and the Oodinychus mites collected from them. Ophiostoma phasma was collected from various Protea and mite species. Ophiostoma gemellus and Sporothrix variecibatus were initially only isolated from mites, but have subsequently also been isolated from Protea spp. The present study clarifies many aspects pertaining to the phylogeny and ecology of the interesting members of Ophiostoma associated with Protea hosts. As such this study will form the platform for further studies on the co-evolution of these insect / mite / fungi / plant associations.

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