The fungal rhizosphere communities associated with soil surrounding Aizoaceae plants in the Namaqua National Park in the Succulent Karoo biodiversity hotspot, South Africa, were analysed by culture-dependent and culture-independent methods. The keratinophilic fungal Order, Onygenales, was the most abundant Order observed in the total fungal population. Fungal communities observed during the dry and flowering seasons differed significantly. The Orders Ascosphaerales and Chaetothyriales were the major contributors to the observed seasonality of fungal communities. No isolates belonging to these Orders were obtained in the culture-dependent experiments, while isolates belonging to the Orders Hypocreales and Eurotiales were abundant in culture. Seasonal differences were most pronounced for the Order Ascosphaerales, which contributed less than 0.1% to the total population during the dry season, but more than 20% to the flowering season population. This study provides the first next generation sequencing analysis of the remarkable soil-associated fungal richness and diversity in the Succulent Karoo, the most biodiverse arid ecoregion in the world.