Fungi are responsible for many plant infections. Indeed, fungi are responsible for most plant spoilage that occurs in the field or in storage, causing symptoms of dieback, leaf spot, rot, rust and wilt. A more serious issue is that fungi produce mycotoxins, which can contaminate crops. Samples of leaf, fruit and root tissues were collected from sick crops, pre- and post-harvest within Al-Baha region. Using molecular markers, twenty fungal isolates were identified that represent eight genera and thirteen species. The most common species were Penicillium spp. (45%), followed by Aspergillus spp. (25%). The remaining 30% were comprised of multiple species (Alternaria alternata, Fusarium oxysporum, Geotrichum candidum, Lasiodiplodia theobromae, Megasporoporia minor and Rhizopus arrhizu). The majority of these isolates are recognised as spoilers of crops and producers of mycotoxins. The study evaluated which temperatures optimised fungal growth. Half of the isolates thrived best at 25 °C; six isolates favoured 30 °C, whilst four isolates grew best at 20 °C. The study also evaluated the optimal water activity (aw) for fungal growth, at four points (0.995, 0.99, 0.90 and 0.85 aw). The isolates were categorised as being either xerophilic, mesophilic or hydrophilic. The majority of isolates (70%) were xerophilic (Aspergillus and Penicillium spp.), 10% were mesophilic (Alternaria alternata and Rhizopus oryzae spp.) and 20% were hydrophilic (Lasiodiplodia theobromae, Geotrichum candidum, Megasporoporia minor and Fusarium oxysporum spp). The information derived from this study will assist in building effective strategies to control these pathogens, minimising crops losses and mycotoxin contamination.