Phytohormone abscisic acid (ABA) plays significant role in many physiological processes and response of plants to abiotic and biotic stresses. Phytopathogenic fungi also produce ABA, but the role of this trait in interactions with host plants is poorly understood. In this work 65 collection strains of phytopathogenic fungi (13 genera, 25 species) were screened for ABA production in batch culture using a modified potato dextrose (MPD) and original chemically defined (OCD) media. Analysis of ABA content was carried out by ultra-performance liquid chromatography. Thirty-four strains belonging total of 13 species produced ABA growing on MPD medium, and among them nineteen strains also produced ABA growing on OCD medium. A maximum ABA concentration was detected in MPD culture fluid of strain Apiospora montagnei MF-R13.8 (56.5 ± 0.1 µg L-1), whereas strain MF-S41.5 of the same species was the most active ABA producer (13.4 ± 1.1 µg L-1) growing on OCD medium. For the first time ABA was detected in species Alternaria tenuissima, Apiospora montagnei, Bipolaris sorokiniana, Fusarium avenaceum, F. solani, Pythium ultimum, Sclerotinia sclerotiorum, and Sclerotium varium. No correlation between the ability to produce ABA and host plant, plant organ of isolation or region of strain origin was found. In agar dish culture three tomato cultivars were inoculated with strains of Fusarium solani or F. oxysporum differing in ABA production in vitro to test relationship between the ability of fungi to produce ABA and to appear negative effects on plants. Generally, ABA production didn’t correlate with the effects of fungi of tomato roots, with one exception that ABA production by F. solani strains negatively correlated (r = -0.82, P = 0.046, n = 6) with root length of cultivar Ailsa-Craig. The results suggest possibility for the role of fungal ABA as a positive modulator of pathogenesis, but manifestation of this effect depends on plant genotype and fungus species. The selected ABA-producing strains can be used to study mechanisms underlying involvement of fungal ABA in plant-microbe interactions.