First Report of Verticillium dahliae Race 3 Causing Verticillium Wilt of Tomato ( Solanum lycopersicum ) in Uruguay

Verticillium dahliae is a soilborne vascular pathogen that causes Verticillium wilt (VW) of tomato. Three races of V. dahliae have been described; race 3 has recently been reported in tomato, potato, and other crops (Lerohl et al., 2025; Mandal et al., 2024; Usami et al., 2017). In July 2025, tomato plants cv. ‘Tronus’ exhibiting wilting, leaf chlorosis, and vascular discoloration were observed in two commercial greenhouses in Canelones Department, southern Uruguay. Disease incidence was estimated at 70% within an area of approximately 1,000 m 2 . Stem samples were collected from symptomatic plants, surface-sterilized, and longitudinally sectioned. Small fragments taken from the advancing margin of necrotic vascular tissue were plated on PDA and incubated at 25°C under a 12-h photoperiod for 10 days. Nine isolates were obtained, all exhibiting similar colony morphology and were identical in the identification of race 3. Therefore, three isolates (V3, V6, and V9) were selected for morphological characterization and additional molecular analyses, and pathogenicity assays. On PDA, colonies were initially white, turning dark with age, and had lobulated margins with abundant aerial mycelium. Microsclerotia measuring 31.79–74.88 µm in diameter (mean = 47.46 µm; n = 50) were observed after 7 days. Conidia were hyaline, unicellular, measuring 3.6–6.5 × 2.0–3.5 µm (mean = 4.96 × 2.75 µm; n = 100). Species identity was confirmed by PCR amplification and sequencing of the ITS region (primers ITS1/ITS4; GenBank acc. nos. PZ122748–PZ122750) and the GAPDH gene (primers GDF/GDR; GenBank acc. nos. PZ121736–PZ121738) following Suaste-Dzul et al., (2022). The sequences showed 100% identity to V. dahliae isolate PD322 (NR_126124.1for ITS and HQ414719 for GAPDH). Race identification was performed using specific primers targeting the three known effectors, Ave1 (VdAve1-F/VdAve1-R), VdR2e (VdR2e-FAM-F/VdR2e-R), and VdR3e (VdR3e-F/VdR3e-R) (Mandal et al., 2024). Only the VdR3e primer pair produced an amplicon (~350 bp), and the sequence (GenBank acc. nos. PZ121739 –PZ121741) showed 100% identity to race 3 isolate VdPN15.23 (PQ662629). To fulfill Koch’s postulates, the three representative isolates were each used to inoculate five 60-day-old tomato plants (cv. ‘Tronus’). For inoculation, roots were wounded and immersed in a 1 × 10 7 conidia mL –1 suspension. The plants were then transplanted into pots containing sterile peat substrate and maintained under high humidity for 48 h at 24°C with a 12-h photoperiod. Symptoms were first observed at 13 days after inoculation, followed by plant collapse at 16 days. By 30 days after inoculation, a final wilt incidence of 78% was recorded. Control plants remained asymptomatic. The pathogen was reisolated from symptomatic tissues and confirmed as V. dahliae race 3 by molecular analysis. To our knowledge, this is the first report of V. dahliae race 3 causing VW of tomato in Uruguay. The presence of V. dahliae race 3 has a serious risk to tomato crop health in Uruguay, because current commercial varieties are susceptible due to the absence of effective resistance genes against this pathotype.