First report of <i>Fusarium oxysporum</i> f. sp. <i>lycopersici</i> race 3 on tomato in Spain

Fusarium wilt of tomato (Solanum lycopersicum), caused by Fusarium oxysporum f. sp. lycopersici (Fol), threatens global tomato production for both processing and the fresh-market (Chitwood-Brown et al., 2021). Three physiological races of Fol (1, 2, and 3) are distinguished by their specific pathogenicity on differential cultivars carrying dominant race-specific resistance genes (Cai et al., 2003). Extremadura is the main industrial tomato-growing region in Spain, with an area of around 20,000 ha (Ministerio de Agricultura, Pesca y Alimentación, 2023) mainly located in Badajoz (southwest Spain), where most of the cultivars grown are resistant to Fol race 2. In 2020, symptoms of Fusarium wilt disease were observed in a commercial plot (38° 59´ 40.42’’ N, 6° 0´ 41.05’’ W) planted with the tomato cv. H1015, resistant to race 2. In 2021, a trial was established in the same plot with the cvs. H1015 and UG16112 (resistant to race 2) and UG29814 (resistant to race 3). The presence of symptoms such as wilting, yellowing and drying of leaves (Fig. 1) and vascular necrosis (Fig. 2) was evaluated in 350 plants per cultivar. Symptoms were observed in H1015 (61.4%) and UG1612 (10.5% of the plants), but not in UG29814. In a subsequent survey in 2024, similar symptoms were observed in seven commercial plots in the same area, all planted with race 2-resistant cultivars. Fusarium oxysporum was consistently isolated from stem sections of diseased plants analysed on potato dextrose agar medium (PDA). Three isolates (one from cv. UG16112 and two from cv. H1015) were identified as Fol race 3 with PCR analysis using the primers P12-F2B/P12-R1 and SIX3-F1/SIX3-R2 for the identification of the forma specialis lycopersici, and three forward primers (SIX3-G121A-F2, SIX3-G134A-F2 and SIX3-G137C-F1) combined with the reverse primer SIX3-R3 for differentiating race 2 and race 3 isolates (Lievens et al., 2009) (Table 1). Pathogenicity tests were done to confirm the identification of Fol race 3, using five differential tomato cultivars: ACE55 and Marmande (susceptible to races 2 and 3), H1015 and Pasadena (resistant to race 2 and susceptible to race 3) and UG29814 (resistant to all races). Plants at the two-to three-true-leaf stage (10 plants per cultivar and isolate, plus 10 control plants per cultivar) were transplanted into one-litre pots containing a sterilised substrate (a vermiculite and peat mixture [1:1, v:v]), which was inoculated by pouring 10 mL of a conidial suspension (1 × 106 CFU/mL) into the substrate per plant and grown in a growth chamber at 27°C day/23°C night and a16 hr photoperiod. Fusarium wilt symptoms were observed on cv. ACE55 and Marmande and on cv. H1015 and Pasadena (90%, 100%, 100% and 80% of the plants, respectively) but no symptoms were observed in cv. UG29814 or in the controls. Fusarium oxysporum was isolated consistently on PDA (100% of the plants) from vascular tissues of the four cultivars susceptible to race 3 (Fig. 3). This is the first formal report of F. oxysporum f. sp. lycopersici race 3 in Spain. The spread of race 3 threatens industrial tomato production in Extremadura and Europe, especially where race 2-resistant cultivars are common. Monitoring and using race 3-resistant cultivars are essential to prevent further spread. This work was supported by Operational Programme ERDF Extremadura 2021–2027 Action 1A1103 and by PID2021-123594OR-C22 project (MCIN/AEI/10.13039/501100011033/FEDER, UE)