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Spain is the second largest onion-producing country in Europe with 1,465,430 tons and an export value of 584 million € in 2021 (MAPA 2022). In summer 2022 rot bulb symptoms were observed in five commercial fields and during the storage of cultivars 'Orlenda', 'Veleta', 'Mallory', 'Citation' and 'Pantano' from La Roda in Albacete (Castilla-La Mancha, Spain). Approximately 20% of sampled bulbs (113 bulbs analyzed) were affected with dry scales showing brown to dark brown rot on the top and basal plate of the onion bulbs. Occasionally, white to light pink fungal mycelium was observed between rotten scales and the plate basal. Sections of dry scales (5-10 mm) of the apical and basal plate were cut and placed on potato dextrose agar (PDA) and Komada medium (Komada 1975). From 5-day-old cultures typical white to light pink mycelium with microconidia in chains formed on polyphialides and macroconidia resembling Fusarium proliferatum (Nelson et al. 1983). To confirm the pathogen identity, partial translation elongation factor 1-alpha ( TEF1 ) and RNA polymerase II subunit 2 ( RPB2 ) genes were amplified and sequenced using primers reported in O´Donnell et al. (1998) and Samuels et al. (2002) for TEF1 and Liu et al. (1999) for RPB2 . In BLAST analyses, the sequences showed 100% identity to the corresponding region of F. proliferatum (KP964908 and JF740801). Sequences were submitted to GenBank, and registered accession numbers are OR061014-16 for TEF1 and OR061017-19 for RPB2. Pathogenicity tests were conducted by inoculating healthy onion bulbs (five replicates per treatment) on the apical and basal plate by placing a 7-day old mycelial plug (10 mm diameter) from PDA cultures. Two onion cultivars ('Pandero' by Nunhems USA and 'Mallory' by Bejo The Netherlands) were inoculated separately with three isolates (PRO1, PRO9, PRO12). Control bulbs were inoculated with sterile PDA. The experiment was carried out twice. All bulbs were placed in a moist chamber and incubated at 25°C in the dark. After 15 days, bulbs inoculated with mycelial plugs showed similar symptoms to those of the original diseased bulbs. Browning dry rot was observed on the apical and basal plate of bulbs. When bulbs were cut longitudinally inner progressing rot was observed. Control bulbs remained symptomless. In both experiments, F. proliferatum was successfully re-isolated and morphologically confirmed from symptomatic bulbs to fulfill Koch's postulates. These results confirmed that isolates PRO1, PRO9 and PRO12 were the pathogen causing basal and dry rot on onion bulbs. This pathogen has recently been identified in China on Allium cepa L. var. agrogatum (Liu et al. 2022) and Idaho on onion (Beck et al. 2020) and could become a serious threat to onion production in Spain, reducing the quality and yield of onion.

Spain is the fourth largest lettuce-producing country in the world and the leading European producer. Much of the production, mainly grow in open field, is dedicated to export with a value of 887 million U.S. dollars per year. In summer 2021 wilting symptoms were observed in a commercial field crop on butterhead lettuce 'Amible' in Albacete (Castilla-La Mancha, Spain). Approximately 15% of plants were affected, but losses were even more severe on subsequent crops. Vascular tissue of affected plants showed a brown to red discoloration. Sections of infected vascular tissue (3 to 5 mm long) were surface sterilized in 70% ethanol for 30 s, washed three times with sterile water, and plated on potato dextrose agar (PDA) amended with streptomycin sulfate (100 mg/liter). From 5-day-old cultures typical pale cream to purplish mycelia with microconidia, macroconidia, and chlamydospores of Fusarium oxysporum were observed. Microconidia were abundant on carnation leaf agar and measured 6.1 to 9.2 μm (mean 7.1 ± 0.7 μm; n=50)). Macroconidia were sparse, three-septate, straight to slightly curved, 23.3 to 34.8 × 4 to 5.2 µm (mean 31.5 ± 2.8 × 4.2 ± 0.3 µm; n=50). Chlamydospores were terminal and intercalary, rough walled, and measured 7.2 to 10.1 μm (mean 9.5 ± 0.6 μm; n=50) µm. DNA was extracted from three single-spore isolates using the protocol of Querol et al. (1992) and the translation elongation factor 1-α gene (TEF) was sequenced with exTEF-F/FUexTEF-R primers as described by Taylor et al. (2016). All TEF sequences (GenBank accession no. OP903519) were identical. In BLAST analyses, the isolates showed 100% identity to the corresponding region of Fusarium oxysporum f. sp. lactucae (FOL) race 4 (MK059958). All Spanish isolates were identified as FOL race 4 using a race-specific polymerase chain reaction (PCR) with the primers FPUF/FPUR (Gilardi et al. 2017), and a previously identified FOL race 4 isolate Fus 1.01 as a positive control. Pathogenicity tests were conducted to confirm the positive result of the race 4-specific PCR and to complete Koch's postulates. Three differential lettuce cultivars ('Costa Rica No. 4', 'Banchu Red Fire', and 'Romana Romabella 30 CN') provided by Rijk Zwaan (The Netherlands) were inoculated with three Spanish isolates (Al1A1, Al1D, Al2B) and the Fus 1.01 isolate used as FOL race 4 positive control (Claerbout et al., 2018). Roots of 3-week-old plants (five replicates per treatment) were dipped in a spore suspension (1 × 106 conidia/ml) for 10 min before transplanting into 250-ml pots with sterile substrate. Non-inoculated control plants were dipped in sterile water for 10 min. The experiment was carried out twice. Inoculated lettuce seedlings were planted and maintained in a growth chamber (25°C day, 18°C night). Plants were slightly watered every other day. After 21 days, wilting was observed in the cultivars 'Costa Rica No. 4' and 'Romana Romabella 30 CN'. Moreover, taproots were cut longitudinally, and vascular browning was observed in the taproot. No discoloration could be observed in the taproot of 'Banchu Red Fire' plants, coinciding with the result of isolate Fus 1.01 used as FOL race 4 positive control. Non-inoculated control plants remained healthy and vascular browning was not observed. In both experiments, F. oxysporum f.sp. lactucae was consistently reisolated using PDA medium. These results confirmed that the isolates Al1A1, Al1D and Al2B were FOL race 4. This race has recently been identified in The Netherlands (Gilardi et al. 2017), Belgium (Claerbout et al. 2018), United Kingdom, Ireland (Taylor et al. 2019) and Italy (Gilardi et al. 2019) and could become a serious threat to Spain lettuce production.