First Report of Fusarium Wilt Caused by Fusarium oxysporum f. sp. passiflorae on Passion Fruit in Portugal

Passion fruit, Passiflora edulis Sims f. edulis, is native to Central and South America and is commercialized in most of the tropical and subtropical zones of the world. Many of the producing countries see their expansion limited owing to the presence of various plant pathogens that challenge the development of this crop (Teixeira et al. 2016). Fusarium oxysporum f. sp. passiflorae, the causal agent of vascular wilt, is responsible for significant economic losses (Silva et al. 2013). This disease was first reported in Australia in 1950 and later in South Africa, Malaysia, Brazil, Panama, and Venezuela (Ortiz and Hoyos-Carvajal et al. 2016). In July 2018, severe wilting symptoms were observed in 1-year-old plants of P. edulis in a greenhouse in Aveiro (Central Region of Portugal). Dark brown vascular discoloration was present in the roots and lower stems of the plants, similar to the symptoms reported in North America on this host (Rooney-Latham et al. 2011). Symptoms were seen on approximately 25% of the plants. The surface of the roots and stems from two plants were sterilized in bleach at 10% before placing them on potato dextrose agar (PDA) or malt extract agar (MEA). In these plants, F. oxysporum colonies were consistently isolated from symptomatic roots and stems on PDA and MEA. The morphology of the colonies and conidia was assessed onto PDA and Spezieller-Nährstoffarmer agar (SNA). Colonies were pinkish violet with chlamydospores, and multiseptate macroconidia and microconidia formed in masses on monophialidic conidiophores. One isolate was recovered and used for all further experiments. Molecular characterization of the EF-1α region was performed as previously described (Sharma et al. 2018). The BLAST analysis of the amplicon (MK430070) showed 100% similarity with an isolate of F. oxysporum f. sp. passiflorae from North America (JF332039) and 99% similarity with an isolate from Australia (KX434919). A portion of the β-tubulin gene was amplified as previously described (Glass and Donaldson 1995) and sequenced with primers Bt2a and Bt2b (MK430071). The analysis of the amplicon showed 99% similarity with an isolate from Brazil (AF008540). To fulfill Koch’s postulates, inoculations were performed on four 6-month-old plants of P. edulis with 12 true leaves: one control plant and three inoculated plants. A conidial spore suspension was prepared by inoculating MEA liquid medium with 10 mycelium plugs of F. oxysporum grown on SNA for 5 days. The concentration was adjusted to 1 × 10⁶ spores/ml. Plant roots were washed and cut approximately 5 cm from the terminal apices and submerged in the conidial spore suspension for 20 min. The process with the control plant was the same, but it was submerged in sterile water. All plants were planted in 125-cm³ pots and kept in a 25°C growth chamber with a 12-h photoperiod. Symptoms were observed on the inoculated plants within 3 weeks and included wilt, chlorotic yellow leaves with subsequent defoliation and necrotic roots. No symptoms were observed on the control plant. F. oxysporum reisolated from the symptomatic root and stem tissue from all the inoculated plants was morphologically and genetically identical to the original isolate. F. oxysporum was not isolated from the control plant. F. oxysporum strain MUM 18.58 has been deposited in the culture collection of Micoteca of the Universidade do Minho (Braga, Portugal).