First report of Fusarium odoratissimum causing postharvest kiwifruit rot

In 2019, kiwifruits (Actinidia chinensis cv. ‘Huayou’) with typical soft rot symptoms (54% incidence, n=200) were randomly collected from a commercial orchard in Shaanxi Province, China (107°39′E, 33°42′N). Lesions were round or oval, with yellowish centers and a water-soaked margin between infected and healthy tissue (Fig. 1a). Tissue segments (4×4 mm) taken from lesion margins of five decayed fruits were surface-disinfected (1% NaClO, 30 s; 70% ethanol, 60 s), rinsed with sterile water, air-dried on filter paper, and plated on PDA at 25 °C for 3 days. Hyphal tips were transferred to PDA for pure cultures. A total of 13 fungal isolates were obtained, including two previously unidentified strains (SXHY2-1 and SXHY3-1) and 11 reported strains (two Botryosphaeria dothidea and nine Diaporthe spp.). In 2021, two additional phenotypically similar strains (SXHY2-2 and SXHY3-2) were isolated from the same orchard. On PDA, SXHY2-1 and SXHY2-2 exhibited purple-red central hyphae with white margins, while SXHY3-1 and SXHY3-2 produced abundant white aerial mycelium (Fig. 1b, c). All four isolates produced similar conidia on carnation leaf agar (CLA). Macroconidia were slender, thin-walled, 3-4 septa, measuring 25.1-54.9×3.1-6.2 µm (mean 44.9 × 4.5 µm, n=35). Microconidia were oval, elliptical, or kidney-shaped, 0-1 septum, 4.8-14.4×1.4-4.0 µm (mean 8.6 × 2.8 µm, n=35) (Fig. 1d, e). Chlamydospores were terminal or intercalary, rough-walled, single or paired, 5.6-12.8 µm in diameter (mean 8.1 µm, n=35) (Fig. 1f, g). Conidiogenous cells were short monophialides with 2-5 loci (Fig. 1h, i). These features matched Fusarium odoratissimum (Maryani et al., 2019; Ujat et al., 2021). The four isolates were identified by sequencing RPB1, RPB2, and EF1-α genes using specific primers (O'Donnell et al., 2010). With SXHY3-2 identical to SXHY3-1, the sequences of RPB1 (MW646527, PX596509, MN264748), RPB2 (MW646526, PX596510, MN264750), and EF1-α (MW646525, PX548364, MN264749) were obtained from SXHY2-1, SXHY2-2, and SXHY3-1, respectively. Polyphasic identification with the three sequences by Fusarium MLST, SXHY2-1, SXHY2-2 and SXHY3-1 shared 99.83%, 98.18%, 99.70% identity respectively with the F. odoratissimum strain LC13762. BLASTn analysis confirmed that isolates SXHY2-1, SXHY2-2 and SXHY3-1 shared 99.22 to 100% identity in their EF1-α, RPB1, and RPB2 genes with F. odoratissimum reference sequences (OR865337, PV613661, OR865341). Phylogenetic analysis (Maximum Likelihood, MEGA7) of the combined dataset confirmed all four strains as F. odoratissimum (Fig. 2). For pathogenicity tests, surface-disinfested fruits of ‘Xuxiang’ (A. deliciosa) and ‘Sungold’ (A. chinensis) were rinsed and air-dried. Wounds (3 mm deep) were inoculated with 15 µL of conidial suspension (1×10⁶ cfu/mL); controls received sterile water. Fruits were incubated at 25 °C and 80% RH. Watery soft decay developed on all inoculated fruits within seven days. The mean lesion diameter was 24.5±1.2 mm on 'Xuxiang'(n=15) and 17.5±0.9 mm on 'Sungold' ( n=15 )(Fig.1j-1m). Control fruits remained healthy. The test was conducted twice. Fungi re-isolated from lesions were confirmed as F. odoratissimum based on morphology and RPB2 sequencing. F. odoratissimum (formerly F. oxysporum f.sp. cubense) is a distinct species due to genomic differences (Maryani et al., 2019; Ujat et al., 2021; van Westerhoven et al., 2023). As far as we know, this is the first report of F. odoratissimum causing kiwifruit rot.