First report of Neopestalotiopsis clavispora causing leaf spot disease on banana (Musa acuminate L.) in China

Banana (Musa acuminate L.) is an important tropical fruit in China. In October 2020, a new leaf spot disease was observed on banana plants at an orchard of Zhenkang county (23°45'23.46″ N, 98°48'46.52″ E), Lincang city, Yunnan province, China. The disease incidence was about 1%. The leaf spots occurred sporadically and the percentage of the leaf area covered by lesions was less than 5%. Symptoms on the leaves were initially small, irregular, reddish-brown spots that gradually expanded to fusiform-shaped lesions with a yellow halo and eventually become necrotic, dry, and cracked. To isolate the pathogen, thirty symptomatic leaves (15 mm2) from five plants were surface disinfected in 70% ethanol (10 s) and 0.8% NaClO (2 min), rinsed in sterile water three times, and transferred to potato dextrose agar (PDA) at 28°C for 5 days. Twenty-five colonies formed on the PDA plates were white with cottony aerial mycelium, round with a light orange underside. Abundant black globular acervuli semi-immersed on PDA were observed after a week. Conidia were straight or slightly curved, clavate to spindle, five cells, four septa with dimensions of 17.49 to 34.51 µm × 4.24 to 7.28 µm (avg. 23.83 × 5.62 µm; n=50). The apical and basal cells were hyaline, whereas the three median cells were dark brown. Conidia had a single basal appendage with lengths of 2.95 to 17.7 µm (avg. 7.18 µm; n=50) and two to three apical appendages with lengths of 10.7 to 53.84 µm (avg. 17.36 µm; n=50). These morphological characteristics are consistent with those of Neopestalotiopsis spp. (Maharachchikumbura et al. 2014). To confirm species, single-spore cultures of two representative isolates CATAS-102001 and CATAS-102002 were selected for further identification. The internal transcribed spacer (ITS) region, translation elongation factor 1-α (TEF1-α) and β-tubulin (TUB2) genes of the two isolates were amplified with primers ITS1/ITS4 (White et al. 1990), EF1-728/EF2 (O'Donnell et al. 1998; Carbone and Kohn, 1999) and T1/Bt2b (Glass and Donaldson, 1995; O'Donnell and Cigelnik, 1997), respectively, and sequenced. The sequences were deposited in GenBank (ITS: OM281005 and OM281006; TEF1-α: OM328820 and OM328821; TUB2: OM328818 and OM328819). A maximum likelihood phylogenetic tree was constructed using the MEGA 7.0 (Kumar et al. 2016) based on the concatenated sequences ITS region, EF1-α and TUB2 gene, and the cluster analysis placed the representative isolates CATAS-102001 and CATAS-102002 within a cla