Pengfei He

Maize (Zea mays. L) is cultivated globally as a staple food crop, animal feed, and biofuel. However, persistent diseases in maize have led significant yield losses and a decline in grain quality (Yang et al., 2017). In August 2024, long and narrow lesions were observed on the leaves of maize growing in a field in Luoping County, Qujing City, Yunnan Province, China (24° 94' 87" N, 101° 32' 24" E). The incidence of the disease was investigated to be about 15-30% in different varieties of maize planting areas in seven townships. Initial symptoms appeared as yellow to tan stripes, with the lesions measuring 3 to 48 mm in width and 45 to 560 mm in length. In severe cases, the lesions expanded, leading to complete leaf necrosis, which caused significant damage to maize production. Dark brown sub-epidermal pycnidia and conidia were observed on the necrotic leaf tissue. The pycnidia were globose or sac-like structures, with short and narrow conidiophores. The newly formed conidia are narrow, non-septate, and have transparent cytoplasm. The conidia are subcylindrical to narrowly ellipsoid, straight, curved, or occasionally irregular, measuring 38 to 86 × 7.2 to 12.3 µm. They are 0-3-septate, smooth-walled, pale brown, apex obtuse, base truncate. Therefore, 80 individual conidia were picked under a dissecting microscope, transferred to PDA plates containing antibiotics (10 mg/L rifampicin and ampicillin) followed by incubation at 28°C for 7 days (Fei et al., 2019). After seven days, the colonies of all isolates were turned grey and downy. Based on the maize leaf symptoms and the morphological characteristics of the pycnidia and conidia, the fungus was tentatively identified as Stenocarpella spp. (Luna et al., 2016). To confirm the identity of all isolates, DNA was extracted from 10 randomly selected isolates using the cetyl trimethyl ammonium bromide (CTAB) method. The internal transcribed spacer (ITS) region and the translation elongation factor 1α (TEF-1α) gene were amplified using the primer pairs ITS-1/ITS-4 and EF-1/EF-2, respectively, and then sequenced (Lamprecht et al., 2011). After comparison, the two target sequences of 10 isolates were consistent. BLASTn analysis revealed 99.27% and 100.00% homology with the two target sequences of Stenocarpella macrospora (syn. = Diplodia macrospora) (GQ259128.1, MG934504.1). Based on phylogenetic analysis of two targets, the isolate MS01 was clustered on the same evolutionary branch with S. macrospora. The two target sequences of isolate MS01 was submitted to GenBank under the accession numbers PQ241617.1 and PQ243292.1. To confirm the pathogenicity, a suspension (1 × 106 spores /mL) of isolate MS01 conidial was sprayed onto nine healthy leaves of maize plants in vitro and three pots cultivated with the plants (five plants per pot) in vivo. Plant treated with sterilized water served as control group. The treated plants were incubated at 28°C, 90% relative humidity with alternating light-dark cycles (i.e. 12 h shifts) for 3-10 days. At the same time, the conidial suspension was injected and inoculated onto maize ears, with water used as a blank control. Each treatment contained 3ears, 10 µL of the suspension was applied to each seed treatment. Inoculated leaves developed brown spots that gradually expanded into streak lesions extending toward the ends of the leaves, similar to those observed in the field. Additionally, the maize seeds become also moldy, whereas the control leaves and ears remained symptomless. The same pathogen was re-isolated from the infected leaves and ears, confirming Koch's postulates. Previously, S. macrospora and S. maydis have been reported in many countries across North, Central and South America, Africa, Asia, and Oceania, where maize is grown commercially. These pathogens are known to infect maize ears, stalks, and leaves of maize and also can produce mycotoxins (Romero et al., 2015; Snyman et al., 2011). To our knowledge, S. macrospora causing dry rot and ear rot of maize has been reported in China, but it is mostly found in quarantine at the time of importation (Tai et al., 1979; Shan et al., 2024). This is the first report of S. macrospora causing Diplodia leaf streak disease of maize in Yunnan Province, China. Since Stenocarpella spp. is one of the most destructive diseases of maize globally, the results of this study provide a foundation for developing effective prevention, control, and quarantine strategies for managing this disease.

Aloe genus plants are perennial evergreen herb belonging to Liliaceae family which is widely used in food, medicine, beauty, and health care (Kumar et al. 2019). In August 2021, symptoms of root and stem rot was observed in approximately 20% of Aloe vera plantings in Yuanjiang County, Yunnan Province, China (23° 64' 53" N, 101° 99' 84" E). The most typical symptoms were stem and root rot, browning and necrosis of vascular tissues, gradual greening, and reddish-browning of leaves from bottom to top, abscission, and eventual plant death (Fig. S1). Therefore, to isolate and identify the pathogen, the plants showing the above symptoms were collected. The plant tissues were cut from the edges of root and stem lesions, followed by disinfection with 75% ethanol for 1 min, rinsed three times with sterilized distilled water, and cut into 3 × 3 mm small squares after excision of marginal tissues. The tissues were transferred to the oomycetes selective medium (Liu et al. 2022) and incubated at 28 °C in the dark for 3~5 days, and suspected colonies were purified. The colonies were then inoculated onto potato dextrose agar (PDA), V8-juice agar (V8), and oatmeal agar (OA) medium plates for morphological characteristics. Finally, 18 isolates with the same colonial and morphological characteristics were obtained from 30 lesioned tissue and one of them was named as ARP1. On PDA, V8 and OA medium plates, the ARP1 colonies were white. On PDA plate, the mycelia were dense and the colonies were petal-like; on V8 plate, the mycelia were cashmere and the colonies were radial or star-like. Whereas, on OA plate, the mycelia were cotton-like and the colonies were fluffy and radial (Fig. S2 A~C). Mycelium did not have septum with high branching and swelling. Sporangia were abundant, semi-papillate, varying in shape from ovoid-ellipsoid to long-ellipsoid, 18-26 × 45-63 μm (average: 22 × 54 μm, n = 30), sporangia released numerous zoospores from the papillate after maturation. The chlamydospores were spherical, 20-35 μm in diameter (average: 27.5 μm, n = 30) (Fig. S2 D~F). These morphological features were like those of the pathogenic species of the oomycetes (Chen et al. 2022). For the molecular characterization, the genomic DNA of the isolate was extracted using the cetyl trimethyl ammonium bromide method, and the translation elongation factor 1α (tef-1α) (Stielow et al. 2015), β-tubulin (β-tub) (Kroon et al. 2004) and internal transcribed spacer (ITS) (White et al. 1990) of isolated strain ARP1 were amplified using primer pairs EF1-1018F/EF1-1620R, TUBUF2/TUBUR1 and ITS1/ITS4, respectively. The tef-1α, β-tub genes and ITS region of ARP1 were directly sequenced and their sequence information was deposited in GenBank under accession numbers OQ506129, OQ506127 and OQ449628. ARP1 was clustered on the same evolutionary branch with Phytophthora palmivora (Fig. S3). To confirm the pathogenicity of ARP1, the main root of A. vera was wounded to 1 cm long and 2 mm deep with a scalpel blade followed by inoculation with 50 ml suspension of ARP1 zoospores at a concentration of 1 × 106 spores / ml per potted plant, and an equal volume of water as control. All inoculated plants were placed in the greenhouse at 28°C, 12 h / 12 h light / dark. After 15 dpi, the inoculated plants showed typical symptoms of wilted and drooping leaves and stem and root rot, same as observed in the field condition (Fig. S4). After inoculation with ARP1, a strain with the same morphological and molecular characteristics as the original isolate was re-isolated, confirming Koch's postulates. To our knowledge, this is the first report of P. palmivora causing root and stem rot of A. vera in the study region. This disease could be a potential risk for aloe production and therefore appropriate management measures should be taken.

Soft rot caused by Pectobacterium carotovorum subsp. carotovorum (Pcc) is a major constraint in the production of Chinese cabbage. The objective of this study was to demonstrate that the causative agent Pcc may be successfully managed by Bacillus amyloliquefaciens KC-1, both in vitro and in vivo. Chinese cabbage seedlings were cultivated in organic substrate termed bio-organic substrate using a floating-seedling system with B. amyloliquefaciens KC-1. This approach was applied in a greenhouse to evaluate the management of soft rot. The results showed that the extent of soft rot, as well as the transmission of Pcc to the stem progeny and its survival in the rhizosphere, was reduced following inoculation with B. amyloliquefaciens KC-1. In contrast, the population diversity of B. amyloliquefaciens KC-1 persisted in the Chinese cabbage stems after germination. These findings revealed that B. amyloliquefaciens KC-1 was able to survive and suppress the growth of Pcc in Chinese cabbage and its rhizosphere, protecting the host from the pathogen. The use of B. amyloliquefaciens KC-1 throughout the growth period of plants may be an effective strategy for the prevention of soft rot in Chinese cabbage.