The present study was aimed at isolating endophytic fungi from the Asian culinary and medicinal plant Lilium davidii and analyzing its antifungal and plant growth‐promoting effects. In this study, the fungal endophyte Acremonium sp. Ld‐03 was isolated from the bulbs of L. davidii and identified through morphological and molecular analysis. The molecular and morphological analysis confirmed the endophytic fungal strain as Acremonium sp. Ld‐03. Antifungal effects of Ld‐03 were observed against Fusarium oxysporum, Botrytis cinerea, Botryosphaeria dothidea, and Fusarium fujikuroi. The highest growth inhibition, i.e., 78.39 ± 4.21%, was observed for B. dothidea followed by 56.68 ± 4.38%, 43.62 ± 3.81%, and 20.12 ± 2.45% for B. cinerea, F. fujikuroi, and F. oxysporum, respectively. Analysis of the ethyl acetate fraction through UHPLC‐LTQ‐IT‐MS/MS revealed putative secondary metabolites which included xanthurenic acid, valyl aspartic acid, gancidin W, peptides, and cyclic dipeptides such as valylarginine, cyclo‐[L‐(4‐hydroxy‐Pro)‐L‐leu], cyclo(Pro‐Phe), and (3S,6S)‐3‐benzyl‐6‐(4‐hydroxybenzyl)piperazine‐2,5‐dione. Other metabolites included (S)‐3‐(4‐hydroxyphenyl)‐2‐((S)‐pyrrolidine‐2‐carboxamido)propanoic acid, dibutyl phthalate (DBP), 9‐octadecenamide, D‐erythro‐C18‐Sphingosine, N‐palmitoyl sphinganine, and hydroxypalmitoyl sphinganine. The strain Ld‐03 showed indole acetic acid (IAA) production with or without the application of exogenous tryptophan. The IAA ranged from 53.12 ± 3.20 μg ml^-1 to 167.71 ± 7.12 μg ml^-1 under different tryptophan concentrations. The strain was able to produce siderophore, and its production was significantly decreased with increasing Fe(III) citrate concentrations in the medium. The endophytic fungal strain also showed production of organic acids and phosphate solubilization activity. Plant growth‐promoting effects of the strain were evaluated on in vitro seedling growth of Allium tuberosum. Application of 40% culture dilution resulted in a significant increase in root and shoot length, i.e., 24.03 ± 2.71 mm and 37.27 ± 1.86 mm, respectively, compared to nontreated control plants. The fungal endophyte Ld‐03 demonstrated the potential of conferring disease resistance and plant growth promotion. Therefore, we conclude that the isolated Acremonium sp. Ld‐03 should be further investigated before utilization as a biocontrol agent and plant growth stimulator.

Lily (Lilium spp.) is a popular ornamental plant. Traditional genetic transformation methods have low efficiency in lily, thus development of a high-efficiency genetic transformation system is important. In this study, a novel transient transformation method involving pollen magnetofection was established and optimized pollen viability, and exogenous gene expression in magnetofected pollen and that of different germplasm were assessed. The highest germination percentage of Lilium regale pollen was 85.73% in medium containing 100 g/L sucrose, 61.5 mg/L H₃BO₃, and 91.5 mg/L CaCl₂. A 1:4 ratio of nanomagnetic beads to DNA plasmid and transformation time of 0.5 h realized the highest transformation efficiency (88.32%). The GFP activity in transformed pollen averaged 69.66%, while that of the control pollen was 0.00%. In contrast to the control, transgenic seedlings obtained by pollination with magnetofected pollen showed strong positive GUS activity with 56.34% transformation efficiency. Among the lily germplasm tested, ‘Sweet Surrender’ and L. leucanthum had the highest transformation efficiency (85.80% and 54.47%), whereas L. davidii var. willmottiae was not successfully transformed. Transformation efficiency was positively correlated with pollen equatorial diameter and negatively correlated with polar axis/equatorial diameter ratio. The results suggest that pollen magnetofection-mediated transformation can be applied in Lilium but might have species or cultivar specificity.

Lily bulbils, which serve as advantageous axillary organs for vegetative propagation, have not been extensively studied in terms of the mechanism of bulbil initiation. The functions of auxin and sucrose metabolism have been implicated in axillary organ development, but their relationship in regulating bulbil initiation remains unclear. In this study, exogenous indole-3-acetic acid (IAA) treatment increased the endogenous auxin levels at leaf axils and significantly decreased bulbil number, whereas treatment with the auxin polar transport inhibitor N-1-naphthylphthalamic acid (NPA), which resulted in a low auxin concentration at leaf axils, stimulated bulbil initiation and increased bulbil number. A low level of auxin caused by NPA spraying or silencing of auxin biosynthesis genes YUCCA FLAVIN MONOOXYGENASE-LIKE 6 (LlYUC6) and TRYPTOPHAN AMINOTRANSFERASERELATED 1 (LlTAR1) facilitated sucrose metabolism by activating the expression of SUCROSE SYNTHASES 1 (LlSusy1) and CELL WALL INVERTASE 2 (LlCWIN2), resulting in enhanced bulbil initiation. Silencing LlSusy1 or LlCWIN2 hindered bulbil initiation. Moreover, the transcription factor BASIC HELIX-LOOP-HELIX 35 (LlbHLH35) directly bound the promoter of LlSusy1, but not the promoter of LlCWIN2, and activated its transcription in response to the auxin content, bridging the gap between auxin and sucrose metabolism. In conclusion, our results reveal that an LlbHLH35-LlSusy1 module mediates auxin-regulated sucrose metabolism during bulbil initiation.

Dynamic miRNA detection using the qRT-PCR technique requires appropriate reference genes to ensure data reliability. Previous studies have screened internal reference genes in plants during embryonic development and various stress treatment, involving relatively few tissues and organs. There is no relevant miRNA study in Lilium henryi Baker and limited research on the optimal miRNA reference genes in lilies, such as 5S, 18S, U6 and Actin. Twelve genes were selected as candidate reference genes whose expression stability was analyzed in petals at different developmental stages and other tissues using various algorithms, such as geNorm, NormFinder, BestKeeper, and Delta CT. The results revealed that the optimal combination of reference genes for Lilium henryi Baker petals at different developmental stages was osa-miR166m and osa-miR166a-3p, while that for different tissues of Lilium henryi Baker was osa-miR166g-3p and osa-miR166a-3p.Four important genes related to growth and development regulation, namely, osa-miR156a, osa-miR395b, osa-miR396a-3p, and osa-miR396a-5p, were selected for validation. The findings of the present study could contribute to future investigations onmiRNA expression and the related functions in Lilium henryi Baker while providing important references for the normalization of the miRNA expression in other varieties of lily.

Lilium davidii var. willmottiae, known as Lanzhou lily, is a famous edible crop that is mostly distributed in the middle area of Gansu Province in China. In the winter of 2019, symptoms of bulb rot were observed on Lanzhou lilies harvested from Lanzhou, Gansu Province, during storage at the Institute of Grassland, Flowers and Ecology (39°57'55.984" N, 116°20'8.124" E), Beijing Academy of Agriculture and Forestry Sciences, at an incidence of nearly 50%. The decayed bulb (Fig.1a)was washed under tap water and surface disinfested with 75% ethanol for 1 min, followed by 2.5% sodium hypochlorite for 5 min, and washed with sterile distilled water three times. The 5 mm×5 mm tissue pieces from the junction of the diseased part and the healthy part were clipped, placed on potato dextrose agar (PDA) medium and subsequently incubated at 25 °C. Thirteen dominant pure fungal isolates with the same morphological characteristics were obtained by the hyphal-tip method. Three representative isolates LZ-8, LZ-9-2 and LZ-10 were chosen for phylogenetic analyses. The internal transcribed spacer (ITS), translation elongation factor 1-alpha (TEF-1a), and RNA polymerase II second largest subunit (RPB2) sequences were PCR amplified using the primer pairs ITS1/ITS4 (White et al. 1990), EF1-728F/EF1-986R (Carbone and Kohn 1999), and RPB2-5F2/RPB2-7cR (O'Donnell et al. 2022), respectively. BLAST analysis showed that the ITS,TEF-1a, and RPB2 sequences of the isolates LZ-8 (GenBank accession nos. PP422096, PP447248, and PP447251), LZ-9-2 (GenBank accession nos. PP422098, PP447249, and PP447252) and LZ-10 (GenBank accession nos. PP422099, PP447250, and PP447253) had 99.27 to 99.71% identity with multiple GenBank sequences of Trichoderma hamatum, and the three DNA fragments of the three isolates showed 100% sequence identity. A phylogenetic tree based on concatenated sequences of the three genes using maximum -likelihood analyses revealed that the three isolates LZ-8, LZ-9-2 and LZ-10 were in the same clade with T. hamatum strains (Fig.2). One representative isolate, LZ-10, was chosen for morphological studies and test of the pathogenicity. The colony of LZ-10 on PDA appeared white with cotton-shaped aerial hyphae early, which later turned light green to green and formed concentric rings (Fig.1d-1f). At the end of conidiophores, three to six pear-shaped branches were irregularly gathered(Fig.1h). Conidia were ellipsoid with the size of 3.1 to 4.4 × 2.2 to 3.1 µm (n =20) (Fig.1g). These morphological characteristics were consistent with the description of Trichoderma hamatum. (Kamala et al. 2015, Han et al. 2017).To test pathogenicity, healthy bulbs were punctured with disposable sterilized needles and soaked in equal amounts of sterile water and conidial suspension (1×107 conidia/mL) for 30 min respectively. The pathogenicity experiment was repeated three times. After 6 days of inoculation at 25 °C and 80% relative humidity, the surface of the inoculated bulbs produced water-stained spots and mycelium layers(Fig.1b-1c) consistent with the symptoms exhibited by Lilium davidii var. willmottiae bulbs during storage, meanwhile the uninoculated lily bulbs remained symptomless. Trichoderma hamatum was reisolated from the infected bulbs and identified based on morphological and molecular characteristics, fulfilling Koch's postulates. To our knowledge, this is the first report of bulb rot on Lilium davidii var. willmottiae caused by Trichoderma hamatum in China. This study will contribute to a better understanding and controlling of this postharvest disease in Lilium davidii var. willmottiae.