First Report of Leptosphaerulina Leaf Spot Caused by <i>Leptosphaerulina trifolii</i> on Alfalfa in Heilongjiang Province, China

Liu, Xiangping; Xia, Jing; Huang, Yan; Li, G. L.; Luo, Yiqi

doi:10.1094/pdis-04-19-0693-pdn

Cited by 4 publications

(6 citation statements)

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“…This finding is supported by evidence from observations of alfalfa plant symptoms, morphological identification, multigene phylogenies, and pathogenicity tests. This marks the first recorded instance of L. trifolii infecting alfalfa in these regions, which had not been documented in China before 2015 [21]. Therefore, Inner Mongolia, China is a new territory of Leptosphaerulina leaf spot of alfalfa.…”

Section: Discussionmentioning

confidence: 74%

“…Ascospores were oval or obovate-elliptic, with two to four horizontal septa and zero to two vertical septa, ranging in size from 18.90 to 40.47 × 8.06 to 17.54 µm. Based on morphological characteristics, the nine isolates were tentatively identified as L. trifolii [21,25,28,29]. These isolates thrived well when grown on V8J medium at 25 • C, as depicted in Figure 2c,d.…”

Section: Morphological Identification Of Isolatesmentioning

confidence: 97%

“…The morphology, color, ascomata, asci, and ascospores were observed, measured, and photographed across 20 replicates. Isolates were identified based on morphological characterization of L. trifolii [21,25,28,29].…”

Section: Morphological Identificationmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

“…Leptosphaerulina leaf spot on alfalfa can be caused by different Leptosphaerulina species, impacting both the alfalfa plant and the nutrient content of its leaves [18][19][20][21][22][23]. Liu et al [21] first reported Leptosphaerulina leaf spot caused by L. trifolii on alfalfa in Heilongjiang Province, China, in 2019. Furthermore, this pathogen has been found to infect other plants, including taro (Colocasia esculenta), various annual Medicago species, subterranean clover (Trifolium subterraneum), and white clover (Trifolium repens), confirming cross-infectivity between alfalfa and white clover [9,[24][25][26].…”

Section: Introductionmentioning

confidence: 99%

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Isolation, Identification and Characterization of Leptosphaerulina trifolii, the Causative Agent of Alfalfa Leptosphaerulina Leaf Spot in Inner Mongolia, China

Huo,

Huangfu,

Song

et al. 2024

Agronomy

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Leptosphaerulina leaf spot, caused by Leptosphaerulina trifolii, is a major disease of alfalfa (Medicago sativa), leading to noticeable losses. From 2022 to 2023, we collected samples of alfalfa with symptoms of the disease from different locations in Inner Mongolia, China. Nine fungal isolates recovered from these samples were identified through morphological traits and a maximum likelihood phylogeny based on concatenated partial sequences of ITS, 28S, and rpb2. A pathogenicity test on alfalfa confirmed the pathogenicity of the isolates on alfalfa. Analysis of physiological traits of L. trifolii revealed optimal mycelium growth at 20 °C and a pH range of 5 to 7, with soluble starch as the preferred carbon source and yeast extract as the optimal nitrogen source. The pathogen thrived in V8-juice agar and oat agar media. This study confirms L. trifolii as the causative agent of Leptosphaerulina leaf spot of alfalfa in Inner Mongolia and provides valuable insights into its optimal growth conditions. These findings enhance the understanding and management of this disease in alfalfa fields.

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Section: Discussionmentioning

confidence: 74%

Section: Morphological Identification Of Isolatesmentioning

confidence: 97%

Section: Morphological Identificationmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Isolation, Identification and Characterization of Leptosphaerulina trifolii, the Causative Agent of Alfalfa Leptosphaerulina Leaf Spot in Inner Mongolia, China

Huo,

Huangfu,

Song

et al. 2024

Agronomy

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First Report of Leptosphaerulina Leaf Spot Caused by Leptosphaerulina trifolii on Trifolium repens in Brazil

et al. 2020

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First Report of Alfalfa Leaf Spot Caused by Leptosphaerulina australis in China

Zhang

2021

Plant Disease

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A disease was observed on alfalfa cultivar WL168 characterized by white to brown leaf spots of regular to round shapes, in Aluhorqin County, Inner Mongolia, China (120°13′23″ to 120°29′14″ E, 43°27′52″to 43°35′16″ N, 281.71m to360.13 m Altitude) during 2019 to 2020. The disease mainly presented in spring one month after re-greening and the incidence was 78.30% in this field. Twenty alfalfa plants with severe symptoms were used for pathogen isolation. The infected tissue was cut into 2 × 2 mm pieces, surface-sterilized (in 75% ethanol and 5% commercial bleach (NaClO) for 30 s and 2 min, respectively), rinsed five times with sterilized distilled water, and dried between sterile filter paper (Wang et al. 2019). The diseased tissue from each plant sample were cultured on potato dextrose agar (PDA) and incubated at 25 °C with 12 h light/day for ten days. A fungus was isolated from the diseased leaves at a 100% frequency. Fungal growth on PDA was round with a black surface, radial edge, and a dirty white center. The ascocarps were moved to a clean microscope slide to release asci and ascospores. Ascocarps were spheroidal, subglobose brown, 120 to 160 µm × 160 to 180 µm, which contain several ascus. The size of ascus were 31.0 to 41.6 μm × 75.0 to 87.5 μm and each asci having eight ascospores. Ascospores were ellipsoid to oblong with a gelatinous sheath, brown, 8.8 to 15.0 µm × 29.9 to 43.0 µm with 2 to 3 horizontal septums, and 0 to 2 vertical septums. A phylogenetic tree was constructed after DNA extraction, PCR with primers to amplify the ITS (VG9: 5′- TTACGTCCCTGCCCTTTGTA-3′ and ITS4: 5′-TCCTCCGCTTATTGATATGC-3′) and LSU (LR7: 5′-TACTACCACCAAGATCT-3′ and LROR: 5′- GTACCCGCTGA ACTTAAGC -3′) regions. The LSU (SUB8273071) and ITS (SUB8218291) amplicons showed 99% similarity with L. australis (EU754166.1) in the GenBank. To verify the pathogenicity, fungs plugs were inverted on three compound leaves of 20 alfalfa WL168 for two days. Agar plugs (PDA) were inverted on another 20 alfalfa WL168 three compound leaves which were control. All plants were maintained at 22 °C and 44% relative humidity in a growth chamber. Similar disease symptoms were observed on infected leaves ten days after inoculation, while control plants showed no symptoms. The same fungus was re-isolated from the lesions, and further morphological characterization and molecular assays, as described above. L. australis has been reported on various plants, including Prunusarmeniaca, Dolichos, Poa, Lolium, and Vitis in Australia (Graham and Luttrell., 1961), and also from Korean soil in 2018 (Weilan et al., 2018). Additionally, L. briosiana, which is common in the USA, China, and other countries, causes Leptosphaerulina leaf spot (Samacet al., 2015). L. trifolii is newly reported to occur in China (Liu et al., 2019). To the best of our knowledge, this is the first report of L. australis infecting alfalfa in China. Considering the large planting area in Inner Mongolia, this pathogen may losses to alfalfa cultivation. Hence, future studies should explore aspects of effective management of this disease.

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First Report of Leptosphaerulina Leaf Spot Caused by Leptosphaerulina trifolii on Alfalfa in Heilongjiang Province, China

Cited by 4 publications

References 6 publications

Isolation, Identification and Characterization of Leptosphaerulina trifolii, the Causative Agent of Alfalfa Leptosphaerulina Leaf Spot in Inner Mongolia, China

Isolation, Identification and Characterization of Leptosphaerulina trifolii, the Causative Agent of Alfalfa Leptosphaerulina Leaf Spot in Inner Mongolia, China

First Report of Leptosphaerulina Leaf Spot Caused by Leptosphaerulina trifolii on Trifolium repens in Brazil

First Report of Alfalfa Leaf Spot Caused by Leptosphaerulina australis in China

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