中国首次报道玫瑰毛霉引起茶叶腐烂。

IF 4.4 2区 农林科学 Q1 PLANT SCIENCES Plant disease Pub Date : 2024-10-11 DOI:10.1094/PDIS-07-24-1419-PDN
Jinmei Lei, Bingbing Jiang, Fang Yang, Cunwu Guo, Xiao Fang He, Li Mei Li, Shihao Zhang, Ya Min Wu, Chun Yan Liu, Li Jiao Chen, Bai Juan Wang
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The pieces were placed on acidified potato dextrose agar (PDA) plates and incubated in darkness at 28°C. A total of 15 fungal isolates with identical morphologies were collected. The colonies appeared pale pink with white mycelia initially then turned orange-pink at the center and light white at the edges. After 10-15 days, exhibiting a powdery texture and concentric rings with uniform edges. Conidia were found at the apex peduncle and were inverted pear-shaped or oval, either non-septate (15.3 ± 2 × 7.8 ± 1.8 μm in size, n = 60) or septate, with a slightly constricted spore base featuring papillary projecvtions (14.8 ± 1.5 × 7.4 ± 0.7 μm in size, n = 60). The morphology closely resembled Trichoderma roseum (Oh et al. 2014). To confirm the species, the strain CYB5 was selected for identification by sequencing the ribosomal internal transcribed spacer (ITS) and large subunit (LSU) genes using polymerase chain reaction (PCR) (White et al.1990). 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引用次数: 0

摘要

茶叶(Camellia sinensis)因其诸多益处而被全世界消费,中国的茶叶产量居世界首位。2023 年 3 月,在中国云南勐海(北纬 21°46'13",东经 100°30'6")一个 50 公顷的商业茶园中,约 10% 的茶树出现叶斑病。最初的症状表现为小斑点,随后逐渐扩大并遍布整个叶面。随后,病斑上出现淡粉色霉层(图 S1)。为了分离病原体,从病斑边缘剪下小叶片(3 × 3 毫米),用 75% 的乙醇消毒 30 秒,再用 0.5% 的 NaClO 消毒 30 秒,然后用无菌水冲洗三次。将切片放在酸化的马铃薯葡萄糖琼脂(PDA)平板上,28°C 黑暗培养。共收集到 15 个形态相同的真菌分离物。菌落最初呈淡粉色,带有白色菌丝,然后中心变为橙粉色,边缘变为浅白色。10-15 天后,菌落呈现粉状质地和边缘一致的同心环。分生孢子出现在先端花序梗上,呈倒梨形或椭圆形,要么无隔膜(大小为 15.3 ± 2 × 7.8 ± 1.8 μm,n = 60),要么有隔膜,孢子基部略微收缩,具有乳头状突起(大小为 14.8 ± 1.5 × 7.4 ± 0.7 μm,n = 60)。其形态与玫瑰毛霉非常相似(Oh 等人,2014 年)。为确认菌种,利用聚合酶链式反应(PCR)(White et al.1990)对核糖体内部转录间隔(ITS)和大亚基(LSU)基因进行测序,选择了菌株 CYB5 进行鉴定。ITS(GenBank登录号OR889657)和LSU(PQ270526)基因序列与NCBI数据库中玫瑰毛霉序列KP317992的相似度为98%。根据菌株 CYB5 和参考菌株的序列(ITS 和 LSU),使用 MEGA 11(Felsenstein,1981 年)构建了一棵系统发生树(图 S2)。分析结果证实 CYB5 是 T. roseum(Inácio 等人,2011 年)。致病性试验在培养皿中置于湿滤纸上的健康茶叶上进行。用灭菌针在叶片上刺出微小伤口,然后接种 6 毫米的 CYB5 栓塞。对照叶片接种无真菌的琼脂盘。培养皿在 25°C 黑暗中培养 7 天。接种了 CYB5 的叶片在接种部位周围出现了红褐色至黑褐色的病斑,而对照叶片仍无症状。从病变处重新分离出真菌,分离物在形态上与原始培养物完全相同。第二次致病性试验是在栽培品种为 "云冈 10 号 "的盆栽茶树上进行的。用 20 毫升 CYB5 孢子悬浮液(105 个孢子/毫升)喷洒三株用针划伤的植株和三株未受伤的植株。喷洒无菌水的植物作为对照。所有植物均在 28°C 和 70% 相对湿度下的生长室中生长。病斑在接种后 3 天出现,10 天后仅在孢子接种的植株上出现典型症状。根据形态学和分子分析,我们重新分离并鉴定了蔷薇真菌,从而实现了科赫假说。据我们所知,这是中国首次报道 T. roseum 引起茶叶腐烂病。
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First report of Trichoderma roseum causing tea leaf rot in China.

Tea (Camellia sinensis) is consumed worldwide for its numerous benefits and China lead the world production. In March 2023, leaf spots were observed on approximately 10% of tea plants in a 50-ha commercial tea plantation in Menghai (21°46'13"N, 100°30'6"E), Yunnan, China. Initial symptoms appeared as small spots, which progressively expanded and spread over the entire leaf surface. Subsequently, pale pink mold layers developed from the lesions (Fig. S1). To isolate the pathogen, small leaf pieces (3 × 3 mm) were cut from the margins of the lesions, sterilized with 75% ethanol for 30 sec and 0.5% NaClO for another 30 sec, and rinsed three times with sterile water. The pieces were placed on acidified potato dextrose agar (PDA) plates and incubated in darkness at 28°C. A total of 15 fungal isolates with identical morphologies were collected. The colonies appeared pale pink with white mycelia initially then turned orange-pink at the center and light white at the edges. After 10-15 days, exhibiting a powdery texture and concentric rings with uniform edges. Conidia were found at the apex peduncle and were inverted pear-shaped or oval, either non-septate (15.3 ± 2 × 7.8 ± 1.8 μm in size, n = 60) or septate, with a slightly constricted spore base featuring papillary projecvtions (14.8 ± 1.5 × 7.4 ± 0.7 μm in size, n = 60). The morphology closely resembled Trichoderma roseum (Oh et al. 2014). To confirm the species, the strain CYB5 was selected for identification by sequencing the ribosomal internal transcribed spacer (ITS) and large subunit (LSU) genes using polymerase chain reaction (PCR) (White et al.1990). The ITS (GenBank accession OR889657) and LSU (PQ270526) gene sequences exhibited 98% similarity with the Trichoderma roseum sequence KP317992 from NCBI database. A phylogenetic tree was constructed using MEGA 11 (Felsenstein 1981) based on the concatenated sequences (ITS and LSU) of the strain CYB5 and reference strains (Fig. S2). The analysis confirmed that CYB5 is T. roseum (Inácio et al. 2011). Pathogenicity tests were conducted on detached healthy tea leaves placed on wet filter paper in petri dishes. Micro-wounds were made on leaves using a sterilized needle, followed by inoculation with a 6-mm plug of CYB5. Control leaves were inoculates with fungus-free agar disks. The dishes were incubated at 25°C in the dark for 7 days. The leaves inoculated with CYB5 developed reddish brown to dark brown lesions around the inoculated sites, while control leaves remained asymptomatic. The fungus was reisolated from the lesion, and the isolates were morphologically identical to the original cultures. A second pathogenicity test was conducted on potted tea plants of the cultivar 'Yunkang No. 10.' Three plants scratched with a needle and three non-wounded plants were inoculated by spraying 20 ml of a spore suspension (105 spores/ml) of CYB5. Plants sprayed with sterile water served as controls. All plants were maintained in a growth chamber at 28°C, and 70% relative humidity. The lesions developed three days post-inoculation, and typical symptoms appeared after 10 days on spore-inoculated plants only. T. roseum was reisolated and reidentified based on the morphology and molecular analyses, thus fulfilling Koch's postulates. To our best knowledge, this is the first report of T. roseum causing tea leaf rot in China.

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来源期刊
Plant disease
Plant disease 农林科学-植物科学
CiteScore
5.10
自引率
13.30%
发文量
1993
审稿时长
2 months
期刊介绍: Plant Disease is the leading international journal for rapid reporting of research on new, emerging, and established plant diseases. The journal publishes papers that describe basic and applied research focusing on practical aspects of disease diagnosis, development, and management.
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