艾蒿是一种原产于东亚的多年生草本植物。它是一种重要的传统中药植物,以其浓郁的风味和药用作用而闻名。它含有丰富的活性成分,具有广泛的生物活性,包括消炎药,抗氧化剂,和免疫调节特性。2023年5月至7月,唐河县几个农场(约200亩)的艾吉发生了严重的叶腐病暴发(北纬32°46\'44\”,112°43'13''E),河南省,中国。发病率达到65%(n=200)。淡黄色斑点(直径1-2厘米)首先出现在叶子上,然后扩张形成不规则的黄褐色病变,最终导致整个叶子枯萎。收集患病的叶(30),并在感染和健康组织之间的区域中切成5×5mm2的片。将切下的植物组织在75%乙醇和1%次氯酸钠溶液中灭菌30秒零1分钟,分别。然后用无菌水冲洗组织并置于马铃薯葡萄糖琼脂(PDA)上,然后在25°C下孵育3天。分离的菌株属于镰刀菌属和链格孢属。致病性验证后,获得了25个纯化的镰刀菌菌株。三个代表性菌株(AC-Q,AC-X,来自不同地区的AC-Y)用于进一步研究。每个菌株都形成了丰富的气生菌丝体,最初是白色的,后来发展成紫色的色素。空中分生孢子稀疏分枝,以轮生phialides终止。大分生孢子纤细,直,测量为21.8至47.5×3.1至4.4μm,有两到四个隔片。微分生孢子为棒酸盐,测量值为8.31至11.6×2.1至3.5μm。形态特征与镰刀菌的物种描述一致(Sacc。)尼伦伯格1976年(莱斯利和萨姆梅雷尔,2006).rDNA内部转录间隔区(ITS),β-微管蛋白基因(tub2),翻译延伸因子1-α基因(tef1),calmodulin(cmdlet),扩增RNA聚合酶II最大亚基(rpb1)和RNA聚合酶II第二最大亚基(rpb2)以进行分子鉴定(O'Donnell等人。,2022年)。序列被保存在GenBank中,登录号为OR960548,OR960552,OR960555(ITS),OR972413,OR972414,OR972415(tub2),OR797685,OR797686,OR797687(tef1),OR972410,OR972411,OR972412(cmdlet),PP035106,PP035107,PP035108(rpb1),和PP035109、PP035110、PP035111(rpb2)。AC-Q序列的BLASTn分析显示出99%至100%的相似性与轮虫的序列(菌株CBS576.78)MT010888,rpb1的MT0109566和rpb2的MT010972。用串联序列构建了一个系统发育树(tub2,tef1,cmdlet,rpb1,rpb2),与使用邻居连接方法的类型菌株的序列一起。这三个菌株与轮虫的CBS576.78型菌株形成了进化枝,并与其他镰刀菌分离。这些形态学和分子鉴定表明该病原体为轮虫。通过用分生孢子悬浮液(106分生孢子ml-1)喷洒10个健康的2个月大的盆栽幼苗来测试致病性,和5个幼苗用灭菌水喷雾作为对照。将植物置于28°C和约90%的相对湿度的气候培养箱中。幼苗接种后十天,在处理过的植物上观察到典型的病变,除了对照组。通过形态学和分子鉴定,将重新分离的菌株鉴定为轮虫。履行科赫的假设。已知轮虫会导致玉米镰刀菌穗腐烂,以及中国其他植物的疾病,如芸苔属植物(Akram等人。,2020年)和荆棘(李等人。,2024).这是在世界范围内有关黄曲霉引起叶腐病的第一份报告。病原体的鉴定对于实施减少产量损失的管理方法至关重要。
Artemisia argyi is a perennial herb native to East Asia. It is an important traditional Chinese medicinal plant known for its strong flavor and medicinal effects. It is rich in active ingredients and has a wide range of biological activities, including anti-inflammatory, antioxidant, and immune regulation properties. From May to July in 2023, a serious leaf rot outbreak occurred on A. argyi in several farms (approximately 200 acres) in Tanghe county (32°46\'44\" N, 112°43\'13\" E), Henan Province, China. The incidence rate reached 65% (n=200). Pale yellow spots (1-2 cm in diameter) first appeared on the leaves, then expanded to form irregular yellowish-brown lesions, eventually causing the entire leaves to wither. Diseased leaves (30) were collected and cut into 5 x 5 mm2 pieces in the areas between infected and healthy tissues. The excised plant tissues were sterilized in 75% ethanol and 1% sodium hypochlorite solution for 30 seconds and one minute, respectively. The tissues were then rinsed with sterile water and placed on potato dextrose agar (PDA) followed by incubating at 25 °C for 3 days. The isolated strains belonged to the genera Fusarium and Alternaria. After pathogenicity verification, 25 purified Fusarium strains were obtained. Three representative strains (AC-Q, AC-X, AC-Y) from different regions were used for further studies. Each strain formed abundant aerial mycelium that was initially white and later developed into purple pigments. Aerial conidiophores were sparsely branched, terminating with verticillate phialides. Macroconidia were slender, straight, and measured 21.8 to 47.5 × 3.1 to 4.4 μm, with two to four septa. Microconidia were clavate and measured 8.31 to 11.6 × 2.1 to 3.5 μm. Morphological characteristics were consistent with the species description of Fusarium verticillioides (Sacc.) Nirenberg 1976 (Leslie and Summerell, 2006). The rDNA internal transcribed spacer (ITS), β-tubulin gene (tub2), translation elongation factor 1-alpha gene (tef1), calmodulin (cmdA), RNA polymerase II largest subunit (rpb1) and RNA polymerase II second largest subunit (rpb2) were amplified for molecular identification (O\'Donnell et al., 2022). The sequences were deposited in GenBank with accession Nos. OR960548, OR960552, OR960555 (ITS), OR972413, OR972414, OR972415 (tub2), OR797685, OR797686, OR797687 (tef1), OR972410, OR972411, OR972412 (cmdA), PP035106, PP035107, PP035108 (rpb1), and PP035109, PP035110, PP035111 (rpb2). BLASTn analysis of AC-Q sequences exhibited 99 to 100% similarity with F. verticillioides sequences (strains CBS 576.78) MT010888 of cmdA, MT0109566 of rpb1, and MT010972 of rpb2. A phylogenetic tree was constructed with concatenated sequences (tub2, tef1, cmdA, rpb1, rpb2), alongside the sequences of the type strains using the neighbor-joining method. The three strains formed a clade with the type strain CBS 576.78 of F. verticillioides, and were separated from
other Fusarium spp. These morphological and molecular identifications indicated that the pathogen was F. verticillioides. Pathogenicity was tested on 10 healthy 2-month-old potted seedlings by spraying them with a conidial suspension (106 conidia ml-1), and 5 seedlings were sprayed with sterilized water as a control. The plants were placed in a climate incubator at 28°C and a relative humidity of approximately 90%. Ten days after seedling inoculation, typical lesions were observed on the treated plants, except in the control group. The reisolated strains were identified as F. verticillioides by morphological and molecular characterization, fulfilling Koch\'s postulates. F. verticillioides is known to cause Fusarium ear rot on maize, as well as diseases on
other plants in China such as Brassica rapa (Akram et al., 2020) and Schizonepeta tenuifolia (Li et al., 2024). This is the first report of F. verticillioides causing leaf rot on A. argyi worldwide. Identification of the pathogen is crucial for implementing management approaches to reduce yield losses.