Liu Qiuping, Wang Renfen, Song Qin, Wang Jiahua, Shi Yiming, Ding Wei, Tan Leitao
Guizhou Normal University, 12686, School of Life Sciences, Guiyang, Guizhou, China;
Southwest University, 26463, College of Plant Protection, Chongqing, China;
Plant Dis. 2022 Mar 12. doi: 10.1094/PDIS-12-21-2629-PDN.
is a plant with colorful leaves that is widely used in landscaping. Due to its shade tolerance and cold resistance (Li et al. 2013), has become a dominant plant species in Guiyang city, China. From 2018 to 2021, an anthracnose disease was observed on in Huaxi District (26°23'03'' N, 106°37'58'' E), Guiyang, Guizhou Province. The incidence of anthracnose in newly planted reached 90%, resulting in a mortality rate of 30%. This has become a major disease for in Guiyang. Typical symptoms include faded spots that initially appear at the infection center, followed by black-brown lesions with irregular edges. At the latter stage, slightly raised black spots are seen arranged in a wheel pattern. In severe cases, diseased leaves fall off. To identify the pathogen, leaf pieces (5 mm × 3 mm) containing symptomatic tissue with healthy margins were excised. The pieces were sterilized with 75% ethanol for 1 min, rinsed three times with sterile water, and cultured on potato dextrose agar (PDA) in Petri dishes at 26°C. White to cream colonies were developed after growth on PDA for 2 days. Mycelial growth ranged from 4.8 to 5.0 mm/day. Conidia were cylindrical, obtuse apex, and protruding base, with dimensions of 18.5 to 20.0 × 7.0 to 9.0 μm (n = 20). Conidial length:breadth ratio was 2-3 (n = 20). Acervuli and sclerotia had not been seen on PDA. To confirm the species of the isolate, PCR was performed on extracted DNA to amplify the ribosomal DNA internal transcribed spacer (ITS), a 200-bp intron of the glyceraldehyde-3-phosphate dehydrogenase (GAPDH), a partial sequence of the calmodulin (CAL) gene, and the large subunit (LSU) region by using the primer pairs ITS1/ITS4 (Schoch et al. 2012), GDF1/GDR1 (Guerber et al. 2003), CAL228F/CAL737R (Carbone & Kohn 1999), and NL1/NL4 (Ziemiecki et al. 1990), respectively. The obtained 553 bp ITS sequence (Genbank accession no. MZ424503) showed 99.82% similarity with several sequences (MN429163, MN542218, KM520024, and KM520014). The obtained 233 bp GAPDH sequence (OM643394) showed 99.07% similarity with (MT602277, GQ221769, MK544890, and MK544888). The obtained 763 bp CAL sequence (OM313386) showed 99.32% similarity with (MK569127). The obtained 584 bp LSU sequence (OK513272) showed 99.83% similarity with (MH876452, MH877053, and MH876450). Phylogenic trees based on the sequences of ITS, GAPDH, and CAL (Damm et al. 2012), the species of isolate was confirmed as . To confirm pathogenicity and to fulfill Koch's postulates, leaves of five plants were wounded with a sterilized hypodermic needle and inoculated with 1 μL of a conidial suspension (106 conidia/mL). As control, 5 plants were inoculated with 1 μL of sterile water. After 7 days in a moist climate chamber at 26°C under a 16 h/8 h light/dark cycle, symptoms appeared on all inoculated leaves, while the control leaves remained healthy. The pathogen was reisolated from the inoculated leaves, and was confirmed to be using the same morphological and molecular methods as before. has been reported to infect many plant species, including , , and (Ding et al. 2021; Tozze et al. 2009; Zhang et al. 2018). Another pathogen, , was reported to cause anthracnose on in China (Gong et al. 2016). To our knowledge, this is the first report of anthracnose disease caused by in Guizhou province, China. Identifying this pathogen provides a foundation to prevent this complex disease and to reduce economic loss.
是一种叶片色彩丰富的植物,广泛用于园林绿化。由于其耐荫性和抗寒性(Li等人,2013年),已成为中国贵阳市的优势植物物种。2018年至2021年期间,在贵州省贵阳市花溪区(北纬26°23′03″,东经106°37′58″)观察到上发生炭疽病。新种植的上炭疽病发病率达到90%,导致死亡率为30%。这已成为贵阳市上的一种主要病害。典型症状包括最初出现在感染中心的褪色斑点,随后是边缘不规则的黑褐色病斑。在后期,可见略微凸起的黑点呈轮状排列。严重时,病叶脱落。为了鉴定病原菌,切取含有症状组织且边缘健康的叶片小块(5毫米×3毫米)。将小块用75%乙醇消毒1分钟,用无菌水冲洗三次,并在26℃下在培养皿中的马铃薯葡萄糖琼脂(PDA)上培养。在PDA上生长2天后形成白色至奶油色菌落。菌丝生长速度为每天4.8至5.0毫米。分生孢子呈圆柱形,顶端钝圆,基部突出,尺寸为18.5至20.0×7.0至9.0微米(n = 20)。分生孢子长度与宽度之比为2 - 3(n = 20)。在PDA上未观察到分生孢子盘和菌核。为了确认分离株的种类,对提取的DNA进行PCR,以扩增核糖体DNA内部转录间隔区(ITS)、甘油醛-3-磷酸脱氢酶(GAPDH)的200碱基对内含子、钙调蛋白(CAL)基因的部分序列以及大亚基(LSU)区域,分别使用引物对ITS1/ITS4(Schoch等人,2012年)、GDF1/GDR1(Guerber等人,2003年)、CAL228F/CAL737R(Carbone和Kohn,1999年)以及NL1/NL4(Ziemiecki等人,1990年)。获得的553碱基对ITS序列(Genbank登录号MZ424503)与几个序列(MN429163、MN542218、KM520024和KM520014)显示出99.82%的相似性。获得的233碱基对GAPDH序列(OM643394)与(MT602277、GQ221769、MK544890和MK544888)显示出99.07%的相似性。获得的763碱基对CAL序列(OM313386)与(MK569127)显示出99.32%的相似性。获得的584碱基对LSU序列(OK513272)与(MH876,452、MH877053和MH876450)显示出99.83%的相似性。基于ITS、GAPDH和CAL序列构建的系统发育树(Damm等人,2012年),确认分离株的种类为。为了确认致病性并满足科赫法则,用无菌皮下注射针划伤五株植物的叶片,并接种1微升分生孢子悬浮液(10^6个分生孢子/毫升)。作为对照,给5株植物接种1微升无菌水。在26℃、16小时光照/8小时黑暗周期的潮湿气候箱中培养7天后,所有接种叶片均出现症状,而对照叶片保持健康。从接种叶片中重新分离出病原菌,并使用与之前相同的形态学和分子方法确认其为。据报道,可感染许多植物物种,包括、和(Ding等人,2021年;Tozze等人,2009年;Zhang等人,2018年)。另一种病原菌,据报道在中国导致上的炭疽病(Gong等人,2016年)。据我们所知,这是中国贵州省由引起的炭疽病的首次报道。鉴定这种病原菌为预防这种复杂病害和减少经济损失提供了基础。