Toporek Sean M, Keinath Anthony P
Clemson University College of Agriculture Forestry and Life Sciences, 114625, Plant and Environmental Sciences, 2700 Savannah Hwy, Charleston, Charleston, South Carolina, United States, 29412;
Clemson University, Coastal REC, 2700 Savannah Hwy, Charleston, South Carolina, United States, 29414-5329;
Plant Dis. 2020 Nov 23. doi: 10.1094/PDIS-08-20-1656-PDN.
Anthracnose fruit rot caused by various Colletotrichum spp. is a serious disease for pepper (Capsicum annuum) growers, resulting in extensive fruit loss (Harp et al. 2008). Samples of five pepper fruits were obtained from two commercial farms in Lexington and Pickens counties, South Carolina, in August and September 2019, respectively. All fruits had two or more soft, sunken lesions covered with salmon-colored spore masses. Pieces of diseased tissue cut from the margins of lesions were surface disinfested in 0.6% sodium hypochlorite, rinsed in sterile deionized water, blotted dry, and placed on one-quarter-strength potato dextrose agar (PDA/4) amended with 100 mg chloramphenicol, 100 mg streptomycin sulfate, and 60.5 mg mefenoxam (0.25 ml Ridomil Gold EC) per liter. Two isolates of Colletotrichum sp. per fruit were preserved on dried filter paper and stored at 10º C. One additional isolate of Colletotrichum sp. had been collected from a jalapeño pepper fruit on a farm in Charleston County, South Carolina, in 1997. Colony morphology of three isolates, one per county, on Spezieller Nährstoffarmer Agar (SNA) was pale grey with a faint orange tint. All isolates readily produced conidia on SNA with an average length of 16.4 μm (std. dev. = 1.8 μm) and a width of 2.2 μm (std. dev. = 0.2 μm). Conidia were hyaline, smooth, straight, aseptate, cylindrical to fusiform with one or both ends slightly acute or round, matching the description of C. scovillei (Damm et al. 2012). The glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and beta-tubulin (TUB2) genes from three isolates were amplified and sequenced with the primer pairs GDF1/GDR1 and T1/Bt2b, respectively. Species within the C. acutatum clade can be readily distinguished with GAPDH or TUB2 (Cannon et al. 2012). The GAPDH and TUB2 sequences for all three isolates were 100% similar to each other and strain CBS 126529 (GAPDH accession number JQ948597; TUB2 accession number JQ949918) of C. scovillei (Damm et al. 2012). GAPDH and TUB2 sequences for each isolate were deposited in GenBank under the accessions MT826948-MT826950 and MT826951-MT826953, respectively. A pathogenicity test was conducted on jalapeño pepper fruits by placing a 10-ul droplet of a 5 x 105 conidial suspension of each isolate onto a wound made with a sterile toothpick. Control peppers were mock inoculated with 10 ul sterile distilled water. A humid chamber was prepared by placing moist paper towels on the bottom of a sealed crisper box. Inoculated peppers were placed on upside-down 60 ml plastic condiment cups. Three replicate boxes each containing all four treatments were prepared. The experiment was repeated once. After 7 days in the humid chamber at 26ºC, disease did not develop on control fruits, whereas soft, sunken lesions covered with salmon-colored spores developed on inoculated fruits. Lesions were measured and C. scovillei was re-isolated onto amended PDA/4 as previously described. Lesion length averaged 15.6 mm (std dev. = 4.1 mm) by 11.5 mm (std dev. = 2.0 mm). Colletotrichum sp. resembling the original isolate were recovered from all inoculated fruit, but not from non-inoculated fruit. C. scovillei has been reported in Brazil in South America and in China, Indonesia, Japan, Malaysia, South Korea, Taiwan, and Thailand in Asia (Farr and Rossman 2020). This is the first report of C. scovillei as the casual organism of anthracnose fruit rot on pepper in South Carolina and the United States.
由多种炭疽菌属(Colletotrichum spp.)引起的炭疽病果腐病是辣椒(Capsicum annuum)种植者面临的一种严重病害,会导致大量果实损失(Harp等人,2008年)。2019年8月和9月,分别从南卡罗来纳州列克星敦县和皮肯斯县的两个商业农场采集了五个辣椒果实样本。所有果实都有两个或更多柔软、凹陷的病斑,上面覆盖着鲑鱼色的孢子团。从病斑边缘切下的病组织块在0.6%次氯酸钠中进行表面消毒,用无菌去离子水冲洗,吸干水分,然后放在每升添加了100毫克氯霉素、100毫克硫酸链霉素和60.5毫克甲霜灵(0.25毫升金雷多米尔乳油)的四分之一强度马铃薯葡萄糖琼脂(PDA/4)上。每个果实的两个炭疽菌属分离株保存在干燥滤纸上,并在10℃下储存。1997年从南卡罗来纳州查尔斯顿县一个农场的墨西哥胡椒果实上采集了另外一个炭疽菌属分离株。在特殊营养琼脂(SNA)上,来自三个县的各一个分离株的菌落形态为浅灰色,带有微弱的橙色色调。所有分离株在SNA上都能轻易产生分生孢子,平均长度为16.4微米(标准差 = 1.8微米),宽度为2.2微米(标准差 = 0.2微米)。分生孢子无色、光滑、直、无隔膜,圆柱形至梭形,一端或两端稍尖或圆形,与斯氏炭疽菌(C. scovillei)的描述相符(Damm等人,2012年)。分别用引物对GDF1/GDR1和T1/Bt2b对三个分离株的甘油醛-3-磷酸脱氢酶(GAPDH)和β-微管蛋白(TUB2)基因进行扩增和测序。尖孢炭疽菌(C. acutatum)分支内的物种可以通过GAPDH或TUB2轻易区分(Cannon等人,2012年)。所有三个分离株的GAPDH和TUB2序列彼此100%相似,并且与斯氏炭疽菌的菌株CBS 126529(GAPDH登录号JQ948597;TUB2登录号JQ949918)相似(Damm等人,2012年)。每个分离株的GAPDH和TUB2序列分别以登录号MT826948 - MT826950和MT826951 - MT826953保存在GenBank中。通过将每个分离株的5×105分生孢子悬浮液的10微升液滴滴在无菌牙签造成的伤口上,对墨西哥胡椒果实进行致病性测试。对照辣椒用10微升无菌蒸馏水进行模拟接种。通过在密封保鲜盒底部放置湿纸巾来制备湿度箱。接种的辣椒放在倒置的60毫升塑料调味品杯上。制备了三个重复的盒子,每个盒子包含所有四种处理。该实验重复一次。在26℃的湿度箱中放置7天后,对照果实上没有发病,而接种果实上出现了覆盖着鲑鱼色孢子的柔软、凹陷病斑。测量病斑长度,并且如前所述将斯氏炭疽菌重新分离到改良的PDA/4上。病斑长度平均为15.6毫米(标准差 = 4.1毫米)×11.5毫米(标准差 = 2.0毫米)。从所有接种果实中都分离出了与原始分离株相似的炭疽菌属,但未从未接种果实中分离出。在南美洲的巴西以及亚洲的中国、印度尼西亚、日本、马来西亚、韩国、台湾和泰国都报道过斯氏炭疽菌(Farr和Rossman,2020年)。这是斯氏炭疽菌作为南卡罗来纳州和美国辣椒炭疽病果腐病致病病原体的首次报道。