番石榴(PsidiumguajavaL.)是一种流行的水果作物,在泰国广泛种植。2023年11月,在22至31°C和70至75%相对湿度的采后储存期间,在方区3至7天的时间内观察到番石榴上的褐斑病,清迈省,泰国。每个托盘箱100个水果的发病率约为20%。每个水果的疾病严重程度为受病变影响的表面积的40%至70%。症状表现为圆形至不规则棕色至深棕色斑点,直径范围从5到30毫米。使用单一分生孢子分离方法从病变中分离真菌(Choi等人。1999).获得了两种形态相似的真菌分离株(SDBR-CMU497和SDBR-CMU498)。马铃薯葡萄糖琼脂(PDA)和麦芽提取物琼脂(MEA)上的菌落直径为65至67和29至38mm,分别在25°C孵育1周后。PDA和MEA上的菌落是平的,轻微起伏,中间呈绿色灰色,边缘灰绿色;反向黑色。两种分离物都产生无性结构。毕尼迪亚是黑色的,颗粒状,并分组。分生孢子细胞是透明的,亚圆柱形到圆柱形,8.5至17.5×3至5.5µm。分生孢子是单细胞的,透明玻璃,卵圆形到椭圆形,5.2至9.4×3.6至7.5µm(n=50),光滑的壁,有一个顶端附肢。形态学上,这两个分离株都类似于资本毛竹(Wikee等人。2013).内部转录间隔区(ITS),大亚基(nrLSU),翻译延伸因子1-α(tef1-α),肌动蛋白(行为),使用引物对ITS5/ITS4,LROR/LRO5,EF1-728F/EF2,ACT-512F/ACT-783R扩增甘油醛-3-磷酸脱氢酶(GAPDH)基因,和GPD1-LM/GPD2-LM,分别(怀特等人。1990年;Zhang等人。2022年)。序列存放于GenBank(ITS:PP946770,PP946771;nrLSU:PP948677,PP948678;tef1-α:PP948012,PP948013;行为:PP948014,PP948015;GAPDH:PP948016,PP948017)。对连接的五个基因的最大似然系统发育分析将两个分离株鉴定为P。因此,形态学和分子数据均证实了该真菌为Capatalensis。为了确认致病性,健康的商业番石榴果实品种KimJu用0.1%NaClO表面消毒3分钟,用无菌蒸馏水冲洗三次,并受伤(Cruz-Lagunas等人。2023年)。在PDA上从2周龄培养物中收集分生孢子并悬浮在无菌蒸馏水中。将15微升1×106分生孢子/ml悬浮液滴在受伤的果实上。模拟接种用作无菌蒸馏水的对照。对于每种处理进行10次重复并重复两次。将接种的果实在25°C和80至90%相对湿度下储存在单独的无菌塑料箱中。7天后,所有接种的水果都表现出棕色到深棕色的病变,而对照水果无症状。一直从PDA上的接种组织中重新分离出毛竹,以完成Koch的假设。在这项研究之前,已知在中国田间种植的番石榴果实上引起褐色或黑斑病(Liao等人。2020),埃及(阿拉法特2018),和墨西哥(Cruz-Lagunas等人。2023年)。据我们所知,这是泰国番石榴果实采后褐斑病的首次报道。结果将为流行病学调查和未来的治疗方法提供信息。
Guava (Psidium guajava L.) is a popular fruit crop that is widely cultivated in Thailand. In November 2023, brown spot disease on guava was observed during postharvest storage at 22 to 31°C and 70 to 75% relative humidity over a period of 3 to 7 days in Fang District, Chiang Mai Province, Thailand. The disease incidence was ~20% of 100 fruits per pallet box. The disease severity on each fruit ranged from 40 to 70% of the surface area affected by lesions. The symptoms appeared as circular to irregular brown to dark brown spots, ranging from 5 to 30 mm in diameter. Fungi were isolated from lesions using a single conidial isolation method (Choi et al. 1999). Two fungal isolates (SDBR-CMU497 and SDBR-CMU498) with similar morphology were obtained. Colonies on potato dextrose agar (PDA) and malt extract agar (MEA) were 65 to 67 and 29 to 38 mm in diameter, respectively after incubation for 1 week at 25°C. Colonies on PDA and MEA were flat, slightly undulate, greenish gray in the center, greyish green at the margin; reverse black. Both isolates produced asexual structures. Pycnidia were black, granular, and grouped. Conidiogenous cells were hyaline, subcylindrical to cylindrical, 8.5 to 17.5 × 3 to 5.5 µm. Conidia were single-celled, hyaline, obovoid to ellipsoid, 5.2 to 9.4 × 3.6 to 7.5 µm (n = 50), smooth-walled, with a single apical appendage. Morphologically, both isolates resembled Phyllosticta capitalensis (Wikee et al. 2013). The internal transcribed spacer (ITS) region, large subunit (nrLSU), translation elongation factor 1-alpha (tef1-α), actin (act), and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) genes were amplified using primer pairs ITS5/ITS4, LROR/LRO5, EF1-728F/EF2, ACT-512F/ACT-783R, and GPD1-LM/GPD2-LM, respectively (White et al. 1990; Zhang et al. 2022). Sequences were deposited in GenBank (ITS: PP946770, PP946771; nrLSU: PP948677, PP948678; tef1-α: PP948012, PP948013; act: PP948014, PP948015; GAPDH: PP948016, PP948017). Maximum likelihood phylogenetic analyses of the concatenated five genes identified both isolates as P. capitalensis. Thus, both morphology and molecular data confirmed the fungus as P. capitalensis. To confirm pathogenicity, healthy commercial guava fruits cultivar Kim Ju were surface disinfected by 0.1% NaClO for 3 min, rinsed three times with sterile distilled water, and wounded (Cruz-Lagunas et al. 2023). Conidia were collected from 2-week-old cultures on PDA and suspended in sterile distilled water. Fifteen microliters of a 1 × 106 conidia/ml suspension were dropped onto the wounded fruits. Mock inoculations were used as a control with sterile distilled water. Ten replications were conducted for each treatment and repeated twice. The inoculated fruits were stored in individual sterile plastic boxes at 25°C with 80 to 90% relative humidity. After 7 days, all inoculated fruits exhibited brown to dark brown lesions, while control fruits were asymptomatic. Phyllosticta capitalensis was consistently reisolated from the inoculated tissues on PDA to complete Koch\'s postulates. Prior to this study, P. capitalensis was known to cause brown or black spot disease on guava fruits cultivated in fields in China (Liao et al. 2020), Egypt (Arafat 2018), and Mexico (Cruz-Lagunas et al. 2023). To our knowledge, this is the first report of P. capitalensis causing postharvest brown spot disease on guava fruit in Thailand. The results will inform epidemiological investigations and future approaches to managing this disease.