畜牧兽医学报 ›› 2024, Vol. 55 ›› Issue (5): 2206-2213.doi: 10.11843/j.issn.0366-6964.2024.05.038

• 基础兽医 • 上一篇    下一篇

花生茎源茉莉炭疽菌的分离鉴定及生物学特性研究

郑芮1, 刘紫石1, 张康友2, 颜勇2, 魏玲2, 泽仁翁姆2, 丁则德米2, 黄建钧2, 王利1*, 魏勇3*   

  1. 1. 西南民族大学 青藏高原动物遗传资源保护与利用教育部和四川省重点实验室, 成都 610041;
    2. 甘孜州雅江县农牧农村和科技局, 甘孜 626700;
    3. 四川省畜牧科学研究院, 成都 610066
  • 收稿日期:2023-08-15 出版日期:2024-05-23 发布日期:2024-05-27
  • 通讯作者: 王利,主要从事动物病理学研究,E-mail:qinxin916@aliyun.com;魏勇,主要从事动物疫病防控,E-mail:veishangyan@163.com
  • 作者简介:郑芮(1999-),女,云南昆明人,硕士生,主要从事动物病理学研究,E-mail:zryjsyx@163.com
  • 基金资助:
    四川省科技厅科研基金项目(2023YFQ0077);四川省肉羊创新团队防疫岗位(sccxtd-2020-14);西南民族大学2023年中央高校优秀学生培养工程项目(2023NYXXS116)

Isolation, Identification and Biological Characterization of Colletotrichum jasminigenum in Stems of Peanuts

ZHENG Rui1, LIU Zishi1, ZHANG Kangyou2, YAN Yong2, WEI Ling2, ZEREN Wengmu2, DINGZE Demi2, HUANG Jianjun2, WANG Li1*, WEI Yong3*   

  1. 1. Southwest University for Nationalities, Qinghai Tibet Plateau Animal Genetic Resources Protection and Utilization Key Laboratory of the Ministry of Education and Sichuan Province, Chengdu 610041, China;
    2. Agriculture, Animal Husbandry, Rural Affairs and Science and Technology Bureau of Yajiang County, Ganzi 626700, China;
    3. Sichuan Academy of Animal Husbandry Sciences, Chengdu 610066, China
  • Received:2023-08-15 Online:2024-05-23 Published:2024-05-27

摘要: 旨在探究首次从花生茎中分离出茉莉炭疽菌(Colletotrichum jasminigenum)的生物学特性。通过三点接种法,经马铃薯葡萄糖琼脂(potato dextrose agar, PDA)培养基分离培养,分别用乳酸酚棉蓝染色和扫描电镜观察其形态,提取分离真菌DNA,用PCR方法扩增ITS区并测序,比对该基因同源性及遗传进化关系。将孢子悬液以5×107 CFU·mL-1剂量感染昆明小鼠,并观察小鼠的临床症状、血清生化指标和剖检病变。用K-B纸片法探究其耐药表型。结果显示,分离株分生孢子长29.6 μm±0.87 μm,呈月牙形;菌丝透明无性,有隔和分枝,无刚毛;附着孢呈棍棒状或者卵球形。PCR扩增得到该菌ITS区序列长度为586 bp,序列提交GenBank(OR472966)。根据形态学结合ITS区序列分析结果确定分离株为茉莉炭疽菌,命名为SLY01。用SLY01菌株攻毒小鼠14 d后导致血清碱性磷酸酶含量极显著升高(P<0.01),肝脏器指数显著下降(P<0.05),并从感染小鼠体内再次分离鉴定出茉莉炭疽菌。组织病理学结果显示,试验组小鼠肺泡壁水肿伴有大量淋巴细胞浸润;肝细胞出现大面积颗粒变性,肝小叶内见出血和肝细胞坏死;肾可见肾小管上皮发生颗粒变性。药敏试验显示,SLY01菌株对卡泊芬净和伏立康唑耐药,对两性霉素B敏感,对伊曲康唑中度敏感。本研究分离鉴定得到茉莉炭疽菌,感染小鼠主要损伤靶器官为肝、肺,并且对多种药物耐药。这为进一步研究茉莉炭疽菌的防治提供理论依据。

关键词: 茉莉炭疽菌, 分离鉴定, 致病性, 耐药性

Abstract: The objective of this study was to investigate the biological properties of Colletotrichum jasminigenum. The fungi were cultivated in PDA using the three-spot inoculation method, followed by observation of their morphology through lactophenol cotton blue staining and scanning electron microscopy. Fungal DNA was extracted via PCR to amplify the ITS region of the gene and sequenced for comparison of gene homology and genetic evolutionary relationships. The concentration of the spore suspension was adjusted to 5×107 CFU·mL-1 for mouse infection, and clinical symptoms, serum biochemical indexes, and anatomical lesions were observed. The Kirby-Bauer method was employed to investigate fungal drug resistance phenotypes. The results showed that the isolated conidia were 29.6 μm±0.87 μm in length and crescent-shaped. The mycelium was hyaline, asexual, septate, and branched without bristles. Additionally, the adherent spores were clavate or ovoid in shape. The PCR amplification yielded a sequence length of 586 bp for the ITS region of the fungi, which was subsequently submitted to GenBank (OR472966). Based on both morphology and sequence analysis of the ITS region, it was determined that the isolate belonged to Colletotrichum jasminigenum and was named SLY01. Attacking mice with strain SLY01 for 14 days resulted in a highly significant increase in serum alkaline phosphatase levels (P<0.01) and a significant decrease in liver organ index (P<0.05). Colletotrichum jasminigenum was reisolated from the infected mice. Histopathological results showed edema of the alveolar walls with massive lymphocytic infiltration, extensive granular degeneration with hemorrhage and hepatocellular necrosis in the lobules of hepatocytes, and granular degeneration of the renal tubular epithelium in the kidneys. The results of the drug sensitivity test showed that SLY01 was resistant to caspofungin and voriconazole, sensitive to amphotericin B and moderately sensitive to itraconazole. In this study, Colletotrichum jasminigenum was isolated and characterized. The liver and lungs were identified as the main target organs affected in infected mice, and the fungus showed resistance to various drugs. These findings provide a theoretical basis for further research on fungal infections originating from food sources.

Key words: Colletotrichum jasminigenum, isolation and identification, pathogenicity, drug resistance

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