香蕉炭疽菌菌株亲缘关系的RAPD分析

应用筛选的8个Operon随机引物对供试菌株的DNA进行扩增,所产生的RAPD图谱揭示了香蕉炭疽菌具有丰富的种内遗传多样性。UPGMA聚类分析的结果表明：26个香蕉炭疽菌被分为两个与地理来源相关的RAPD聚类群,它们分别由广东菌株和海南菌株为主组成。在两个香蕉菌群的外侧,两个胶胞炭疽菌菌株聚为一个小的外群。这些结果表明香蕉炭疽菌存在有与地域相关的种下类群分化。     

Seeds as Vehicles for Pathogen Importation

Since the 1900s, consumer demand for new plant products gave opportunity for many plant pathogens to disseminate to new areas on imported seeds. New markets for plant commodities encouraged plant breeders to begin collecting seed stocks from abroad. The birth of new seed companies extend their markets to new area. These events began the global dissemination of many seedborne pathogens. Many seedborne pathogens gained entry and escaped detection by specific traits that favored their dissemination. Three recent case scenarios are presented that illustrate how plant pathogens that passively employ the seed coats of their host achieved global dissemination and permanence in each patho-system. Evidence is presented to show that asparagus (Asparagus officinalis) seed produced in the US acted as a vehicle for disseminating one vegetatively compatible group (VCG) of a pathogenic fungus on asparagus called Fusarium proliferatum throughout new plantings in Australia. Similarly, public demand for Mediterranean cuisine in the US and abroad during the last 20 years led to an increase in the importation of basil (Ocimum basilicum) seed along with an inconspicuous fungus called Fusarium oxysporum. The fungus caused a destructive disease called Fusarium wilt of basil that appeared in over 25 separate locals spanning three continents. The third example demonstrated how new developments in lupine (Lupinus spp.) cultivars and increased public demand led to the global dispersal of a seedborne pathogen called Colletotrichum gloeosporioides. Each case highlights how these pathogens use seeds, humans, and particular traits to disperse globally in short period of time.     

基于CRISPR-Cas9技术构建橡胶树胶孢炭疽菌的基因敲除系统

[背景]CRISPR-Cas9基因组编辑技术为病原真菌的基因敲除、敲入及定点编辑提供了新的思路。[目的]建立适用于橡胶树胶孢炭疽菌的CRISPR-Cas9基因敲除系统。[方法]通过大肠杆菌原核表达系统合成含有细胞核定位信号的Cas9蛋白;以URA5为靶标基因,预测该基因中Cas9的切割位点,并在体外转录合成相应的SgRNA;体外构建Cas9-SgRNA复合体,并将该复合体转入橡胶树胶孢炭疽菌原生质体;通过表型筛选及测序鉴定,筛选URA5的敲除突变体菌株。[结果]体外表达的Cas9蛋白与SgRNA能够形成复合体,并在体外对目标基因URA5的DNA序列进行切割;Cas9-SgRNA复合体能够成功转入橡胶树胶孢炭疽菌原生质体,并完成对URA5的敲除;敲除突变株表现出尿嘧啶缺陷表现型。[结论]建立了适用于橡胶树胶孢炭疽菌的基因敲除系统。     

Morphological and molecular identification of Colletotrichum siamense,a novel leaf pathogen associated with Sterculia lanceolata recorded in China

Sterculia lanceolata, an important tropical woody plant, has high ornamental and medicinal value. To our knowledge, only brown root disease in this plant has been reported. In Nanning, Guangxi, China, an outbreak of leaf blight disease was observed on S. lanceolata in June 2019, with the leaf infection rate ranging from 80% to 100%. The disease seriously affected the leaves of trees and caused economic loss. Eight isolates were recovered from the infected leaves of different trees, and the pathogenicity was then determined by the methods of mycelial disc and conidial suspension, fulfilling Koch's postulates. According to the morphological and molecular biological characteristics of isolates, the pathogen causing leaf blight on S. lanceolata was identified as Colletotrichum siamense. Accurate identification of the pathogen provides a reliable basis for the control of the disease.     

Responses of embryogenic mango cultures and seedling bioassays to a partially purified phytotoxin produced by a mango leaf isolate of Colletotrichum gloeosporioides penz

Summary Colletotrichum gloeosporioides Penz., the causal agent of mango anthracnose, produces a phytotoxin in vitro. The partially purified phytotoxin, presumably colletotrichin, caused anthracnose-like symptoms on young mango leaves, was toxic to embryogenic suspension cultures of two mango cultivars, ‘Hindi’ and ‘Carabao,’ and inhibited in vitro seed germination of two nonhosts, lettuce and tobacco. There were linear relationships between concentration of the partially purified phytotoxin and mortality of mango embryogenic cultures. Embryogenic cultures grown in the presence of the partially purified phytotoxin showed significantly lower growth rates than the controls. Similarly, embryogenic cultures grown in the presence of 40% (vol/vol) fungal culture filtrate showed significantly lower growth rates than unchallenged controls. Medium containing 40% (vol/vol) Czapek-Dox fungal broth did not reduce growth of embryogenic cultures, indicating the production of phytotoxin in vitro. The results suggest that either fungal culture filtrate or purified phytotoxin can be used as in vitro selection agents to screen for resistance to this fungus.     

Live-cell imaging of conidial fusion in the bean pathogen, Colletotrichum lindemuthianum

Fusion of conidia and conidial germlings by means of conidial anastomosis tubes (CATs) is a common phenomenon in filamentous fungi, including many plant pathogens. It has a number of different roles, and has been speculated to facilitate parasexual recombination and horizontal gene transfer between species. The bean pathogen Colletotrichum lindemuthianum naturally undergoes CAT fusion on the host surface and within asexual fruiting bodies in anthracnose lesions on its host. It has not been previously possible to analyze the whole process of CAT fusion in this or any other pathogen using live-cell imaging techniques. Here we report the development of a robust protocol for doing this with C. lindemuthianum in vitro. The percentage of conidial germination and CAT fusion was found to be dependent on culture age, media and the fungal strain used. Increased CAT fusion was correlated with reduced germ tube formation. We show time-lapse imaging of the whole process of CAT fusion in C. lindemuthianum for the first time and monitored nuclear migration through fused CATs using nuclei labelled with GFP. CAT fusion in this pathogen was found to exhibit significant differences to that in the model system Neurospora crassa. In contrast to N. crassa, CAT fusion in C. lindemuthianum is inhibited by nutrients (it only occurs in water) and the process takes considerably longer.     

Biochemical aspects of bean resistance to anthracnose mediated by silicon

This study investigated the effect of silicon (Si) on resistance of bean plants (cv. ‘Peróla’) to anthracnose, caused by Colletotrichum lindemuthianum, grown in a nutrient solution containing 0 (?Si) or 2 mmol Si L^?1 (+Si). The concentration of Si in leaf tissue and the incubation period increased by 55.2% and 14.3%, respectively, in +Si plants in relation to ?Si plants. The area under anthracnose progress curve and the severity estimated by the software QUANT significantly decreased by 32.9% and 27%, respectively, for +Si plants. Si did not affect the concentration of total soluble phenolics. Chitinases activity was higher in the advanced stages of infection by C. lindemuthianum for leaves of ?Si plants. β‐1,3‐Glucanase activity increased after C. lindemuthianum infection, but it was not enhanced by Si. Peroxidase and polyphenoloxidase activities had no apparent effect on the resistance of bean plants to anthracnose, regardless of the presence of Si. The increase in lignin concentration as well as on the phenylalanine ammonia‐lyase and lipoxygenase activities were important for the resistance of +Si plants against anthracnose. The results of this study suggest that Si may increase resistance to anthracnose in bean plants by enhancing certain biochemical mechanisms of defence as opposed to just acting as a physical barrier to penetration by C. lindemuthianum.