Characterization of Colletotrichum truncatum CP2 and its interaction with chillies (Capsicum annuum L.) during pathogenesis

• Main Author: Mohamad Remli, Nurul Atika
• Format: Thesis
• Language: English
• Published: 2018
• Subjects: Peppers - Case studies; Colletotrichum
• Online Access: http://psasir.upm.edu.my/id/eprint/75713/1/FBSB%202018%2044%20IR.pdf; http://psasir.upm.edu.my/id/eprint/75713/
• Institution: Universiti Putra Malaysia

Anthracnose caused by Colletotrichum species is the most destructive disease of chilli worldwide. It is responsible for worldwide yield losses and could be even more severe without a successful control that still relies on the use of fungicides. Due to the growing concern about environmental and health damages caused by this control, an understanding of the mechanisms leading to the fungal pathogenicity in a particular host is essential for the implementation of effective disease control. This study aimed to investigate the mechanism leading to pathogenesis of Colletotrichum species in chilli fruit as little is known about the pathogenicity factor involved in this interaction. Thirty five fungal isolates were isolated from chilli lesions of anthracnose from different geographic locations in Malaysia. The ability of fungal isolates to produce cell wall-degrading enzymes was screened and the best cell wall-degrading enzymes producer was selected for further study. Based on its morphological, biochemical and molecular identification, fungal isolate CP2 was identified as Colletotrichum truncatum. Successful inoculation of the C. truncatum CP2 on detached chilli fruits proved its pathogenicity and was confirmed to be a primary pathogen of chilli when it successfully infected the chilli fruits. In order to illustrate the infection strategy adopted by C. truncatum CP2, the infection process of this fungus in the chilli fruit was characterized using light, scanning and transmission microscope. C. truncatum CP2 exhibited a prolonged biotrophic phase of about 48 hour, before switched to necrotrophic phase at approximately 72 hour after inoculation. The first phase of necrotrophy in C. truncatum CP2 was characterized by formation of germ tube, appresorium and infectious hyphae. The destructive necrotrophic phase was characterized by formation of sunken lesions and production of numerous acervuli. The role of cell wall-degrading enzymes in facilitating the C. truncatum CP2 to colonize the host cell was investigated taking into consideration changes in the morphological and chemical compositions of the chilli fruits. The results of enzymatic activity experiment indicated that polygalacturonase (PG) was the first cell wall-degrading enzymes detected and the activities obtained were higher (0.24±0.10 U/mL) than other enzymes, which appeared later and in lower amount. Significant changes in the pectin (total uronide content increased up to 50.33% - 71.85%) and cellulose contents (decreased to 11.45% - 12.32%) in chilli treated with PG and combination of PG and cellulases showed the main role of these enzymes in facilitating the C. truncatum CP2 during pathogenesis in chilli fruits. According to Fourier transform infrared analysis, there were remarkable changes in the vibration side of cellulose (3290 cm-1 and 2924 cm-1) and ring and vibration side of pectin (1581, 1337 and 1029 cm-1) in the cell wall of chilli treated with PG and mixture of both enzymes. In order to understand the exact role of PG enzymes in pathogenesis, PG enzymes from C. truncatum CP2 was purified using aqueous two phase system. The optimum purification condition of PG was achieved using 22% (w/w) polyethylene glycol and 15% (w/w) sodium citrate comprising crude load of 16% (w/w) at pH 7.0 with addition of 1.0% (w/w) sodium chloride. The necrotizing activity of the crude and purified PG from C. truncatum CP2 was then tested on detached chilli fruits. The faster lesion formation on the chilli treated with purified PG had confirmed the involvement of this enzyme in anthracnose of chilli. In conclusion, C. truncatum CP2 possess all the features to be termed as a serious anthracnose pathogen with the presence of pathogenicity factors such as PG enzymes. The results from this study provide a better insight into the interaction of C. truncatum CP2 and chilli fruits and these findings may be used in the development of efficient disease management strategies in Malaysia.