การคัดเลือกและทดสอบประสิทธิภาพของเอนโดไฟติกแอคติโนแบคทีเรียในการควบคุมโรครากเน่าของหญ้าหวาน

Abstract

The symptoms of stevia root rot were examined. The samples were taken from stevia plantation fields in Chiang Mai Province, Thailand. The pathogens were isolated from infected tissue using the tissue transplanting method, yielding 9 fungal isolates. The baiting method was also employed to isolate the pathogen from the rhizosphere soil, yielding 11 fungal isolates. Pathogenicity testing of the total fungal isolates on stevia seedling was done, and the results revealed that two isolates, CSP1 and CSP2, had the highest disease severity, exhibiting rapid wilt symptoms, severe root rotting, and plant death within 3 days after inoculation. The fungal pathogens were identified based on their morphological and molecular characteristics, and the isolates CSP1 and CSP2 were proven to be Pythium aphanidermatum and Phytopythium helicoides, respectively. The efficacy of endophytic actinobacteria to inhibit the growth of pathogenic fungi was done using dual culture method. Eleven isolates of endophytic actinobacteria which were isolated from roots of Kariyat (Andrographis paniculata) and 14 isolates of endophytic actinobacteria obtaining from the previous experiments were tested against P. aphanidermatum and P. helicoides fungi. The results revealed that the 3 isolates of AND2, AND4, and AND5 effectively inhibited P. helicoides, with radial growth inhibition more than 75%. While, the 0% inhibition results revealed that none trial of the endophytic actinobacteria isolate successfully inhibited the growth of P. aphanidermatum. The identification of the selected endophytic actinobacteria isolates based on the morphological and molecular characteristics confirmed that all of the isolates were Streptomyces albus. The efficacies of culture filtrates obtaining from 5 liquid media after culturing of the selected endophytic actinobacteria isolate were tested to inhibit the growths of the fungal pathogens using the agar well diffusion method. The results showed that ISP-2 medium was the best to promote the production of secondary metabolites to inhibit the growth of P. helicoides by showing the percent inhibition of radial growth at 100%, followed by GYM and YMD media, respectively. In the case of P. aphanidermatum, none of the media was effectively promoted secondary metabolite to inhibit the fungus. The in vitro trials to control root rot disease were performed. Cell suspensions of selected endophytic actinobacteria were applied to the crown of a one-month-old stevia tissue culture seedling for two days before the plants were inoculated with fungal pathogens. The results showed a significant reduction in P. helicoides severity in all of the isolate trials. The disease severity scores showed non-statistical variations amongst the isolates, with 13.76 % of isolate AND5, 15.20% of isolate AND4, and 32.67% of isolate AND2. Whereas, the trials of P. aphanidermatum discovered that the disease severity index showed non-statistical variations amongst the isolates as well, with 42.10% of isolate AND5, 46.87% of isolate AND4, and 71.17% of isolate AND2. The disease control studies in greenhouse circumstances were carried out by pouring spore suspensions of selected endophytic actinobacteria over the crown of a one-month-old stevia seedling every three days for three times before inoculating pathogenic fungi. All of the isolate trials yielded a considerable reduction in P. helicoides severity. The disease severity scores varied non-statistically among the isolates, with 22.43% of isolate AND5, 24.27% of isolate AND4, and 36.04% of isolate AND2. In the P. aphanidermatum trials, the disease severity index indicated non-statistical variances across the selected isolates, with 40.96% of isolate AND5, 48.40% of isolate AND4, and 67.93% of isolate AND2. These outcomes were repeatedly revealed throughout laboratory studies.