Yulmira Yanti

Screening of Rhizobacterial Indigenous as Biocontrol Agents Against Bacterial Wilt on Chili Using In Planta Technique

Bacterial wilt caused by Ralstonia solanacearum is one of the severe diseases on chili. Until now this disease is difficult to control, while these bacteria attack the xylem vessel and are unreachable by any bactericide. One group of important biocontrol agents are the plant growth-promoting rhizobacteria (PGPR). The PGPR have also the ability to increase growth and yield of plants. The aim of this research was to obtain rhizobacterial isolates, which have the ability to control bacterial wilt on chili and to increase growth and yield. Source of ​of rhizobacterial isolates were from healthy chili rhizosphere at endemic area of bacterial wilt diseases in two district (Agam and Solok) West Sumatera Province, Indonesia. Screening method based on in planta selection of root-colonizing bacterial isolates. This approach focuses on indirect mechanisms (systemic induced resistance). This technique has the possibility to find new, easy, and cheap biocontrol organisms. We have isolated 42 rhizobacteria from healthy chili’s rhizosphere. Only 17 rhizobacterial isolates could increase seedling growth of chili compared to control plants; the rest of isolates reduced growth of chili seedlings. There were no bacterial wilt symptom on 13 rhizobacterial isolate introduced chilies and lower infection (33.3%) on two rhizobacterial isolate introduced chilies, compared with control plants (100% wilt and die). We have obtained also two rhizobacterial isolates which could control bacterial wilt diseases and increase growth of chili. They were RZ.2.1.AG1 identified as Bacillus cereus strain INACH001 and RZ.1.3.AP1 as Bacillus subtilis strain BSn5.

Identification and Characterizations of Potential Indigenous Endophytic Bacteria which had Ability to Promote Growth Rate of Tomato and Biocontrol Agents of Ralstonia solanacearum and Fusarium oxysporum fsp. solani

The research aimed to isolate and characterize indigenous yeast strain from Tacca leontopetaloides with respect to the ethanol and glucose tolerance ability. Research done experimentally and the data analyzed descriptively. Yeasts isolated from 1g Tacca leontopetaloides grown at modified Potato Dextrose Agar/PDA (Oxoid Ltd.) with 3% Yeasts Extract/YE (Kraft Foods) and 10 ppm amoxicillin addition. Yeasts-like colony was tested in the ability to tolerate ethanol and glucose contents by grown on modified Nutrient Broth/NB (Oxoid Ltd.) with 3% Yeasts Extract/YE (Kraft Foods) and 10 ppm amoxicillin then added with glucose monohydrate (10%, 20%, 30%) or ethanol (10%, 20%, 30%) and incubated for 72h at ambient (23-28°C). Optical density (OD) was read for UV absorbance at 600 nm using UV-Vis spectrophotometer every 24h until 72h. The strain of best isolate with the ability to tolerate high ethanol and glucose contents were identified by the sequence analysis of ITS (Internal Transcribed Spacer) region using primers ITS1 (5′-TCCGTAGGTGAACCTGCGG-3′) and ITS4 (5′-TCCTCCGCTTATTGATATGC-3′). The sequencing was performed at Macrogen Inc. (Seoul, South Korea), and the sequences was compared with the GenBank database using BLAST (Basic Local Alignment Search Tools) algorithm (http//:www.ncbi.nlm.nih.gov/BLAST/). Results shown there are four yeasts-like isolate and the TK1 isolate showed the best ethanol tolerance ability with highest OD at 30% ethanol concentration (0.486) and the highest OD at 30% glucose concentration (1.732). The species identification identified the TK1 isolate as 99% identical with Candida natalensis (ITS1) and 100% identical with Candida quercitrusa (ITS4).Among Plant Growh Promoting Rhizobacteria (PGPR) groups, endophytic bacteria considered as one of the options to control vascular wilt disease because of its ability to live and colonized internal roots of plants without causing any damages. Our previous research had screened 9 isolates which had best ability to promote growth rate and increase yields of tomato and biocontrol agents of R. solanacearum and Fusarium oxysporum f.sp solani in planta condition. In order to know its abilities, those isolates need to be characterized. This research purposed to characterize those isolates abilities to produce IAA, phosphate solubilizing, siderophore production, cyanide production, NH3 production, and ability to colonize endophytically and identified the isolates using 16S rRNA. Result shown that all isolates can produce IAA, where TLE1.1 produce highest IAA concentration (42.5 ppm). Isolates E1AB1.3, TLE 1.1 and TLE2.2 can dissolved phosphate. None of the isolates produced HCN and NH3. Only TLE 2.3 isolate can produce siderophore. All of 9 isolates were identified using 16S rRNA gene using 27F and 1492R primers. All isolates were identified as different species, i.e. Bacillus toyonensis strain BCT-7112 (EPL1.1.3), Serratia nematodiphila strain DZ0503SBS1 (TLE2.3), Bacillus anthracis strain ATCC 14578 (EPL1.1.4), Bacillus cereus ATCC 14579 (TLE1.1), Bacillus cereus strain JCM 2152 (SNE2.2), Enterobacter cloacae subsp. dissolvens strain ATCC 23373 (E1.AB1.2), Serratia marcescens strain NBRC 102204 (E1AB2.1), Klebsiella michiganensis strain W14 (TLE2.2), and Chryseobacterium rhizoplanae strain JM-534 (KLE3.3).