Characterization of arbuscular mycorrhizal fungi and endophytic bacillus in tomato roots and their antagonism against Fusarium Wilt.
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Date
2021-07
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College of Post Graduate Studies in Agricultural Sciences, CAU-Imphal, Umiam
Abstract
Tomato (Solanum lycopersicum M.) is one of the most popular vegetable ranking next to potato and widely grown in Meghalaya. Among the soil borne diseases, Fusarium wilt caused by Fusarium oxysporum f. sp. lycopersici (FOL) is a devastating disease causing wilting and death of tomato thereby inflicting major yield losses. Arbuscular mycorrhizal fungi (AMF) are found to be associated with 80-90% of plants including tomato. Bacillus endophytes and AMF are known to promote plant growth and suppress soil borne pathogens. In the present investigation, AMF spores were isolated from 20 sample sites coming under six districts and four agroclimatic zones of Meghalaya with highest number of spores/50 g soil recovered from Umsning (680.67 spores). A total of 41 AMF species were recovered and identified from 20 sample sites. Daistong with 15 AMF species had the highest species richness. Funneliformis mosseae, Glomus aggregatum, Glomus intraradices and G. fasciculatum were the dominating AMF species having isolation frequencies (IF) of more than 60%, whereas G. australe, F. badium, A. melleae and A. scrobiculata had IF of more than 45%. All the tomato roots collected from the 20 sample sites were colonized by AMF with Umiet (41%) recorded the highest colonization per cent. From trap culture, Umiet had the maximum spore density (1103 spore/50 g soil). All 41 AMF species isolated from field soil were also recovered from trap culture inoculum along with an additional 6 species viz. Archaeospora trappie, Acaulospora longula, Appendisphaera centroreticulata, F. geosporum, G. boreale, and G. viscosum from trap culture soil. Umiet with 19 AMF species had the maximum species richness. G. intraradices had the highest IF of 70% followed by F. mosseae, G. aggregatum, and G. fasciculatum, while AMF species with more than 50% IF were F. badium, A. mellae, A. scrobiculata and G. australe. AMF root colonization in trap plant maize was maximum in Umiet derived inoculum (92%). All the 47 AMF species showed variation in shape, size, colour and hyphal form attached to the spore walls. Out of the 12 dominating AMF species screened for growth promotion of tomato plant in pot experiment, F. mosseae and G. fasciculatum performed best in terms of plant and mycorrhizal parameters. A total of 130 endophytic Bacillus were isolated from tomato roots collected from 20 sample sites of Meghalaya. Based on biochemical tests, morphological character and bacterial identification software ABIS online all the 130 isolates were tentatively identified up to species level. The isolates belonged to the genera Bacillus, Paenibacillus and Viridibacillus with maximum of 17 isolates each tentatively identified as Bacillus amyloliquefaciens and B. thuringiensis. Out of 130 Bacillus isolates, 12 isolates showed more than 27.44% inhibition of growth of FOL in dual culture method of which isolates ERBS51 (58.43%) and ERBS10 (55.68%) recorded the maximum inhibition of FOL. Further, antagonistic test of the 12 potential isolates revealed ERBS51 (55.83% and 91.52%) followed by ERBS10 (43.37% and 87.28%) as the best one with highest FOL inhibition in sealed plate method and antagonistic activity in liquid media respectively. The 12 potential isolates were identified as Bacillus sp. (ERBS4, ERBS10, ERBS29, ERBS47, ERBS75, ERBS118), while other as B. velezensis (ERBS51), B. cereus (ERBS69 and ERBS119) and B. subtilis (ERBS80), B. swezeyi (ERBS115) and B.subtilis sub sp. spizizenii (ERBS130) using 16s rRNA gene with bacterial universal primer(27F/1492R) and Bacillus specific primers (BCF1/BCR2). All the 12 isolates were grouped together in Neighbour-joining phylogenetic tree. Majority of the isolates were positive for iturin, surfactin, and bacillomycin genes. The 12 isolates were further screened for hydrolytic enzyme production, antimicrobial and plant growth promotion (PGP) activities and found to have varying results. Isolate ERBS51 was found to be positive for majority of the attributes such as siderophore, ammonia production, phosphate, ZnCO3, ZnCl solubilisation, alpha amylase, cellulase, protease, pectinase and lipase production. ERBS51 (95% and 1472.50) followed by ERBS10 (88% and 1287.54) treated tomato seeds had the maximum germination and vigour index respectively. Pot and field experiment were conducted with F. mosseae, G. fasciculatum, ERBS51 (Bacillus velezensis) and ERBS10 (Bacillus sp.) both individual and combinations as treatments. The combined application gave better result than individual microbes as treatment. Among all the treatments given as single inoculum and all the possible combinations of the four microbes,F. mosseae+G. fasciculatum+B. velezensis+Bacillus sp. treatment as consortium with recorded highest disease reduction in wilt severity in both pot (77.44%) and field (66.74%) experiments respectively. F. mosseae+G. fasciculatum+B. velezensis+Bacillus sp. treatment also recorded high plant and yield attributes in both the conditions. F. mosseae, G. fasciculatum, B. velezensis and Bacillus sp. as a consortium could be used as a part of integrated management of Fusarium wilt of tomato in Meghalaya.
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Keywords
Fusarium oxysporum, Biological control