Isolation and characterization of endophytic bacteria from endorhizosphere of sugarcane and ryegrass

摘要

The present study was conducted with a view to isolate and characterize endophytic bacterial diversity from endorhizosphere of sugarcane (Saccharum sp.) and rye grass (Lolium perenne). Eight bacterial isolates from sugarcane and seven from rye grass were obtained and identified as Azospirillum, Bacillus, E. coli and Pseudomonas. S1 and R1 (Azospirillum) sugarcane and ryegrass isolate exhibited maximum nitrogenase activity of 1221.0 and 1168.3 nM C2H4h-1 mg-1 protein respectively while the maximum IAA production was recorded in S5 (E. coli) and R7 (Bacillus) i.e. 19.3 and 20.0 mg ml-1 respectively. The maximum siderophore production was observed in S6 (E.coli) and R6 (Pseudomonas) i.e. 2.4 and 3.0 mg ml-1 respectively. Only S5 (E.coli) was observed to be solubilize phosphate (21 mg P). The endorhizosphere of sugarcane and rye grass exhibited endophytic bacterial diversity not only in terms of different types of isolates but also in terms of functional diversity. Introduction Endophytes are of agronomic interest as they can enhance plant growth in non-leguminous crops and improve their nutrition through nitrogen fixation (Boddey et al 2003). Endophyte being a broader term includes fungal, actinomycetal as well as bacterial forms. Endophytic bacteria reside within the interior of plants without causing disease or forming symbiotic structures. Thus, they inhabit various tissues of seeds, roots, stem and leaves (Johri 2006). Majority of them are non-specific regarding their host preference which holds greater promises for plant growth promotion and increased yield in agriculturally important grasses such as sugarcane (Saccharum sp), rice (Oryza sativa), wheat (Triticum aestivum), sorghum ((Sorghum bicolor), maize (Zea mays), Panicum maximum, Brachiaria spp and Pennisetum purpureum. Endophyte-infected plants often grow faster than non-infected ones which may partly by due to endophytic production of phytohormones such as indole-3-?acetic acid (IAA), cytokinin, and other plant growth-promoting substances such as production and secretion of siderophores (Fe chelating ligands) and/or partly owing to the fact that endophytes could have enhanced the host’s uptake of nutritional elements such as nitrogen and phosphorus. Thus, study was conducted with a view to isolate endophytic bacteria and to asses functional potentialities in relation to plant growth promoting activities i.e. IAA, phosphate solubilizer, nitrogenase activity and siderophore production. These can be recommended as bioinoculants for non-leguminous crops, which can help to reduce dependence on chemical fertilizers and provide a step forward towards sustainable agriculture. Materials And Methods Sugarcane and ryegrass root samples (eight each) were collected from different areas of Ludhiana i.e Jagraon, Gujjarwal, Laddowal, Samrala and Punjab Agricultural University. Isolation of bacterial diversity was done by using Reis method (Reis et al 1994). Morphological characterization of all the isolates was done on the basis of colony colour, appearance, motility and gram staining. Biochemical characterization includes acid production, catalase production and sugar utilization tests. The intrinsic antibiotic resistance spectra was deciphered using 7 antibiotics Amikacin, Ampicillin, Choramphenicol, Gentamycin, Oxytetracycline, Pencillin G and Streptomycin in a disc assay. Commercially available discs impregnated with known ppm of antibiotic were used and zone of inhibition around the disc was observed to infer the results. Functional characterization was done by performing acetylene reduction assay for nitrogenase activity (Hardy et al 1973). Qualitative screening of phosphate solubilizing isolates was done by using NBRI-BPB medium (Nautiyal 1999). Quantitative characterization of isolates for phosphate solubilization was done by Jackson’s method (1973) by using Ammonium molybdate-Ammonium vandate reagent. Siderophore production was estimated by using Arnow’s method. IAA producti