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Current Trends in Biotechnology and Pharmacy Vol. 5 (3) 1282-1297 July 2011, ISSN 0973-8916 (Print), 2230-7303 (Online) 1287 Table 1 The antimicrobial activities of actinomycetes isolate No.9 Tested microbial pathogen Diameter of inhibition zone (mm) Escherichia coli ATCC 25922 20 Bacillus subtilis NRRL B-4219 30 Pseudomonas aeruginosae ATCC 27953 - Staphylococcus aureus ATCC 29213 20 Candida albicans ATCC 10231 30 Saccharomyces cerevisiae ATCC 10275 25 Aspergillus niger NRRL 599 30 Aspergillus flavus NRRL 1957 - Alcaligenes faecalis NRRL B-170 30 Micrococcus luteus NRRL B-287 10 Trycophyton mentagrophytes ATCC 9533 14 Table 2 Identification criteria of actinomycetes isolate no. 9 (Nocardioides luteus) Experimental Parameter Character Experimental Parameter Character Aerial mycelia color Grey L-diaminopimelic acid in whole cell hydrolysate + Vegetative mycelia color Yellow-Brown Growth on sole carbon sources (1.0 % w/v) Melanin on tyrosine agar + Sucrose + Degradation of - Lactose + Casein + Salicin + Xanthene - D-Fructose + Hypoxanthene- Galactose + Urea + D-Mannose + Growth at D-Mannitol + 45 o C - Rafinose + 10 o C + D-Xylose + pH 4.3 + L-Arabinose + Growth in the presence of (%w/v) Cellobiosem NaCl (4) + Inositol + NaCl (7) + Rhamnose + NaCl (10) + Glucose + Crystal violet (0.0001) - Sodium acetate (0.1 %) + Phenol (0.1) Sodium citrate (0.1%) + Resistance to antibiotics Sodium propionate (0.1 % ) + Gentamicin (10 µg) Growth on sole nitrogen source (0.1 %) Streptomycin (10 µg) - L-Arginine + Neomycin (10 µg) - L-Histidine + Penicillin G (10 units) - L-Cysteine + Rifampicin (5 µg) - L-Valine + Tetracycline (30 µg) - L-Phenyl alanine + Tobramycin (10 µg) + Potassium nitrate + Erythromycin (15 µg) - L-Threonine + Bacitracin (10 µg) - L-Serine + Chloramphenicol (30 µg) - L-Methionine + Novobiocin (5 µg) - El-Refai et al
Current Trends in Biotechnology and Pharmacy Vol. 5 (3) 1282-1297 July 2011, ISSN 0973-8916 (Print), 2230-7303 (Online) 1288 used in the identification were listed in materials and methods, results of which are given in Table 2. The genus Nocardioides is easily differentiated from all other actinomycetes on the basis of the diaminopimelic acid type in the cell wall peptidoglycan, menaquinone profile, and the cellular fatty acid profile (19,20). The genus Nocardioides is one of a few genera that have LL-2,6-diaminopimelic acid (LL-DAP) as the diagnostic diamino acid in the cell wall peptidoglycan layer(21). This feature, together with glycine in the cell wall peptidoglycan, indicates that this genus is wall chemotype I (22). This chemical marker distinguishes the genus Nocardioides from many other actinomycetes genera that possess other types of diamino acids in the cell wall peptidoglycan. Regarding the bioactivities of the Nocardioides spp., only a few Nocardioides strains were found to produce antibiotics (23,24,25). Strain ATCC 39419, which produced a novel anti-tumor antibiotic, sandramycin, was originally assigned to Nocardioides (26) but has later been classified as a member of a new genus Kribbella (27). However, to our knowledge, this is the first study to investigate the ability of a Nocardioides sp. to produce antibiotics in detail; regarding the physiological parameters that affect the production followed by an attempt to isolate and identify the active compound(s). Cultivation parameters : Since antibiotics are secondary metabolites synthesized by pathways, which are often connected and influenced by primary metabolism, thus, frequently an intermediate metabolite from primary metabolism serves as precursor for the biosynthesis of the antibiotic. Therefore, the composition of the culture medium, closely connected with the metabolic capacities of the producing organism, greatly influences the biosynthesis of antibiotics. Changes in the nature and concentration of carbon and nitrogen sources, phosphorus concentration and trace elements have been reported to affect antibiotic biosynthesis in different organisms. Results (Fig. 1) showed that activity and growth gradually increased with increasing the incubation period till the activity reached maximum after 7 days Fig. 1. Effect of different fermentation periods on anticandidal activity of Nocardioides luteus of incubation then it began to decline with time while the growth nearly remained constant after 7 days. These results revealed that the anticandidal metabolite was early produced and reached maximum at the stationary phase of growth. The cessation of growth in the stationary phase is most commonly caused by the exhaustion of the essential nutrients of the medium as well as the accumulation of undesirable metabolites. Similarly, the optimal incubation period required for maximum growth and antibiotic yields by Streptomyces sp. 201 was six days (28). On the other hand, the highest biomass and antibiotic production of Streptomyces sp. KEH23 is observed after 96 h of incubation. However, after 108 h, the diameter of the inhibition zones dropped gradually (29). Studies on the production of actinomycin by Nocardioides luteus
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(December 7-9, 2011) Venue: Karunya
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