Molecular, chemical and biological screening of soil actinomycete isolates in seeking bioactive peptide metabolites
Abstract
Background and Objective: Due to the evolution of multidrug-resistant strains, screening of natural resources, especially actinomycetes, for new therapeutic agents discovery has become the interests of researchers. In this study, molecular, chemical and biological screening of soil actinomycetes was carried out in order to search for peptide-producing actinomycetes.
Materials and Methods: 60 actinomycetes were isolated from soils of Iran. The isolates were subjected to molecular screening for detection NRPS (non-ribosomal peptide synthetases) gene. Phylogenic identification of NRPS containing isolates was performed. Chemical screening of the crude extracts was performed using chlorine o-dianisidine as peptide detector reagent and bioactivity of peptide producing strains was determined by antimicrobial bioassay. High pressure liquid chromatography- mass spectrometry (HPLC-MS) with UV-visible spectroscopy was performed for detection of the metabolite diversity in selected strain.
Results: Amplified NRPS adenylation gene (700 bp) was detected among 30 strains. Phylogenic identification of these isolates showed presence of rare actinomycetes genera among the isolates and 10 out of 30 strains were subjected to chemical screening. Nocardia sp. UTMC 751 showed antimicrobial activity against bacterial and fungal test pathogens. HPLC-MS and UV-visible spectroscopy results from the crude extract showed that this strain has probably the ability to produce new metabolites.
Conclusion: By application of a combined approach, including molecular, chemical and bioactivity analysis, a promising strain of Nocardia sp. UTMC 751 was obtained. This strain had significant activity against Staphylococcus aureus and Pseudomonas aeruginosa. Strain Nocardia sp. UTMC 751 produce five unknown and most probably new metabolites with molecular weights of 274.2, 390.3, 415.3, 598.4 and 772.5. This strain had showed 99% similarity to Nocardia ignorata DSM 44496 T.
. Dubin A, Mak P, Dubin G, Rzychon Mg, Stec-Niemczyk J, Wladyka B et al. New generation of peptide antibiotics. Acta Biochimica Polonica 2005; 52:633.
. Mkrtchyan H, Gibbons S, Heidelberger S, Zloh M, Limaki HK.Purification, characterisation and identification of acidocin LCHV, an antimicrobial peptide produced by Lactobacillus acidophilus nv Er 317/402 strain Narine. Int J Antimicrob agents 2010; 35:255-260.
. Dashper SG, Liu SW, Reynolds EC. Antimicrobial peptides and their potential as oral therapeutic agents. Int J Pept Res Ther 2007; 13:505-516.
. Challis GL, Naismith JH. Structural aspects of non-ribosomal peptide biosynthesis. Curr Opin Struct Biol 2004; 14: 748-756.
. Reddy KVR, Yedery RD, Aranha C. Antimicrobial peptides:premises and promises. Int J Antimicrob agents 2004; 24:536-547.
. Rajanbabu V, Chen JY. Applications of antimicrobial peptides from fish and perspectives for the future. Peptides 2011; 32:415-420.
. Olano C, Méndez C, Salas JA. Antitumor compounds from marine actinomycetes. Mar Drugs 2009; 7(2): 210-248.8 . Mann J. Natural products as immunosuppressive agents. Nat Prod Rep 2001; 18: 417-430.
. Sarmadi BH, Ismail A. Antioxidative peptides from food proteins: a review. Peptides 2010; 31: 1949-1956.
. Erdmann K, Cheung BW, Schröder, H. The possible roles of food-derived bioactive peptides in reducing the risk of cardiovascular disease. J Nutr Biochem 2008; 19: 643-654.
. Gauthier SF, Pouliot Y, Saint-Sauveur D. Immunomodulatory peptides obtained by the enzymatic hydrolysis of whey proteins. Int Dairy J 2006; 16: 1315-1323.
. St Georgiev V. Immunomodulatory activity of small peptides.Trends Pharmacol Sci 1990; 11: 373-378.
. Jiang Y, Han L, Chen X, Yin M, Zheng D, Wang Y et al. Diversity and Bioactivity of Cultivable Animal Fecal Actinobacteria. Adv Microbiol 2013; 3:1.
. González I, Ayuso Sacido A, Anderson A, Genilloud O.González. Actinomycetes isolated from lichens: evaluation of their diversity and detection of biosynthetic gene sequences. FEMS microbiol Ecol 2005; 54: 401-415.
. Busti E, Monciardini P, Cavaletti L, Bamonte R, Lazzarini A, Sosio M et al. Antibiotic-producing ability by representatives of a newly discovered lineage of actinomycetes. Microbiology 2006; 152(3): 675-683.
. Grünewald J, Marahiel MA. Chemoenzymatic and template- directed synthesis of bioactive macrocyclic peptides.Microbiol Mol Biol Rev 2006; 70: 121-146.
. Dharmaraj S. Marine Streptomyces as a novel source of bioactive substances. World J Microbiol Biotechnol 2010; 26):2123-2139.
. Bredholdt H, Galatenko OA, Engelhardt K, Fjærvik E, Terekhova LP, Zotchev SB. Rare actinomycete bacteria from the shallow water sediments of the Trondheim fjord, Norway: isolation, diversity and biological activity. Environ Microbiol 2007; 9: 2756-2764.
. Jork H, Funk W, Fischer W, Wimmer H, Burns DT (1990).Thin-layer chromatography. Reagents and detection methods. Volume 1a: VCH, Weinheim.
. Taddei A, Valderrama M, Giarrizzo J, Rey M, Castelli C. Chemical screening: A simple approach to visualizing Streptomyces diversity for drug discovery and further research. Res Microbiol 2006; 157: 291-297.
. Wilson ID, Plumb R, Granger J, Major H, Williams R, Lenz EM. HPLC-MS-based methods for the study of metabonomics. J Chromatogr A 2005; 817: 67-76.
. Arifuzzaman M, Khatun MR, Rahman H. Isolation and screening of actinomycetes from Sundarbans soil for antibacterial activity. Afr J Biotechnol 2010; 9: 4615-4619.
. Nikodinovic J, Barrow KD, Chuck JA. High yield preparation of genomic DNA from Streptomyces. Biotechniques 2003; 35:932-936.
. Ayuso-Sacido A, Genilloud O. New PCR primers for the screening of NRPS and PKS-I systems in actinomycetes: detection and distribution of these biosynthetic gene sequences in major taxonomic groups. Microb Ecol 2005; 49:10-24.
. Kumar V, Bharti A, Gusain O, Bisht GS. An improved method for isolation of genomic DNA from filamentous actinomycetes. J Sci Engg Tech Mgt 2010; 2:10-13.
. Chun J, Lee JH, Jung Y, Kim M, Kim S, Kim BK et al. EzTaxon: a web-based tool for the identification of prokaryotes based on 16S ribosomal RNA gene sequences. Int J Syst Evol Microbiol 2007;57: 2259-2261.
Kieser T, Bibb MJ, Buttner MJ, Chater KF, Hopwood DA (2000). Practical Streptomyces genetics. John Innes foundation, UK.28 . Ding D, Chen G, Wang B, Wang Q, Liu D, Peng M et al.Culturable actinomycetes from desert ecosystem in northeast of Qinghai-Tibet Plateau. Annals Microbiol 2013; 63: 259-266.
. Deepika L, Kannabiran K. Antibacterial and antifungal activity of Streptomyces sp. VITDDK3 isolated from Ennore coast, Tamil Nadu, India. Pharma. Res. Health Care 2010; 2:188-196.
. Hemashenpagam N. Purification of secondary metabolites from soil actinomycetes. Intl J Microbio Res 2011; 3:148-156.
. Nedialkova D, Naidenova M. Screening the antimicrobial activity of actinomycetes strains isolated from Antarctica. J Culture Collections 2005; 4: 29-35.
. Anibou M, Chait A, Zyad A, Taourirt M, Ouhdouch Y, Benherref A. Actinomycetes from Moroccan habitats: isolation and screening for cytotoxic activities. World J Microbiol Biotechnol 2008; 24: 2019-2025.
. Okudoh VI, Wallis FM. Antimicrobial activity of rare actinomycetes isolated from natural habitats in KwaZulu- Natal, South Africa. S Afr J Sci 2007; 103 (5-6): 216-222.
. Chun J, Lee JH, Jung Y, Kim M, Kim S, Kim BK et al. EzTaxon: a web-based tool for the identification of prokaryotes based on 16S ribosomal RNA gene sequences. Int J Syst Evol Microbiol 2007;57: 2259-2261.
. Wélé A, Zhang Y, Caux C, Brouard JP, Dubost L, Guette C et al. Isolation and structure of cyclosenegalins A and B, novel cyclopeptides from the seeds of Annona senegalensis. J. Chem Soc Perkin Trans 1 2002; : 2712-2718.
. Agyei D, Danquah MK. Industrial-scale manufacturing of pharmaceutical-grade bioactive peptides. Biotechnol Advances 2011; 29: 272-277.
. Hancock RE, Sahl HG. Antimicrobial and host-defense peptides as new anti-infective therapeutic strategies. Nature Biotechnol 2006; 24:1551-1557.
. Konno M. Nosocomial infections caused by methicillin- resistantStaphylococcus aureus in Japan. J Infect Chemother 1995;1: 30-39.
.Ahmed Z, Saeed Khan S, Khan M. Invitro trials of some antimicrobial combinations against Staphylococcus au- reus and Pseudomonas aeruginosa. Saudi J Biol Sci 2013;20:79-83.
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Issue | Vol 7 No 1 (2015) | |
Section | Original Article(s) | |
Keywords | ||
Actinomycetes Molecular screening Non-ribosomal peptide synthetases Peptide |
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