RESEARCH ARTICLE ISOLATION and CHARACTERIZATION of MARINE ACTINOMYCETES from MUTHUPPETTAI MANGROVES , TAMILNADU, INDIA Prabhahar.C*,K.Saleshrani** and Enbarasan.R *1

Available Online at http://www.recentscientific.com International Journal of Recent Scientific International Journal of Recent Scientific Research Research Vol. 5, Issue, 4, pp.906-910, April, 2014 ISSN: 0976-3031 RESEARCH ARTICLE ISOLATION AND CHARACTERIZATION OF MARINE ACTINOMYCETES FROM MUTHUPPETTAI MANGROVES , TAMILNADU, INDIA Prabhahar.C*,K.Saleshrani** and Enbarasan.R *1 *.Department of zoology, Annamalai University, Tamil Nadu, India ** ARTICLE INFO Department of zoology, ManonmaniamABSTRACT Sundaranar University, Thirunelveli, Tamil Nadu Article History: Microorganisms from Muthuppettai Mangrove areas are play an important role in Received 17th, March, 2014 biodegradation of dead plant material. This area is rich in Avicennia officinalis, Received in revised form 24th, March, 2014 Rhizophora mucaronata. Acanthus illicifolius and Excoecaria agallocha plants. Accepted 12th, April, 2014 Mangrove has a salty ecosystem and it is known to be large sources of organic matter Published online 28th, April, 2014 due to various microbial enzymatic and metabolic activities . Actinomycetes are all Gram-positive, filamentous and are facultatively anaerobic. All species grow best Key words: under anaerobic conditions. Actinomycetes play an important role among the Actinomycetes, Muthuppettai Mangrove sediments, mangroves bacterial communities, because of its diversity and ability to produce novel marine ecosystem, Antibacterial, antifungal activity chemical compounds of high commercial value A total ten isolates of Actinomycetes were collected from sediments of Muthuppettai mangroves These isolates were designated as PAct-1 to PAct-10. Out of ten isolates, eight isolates exhibited significant antibacterial activity. All the ten isolates were charecterised for biochemical and cultural aspects. These isolates were identified as Streptomyces sp. Among the ten isolates, PAct-1 exhibited the maximum zone of inhibition against bacterial activity. © Copy Right, IJRSR, 2014, Academic Journals. All rights reserved. INTRODUCTION carried out to sea in the form of resistant spores (Ravel et al., 1998; Ramya and Vijayakumar, 2008; Deepika and Marine environments are largely untapped source for the Kannabiran, 2010). Microorganisms found in marine isolation of new microorganisms with potentiality to produce environments have attracted a great deal of attention, due to active secondary metabolites. Among such microorganisms, the production of various natural compounds and their Actinomycetes are of special interest, since they are known to specialized mechanisms for adaptation to extreme environment produce chemically diverse compounds with a wide range of (Solingen et al., 2001). The pre-treatment including biological activities (Bredholt et al., 2008; Kalyani et al., enrichment, physical and selective media may be used to study 2012). The demand for new antibiotics continues to grow due the ecology of actinomycetes in natural habitats such as soil or to the rapid emerging of multiple antibiotic resistant pathogens water samples (Jensen et al., 2005; Gebreyohannes et al., causing life threatening infection. Although, considerable 2013). Since marine sediments represent an environment progress is being made within the fields of chemical synthesis which is markedly different from that associated with soil and engineered biosynthesis of antibacterial compounds, samples, it is not clear how effective the pre-treatment of such nature still remains the richest and the most versatile source for sediments would be for the recovery of bioactive new antibiotics (Baltz, 2006; Pelaez, 2006). Traditionally, actinomycetes. Marine sediment is an inexhaustible resource actinomycetes have been isolated from terrestrial sources that has not been properly exploited. Reports from the East although, the first report of mycelium forming Actinomycetes Coast of India, suggests that soil is a major source of being recovered from marine sediments appeared several actinomycetes (Sivakumar et al., 2005; Vijayakumar et al., decades ago (Weyland, 1969). Recently, the marine derived 2007; Dhanasekaran et al., 2008; Vijayakumar et al., 2009; actinomycetes have become recognized as a source of novel Deepika and Kannabiran, 2010; Kalyani et al., 2012; antibiotic and anticancer agent with unusual structure and Gebreyohannes et al., 2013). Correspondingly, the Muthuppettai properties (Jensen et al., 2005; Gebreyohannes et al., 2013). mangroves ecosystem is largely unexplored, and may provide Actinomycetes represent a ubiquitous group of microbes a rich source of the microorganisms producing novel and widely distributed in natural ecosystems around the world and efficient antimicrobial compounds. Hence, the present study especially significant for their role on the recycling of organic was undertaken to isolate the Actinomycetes from marine matter (Srinivasan et al., 1991). The literatures suggested that, sediments of Muthuppettai mangroves region. marine sediment sources are voluble for the isolation of novel actinomycetes with the potential to yield useful new products MATERIALS AND METHODS (Goodfellow and Haynes, 1984). However, it has been Collection of sample resolved whether Actinomycetes form part of the autochthonous marine microbial community of sediment The sediments sample were collected from Muthuppettai samples originated from terrestrial habitats and were simply mangroves area Muthupet mangrove environment (Lat.10’ * Corresponding author: Prabhahar.C Department of zoology, Annamalai University, Tamil Nadu, India International Journal of Recent Scientific Research, Vol. 5, Issue, 4, pp.906-910, 2014 20’N and 79’ 35’E) is known to be very rich microbial of oxygen. Type of carbon source utilized by microorganism diversity due to high amount of dissolved and particulate was identified by change in pH of the carbon utilization agar organic matter and therefore, different types of medium. Positive assimilation of growth indicated by color microorganisms are found in this type of environment .For the change from purple to yellow induced by bromocresol purple isolation of actinomycetes from different places at a depth of dye present in the medium. Sterile carbohydrate utilization 10 m using a core sampler. The central portions of the agar with bromocresol purple dye was prepared. It was, then, sediments were ascetically transferred to the sterile bottles. inoculated with 1 ml of soil isolates and poured into sterile The sediment sample was blackish brown color and of a sandy petri dishes. After solidification, sterile discs containing 3% of texture. different carbon sources such as dextrose, sucrose, starch, lactose, and maltose were placed aseptically on the surface of Isolation of Actinomycetes colonies from the marine the medium and incubated at 27°C for 8-10 days. sediments Starch hydrolysis Isolation and enumeration of Actinomycetes from sediments were performed by the soil dilution plate technique (Ellaiah et The soil isolates were streaked across starch – peptone agar al., 1996) using starch casein agar medium (g/;L: starch-10, plates using loopful of spores and incubated for 5 days at 28°C casein- 0.3, KNO3- 2, NaCl- 2, K2HPO4-2, MgSO4.7H2O- 0.05, (Kokare et al., 2003). At the end of incubation, the plates were CaCO3- 0.02, FeSO4.7H2O- 0.01 and agar- 18). 50 ml of starch flooded with weak iodine solution. The width of the casein agar media in 250 ml flask were sterilized at 121°C for hydrolysed zone around the growth and the width of the 20 min by autoclaving. The media was prepared by using 50% growth were measured. (v/v) sea water. 1 g each of the marine sediment sample was Coagulation and peptonization of milk taken in 250 ml Erlenmeyer flask containing 50 ml of sterile Skimmed milk was prepared by centrifuging the milk water. The flasks were shaken on rotary shaker for 30 min for contained in 50 ml plastic conical centrifuge tubes (Tarsons, the detachment of spore chains. The flasks were kept aside for Calcutta) at 2000 rpm for 45 min (Kokare et al., 2003). It was 15 min to settle down the particulate matter. The suspension transferred in 10 ml portions aseptically into the sterile boiling was serially diluted up to 6 fold. 1 ml each of these dilutions tubes. These boiling tubes were tyndallized at 80°C for 1 h for were added to each of 50 ml of the aforementioned sterile 3 consecutive days. The skimmed milk tubes were inoculated molten media maintained between 40 to 45°C, thoroughly with test cultures and incubated at 28°C. The extent of mixed and poured into Petri plates and incubated at 28°C. The coagulation and peptonization were recorded after 5 days of incubated Petri plates were observed from one week onwards incubation. for three weeks or colonies of Actinomycetes. The starch casein agar media was supplemented with 2.5 μg/ml of Nitrate reduction test rifampicin and 75 μg/ml of flucanozole to minimize bacterial Organic nitrate broth of 10 ml was inoculated with 50 µl of and fungal contaminations, respectively. Actinomycetes spore suspension and incubated at 28°C for 7 days (Kokare et colonies were marked, thus identical colonies were scored out al., 2003). Controls were run also without inoculation. On 7th and the selected colonies were sub cultured on SCA slants and day, the clear broth was tested for the presence of nitrite. To 1 incubated at 28°C for one week. ml of broth under examination and 1ml of control, two drops of sulphanilic acid solution followed by 2 drops of α - Characterization of Actinomycetes napthylamine solution were added. The presence

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