Starch Casein Agar Medium Research Articles

Promising bioactive metabolites of mangrove inhabitant Streptomyces tauricus and prostate cancer PC3 cell inhibition by antimicrobial peptides.

Streptomyces is a group of microbes known for antibiotic production and has contributed to more than 70% of present commercially available antibiotics. These antibiotics are important in the management, protection, and treatment of chronic illnesses. In the present study, the isolated S. tauricus strain from mangrove soil in Mangalore, India (GenBank accession number: MW785875) was subjected for differential cultural characterization, phenotype involving brown pigmentation, filamentous mycelia, and ash-colored spore production was observed using field emission scanning electron microscopy (FESEM) analysis revealing filamentous mycelia possessing a straight spore chain. Spores were visualized as elongated, rod-shaped, smooth surfaces with curved edges. After optimized growth conditions for S. tauricus on starch-casein agar medium, the GC/MS analysis of S. tauricus intracellular extract detected bioactive compounds reported for pharmacological applications. Analyzed using the NIST library, most of the bioactive compounds identified in intracellular extract had molecular weights of less than 1 kDa. On the PC3 cell line, the Sephadex G-10 partially purified eluted peak protein fraction demonstrated significant anticancer activity. The LCMS analysis revealed the presence of Tryprostatin B, Fumonisin B1, Microcystin LR, and Surfactin C with molecular weights below 1 kDa. This study found that small molecular weight microbial compounds are more effective in a variety of biological applications.

Exploring the antimicrobial, antiviral, antioxidant, and antitumor potentials of marine Streptomyces tunisiensis W4MT573222 pigment isolated from Abu-Qir sediments, Egypt

Due to the therapeutic importance of microbial pigments, these pigments are receiving the attention of researchers. In this present study 60 isolates were isolated from sediments of Abu-Qir coast of the Mediterranean sea, Alexandria, Egypt, out of which 12 were considered as pigmented actinomycetes. Streptomyces sp. W4 was characterized by small round green pigmented colonies when grown on starch-casein agar medium. The green pigment was extracted using a mixture of acetone-methanol (7:3 v/v). The antimicrobial, antioxidant, antiviral, and anticancer activities of the green pigment produced by Streptomyces sp.W4 were investigated. The pigment was characterized using FTIR, Raman spectroscopy, EDX and GC–MS. The results revealed that the pigment has antibacterial and antifungal activity and also showed inhibition of HAV 78% but its antiviral activity against the Adenovirus was weak. The results proved the safety of the pigment toward normal cells and anticancer activity against three different cancer cell lines HepG-2 (liver cancer cell line), A549 (lung cancer cell line), and PAN1 (pancreas cancer cell line). The pigment was combined with 9 antibiotics and then tested against the Gram-negative bacterium Enterococcus faecalis using disc diffusion bioassay. LEV showed an antagonistic effect, while CXM and CIP showed a synergistic effect.

Ceropegia rupicola, Acokanthera schimperi, Jatropha variegate Antimicrobial, Cytotoxicity and Phytochemical

In Asian countries, a variety of medicinal plants are utilized in folk medicine for hemostatic and antiseptic. The methanolic extracts of Jatropha variegata, Acokanthera schimperi, and Ceropegia rupicolawere tested for the following purposes: first, isolation of endophytic fungi, second, antimicrobial and cytotoxicity activity, and third, phytoconstituents of the methanolic extract using the GC-Mass technique. The plants came from a variety of Yemeni locations. Endophytic fungus strains were isolated and identified using starch casein and potato dextrose agar medium. Methanol was used to extract the stem and leaves of the plants. The agar well diffusion method was used to test the antibacterial properties of the produced methanolic extracts against four Gram-negative bacteria, three Gram-positive bacteria, and one species of yeast. The IC50 values for the most promising extracts are calculated. The methanolic extracts were also subjected to a phytochemical screening by GC-Mass. A total of forty fungal endophyte isolates were isolated. Fusarium spp. (15), Phomaspp. (6), Alternaria alternata (5), White Sterile mycelia (6), Dark Sterile mycelia (6), Cochlioboluslunatus (1), and Mucor spp. (1). were found throughout the identification process. The highest yield extract was obtained from Jatropha variegate (45.4%), followed by Acokanthera schimperi (32.3%), and finally Ceropegia rupicola (20.5%). Only Gram-negative bacteria were found to have antibacterial activity, and the most effective extract was Acokanthera schimperi against the majority of microorganisms tested (8–18mm). The methanolic extracts of Jatropha variegate had the best antibacterial activity against Klebsiella pneumoniae ATCC 13883015, with a zone of inhibition (45 mm). Furthermore, Ceropegia rupicola extract had the highest inhibitory zone against Candida albicans ATCC 700012 (20mm). In addition, the IC50 results showed that the Jatropha variegate extract on A549 had an IC50 of 84.1ug/ml and a PC3 of 39.4ug/ml. The IC50 for Ceropegia rupicola extract was 49.4ug/ml in A549 cells and 34ug/ml in PC3 cells. Also, the screening for phytochemicals of the three plant extracts revealed the presence of many chemicals such as ethanol, formic acid, and others that may be responsible for the observed actions.

Antiproliferative activity of antimicrobial peptides and bioactive compounds from the mangrove Glutamicibacter mysorens.

The Glutamicibacter group of microbes is known for antibiotic and enzyme production. Antibiotics and enzymes produced by them are important in the control, protection, and treatment of chronic human diseases. In this study, the Glutamicibacter mysorens (G. mysorens) strain MW647910.1 was isolated from mangrove soil in the Mangalore region of India. After optimization of growth conditions for G. mysorens on starch casein agar media, the micromorphology of G. mysorens was found to be spirally coiled spore chain, each spore visualized as an elongated cylindrical hairy appearance with curved edges visualized through Field Emission Scanning Electron Microscopy (FESEM) analysis. The culture phenotype with filamentous mycelia, brown pigmentation, and ash-colored spore production was observed. The intracellular extract of G. mysorens characterized through GCMS analysis detected bioactive compounds reported for pharmacological applications. The majority of bioactive compounds identified in intracellular extract when compared to the NIST library revealed molecular weight ranging below 1kgmole-1. The Sephadex G-10 could result in 10.66 fold purification and eluted peak protein fraction showed significant anticancer activity on the prostate cancer cell line. Liquid Chromatography-Mass Spectrometry (LC-MS) analysis revealed Kinetin-9-ribose and Embinin with a molecular weight below 1 kDa. This study showed small molecular weight bioactive compounds produced from microbial origin possess dual roles, acting as antimicrobial peptides (AMPs) and anticancer peptides (ACPs). Hence, the bioactive compounds produced from microbial origin are a promising source of future therapeutics.

Aktivitas Daya Hambat Isolat Actinomycetes dengan Lama Fermentasi yang Berbeda terhadap Pertumbuhan Klebsiella pneumoniae

Actinomycetes are filamentous, Gram-positive bacteria widely distributed in soil and can produce secondary metabolites useful as antibiotics. The synthesis of secondary metabolites of Actinomycetes is influenced by the growth conditions of each isolate, including the fermentation period. Klebsiella pneumoniae is an opportunistic bacterial pathogen causing lower respiratory tract infection. Objectives: To determined the inhibition activity of Actinomycetes with different fermentation periods against the growth of K.pneumoniae. Methods: Actinomycetes were isolated from soil in Kebun Raya Bogor and grown on Starch Casein Agar (SCA) medium before being fermented in liquid media consisting of 2% of mannitol, 2% of peptone, and 1% of glucose with an incubation time of 6,7, and 8 days. An antibacterial activity test was conducted on Mueller Hinton Agar (MHA) medium using the well diffusion method. Results: The activity of Actinomycetes isolates with fermentation duration of 6,7 and 8 were able to inhibit the growth of K. pneumoniae with an average inhibition zone of 4,46 mm; 4,94 mm; and 5,04 mm, respectively. Actinomycetes' inhibitory mechanism ranges from inhibition of cell wall synthesis to nucleic acid and protein synthesis. One Way ANOVA test showed significant differences between the inhibition zone produced by each fermentation group. Conclusion: Actinomycetes isolates have antibacterial activity against the growth of K. pneumoniae with the highest activity achieved by Actinomycetes with a fermentation time of 8 days.Keywords: Actinomycetes, antibacterial, fermentation, Klebsiella pneumoniae

Presence of Actinomycetes in agarwood tissues of Aquilaria crassna: A preliminary study

Agarwood is a valuable resin produced inside certain tree species of the family Thymalaeaceae distributed in the Asian region. Agarwood production occurs as a defense mechanism when the trees are under physical or biological stresses. However, the formation of agarwood resins in significant extractable quantities due to natural stress conditions is rare; therefore, the farmers use various methods to induce its formation artificially. Certain fungal species such as Fusarium and Aspergillus become more popular among them to produce high-quality agarwood. However, studies are rare on using other microbial organisms such as Actinomycetes, which exhibit properties of both bacteria and fungi. Among the agarwood-producing species, Aquilaria crassna is one of Asia's most commonly planted species for agarwood production. This species was introduced to Sri Lanka in 2012 for mid and lower elevations of the wet zone. Due to the lack of studies on agarwood resin formation by non-fungal microbial methods, the present study attempted to identify the presence of Actinomycetes species in agarwood resinous tissues of A. crassna. Agarwood resinous tissue samples were collected from four plantations in the wet zone of the country. Surface sterilized, small sized tissues were placed on starch casein agar medium and incubated at room temperature for ten days. Using the morphological and microscopic characteristics, it was possible to identify Nocardia, Psuedonocardia, and three Streptomyces species with varying abundance. The species level should be confirmed using molecular analysis, and their potential for agarwood resin formation inducement should be tested by re-inoculating to the healthy A. crassna trees.

Antimicrobial Potential of Streptomyces spp. Isolated from the Rift Valley Regions of Ethiopia

The study was undertaken to isolate, screen, and identify actinomycetes with antimicrobial metabolites. Twenty-one composite soil samples were randomly collected from various unique agroecological niches in the Rift Valley of Ethiopia. The soil samples were serially diluted and spread on starch casein agar medium supplemented with 50 μg/ml cycloheximide and 25 μg/ml nalidixic acid. Two hundred and forty-nine (249) actinomycetes cultures were isolated and screened by cross streaking against various human pathogens. Twenty-four isolates with pronounced antimicrobial activity were selected for identification and further screening. Among the isolates, 172 (69.1%) showed antimicrobial activities against tested pathogens. The inhibition zone of the isolates ranged from 5 ± 0.31 to >40 mm during primary screening. The antimicrobial activity of the crude extracts of promising isolates showed a statistically significant difference (P < 0.05) between them and the control. The isolates RVE129 and RVE217 showed the maximum zone of inhibition at 27 ± 0.6 mm and 26 ± 0.6 mm, respectively, against S. aureus, and the results were higher than the standard drug streptomycin (25 ± 0.58 mm). The inhibition zone of crude extracts from RVE129 was at the maximum of 22 ± 0.0 mm against P. aeruginosa, almost comparable to the standard drug streptomycin (24 ± 0.58 mm). Crude extract from the isolates RVE129 and RVE187 showed higher inhibition zones of 22 ± 0.6 mm and 16 ± 0.33 mm against A. niger ATCC10535, which, however, were smaller than those obtained with the standard drug amphotericin B (29 ± 0.6 mm). Twenty-four actinomycete strains with remarkable bioactivity were characterized using various cultural, morphological, physiological, and biochemical characteristics and assigned under the genus Streptomycetes. The finding of the current study indicates that Streptomyces sp. isolated from the Rift Valley of Ethiopia was found to possess a broad spectrum of bioactivity against a range of human pathogens.

Antimicrobial, Antigenotoxicity, and Characterization of Calotropis procera and Its Rhizosphere-Inhabiting Actinobacteria: In Vitro and In Vivo Studies.

Calotropis procera (C. procera) is a wild shrub that is a medicinal plant found in abundance throughout Saudi Arabia. In this study, we investigated the phytochemical composition and antigenotoxic properties of the ethanolic extract of C. procera, in addition to the antimicrobial activity of the plant and its rhizospheric actinobacteria effects against pathogenic microorganisms. Soil-extract medium supplemented with glycerol as a carbon source and starch–casein agar medium was used for isolation of actinobacteria from rhizosphere. From the plant, a total of 31 compounds were identified using gas chromatography/mass spectrometry (GC–MS). The main components were α-amyrin (39.36%), lupeol acetate (17.94%), phytol (13.32%), hexadecanoic acid (5.55%), stigmasterol (3.16%), linolenic acid (3.04%), and gombasterol A (2.14%). C. procera plant extract’s antimicrobial activity was investigated using an agar well-diffusion assay and minimum inhibitory concentration (MIC) against six pathogenic microbial strains. The plant extract of C. procera was considered significantly active against Staphylococcus aureus, Klebsiella pneumonia, and Escherichia coli, with inhibition zones of 18.66 mm, 21.26 mm, and 21.93 mm, respectively. The plant extract was considered to be a moderate inhibitor against Bacillus subtilis, with MIC ranging from 0.60–1.50 mg/mL. On the other hand, the isolated actinobacteria were considered to be a moderate inhibitor against S. aureus (MIC of 86 µg/mL), and a potent inhibitor, strain CALT_2, against Candida albicans (MIC of 35 µg/mL). The 16S rRNA gene sequence analysis showed that the potential strains belonged to the genus Streptomyces. The effect of C. procera extract against cyclophosphamide (CP)-induced genotoxicity was examined by evaluating chromosome abnormalities in mouse somatic cells and DNA fragmentation assays. The current study revealed that oral pretreatment of C. procera (50, 100, and 200 mg/kg b.w.) for 1, 7, and 14 days to cyclophosphamide-treated animals significantly reduced chromosomal abnormalities as well as DNA fragmentation in a dose-dependent manner. Moreover, C. procera extract had antimicrobial and antigenotoxic effects against CP-induced genotoxicity.

Production, Characterization, and Antimicrobial Activity of Pigment from Streptomyces Species

Production of potential pigments using the bacterial source can be an important area of research to disclose its possible value in diverse industrial applications. Soil samples from various terrestrial, rhizosphere, and forest habitats were collected from Karnataka, and pigment‐producing actinomycetes were recovered. In the present investigation, 25 strains were isolated using starch casein agar medium; further, phenotypic, biochemical, and morphological and the 16S rRNA gene sequence studies have suggested that the strain belonged to the Streptomyces species, strain BJZ10, an actinomycetes. With higher production of pigments, starch casein broth medium was used, and the extraction of actinobacterial pigments was done by exclusively using methanol solvent. The pigment was characterized by UV‐visible spectroscopy; absorption spectra range from 220 to 250 nm. The FTIR characterization was carried out, and the spectrum obtained for the strain BJZ10 indicated alkane, alkyls, alkynes, alcohols, esters, and sulfate functional groups. Antibacterial activity of the pigments was tested against Bacillus cereus (Gram‐positive) and Escherichia coli (Gram‐negative), and significant results were compared. The present study revealed the brown pigment synthesized by Streptomyces sp. strain BJZ10, known for its potential antimicrobial activity.

Isolation and Characterization of Actinobacteria Involved in Biodegradation of Herbicide Sulfosulfuron (Apyros 75 WG)

68 Actinobacteria strains were isolated and in vitro tested for their abilities to grow in the presence of sulfosulfuron (Apyros 75 WG) in starch casein agar medium supplemented at a concentration 10 fold of the recommended dose. The results revealed, that only 5 isolates showed abundant growth, 30 isolates moderate growth, 6 isolates a weak growth and 27 no growth. The ten most tolerant Actinobacteria were tested for their abilities to utilize sulfosulfuron (Apyros75 WG) herbicide as a sole source of carbon and energy. From these, only two strains (A1 and A12) showed an abundant growth on ISP9 agar medium containing sulfosulfuron (Apyros 75 WG) as a sole source of carbon and energy. One strain designated A1 tolerant and able to utilize sulfosulfuron (Apyros 75 WG) as sole source of carbon and energy, was selected for biodegradation assays of this herbicide, on ISP9 broth supplemented with sulfosulfuron (Apyros 75 WG) (100 mg L−1) as a sole source of carbon and energy. Strain A1 reduced the concentration of sulfosulfuron from 75 mg L−1 to 39.8, 2.7, 2.3 and 1.3 mg L−1, respectively, on first, fifth, tenth and fifteenth day of incubation, when it had degraded 98% of initial amount and had reached its highest cell mass of 0.074. Morphological characters and chemotaxonomic parameters indicated that strain A1 belong to the genus of Streptomyces.

Isolation, characterization and antibacterial activity of actinomycetes from soils of the Langun-Gobingob Cave Complex in Calbiga, Samar

The discovery of rare Actinomycetes from unexplored areas has proven to be a promising source of new antimicrobials. The Langun-Gobingob cave complex in Calbiga, Samar, Philippines is the second largest cave complex in Asia, yet this is the first reported search for actinomycetes conducted in the area. The main objective of this study was to assess the antibacterial activities of culturable actinomycetes from soil samples from the Langun-Gobingob cave complex. Five actinomycete colonies were isolated in pure culture from soil samples using Starch-Casein Agar medium. The isolates were identified by their morphological characteristics and they were characterized by 16S rRNA sequencing. The sequences were matched to sequences uploaded to the NCBI Basic Local Alignment Search Tool (BLAST). Four of the isolates belonged to the Genus Streptomyces and one belonged to the Genus Nocardia. The antibacterial potential of the isolates was tested against gram-positive and gram-negative test bacteria. One Streptomyces isolate exhibited antibacterial activity against the two gram-positive test bacteria. The limited cave area sampled in this study leaves the cave complex mostly unexplored in the search for antibacterial secondary metabolites.

Antimicrobial and antifungal activity of soil actinomycetes isolated from coal mine sites.

In the current study, twenty-eight soil samples were collected from coalmine sites of Telangana, India. The isolates were purified and identified based on their culture characterization on oatmeal agar, glycerol asparagine agar, yeast extract-malt extract agar, inorganic salt starch agar, and starch casein agar medium. Further, the supernatant of all the isolates were tested for antimicrobial and antifungal activities. The biochemical and microscopic studies of isolated strains results indicates the potential isolate strains belongs to Streptomyces genus. Among all the strains the biological activity of BHPL-KSKU5 showed higher anti-bacterial and anti-funagal activity. The molecular characterization of BHPL-KSKU5 16s rDNA gene sequence and phylogenetic tree showed that is mostly related to the Streptomysis felleus (S. felleus) strain. This isolate was submitted to gene bank NCBI with accession number MH553077. In addition, physiological studies such as utilization of carbon, nitrogen, amino acid sources of potential isolated were studied. Further, optimization, purification and characterization of the novel compound producing strain may be helpful for discovering the new therapeutic microbial agent.

Molecular Identification and Antibacterial Activity of Streptomyces spp. Isolated from Sulaymaniyah Governorate Soil, Iraqi Kurdistan

Soil is play an important role for reserve abundant groups of microorganisms, especially Streptomyces. Streptomyces are recognized as prokaryotes, aerobic and Gram-positive bacteria with high Guanine + Cytosine contents in their DNA. These groups of bacteria show filamentous growth from a single spore and they are normally found in all kinds of ecosystems, including water, soil, and plants. A total of three Streptomyces strains were isolated from soil of the sides of Darband Ranya in Sulaimani governorate. Different approaches were followed for the identification of the isolated stains. Morphological and cultural properties of these isolates have shown that the isolates are belonging to the genus Streptomyces. Desired colonies of the isolates were distinguished and separated from other bacteria on the basis of colony morphology, pigmentation, ability to produce a different color of aerial hyphae, and bottom mycelium on raffinose-histidine agar and starch-casein agar media. In addition, analysis of phylogenetic tree based on 16S rRNA gene sequences, the strains related the genus. KS010 isolates had the highest identity (99.32%) with the type strain of Streptomyces atrovirens, while KS005 and KS007 isolates were most closely related to Streptomyces lateritius by identity 99.32%. The isolated test strains were also active against Gram-negative and Gram-positive pathogenic bacteria following co-cultivation technique. However, further study should be done to identify the types of these antibacterial compounds.

ISOLATION OF ACTINOMYCETES FROM THE SEDIMENTS OF PICHAVARAM MANGROVE FOREST, SOUTH INDIA AND ANALYSING THEIR ANTIBACTERIAL EFFICACY

Objective: The aim of the present investigation is to isolate actinomycetes from the sediments of Pichavaram mangrove forest, South India, and to screen for their antibacterial efficiency.&#x0D; Methods: Actinomycetes were isolated by culturing the samples in Starch Casein Agar medium; they were screened primarily for their antibacterial efficiency against Gram-positive and Gram-negative bacterial organisms. Solvent extraction was done with 50% (percentage) ethyl acetate, crude extracts of actinomycetes were prepared at different concentrations using dimethyl sulfoxide and treated against the bacterial organisms. Antibacterial assay was done in Mueller–Hinton agar medium.&#x0D; Results: Thirteen actinomycetes were isolated; among them, four actinomycete isolates (Pichavaram mangrove actinomycete 2 [PMA2], PMA6, PMA9, and PMA13) exhibited antibacterial activity.&#x0D; Conclusion: Isolate PMA2 exhibited very strong antibacterial activity and isolate PMA13 is weakly active against the tested bacterial organisms.

Antimicrobial potential of Streptomyces sp. to the Gram positive and Gram negative pathogens

BackgroundThe occurrence of drug resistant infectious disease causing microbial pathogens was highly spreaded because of the wide level application of the commercially available antimicrobial agents. However, the eradication of the microbial pathogens was of huge demand. Although, many antimicrobial compounds were commercially available in the market however the spreading of the pathogens were hugely increased. Actinomycetes produce various secondary metabolites against pathogenic bacteria and fungi. The present investigation aimed to study the antimicrobial potential of the Streptomyces sp. towards infectious diseases causing pathogens. MethodsCulture dependable isolation techniques were followed for the isolation of the active actinomycetes isolates and the antimicrobial properties of the actinomcyetes were detected by primary screening techniques using modified starch casein agar medium. The active isolate was confirmed by various biochemical and morphological techniques. ResultsIn this study, 10 actinomycetes were isolated and later five were selected for secondary screening and noted significant activity against Enterobacter aerogenes and Proteus mirabilis. Among the selected Streptomyces sp., ES2 showed potent activity against selected microbes and was identified as Streptomyces sp. The studied isolates were resisitant towards streptomycin (10μg), ampicillin (50μg) and ciprofloxacin (5μg). The organic solvent extracts of the promising isolate ES2 prononunced comparatively better inhibitory properties towards the studied pathogenic bacteria. ConclusionOverall, the present study evidenced that the actinomycetes were promising candidate for the eradication of the pathogenic strains.

Optimization of Fibrinolytic Enzyme from Streptomyces althioticus BN22

Fibrinolytic enzyme plays an important place in the medical field for cardiovascular disease. Only few studies have reported the fibrinolytic enzyme production from marine actinomycetes, and Streptomyces sp. In this study 114 actinomycetes isolates from different marine soil samples were isolated by using starch casein agar media. These 114 actinomycetes strains were tested for fibrinolytic activity by fibrin plate method. 14 isolates showed fibrinolytic activity amongst the 114 actinomycetes strains, and this was identified based on the clear zone of formation around the colonies. The potent strain of Streptomyces althioticus BN22 was isolated and this was confirmed by 16SrRNA sequencing. The optimization of nutritional and physical parameters for fibrinolytic enzyme production from marine soil was studied for the isolate Streptomyces althioticus. The effects of nutritional and physical parameters were studied for the optimum production of a fibrinolytic enzyme from Streptomyces althioticus BN22 by submerged fermentation. The effects of initial pH, temperature and incubation time were studied. The maximum enzymatic activity was obtained with an initial pH7.5 (45,765 Eu/ml), incubation temperature 50°C (50,827 Eu/ml) and incubation time for (41,175 Eu/ml). The effects of various carbon and nitrogen sources were studied. The results indicated that glucose (51,570 Eu/ml) as carbon source and yeast extract (54,625 Eu/ml) as nitrogen source showed the highest fibrinolytic enzyme activity among them. Thus this study showed its originality by focus, that marine Streptomyces althioticus is an important source for fibrinolytic enzyme production.

Strain improvement for the production of antioxidant activity from Bacillus sp. by induced stress

In the present study, water samples were collected from different beaches in Chennai such as Marina beach, Elliott beach, VGP Golden beach and Kovalam beach to evaluate the best source for antioxidants. The bacteria were isolated on Starch Casein Agar media and screened for the antimicrobial activity. Among 11 isolates, 5 isolates showed antimicrobial potential which were further evaluated for the DPPH (2,2-diphenyl-1-picrylhydrazyl) scavenging activity. Among 5 isolates, one sample showed significant DPPH scavenging activity with half minimal inhibitory concentration of (IC50)344.754µg/mL. The maximum antioxidant production was observed at pH 7 and at temperature of 37ºC with an IC50 of 188.66µg/mL and 293.76µg/mL respectively. The potent antioxidant producing strain was subjected for mutagenesis. In physical mutagenesis, the organism exposed for UV light for 25 minutes showed maximum antioxidant production with an IC50 of 133.55µg/mL. This mutant strain was then subjected for chemical mutagenesis with the addition of different concentrations of Ethidium bromide such as 10µL, 20µL, 30µL, 40µL and 50µL. The mutant strain obtained with the addition of 20µL Ethidium bromide (EtBr) showed significant antioxidant activity with an IC50 of 325.4µg/mL. The sample was purified by solvent extraction method and was evaluated for antioxidant production. The analyte was subjected to HPLC (High Performance Liquid Chromatography) analysis to avince the presence of antioxidants. The ethyl acetate extract showed the total phenolic content of 0.892 mg GAE/g of dry extract. It also showed the total flavonoid content of 0.522mg RE/g of dry extract.

Diversity of antibiotic-producing Actinomycetes in mangrove forest of Torosiaje, Gorontalo, Indonesia

Retnowati Y, Sembiring L, Moeljopawiro S, Djohan TS, Soetarto ES. 2017. Diversity of antibiotic-producing actinomycetes in mangrove forest of Torosiaje, Gorontalo, Indonesia. Biodiversitas 18: 1453-1461. Actinomycetes for antibiotic production have been studied at various extreme environments. Mangrove forest of Torosiaje in Gorontalo Province, Indonesia has unique geomorphological conditions where the forest is surrounded by karst ecosystem consisting of fringe and overwash types. Therefore, the objective of this study was to analyze the distribution and diversity of antibiotic-producing Actinomycetes in various rhizosphere including different locations and mangrove species. Samples of rhizosphere soil were collected in the depth of 0-10 cm, which was then subjected to detailed physicochemical analysis. Actinomycetes were collected through heating pre-treatment (60oC for 15 min) followed by culturing in the Starch Casein Agar medium supplemented with cycloheximide and nystatin. The screening process of antibiotic-producing Actinomycetes was based on Agar block method against pathogenic microorganisms. Grouping of Actinomycetes was determined by ARDRA fingerprinting analysis. The diversity of Actinomycetes was analyzed based on sequencing of 16S rDNA. The results showed that the distribution of Actinomycetes was found in overwash type, middle zone and upper zone of fringe type including rhizosphere of 7 species of mangrove. The highest population of Actinomycetes was found in rhizosphere of R. mucronata at the overwash type, and the lowest one found in rhizosphere of R. apiculata at the middle zone of fringe type. A total of 77 isolates amongst 167 isolate collection showed antibacterial activities. Forty seven representatives from 77 antibacterial-activities isolates were selected using ARDRA for partial characterization according to their phylogenetic diversity. Sequencing and analysis of 16S rDNA from selectedrepresentative isolates displayed the presence of members associated with Actinomycetes genera such as Streptomyces, Amycolatopsis, Saccharomonospora, and Nocardiopsis. The member of genus Streptomyces such as Streptomyces qinglanensis and Streptomyces champavatii were distributed across locations. Genus Saccharomonospora and Nocardiopsis were mostly found at the overwash type, while Amycolatopsis was found at the upper zone of fringe type.

ISOLATION AND SCREENING OF NOVEL STREPTOMYCES FROM SEDIMENTS OF BAY OF BENGAL NEAR SRIKAKULAM COAST

Objective: The Objective of this research is to isolate and screen potent antibiotic producing actinomycetes from unexplored regions of Bay of Bengal Near Srikakulam Coast.Methods: The isolation and screening were done on starch casein agar media. Antimicrobial activity screening was done using cross streak method and agar diffusion method. Further, Morphological characterization of the isolate was done using Scanning electron Microscopy (SEM-JSM 6610). Further, the cultural characterization of the isolates was done using standard protocols.Results: A total of sixty-eight isolates of Actinomycetes were collected from marine sediments of coastal line of Srikakulam district of Andhra Pradesh, which is one of the longest coastal lines of Andhra Pradesh. Due to its longest coastal line and geographical distribution, there are promising chance of diversity of the microorganisms and antibiotic producing potential. Each isolate was tested against various bacteria. Among all the isolates twelve isolates were promising and showed good antimicrobial activity. Within twelve isolates the isolate A-10 was very promising and showed antibiotic potential against most of the tested bacteria. This isolate appears to produce high anti-bacterial compounds on potato dextrose agar and nutrient agar medium respectively by using the agar diffusion method. The potent Actinomycete A-10 was further characterised by morphological methods consist of macroscopic and microscopic methods. The mycelium structure, colour and arrangement of conidiophores were observed through the oil immersion (100X). The strain was grown on Starch-Casein agar medium for morphological study using Scanning Electron microscopy. Scanning Electron micrographs of the strain A-10 results that the spore chain was straight and with more than 20 spores per chain. The spores are cylindrical in shape and had a smooth surface. Whirls, sclerotic granules, sporangia and flagellate spores were not observed. Further various biochemical tests performed for the identification of potent isolates are as follows: Melanin reaction, H2S production, tyrosine reaction, starch hydrolysis, casein hydrolysis, gelatin hydrolysis, milk coagulation and peptonization, nitrate reduction, temperature range of growth, pH tolerance and cell wall type by comparing all these results with the Bergey’s manual of Determinative Bacteriology and the organisms were identified. The isolate was identified as Streptomyces sp.Conclusion: Thus the result of antibacterial screening represents that the coastal area of Srikakulam is a potential source of antibiotic producing actinomycetes. Further the phylogenic tree construction and purification of the antibiotic principle from A-10 were under investigation.Keywords: Actinomycetes, Marine Sediments, Starch-Casein agar, SEM

Larvicidal potency of marine actinobacteria isolated from mangrove environment against Aedes aegypti and Anopheles stephensi.

The marine soil samples were collected from different locations of Parangipettai mangrove ecosystem, Vellar estuary, southeast coast of India. Totally 30 different marine actinobacteria were isolated by serial dilution plate technique on starch casein agar medium. The isolated actinobacteria were investigated for their larvicidal activity against Aedes aegypti and Anopheles stephensi mosquitoes. Streptomyces fungicidicus, S. griseus, S. albus, S. alboflavus and S. rochei were identified as potential biocide producers. Based on the antimicrobial activity, five strains were chosen for larvicidal and pupicidal activity. Among the crude extracts tested, the S. alboflavus extract showed significant activity against Ae. aegypti (LC50 1.48±0.09 and LC90 3.33±0.22) and An. stephensi (LC50 1.30±0.09 and LC90 3.13±0.21). Five isolates have shown a most significant mortality rate of the Ae. aegypti and An. stephensi mosquito larvae. This is an ideal eco-friendly approach for the control of Japanese encephalitis vectors, Ae. aegypti and malarial vector An. stephensi.