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Antibacterial Activities of Actinomycete Isolates Collected from Soils of Rajshahi, Bangladesh

Md. Ajijur Rahman
Department of Pharmacy, University of Rajshahi, Rajshahi 6205, Bangladesh

Mohammad Zahidul Islam
Department of Parasitology, Aichi Medical University, Aichi-ken 480-1195, Japan and 3Department of Biology, The Catholic University of America, Washington, DC 20064, USA

Md. Anwar Ul Islam
Department of Pharmacy, University of Rajshahi, Rajshahi 6205, Bangladesh

Abstract/ Executive Summary:

This study was performed to isolate actinomycete colonies having antibacterial activity from soil samples collected from different places around Rajshahi, Bangladesh. Thirty actinomycete colonies were isolated in pure culture from five soil samples using Starchcasein-nitrate-agar medium. The isolates were grouped in five color series based on their aerial mycelia color and screened for their antibacterial activity against a range of test bacteria. Sixteen isolates (53.3%) were found to have moderate to high activity against four gram-positive and four gram-negative bacteria. Since many isolates showed inhibitory activity against indicator bacteria, it is suggestive that Bangladeshi soil could be an interesting source to explore for antibacterial secondary metabolites.

Keywords: Actinomycete, Soil samples collected, Soils, Indicator bacteria,

Location: The different places of Rajshahi District, Bangladesh

Start Date: 00-07-2017

End Date: 00-12-2017

Program Area: Crop-Soil-Water Management

Commodity: Physico-Chemical

Objectives:

To isolate actinomycete colonies having antibacterial activity from soil samples collected from different places around Rajshahi, Bangladesh

Methodology:

Soil Samples. Soil samples were collected from the different places of Rajshahi District, Bangladesh. Samples were collected from various depth of the earth surface, ranging from layers just beneath the upper surface to 1 meter depth. They were collected in the sterile small plastic tubes and properly labeled with the date of collection. Twenty soil samples were collected within a period of six months (July, 2007–December, 2007). The collected soil samples were dried in a hot air oven at 60–65⁰C for 3 hours and stored in 4⁰C until examined. 2.2. Isolation of Pure Culture of Actinomycetes. Thirty actinomycete strains were isolated and obtained as pure culture by using standard microbiological method. From each soil sample, 1 gm of dried soil was suspended in 9 mL sterile water, and successive serial dilutions were made by transferring 1mL of aliquots to 2nd test tube containing 9 mL of sterile water, and in this way dilutions up to 10−4 were prepared. Each time the contents were vortexed to form uniform suspension. An aliquot of 0.1 mL of each dilution was taken and spread evenly over the surface of starch-caseinnitrate-agar medium supplemented with cycloheximide (100 µg/mL) on 16 cm petridishes. Plates were incubated at 32⁰C and monitored for 7 days. The colonies were carefully counted by visual observation and c.f.u per gram of soil was determined. Plates those gave 100–150 colonies were chosen for further isolation in pure culture. Suitable colonies those showed Streptomyces like appearance under light microscope were recultivated several times for purity. The purified actinomycetes were preserved on yeast-extract-glucose-agar slants at 4⁰C for two months and at −20⁰C in the presence of glycerol (15%v/v) for longer periods. Color Grouping of the Isolates. The color of the aerial mycelia and pigment production by the isolates were determined on yeast-extract-glucose-agar plates after 7 days of incubation at 32⁰C. The color of the substrate mycelia and those of the soluble pigment were determined according to the National Bureau of Standards Color Chart. Screening of Antimicrobial Activities of Pure Isolates. Preliminary screening for antibiotic activity of the isolates was done by using streak-plating technique on yeast-extract-glucose-agar medium. Each pure isolates were streaked individually on different agar plates in a single line. The plates were then incubated at 32⁰C for 5 days to allow the isolates to secrete antibiotics into the medium. After the incubation period, the properly diluted test organisms were cross streaked along the line of fully grown isolates. Each streaking was started near the edge of the plates and streaked toward the Streptomyces growth line. The plates were then incubated for 12 hrs at 37⁰C, and the zone of inhibition was measured using a millimeter scale. 2.5. Test Organisms. Eight test organisms were used to test the antibiotic activity of the isolates. Four of them were gram-positive and four were gram-negative bacteria. Grampositive species were Staphylococcus aureus ATCC-259233, Streptococcus agalactiae, Bacillus cereus, and Bacillus megaterium QL-40. Gram-negative strains were Escherichia coli FPFC-1407, Shigella flexneri AL-30372, Shigella dysenteriae AL-35587, and Shigella sonnei. They were maintained in nutrient agar slants at 4⁰C.

Source of Information: Biotechnology Research International; Volume 2011, Article ID 857925, 6 pages

Article Link (Online Journal): doi:10.4061/2011/857925

Funding Source:

Budget:

Total Budget (Tk.) :

Achievement/Findings:

Comparing the above mentioned results with this study, we can conclude that the soil samples of Rajshahi are rich source of actinomycetes which produce metabolites inhibitory to bacterial pathogens. We found that 53.3% of the isolated colonies were active against the test bacteria. MARS17 and MARS-27 were very active and showed very large zone of inhibition. Future studies will be done to identify the active isolates up to the species level. The type of antimicrobial agents produced by these isolates will be investigated as well.

Knowledge Management: Journal