3.3 Collection of Sample The cow dung sample was collected in sterilized Petri plate using sterile spatula from the cow shed at Kandang Kambing N Lembu, Gombak, Kuala Lumpur. Before bacterial isolation, the samples were stored at 4°C in refrigerator for approximately 12hrs.
3.4 Isolation of Bacteria 0.5g of dung sample was suspended with 50ml 0.85% sterile NaCl solution in a 250ml conical flask, which was shaken at 180rpm for 1 hr at 37°C. Serial dilutions from 100 to 10-7 were prepared using sterilized saline solution. An aliquot of 10ul of each dilution was plated on Nutrient agar plates by spread plate method. The plates were incubated at 37°C for 72hrs. The isolated colonies were further purified using streak plate technique.
3.5 Screening…show more content… Several drops of hydrogen peroxide 3% was transferred into a sterile test tube. A colony was picked to be tested carefully with a wooden stick. The wooden stick was dipped into hydrogen peroxide 3%. An immediate bubbling reaction was observed to indicate a catalase positive result while no bubbling reaction indicates a negative result.
3.10.2.2 Starch Hydrolysis Test A single colony was streaked to be tested onto starch plate and incubated at 30˚C for five days. The starch plate was flooded with iodine after five days and observed. A clear zone of hydrolysis that indicates starch hydrolysis shows a positive result while blue color formation without any starch hydrolysis indicates a negative result.
3.10.2.3 Triple Sugar Ion (TSI) Test Colony to be tested in TSI agar was inoculated with an inoculating needle by stabbing the butt and streaking the slant. The tube was incubated at 30˚C for 24 hours. Yellow butt indicates positive reactions for acid production from dextrose. Yellow slant indicates positive reaction for acid production from lactose and sucrose. H2S production will cause blackening precipitate to occur.
3.10.2.4 Motility Test A fresh bacteria colony was stabbed into the solid medium. Growth from stabbed line indicates