Isolation, Characterization, and Identification of Organohalide Degrading Soil Bacteria

• Main Author: Nanda Khoiriyah, Nurul
• Format: Final Project
• Language: Indonesia
• Subjects: Kimia
• Online Access: https://digilib.itb.ac.id/gdl/view/37965
• Institution: Institut Teknologi Bandung

Organohalides are compounds widely used as precursors and solvents in herbisides, pesticides, insecticides, plastics, and other industries. These compounds are of the biggest pollutants harmful to the environment. One way to reduce the organohalide pollutants is bioremediation, utilizing dehalogenase producing soil bacteria to decompose the organohalides. In this study, isolation, characterization, and identification of organohalide degrading soil bacteria were carried out. Isolation of the bacteria was carried by growing the bacteria from soil samples in liquid minimal medium (MM) without NaCl that contain 10 mM of monochloroacetic acid (MCA) as the sole of carbon source at 37oC with 150 rpm agitation for aeration. The growing bacteria were then transferred to the same solid medium and the growing colonies were further purified by streaking on the same solid medium to obtain single colonies. A total of 19 single colonies were isolated and 8 colonies were selected for testing its ability to grow in the liquid MM without NaCl with MCA concentration improved from 5 mM to 50 mM. These 8 colonies showed different growing ability on different MCA concentration, indicated by the increase of OD450 up to a certain concentration and then plummeted when MCA concentration in the medium become toxic to the bacteria. The maximum MCA concentration that can be tolerated by the bacteria varies between 15 mM to 25 mM. Five isolates with different growth characteristic were selected for further analysis on its ability to degrade MCA. This was done by determining the amount of Cl- ions released during the growth of bacteria. The Cl- ion in the medium was measured by Bergmann and Sanik method. Four isolates with best MCA degrading ability are then chosen. These four isolates were identified as gram negative cocci bacteria that capable to degrade MCA between 3.98% to 4.80%. These isolates were suspected as inductively producing dehalogenase as this enzyme is only produced when there is a substrate in the medium. Results from 16S rDNA typing indicated that colonies number 3, 5, and 7 are Cupriavidus pampae, while colony number 6 is Pseudomonas boreopolis.