Investigation of Bacterial Transport in the Large-Block test, A Thermally Perturbed Block of Topopah Spring Tuff
- PDF / 769,734 Bytes
- 8 Pages / 370.26 x 574.38 pts Page_size
- 98 Downloads / 149 Views
1151 Mat. Res. Soc. Svmo. Proc. Vol. 556 c 1999 MatArials ResAarch Societv
EXPERIMENT Two bacterial species, Bacillus subtilis and Arthrobacteroxydans, were isolated from the Yucca Mountain tuff. Neither of the parental strains is resistant to the simultaneous presence of two antibiotics, streptomycin and rifampicin. However, natural mutants of these parents that can grow under the simultaneous presence of streptomycin and rifampicin were selected from these species by laboratory procedures [5]. The characteristics of double-drug resistance distinguish the natural mutants from the indigenous species. The mutants were cultured in nutrient broth that contained 0.5% agar at 30'C for 3 days. At ambient desert conditions, the inoculated agar medium had a jellylike consistency and was injected (800 ml of 1.Ox 107 cells/ml A. oxydans + 1.2x 10' cells/nil B. subtilus per heater hole), using a large syringe, into the 5 heater boreholes of the large block hours before heating was initiated. The procedures for culturing these two double-drug-resistant bacteria and their installation in the large block have been described [4, 5]. To collect bacterial samples from the collection boreholes, a piece of cotton cloth (which can adsorb bacterial cells) was inserted the length of each of the three boreholes (NO 1,N02, and E03) that were approximately 5 ft below the injection (heater) ports. These installed cotton cloth strips stayed in the boreholes until the next sampling, at which time they were removed and replaced with new ones. Sterile collection methods are not necessary because of the double-drug-plating procedures used to screen the samples. Alternatively, bacterial cells were collected with a sterile TM swab (glass fiber or filter paper). A Pyrex glass tube, which was also installed in the collection bore holes, was pulled out of the ports with the pieces of cotton cloth. The tube was swiped with a moistened sterile glass-fiber swab (or filter paper). The swabs and filter papers were subjected to bacterial assays. Cells that were possibly present in the heater boreholes were collected after the entire LBT experiment was completed. The heaters were removed from the block, and their surfaces were swiped with moistened sterile filter paper. Cells on the bacterial samples were enumerated by patting the cotton cloth, glass-fiber swab, or filter paper on freshly prepared, R2A agar plates that contained streptomycin and rifampicin. The concentrations of streptomycin and rifampicin in the agar plates were 30 mg/l and 170 mg/l, respectively. Each sample was cultured in duplicate plates. The plates were then incubated at 30'C for 3 days. Because double-drug-resistant bacteria are uncommon in nature, bacteria thus screened are likely to be the injected species. The screened samples were submitted to tests, described subsequently, to verify the presence of Bacillus and Arthrobacter. RESULTS Hours after injection of the bacteria, the heaters of the large block were turned on, and the temperature of the large block near the heater rose s
Data Loading...
