In situ biosurfactant production by Bacillus strains injected into a limestone petroleum reservoir.

Biosurfactant-mediated oil recovery may be an economic approach for recovery of significant amounts of oil entrapped in reservoirs, but evidence that biosurfactants can be produced in situ at concentrations needed to mobilize oil is lacking. We tested whether two Bacillus strains that produce lipopeptide biosurfactants can metabolize and produce their biosurfactants in an oil reservoir. Five wells that produce from the same Viola limestone formation were used. Two wells received an inoculum (a mixture of Bacillus strain RS-1 and Bacillus subtilis subsp. spizizenii NRRL B-23049) and nutrients (glucose, sodium nitrate, and trace metals), two wells received just nutrients, and one well received only formation water. Results showed in situ metabolism and biosurfactant production. The average concentration of lipopeptide biosurfactant in the produced fluids of the inoculated wells was about 90 mg/liter. This concentration is approximately nine times the minimum concentration required to mobilize entrapped oil from sandstone cores. Carbon dioxide, acetate, lactate, ethanol, and 2,3-butanediol were detected in the produced fluids of the inoculated wells. Only CO(2) and ethanol were detected in the produced fluids of the nutrient-only-treated wells. Microbiological and molecular data showed that the microorganisms injected into the formation were retrieved in the produced fluids of the inoculated wells. We provide essential data for modeling microbial oil recovery processes in situ, including growth rates (0.06 +/- 0.01 h(-1)), carbon balances (107% +/- 34%), biosurfactant production rates (0.02 +/- 0.001 h(-1)), and biosurfactant yields (0.015 +/- 0.001 mol biosurfactant/mol glucose). The data demonstrate the technical feasibility of microbial processes for oil recovery.

Haloanaerobium kushneri sp. nov., an obligately halophilic, anaerobic bacterium from an oil brine.

Three strains, designated VS-751T, VS-511 and VS-732, of a strictly anaerobic, moderately halophilic, Gram-negative, rod-shaped bacterium were isolated from a highly saline (15-20%) brine from an oil reservoir in central Oklahoma, USA. The optimal concentration of NaCl for growth of these three strains was 2 M (12%), and the strains also grew in the presence of an additional 1 M MgCl2. The strains were mesophilic and grew at a pH range of 6-8. Carbohydrates used by all three strains included glucose, fructose, arabinose, galactose, maltose, mannose, cellobiose, sucrose and inulin. Glucose fermentation products included ethanol, acetate, H2 and CO2, with formate produced by two of the three strains. Differences were noted among strains in the optimal temperature and pH for growth, the maximum and minimum NaCl concentration that supported growth, substrate utilization and cellular fatty acid composition. Despite the phenotypic differences among the three strains, analysis of the 16S rRNA gene sequences and DNA-DNA hybridizations showed that these three strains were members of the same genospecies which belonged to the genus Haloanaerobium. The phenotypic and genotypic characteristics of strains VS-751T, VS-511 and VS-732 are different from those of previously described species of Haloanaerobium. It is proposed that strain VS-751T (ATCC 700103T) be established as the type strain of a new species, Haloanaerobium kushneri.

Substitution of DL-methionine for soybean meal as a winter supplement for gestating cows grazing native range.

A winter grazing study was conducted to determine whether DL-methionine could replace soybean meal as a N supplement for gestating beef cows. During two winters (Trial 1, n = 51; Trial 2, n = 60), crossbred beef cows grazed native foothill range. Three treatment groups were supplemented with either none (CON), DL-methionine (7.5 g Trial 1 and 9 g Trial 2) in .5 kg beet pulp carrier (BPM) or .4 kg soybean meal (SBM). Cows were supplemented individually every other day. Small differences were noted in cow BW, condition score and blood metabolites. Unsupplemented cows lost the greatest amount of BW (P less than .01) in both trials and lost more (P less than .05) condition during Trial 1 than cows fed BPM or SBM supplements. Blood samples were obtained on two consecutive days during each trial (45 d and 25 d prepartum) and analyzed for blood urea N, total bilirubin, creatinine, albumin, total protein and cholesterol. A treatment x day preparatum interaction (P less than .05) was noted for blood urea. Blood urea nitrogen declined as gestation length increased for CON and SBM cows, but blood urea of BPM-supplemented cows remained low and unchanged. In situ forage digestion was measured in 12 ruminally cannulated cows (four/treatment). In both trials, in situ rate of NDF disappearance was greater (P less than .05) for SBM than for BPM. In Trial 2, a treatment x sampling hour interaction was detected for purine concentration of whole ruminal contents; SBM maintained greater purine concentrations throughout the 48-h supplementation cycle than BPM did. Principal component analysis suggested that ruminal ammonia limited the microbial growth response to DL-methionine. Therefore, alternate-day supplementation of DL-methionine plus beet pulp did not effectively substitute for soybean meal in these trials.

Influence of body condition and lasalocid during late gestation on blood metabolites, lamb birth weight and colostrum composition and production in Finn-cross ewes.

The objective of this experiment was to examine the effects of body condition (BC) and of lasalocid (L) the last 4 wk of gestation on blood metabolite profiles, lamb birth weight and colostrum composition and production. Twenty-eight 3-yr-old Finn-Targhee ewes (73 kg) were assigned randomly within BC grouping (2.5 or 3.5) and individually fed a diet of 90% alfalfa pellets and 10% of a supplement containing either no L (C = control) or L. Gestation and 24-h postlambing weights were higher (P less than .05) for 3.5 BC versus 2.5 BC ewes. Lasalocid had no effect (P greater than .05) on ewe weight. Average number of lambs born within treatment groups were similar (P greater than .05). Total kilograms of lamb born were greater (P less than .10) for 3.5 BC ewes. Body condition 3.5 ewes had greater concentrations of total protein (P less than .10) and albumin (P less than .05) the last 4 wk of gestation than those in the 2.5 BC group. Feeding L decreased (P less than .05) blood urea-N in comparison with C ewes. Colostrum composition and production were not influenced (P greater than .05) by BC, L or number of lambs born. Serum 3-hydroxybutyrate seemed to be a good indicator of energy metabolism; albumin and blood urea-N concentrations reflected dietary protein intake. Lasalocid had a minimal effect on nutrient metabolism and productivity of ewes fed in excess of the NRC (1985) protein and energy requirements.