Effect of Linoleic Acid Concentration on Conjugated Linoleic Acid Production by Butyrivibrio fibrisolvensA38

ABSTRACT Butyrivibrio fibrisolvens A38 inocula were inhibited by as little as 15 μM linoleic acid (LA), but growing cultures tolerated 10-fold more LA before growth was inhibited. Growing cultures did not produce significant amounts of cis-9, trans-11 conjugated linoleic acid (CLA) until the LA concentration was high enough to inhibit biohydrogenation, growth was inhibited, and lysis was enhanced. Washed-cell suspensions that were incubated anaerobically with 350 μM LA converted most of the LA to hydrogenated products, and little CLA was detected. When the washed-cell suspensions were incubated aerobically, biohydrogenation was inhibited, CLA production was at least twofold greater, and CLA persisted. The LA isomerase reaction was very rapid, but the LA isomerase did not recycle like a normal enzyme to catalyze more substrate. Cells that were preincubated with CLA lost their ability to produce more CLA from LA, and the CLA accumulation was directly proportional (r 2= 0.98) to the initial cell density. Growing cells were as sensitive to CLA as LA, the LA isomerase and reductases of biohydrogenation were linked, and free CLA was not released. Because growing cultures ofB. fibrisolvens A38 did not produce significant amounts of CLA until the LA concentration was high, biohydrogenation was arrested, and the cell density had declined, the flow of CLA from the rumen may be due to LA-dependent bacterial inactivation, death, or lysis.

Postharvest Biological Control of Rhizopus Rot of Nectarine Fruits by Pichia membranefaciens

A new yeast antagonist, Pichia membranefaciens, isolated from wounds of peach fruit, was evaluated for its biocontrol capability against Rhizopus stolonifer on nectarine fruits at different temperatures and with other treatments. P. membranefaciens at 5 × 108 CFU/ml of washed-cell suspension completely inhibited Rhizopus rot in nectarine wounds artificially inoculated with 5 × 104 spores per ml at 25, 15, and 3°C. A culture filtrate of the yeast antagonist failed to provide any protection against Rhizopus rot in nectarine fruits. The yeast mixed with iprodione at 100 μg a.i./ml gave better control of R. stolonifer than either yeast or iprodione alone. A solution of 20 g CaCl2 per liter enhanced the efficacy of P. membranefaciens (107 to 108 CFU/ml) as an aqueous suspension. Rapid colonization of the yeast in wounds was observed during the first 48 h at 25 and 15°C. P. membranefaciens at 5 × 108 CFU/ml was effective when applied 0 to 72 h before the pathogen, while at 1 × 108 CFU/ml, its efficacy was best when applied 24 to 48 h prior to inoculation with R. stolonifer. However, its efficacy was significantly reduced when the yeast was applied simultaneously with the pathogen, with disease incidence of 60% and lesion diameter of 37 mm.

Propachlor Removal by Pseudomonas Strain GCH1 in an Immobilized-Cell System

ABSTRACT A bacterial strain capable of growing on propachlor (2-chloro-N-isopropylacetanilide) was isolated from soil by using enrichment and isolation techniques. The strain isolated, designated GCH1, was classified as a member of the genusPseudomonas. Washed-cell suspensions of strain GCH1 accumulated N-isopropylacetanilide, acetanilide, acetamide, and catechol. Pseudomonas strain GCH1 grew on propachlor with a generation time of 4.2 h and a rate of substrate utilization of 1.75 ± 0.15 μmol h−1. Gene expression did not require induction but was subject to catabolite expression. Acetanilide was a growth substrate with a yield of 0.56 ± 0.02 mg of protein μmol−1. GCH1 strain cells were immobilized by adsorption onto a ceramic support and were used as biocatalysts in an immobilized cell system. Propachlor elimination reached 98%, with a retention time of 3 h and an initial organic load of 0.5 mM propachlor. The viability of immobilized cells increased 34-fold after 120 days of bioreactor operation.

The Periplasmic 9.6-Kilodalton c-Type Cytochrome of Geobacter sulfurreducens Is Not an Electron Shuttle to Fe(III)

ABSTRACT Geobacter sulfurreducens contains a 9.6-kDac-type cytochrome that was previously proposed to serve as an extracellular electron shuttle to insoluble Fe(III) oxides. However, when the cytochrome was added to washed-cell suspensions of G. sulfurreducens it did not enhance Fe(III) oxide reduction, whereas similar concentrations of the known electron shuttle, anthraquinone-2,6-disulfonate, greatly stimulated Fe(III) oxide reduction. Furthermore, analysis of the extracellularc-type cytochromes in cultures of G. sulfurreducens demonstrated that the dominant c-type cytochrome was not the 9.6-kDa cytochrome, but rather a 41-kDa cytochrome. These results and other considerations suggest that the 9.6-kDa cytochrome is not an important extracellular electron shuttle to Fe(III) oxides.

Simultaneous Reduction of Nitrate and Selenate by Cell Suspensions of Selenium-Respiring Bacteria

ABSTRACT Washed-cell suspensions of Sulfurospirillum barnesiireduced selenate [Se(VI)] when cells were cultured with nitrate, thiosulfate, arsenate, or fumarate as the electron acceptor. When the concentration of the electron donor was limiting, Se(VI) reduction in whole cells was approximately fourfold greater in Se(VI)-grown cells than was observed in nitrate-grown cells; correspondingly, nitrate reduction was ∼11-fold higher in nitrate-grown cells than in Se(VI)-grown cells. However, a simultaneous reduction of nitrate and Se(VI) was observed in both cases. At nonlimiting electron donor concentrations, nitrate-grown cells suspended with equimolar nitrate and selenate achieved a complete reductive removal of nitrogen and selenium oxyanions, with the bulk of nitrate reduction preceding that of selenate reduction. Chloramphenicol did not inhibit these reductions. The Se(VI)-respiring haloalkaliphile Bacillus arsenicoselenatis gave similar results, but its Se(VI) reductase was not constitutive in nitrate-grown cells. No reduction of Se(VI) was noted for Bacillus selenitireducens, which respires selenite. The results of kinetic experiments with cell membrane preparations of S. barnesii suggest the presence of constitutive selenate and nitrate reduction, as well as an inducible, high-affinity nitrate reductase in nitrate-grown cells which also has a low affinity for selenate. The simultaneous reduction of micromolar Se(VI) in the presence of millimolar nitrate indicates that these organisms may have a functional use in bioremediating nitrate-rich, seleniferous agricultural wastewaters. Results with75Se-selenate tracer show that these organisms can lower ambient Se(VI) concentrations to levels in compliance with new regulations proposed for release of selenium oxyanions into the environment.

Characterization of Two Novel Propachlor Degradation Pathways in Two Species of Soil Bacteria

ABSTRACT Propachlor (2-chloro-N-isopropylacetanilide) is an acetamide herbicide used in preemergence. In this study, we isolated and characterized a soil bacterium, Acinetobacter strain BEM2, that was able to utilize this herbicide as the sole and limiting carbon source. Identification of the intermediates of propachlor degradation by this strain and characterization of new metabolites in the degradation of propachlor by a previously reported strain ofPseudomonas (PEM1) support two different propachlor degradation pathways. Washed-cell suspensions of strain PEM1 with propachlor accumulated N-isopropylacetanilide, acetanilide, acetamide, and catechol. Pseudomonas strain PEM1 grew on propachlor with a generation time of 3.4 h and aKs of 0.17 ± 0.04 mM.Acinetobacter strain BEM2 grew on propachlor with a generation time of 3.1 h and a Ks of 0.3 ± 0.07 mM. Incubations with strain BEM2 resulted in accumulation of N-isopropylacetanilide,N-isopropylaniline, isopropylamine, and catechol. Both degradative pathways were inducible, and the principal product of the carbon atoms in the propachlor ring was carbon dioxide. These results and biodegradation experiments with the identified metabolites indicate that metabolism of propachlor by Pseudomonas sp. strain PEM1 proceeds through a different pathway from metabolism byAcinetobacter sp. strain BEM2.