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D D Focht

Publications and source records attributed to D D Focht.

At least 37 records · Page 2Linked to original sources
PubMed1990

Degradation of 2-bromobenzoic acid by a strain of Pseudomonas aeruginosa.

A strain of Pseudomonas aeruginosa producing 2-bromobenzoic acid, designated 2-BBZA, was isolated by enrichment culture from municipal sewage. It degraded all four 2-halobenzoates as well as certain 3-halo- and dihalobenzoates, though none of the 4-halobenzoates supported growth of this organism. 3-Hydroxybenzoate and 3-chlorocatechol were respective inhibitors of salicylate and catechol oxidation: when each was added separately to resting cells incubated with 2-bromobenzoate, salicylate and catechol were found. Oxygen uptake data suggest that the same dehalogenase may be involved in the oxidation of 2-bromo-, 2-chloro-, and 2-iodobenzoates.

Bromobenzoates↗
PubMed1989

Identification of Pseudomonas alcaligenes chromosomal DNA in the plasmid DNA of the chlorobenzene-degrading recombinant Pseudomonas putida strain CB1-9.

The recombinant Pseudomonas putida strain CB1-9, which acquired the ability to grow on chlorobenzenes, contains a 33-kilobase (kb) plasmid (pKFL3) which lacked homology to an indigenous 15-kb plasmid (pKFL1) in Pseudomonas alcaligenes C-0 parent but was homologous to a 55-kb plasmid (pKFL2) from the P. putida R5-3 parent. Chromosomal DNA of P. alcaligenes C-0 hybridized to probes prepared from pKFL3 but not to probes prepared from pKFL2. A single clone from a genomic library of P. alcaligenes C-0 hybridized to EcoRI-digested pKFL3. Southern blot hybridization with the insert DNA from that clone identified homology with specific restriction enzyme fragments in pKFL3. The ability of the recombinant to utilize 3-chlorobenzoate, chlorobenzene, and 1,4-dichlorobenzene as well as its loss of utilization of xylenes and methylbenzoates appears to be associated with the transfer and integration of chromosomal DNA from P. alcaligenes into a Tol-like plasmid of P. putida R5-3.

Blotting, Southern↗
PubMed1989

Bacterial dehalogenation of chlorobenzoates and coculture biodegradation of 4,4'-dichlorobiphenyl.

Acinetobacter sp. strain 4CB1 was isolated from a polychlorobiphenyl-contaminated soil sample by using 4-chlorobenzoate as a sole source of carbon and energy. Resting cells of Acinetobacter sp. strain 4CB1 hydrolytically dehalogenated 4-chlorobenzoate under aerobic and anaerobic conditions, but 4-hydroxybenzoate accumulated only under anaerobic conditions. Cell extracts of Acinetobacter sp. strain 4CB1 oxidized 4-hydroxybenzoate by an NADH-dependent monooxygenase to form protocatechuate, which was subsequently oxidized by both ortho- and meta-protocatechuate dioxygenase reactions. When grown on biphenyl, Acinetobacter sp. strain P6 cometabolized 4,4'-dichlorobiphenyl primarily to 4-chlorobenzoate; however, when this strain was grown in a coculture with Acinetobacter sp. strain 4CB1, 4-chlorobenzoate did not accumulate but was converted to inorganic chloride. When resting cells of Acinetobacter sp. strain 4CB1 were incubated anaerobically with 3,4-dichlorobenzoate, they accumulated 4-carboxy-1,2-benzoquinone as a final product. Since 3,4-dichlorobenzoate is a product that is formed from the cometabolism of 3,4-dichloro-substituted tetrachlorobiphenyls by Acinetobacter sp. strain P6, the coculture has a potential application for dehalogenation and mineralization of specific polychlorobiphenyl congeners.

Acinetobacter↗
PubMed1989

Bacterial metabolism of hydroxylated biphenyls.

Isolates able to grow on 3- or 4-hydroxybiphenyl (HB) as the sole carbon source were obtained by enrichment culture. The 3-HB degrader Pseudomonas sp. strain FH12 used an NADPH-dependent monooxygenase restricted to 3- and 3,3'-HBs to introduce an ortho-hydroxyl. The 4-HB degrader Pseudomonas sp. strain FH23 used either a mono- or dioxygenase to generate a 2,3-diphenolic substitution pattern which allowed meta-fission of the aromatic ring. By using 3-chlorocatechol to inhibit catechol dioxygenase activity, it was found that 2- and 3-HBs were converted by FH23 to 2,3-HB, whereas biphenyl and 4-HB were attacked by dioxygenation. 4-HB was metabolized to 2,3,4'-trihydroxybiphenyl. Neither organism attacked chlorinated HBs. The degradation of 3- and 4-HBs by these strains is therefore analogous to the metabolism of biphenyl, 2-HB, and naphthalene in the requirement for 2,3-catechol formation.

Biodegradation, Environmental↗
PubMed1988

Degradation of 2-hydroxybiphenyl and 2,2'-dihydroxybiphenyl by Pseudomonas sp. strain HBP1.

Pseudomonas sp. strain HBP1 was found to grow on 2-hydroxy- and 2,2'-dihydroxy-biphenyl as the sole carbon and energy sources. The first step in the degradation of these compounds was catalyzed by an NADH-dependent monooxygenase. The enzyme inserted a hydroxyl group adjacent to the already existing hydroxyl group to form 2,3-dihydroxybiphenyl when acting on 2-hydroxybiphenyl and to form 2,2',3-trihydroxybiphenyl when acting on 2,2'-dihydroxybiphenyl. To be substrates of the monooxygenase, compounds required a 2-hydroxyphenyl-R structure, with R being a hydrophobic group (e.g., methyl, ethyl, propyl, sec-butyl, phenyl, or 2-hydroxyphenyl). Several chlorinated hydroxybiphenyls served as pseudosubstrates by effecting consumption of NADH and oxygen without being hydroxylated. Further degradation of 2,3-dihydroxy- and 2,2',3-trihydroxybiphenyl involved meta cleavage, with subsequent formation of benzoate and salicylate, respectively.

Biodegradation, Environmental↗
PubMed1988

Cometabolism of polychlorinated biphenyls: enhanced transformation of Aroclor 1254 by growing bacterial cells.

Acinetobacter sp. strain P6 and a soil isolate, Arthrobacter sp. strain B1B, were tested for their ability to transform Aroclor 1254 as washed resting cells and as growing cells with biphenyl as the substrate. Growing cells were far superior to resting-cell suspensions in terms of total polychlorinated biphenyl (PCB) transformation, transformation of specific PCB congeners, and diversity of congeners that were attacked. Growing cells of Acinetobacter sp. strain P6 and Arthrobacter sp. strain B1B transformed 32 and 23% of the [14C]Aroclor 1254, respectively, whereas resting cells of the same respective cultures transformed only 17 and 8%. Transformation was significantly greater with resting cells in only 2 of 39 cases in which congeners were transformed by both growing and resting cells of both cultures. The components of 19 and 12 capillary gas-chromatographic peaks of Aroclor 1254 were transformed by biphenyl-grown resting cells of Acinetobacter sp. strain P6 and Arthrobacter sp. strain B1B, respectively, whereas the components of an additional 6 and 7 peaks were attacked by growing cells of the same respective cultures. Biphenyl oxidation by resting cells of both cultures decreased with time to less than 8% in 28 h. In addition to the normal 2,3-dioxygenase attack on PCBs, Acinetobacter sp. strain P6 also attacked congeners lacking an open 2,3-position. The ability of Acinetobacter sp. strain P6 to transform the components of 25 of the 40 largest peaks of Aroclor 1254 makes it one of the most versatile PCB-transforming organisms yet reported.

Acinetobacter↗
PubMed1987

Construction of chlorobenzene-utilizing recombinants by progenitive manifestation of a rare event.

Separate continuous cultures of Pseudomonas putida R5-3, grown on toluene, and Pseudomonas alcaligenes C-O, grown on benzoate, were concentrated and continuously amalgamated on a ceramic bead column, which was subjected to a continuous stream of chlorobenzene vapors. A recombinant strain, P. putida CB1-9, was isolated in less than 1 month. P. alcaligenes C-0 grew on benzoate and 3-chlorobenzoate but not on toluene, P. putida R5-3 grew on benzoate and toluene but not on 3-chlorobenzoate, and neither strain grew on chlorobenzene or 1,4-dichlorobenzene; however, the recombinant P. putida CB1-9 grew on all of these substrates. Chlorobenzene-utilizing strains were not found in continuous cultures run at the lowest growth rate (0.05/h) or in the absence of the donor strain, P. alcaligenes C-0. Chloride was released in stoichiometric amounts when P. putida CB1-9 was grown on either chlorobenzene or 1,4-dichlorobenzene. The recombinant strain was related to P. putida R5-3, phenotypically and genetically. Restriction enzyme digests of the single 57-kilobase (kb) plasmid in R5-3 and of the single 33-kb plasmid in CB1-9 were similar, but also indicated rearrangement of plasmid DNA. Coincidental or causal to the loss of the 24-kb fragment was the observation that the recombinant--unlike its parent, R5-3--did not grow on xylenes or methylbenzoates. Although both ortho-pyrocatechase (OP) and meta-pyrocatechase (MP) were found in CB1-9 and R5-3, MP activity was 20- to 50-fold higher in R5-3 cells grown on 4-methylbenzoate than in the same cells grown on benzene.(ABSTRACT TRUNCATED AT 250 WORDS)

Benzoates↗
PubMed1987

Nitrous oxide reduction in nodules: denitrification or n(2) fixation?

Detached cowpea nodules that contained a nitrous oxide reductase-positive (Nor) rhizobium strain (8A55) and a nitrous oxide reductase-negative (Nor) rhizobium strain (32H1) were incubated with 1% N(2)O (95 atom% N) in the following three atmospheres: (i) aerobic with C(2)H(2) (10%), (ii) aerobic without C(2)H(2), and (iii) anaerobic (argon atmosphere) without C(2)H(2). The greatest production of N(2) occurred anaerobically with 8A55, yet very little was formed with 32H1. Although acetylene reduction activity was slightly higher with 32H1, about 10 times more N(2) was produced aerobically by 8A55 than by 32H1 in the absence of acetylene. The major reductive pathway of N(2)O reduction by denitrifying rhizobium strain 8A55 is by nitrous oxide reductase rather than nitrogenase.

Journal Article↗
PubMed1987

Growth kinetics of Pseudomonas alcaligenes C-0 relative to inoculation and 3-chlorobenzoate metabolism in soil.

Pseudomonas alcaligenes C-0 was isolated from activated sewage sludge by enrichment with 3-chlorobenzoate (3CB) as the sole carbon source. The carbon balance from [14C]3CB in pure culture could be accounted for in substrate, biomass, and CO2 from all sampling periods and inoculum densities (0.012, 0.092, 0.20, and 0.92 micrograms of dry cells X ml-1), and inorganic chloride was produced stoichiometrically. Monod parameters as determined in culture were compared with the kinetics of 3CB metabolism in soil with decreasing inoculum densities (1.9 X 10(-1), 1.9 X 10(-3), and 1.9 X 10(-5) micrograms of cells X g-1). 3CB was refractile to attack in soil by indigenous microflora, but it was completely metabolized upon inoculation with P. alcaligenes C-0. The saturation constant KS was much higher in soil than in culture, but the yield coefficient Y and the growth rate constant were the same in both systems: mu max = 0.32 h-1; Y = 34 micrograms cells X mumol-1; KS = 0.18 mM in culture and 6.0 mM in soil solution (1.1 mumol X g-1 of soil). The parameter estimates obtained from the highest inoculum density could be used for the lower inoculum densities with reasonable agreement between predicted and observed 3CB concentrations in soil, although the residual sum of squares was progressively higher. Since the growth rate of P. alcaligenes C-0 in soil was comparable to its growth rate in culture, inoculation should be a viable strategy for biodegradation of 3CB in soil if indigenous microflora are unable to exploit this metabolic niche.

Biodegradation, Environmental↗
PubMed1987

Increased Effectiveness of Competitive Rhizobium Strains upon Inoculation of Cajanus cajan.

A field study was conducted in lysimeters containing N-enriched soil to determine the effects of four competitive rhizobium strains upon yield parameters of pigeon peas (Cajanus cajan). The greatest differences observed were in seed yields; strain P132 effected the highest seed yield (121 +/- 20 g per plant), and the control strain (indigenous rhizobia) effected the lowest yield (43.9 +/- 8 g per plant). With the exception of seeds and pods, the dry matter weights were not different. Although there appeared to be no effect by inoculum strains on the fractional content of N derived from biological nitrogen fixation when the total plant biomass was considered, strains P132 and 401 partitioned more of the N derived from fixation into seeds and leaves than did the other strains. Because the seeds comprised the major portion of plant N, more total N and more N derived from biological nitrogen fixation (about half of total N) were found in plants inoculated with P132, whereas the smallest amount was found in the uninoculated controls. P132 was also the best competitor with respect to indigenous rhizobia and acounted for all of the nodules found on the plants in which it was inoculated.

Journal Article↗
PubMed1986

Comparison of hup trait and intrinsic antibiotic resistance for assessing rhizobial competitiveness axenically and in soil.

The competitiveness of dual-strain inocula of cowpea rhizobia for nodulation of Vigna unguiculata (L.) Walp. was studied axenically between one slow-growing strain (P132, HP147, 401, or 22A1) and one fast-growing strain (176A26 or 176A28) at logarithmic inoculum ratios ranging from 10 to 10. Nodule infectivity was determined by multiple intrinsic antibiotic resistance, since both fast-growing strains were sensitive. Different hydrogen uptake (Hup) efficiencies of dual-strain inocula allowed for the comparison of an indirect rapid method. Infectivity data based on antibiotic resistance and Hup efficiency were fit to linearized fractile plots of log-normal distributions to determine C(AB) (percent infectivity at a 1:1 inoculum density) or I(50) (inoculum ratio at 50% infectivity). The slow growers were always better competitors and had I(50) values which ranged from 7 to 160,000 and C(AB) values which ranged from 62 to 97%. P132 was the best competitor of all those tested. Antibiotic resistance and Hup efficiency methods were in agreement with 401 (Hup) and 176A26 (Hup), but the Hup efficiency method overestimated the I(50) index with 22A1 (Hup) and 176A28 (Hup). The competition of each of the four slow-growing strains with indigenous rhizobia was examined in Cajanus cajan from three tropical soils. Nodule infectivity for all strains ranged from 42 to 96%, and P132 was the best competitor in all the soils. Hup efficiency overestimated infectivity by about 2-fold when Hup inocula (P132 and HP147) were used but underestimated infectivity by more than 100-fold when Hup inocula (401 and 22A1) were used. Although the Hup trait has limited quantitative usage axenically, it is only qualitative in soil competition studies and can only be used with Hup inocula.

Journal Article↗
PubMed1985

Kinetics of denitrifying growth by fast-growing cowpea rhizobia.

Two fast-growing strains of cowpea rhizobia (A26 and A28) were found to grow anaerobically at the expense of NO(3), NO(2), and N(2)O as terminal electron acceptors. The two major differences between aerobic and denitrifying growth were lower yield coefficients (Y) and higher saturation constants (K(s)) with nitrogenous oxides as electron acceptors. When grown aerobically, A26 and A28 adhered to Monod kinetics, respectively, as follows: K(s), 3.4 and 3.8 muM; Y, 16.0 and 14.0 g . cells eq; mu(max), 0.41 and 0.33 h. Yield coefficients for denitrifying growth ranged from 40 to 70% of those for aerobic growth. Only A26 adhered to Monod kinetics with respect to growth on all three nitrogenous oxides. The apparent K(s) values were 41, 270, and 460 muM for nitrous oxide, nitrate, and nitrite, respectively; the K(s) for A28 grown on nitrate was 250 muM. The results are kinetically and thermodynamically consistent in explaining why O(2) is the preferred electron acceptor. Although no definitive conclusions could be drawn regarding preferential utilization of nitrogenous oxides, nitrite was inhibitory to both strains and effected slower growth. However, growth rates were identical (mu(max), 0.41 h) when A26 was grown with either O(2) or NO(3) as an electron acceptor and were only slightly reduced when A28 was grown with NO(3) (0.25 h) as opposed to O(2) (0.33 h).

Journal Article↗
PubMed1985

N Kinetic Analysis of N(2)O Production by Nitrosomonas europaea: an Examination of Nitrifier Denitrification.

A series of N isotope tracer experiments showed that Nitrosomonas europaea produces nitrous oxide only under oxygen-limiting conditions and that the labeled N from nitrite, but not nitrate, is incorporated into nitrous oxide, indicating the presence of the "denitrifying enzyme" nitrite reductase. A kinetic analysis of the m/z 44, 45, and 46 nitrous oxide produced by washed cell suspensions of N. europaea when incubated with 4 mM ammonium (99% N) and 0.4 mM nitrite (99% N) was performed. No labeled nitrite was reduced to ammonium. All labeled material added was accounted for as either nitrite or nitrous oxide. The hypothesis that nitrous oxide is produced directly from nitrification was rejected since (i) it does not allow for the large amounts of double-labeled (m/z 46) nitrous oxide observed; (ii) the observed patterns of m/z 44, 45, and 46 nitrous oxide were completely consistent with a kinetic analysis based on denitrification as the sole mechanism of nitrous oxide production but not with a kinetic analysis based on both mechanisms; (iii) the asymptotic ratio of m/z 45 to m/z 46 nitrous oxide was consistent with denitrification kinetics but inconsistent with nitrification kinetics, which predicted no limit to m/z 45 production. It is concluded that N. europaea is a denitrifier which, under conditions of oxygen stress, uses nitrite as a terminal electron acceptor and produces nitrous oxide.

Journal Article↗
PubMed1985

Kinetics of biphenyl and polychlorinated biphenyl metabolism in soil.

The metabolism of C-labeled PCBs (polychlorinated biphenyls), which comprised the Aroclor 1242 mixture, was greatly enhanced by the addition of biphenyl (BP) to soil. After 49 days, only 25 to 35% of the original PCBs remained in the soil, and 48 to 49% was converted to CO(2) (including soil carbonates) in treatments enriched with BP; by contrast, 92% of the PCBs remained and less than 2% was converted to CO(2) in the unenriched control. Although the mineralization of PCBs in soils inoculated with Acinetobacter strain P6 was not greater than that in uninoculated BP-enriched soils, the initial and maximum mineralization rates and the disappearance of more highly chlorinated PCBs were greater with Acinetobacter strain P6. The mineralization of BP was consistent with kinetic models based upon linear-no growth and exponential growth; lower cell densities (<10/g) of BP-oxidizing bacteria gave a better fit for exponential growth, whereas the highest cell density (10/g) gave a better fit for linear-no growth. The numbers of BP-oxidizing bacteria declined exponentially upon depletion of the substrate. Since the mineralization of the chlorinated cometabolites was brought about by microorganisms (commensals) other than BP oxidizers, CO(2) production could not be fit to either of the two growth models. However, CO(2) production from the highest-density inoculum could be fit to a first-order (no-growth) sequential-reaction series. Although the population dynamics of the commensals could not be determined, the rate-limiting step in the cometabolic-commensal metabolism of PCBs to CO(2) had to be the initial oxidation, since the rate of CO(2) production was directly related to the population density of BP oxidizers.

Journal Article↗
PubMed1984

Deterministic three-half-order kinetic model for microbial degradation of added carbon substrates in soil.

The kinetics of mineralization of carbonaceous substrates has been explained by a deterministic model which is applicable to either growth or nongrowth conditions in soil. The mixed-order nature of the model does not require a priori decisions about reaction order, discontinuity period of lag or stationary phase, or correction for endogenous mineralization rates. The integrated equation is simpler than the integrated form of the Monod equation because of the following: (i) only two, rather than four, interdependent constants have to be determined by nonlinear regression analysis, (ii) substrate or product formation can be expressed explicitly as a function of time, (iii) biomass concentration does not have to be known, and (iv) the required initial estimate for the nonlinear regression analysis can be easily obtained from a linearized form rather than from an interval estimate of a differential equation. CO(2) evolution data from soil have been fitted to the model equation. All data except those from irradiated soil gave better fits by residual sum of squares (RSS) by assuming growth in soil was linear (RSS = 0.71) as opposed to exponential (RSS = 2.87). The underlying reasons for growth (exponential versus linear), no growth, and relative degradation rates of substrates are consistent with the basic mechanisms from which the model is derived.

Journal Article↗
PubMed1984

Invalidity of the concept of slow growth and alkali production in cowpea rhizobia.

A total of 103 rhizobial strains representing the cowpea miscellany and Rhizobium japonicum were studied with regard to growth rate, glucose metabolic pathways, and pH change in culture medium. Doubling times ranged from 1.4 +/- 0.04 to 44.1 +/- 5.2 h; although two populations of "fast-growing" and "slow-growing" rhizobia were noted, they overlapped and were not distinctly separated. Twenty-four strains which had doubling times of less than 8 h all showed NADP-linked 6-phosphogluconate dehydrogenase (6-PGD) activity, whereas only one slow-growing strain (doubling time, 10.8 +/- 0.9 h) of all those tested showed 6-PGD activity. Doubling times among fast growers could not be explained solely by the presence or absence of 6-PGD activity (r = 0.14) because the tricarboxylic acid cycle and the Emden-Meyerhoff-Parnas pathway were operative in both 6-PGD-positive and 6-PGD-negative strains. Growth rate and pH change were unrelated to each other. Fast- or slow-growing strains were not associated with any particular legume species or group of species from which they were originally isolated, with the exception of Stylosanthes spp., all nine isolates of which were slow growers. We conclude that 6-PGD activity is a more distinctive characteristic among physiologically different groups of rhizobia than doubling times and that characterization of the cowpea rhizobia as slow-growing alkali producers is an invalid concept.

Journal Article↗
PubMed1983

Comparison of N(2) Fixation and Yields in Cajanus cajan between Hydrogenase-Positive and Hydrogenase-Negative Rhizobia by In Situ Acetylene Reduction Assays and Direct N Partitioning.

Pigeon peas [Cajanus cajan (L.) Millsp.] were grown in soil columns containing (15)N-enriched organic matter. Seasonal N(2) fixation activity was determined by periodically assaying plants for reduction of C(2)H(2). N(2) fixation rose sharply from the first assay period at 51 days after planting to a peak of activity between floral initiation and fruit set. N(2) fixation (acetylene reduction) activity dropped concomitantly with pod maturation but recovered after pod harvests. Analysis of (15)N content of plant shoots revealed that approximately 91 to 94% of plant N was derived from N(2) fixation. The effect of inoculation with hydrogenase-positive and hydrogenase-negative rhizobia was examined. Pigeon peas inoculated with strain P132 (hydrogenase-positive) yielded significantly more total shoot N than other inoculated or uninoculated treatments. However, two other hydrogenase-positive strains did not yield significantly more total shoot N than a hydrogenase-negative strain. The extent of nodulation by inoculum strains compared to indigenous rhizobia was determined by typing nodules according to intrinsic antibiotic resistance of the inoculum strains. The inoculum strains were detected in almost all typed nodules of inoculated plants.Gas samples were taken from soil columns several times during the growth cycle of the plants. H(2) was never detected, even in columns containing pigeon peas inoculated with hydrogenase-negative rhizobia. This was attributed to H(2) consumption by soil bacteria. Estimation of N(2) fixation by acetylene reduction activity was closest to the direct (15)N method when ethylene concentrations in the gas headspace (between the column lid and soil surface) were extrapolated to include the soil pore space as opposed solely to measurement in the headspace. There was an 8-fold difference between the two acetylene reduction assay methods of estimation. Based on a planting density of 15,000 plants per hectare, the direct (15)N fixation rates ranged from 67 (noninoculated) to 134 kilograms per hectare, while grain yields ranged from 540 to 825 kilograms per hectare. Grain yields were not increased with N fertilizer.

Journal Article↗