Poly(3-hydroxybutyrate) synthesis by recombinant Escherichia coli arcA mutants in microaerobiosis

We assessed the effects of different arcA mutations on poly(3-hydroxybutyrate) (PHB) synthesis in recombinant Escherichia coli strains carrying the pha synthesis genes from Azotobacter sp. strain FA8. The arcA mutations used were an internal deletion and the arcA2 allele, a leaky mutation for some of the characteristics of the Arc phenotype which confers high respiratory capacity. PHB synthesis was not detected in the wild-type strain in shaken flask cultures under low-oxygen conditions, while ArcA mutants gave rise to polymer accumulation of up to 24% of their cell dry weight. When grown under microaerobic conditions in a bioreactor, the arcA deletion mutant reached a PHB content of 27% +/- 2%. Under the same conditions, higher biomass and PHB concentrations were observed for the strain bearing the arcA2 allele, resulting in a PHB content of 35% +/- 3%. This strain grew in a simple medium at a specific growth rate of 0.69 +/- 0.07 h(-1), whereas the deletion mutant needed several nutritional additives and showed a specific growth rate of 0.56 +/- 0.06 h(-1). The results presented here suggest that arcA mutations could play a role in heterologous PHB synthesis in microaerobiosis.     

A new purple sulfur bacterium from saline littoral sediments,Thiorhodovibrio winogradskyi gen. nov. and sp. nov.

Two strains of a new purple sulfur bacterium were isolated in pure culture from the littoral sediment of a saline lake (Mahoney Lake, Canada) and a marine microbial mat from the North Sea island of Mellum, respectively. Single cells were vibrioid-to spirilloid-shaped and motile by means of single polar flagella. Intracellular photosynthetic membranes were of the vesicular type. As photosynthetic pigments, bacteriochlorophyll a and the carotenoids lycopene, rhodopin, anhydrorhodovibrin, rhodovibrin and spirilloxanthin were present.Hydrogen sulfide and elemental sulfur were used under anoxic conditions for phototrophic growth. In addition one strain (06511) used thiosulfate. Carbon dioxide, acetate and pyruvate were utilized by both strains as carbon sources. Depending on the strain propionate, succinate, fumarate, malate, tartrate, malonate, glycerol or peptone may additionally serve as carbon sources in the light. Optimum growth rates were obtained at pH 7.2, 33 °C, 50 mol m^-2 s^-1 intensity of daylight fluorescent tubes and a salinity of 2.2–3.2% NaCl. During growth on sulfide, up to ten small sulfur globules were formed inside the cells. The strains grew microaerophilic in the dark and exhibited high specific respiration rates. No vitamins were required for growth. The DNA base composition was 61.0–62.4 mol% G+C.The newly isolated bacterium belongs to the family chromatiaceae and is described as a member of a new genus and species, Thiorhodovibrio winogradskyi gen. nov. and sp. nov. with the type strain SSP1, DSM No. 6702.     

Microaerophilic growth and induction of the photosynthetic reaction center in Rhodopseudomonas viridis.

Rhodopseudomonas viridis was grown in liquid culture at 30 degrees C anaerobically in light (generation time, 13 h) and under microaerophilic growth conditions in the dark (generation time, 24 h). The bacterium could be cloned at the same temperature anaerobically in light (1 week) and aerobically in the dark (3 to 4 weeks) if oxygen was limited to 0.1%. Oxygen could not be replaced by dimethyl sulfoxide, potassium nitrate, or sodium nitrite as a terminal electron acceptor. No growth was observed anaerobically in darkness or in the light when air was present. A variety of additional carbon sources were used to supplement the standard succinate medium, but enhanced stationary-phase cell density was observed only with glucose. Conditions for induction of the photosynthetic reaction center upon the change from microaerophilic to phototrophic growth conditions were investigated and optimized for a mutant functionally defective in phototrophic growth. R. viridis consumed about 20-fold its cell volume of oxygen per hour during respiration. The MICs of ampicillin, kanamycin, streptomycin, tetracycline, 1-methyl-3-nitro-1-nitrosoguanidine, and terbutryn were determined.     

Microaerophilic Cooperation of Reductive and Oxidative Pathways Allows Maximal Photosynthetic Membrane Biosynthesis in Rhodospirillum rubrum

The purple nonsulfur bacterium Rhodospirillum rubrum has been employed to study physiological adaptation to limiting oxygen tensions (microaerophilic conditions). R. rubrum produces maximal levels of photosynthetic membranes when grown with both succinate and fructose as carbon sources under microaerophilic conditions in comparison to the level (only about 20% of the maximum) seen in the absence of fructose. Employing a unique partial O₂ pressure (pO₂) control strategy to reliably adjust the oxygen tension to values below 0.5%, we have used bioreactor cultures to investigate the metabolic rationale for this effect. A metabolic profile of the central carbon metabolism of these cultures was obtained by determination of key enzyme activities under microaerophilic as well as aerobic and anaerobic phototrophic conditions. Under aerobic conditions succinate and fructose were consumed simultaneously, whereas oxygen-limiting conditions provoked the preferential breakdown of fructose. Fructose was utilized via the Embden-Meyerhof-Parnas pathway. High levels of pyrophosphate-dependent phosphofructokinase activity were found to be specific for oxygen-limited cultures. No glucose-6-phosphate dehydrogenase activity was detected under any conditions. We demonstrate that NADPH is supplied mainly by the pyridine-nucleotide transhydrogenase under oxygen-limiting conditions. The tricarboxylic acid cycle enzymes are present at significant levels during microaerophilic growth, albeit at lower levels than those seen under fully aerobic growth conditions. Levels of the reductive tricarboxylic acid cycle marker enzyme fumarate reductase were also high under microaerophilic conditions. We propose a model by which the primary “switching” of oxidative and reductive metabolism is performed at the level of the tricarboxylic acid cycle and suggest how this might affect redox signaling and gene expression in R. rubrum.     

Microaerophilic cooperation of reductive and oxidative pathways allows maximal photosynthetic membrane biosynthesis in Rhodospirillum rubrum

The purple nonsulfur bacterium Rhodospirillum rubrum has been employed to study physiological adaptation to limiting oxygen tensions (microaerophilic conditions). R. rubrum produces maximal levels of photosynthetic membranes when grown with both succinate and fructose as carbon sources under microaerophilic conditions in comparison to the level (only about 20% of the maximum) seen in the absence of fructose. Employing a unique partial O(2) pressure (pO(2)) control strategy to reliably adjust the oxygen tension to values below 0.5%, we have used bioreactor cultures to investigate the metabolic rationale for this effect. A metabolic profile of the central carbon metabolism of these cultures was obtained by determination of key enzyme activities under microaerophilic as well as aerobic and anaerobic phototrophic conditions. Under aerobic conditions succinate and fructose were consumed simultaneously, whereas oxygen-limiting conditions provoked the preferential breakdown of fructose. Fructose was utilized via the Embden-Meyerhof-Parnas pathway. High levels of pyrophosphate-dependent phosphofructokinase activity were found to be specific for oxygen-limited cultures. No glucose-6-phosphate dehydrogenase activity was detected under any conditions. We demonstrate that NADPH is supplied mainly by the pyridine-nucleotide transhydrogenase under oxygen-limiting conditions. The tricarboxylic acid cycle enzymes are present at significant levels during microaerophilic growth, albeit at lower levels than those seen under fully aerobic growth conditions. Levels of the reductive tricarboxylic acid cycle marker enzyme fumarate reductase were also high under microaerophilic conditions. We propose a model by which the primary "switching" of oxidative and reductive metabolism is performed at the level of the tricarboxylic acid cycle and suggest how this might affect redox signaling and gene expression in R. rubrum.     

Isolation and characterization of a new denitrifying spirillum capable of anaerobic degradation of phenol

Two kinds of phenol-degrading denitrifying bacteria, Azoarcus sp. strain CC-11 and spiral bacterial strain CC-26, were isolated from the same enrichment culture after 1 and 3 years of incubation, respectively. Both strains required ferrous ions for growth, but strain CC-26 grew better than strain CC-11 grew under iron-limited conditions, which may have resulted in the observed change in the phenol-degrading bacteria during the enrichment process. Strain CC-26 grew on phenol, benzoate, and other aromatic compounds under denitrifying conditions. Phylogenetic analysis of 16S ribosomal DNA sequences revealed that this strain is most closely related to a Magnetospirillum sp., a member of the alpha subclass of the class Proteobacteria, and is the first strain of a denitrifying aromatic compound-degrading bacterium belonging to this group. Unlike previously described Magnetospirillum strains, however, this strain did not exhibit magnetotaxis. It grew on phenol only under denitrifying conditions. Other substrates, such as acetate, supported aerobic growth, and the strain exhibited microaerophilic features.     

Photoautotrophic high-density cultivation of vegetative cells of Haematococcus pluvialis in airlift bioreactor

This work aimed to investigate the effects of the bioreactor configurations and their design variables on the cultivation of vegetative cells Haematococcus pluvialis to achieve sustainable high cell density. The addition of vitamin B to F1 growth medium could appreciably enhance the final cell density. Employing this medium, the cultivation in the airlift bioreactor was demonstrated to outperform the bubble column at the same operating conditions. Aeration was crucial for a proper growth of the alga in the airlift bioreactor, but it must be maintained at low level to minimize shear stress. The most appropriate aeration velocity (superficial velocity) was at the lower limit of the pump, i.e. 0.4 cm s(-1) and a smaller riser was shown to have positive influence on the cell growth. A 1% CO(2) supplement to the air supply considerably enhanced the growth rate of H. pluvialis and the most suitable light intensity for the growth was at 20 micromol photon m(-2) s(-1). The semi-continuous culture was successfully implemented with the optimal airlift bioreactor design and under optimal conditions the harvest could be performed every four days with the specific growth rate of 0.31 d(-1).     

Culture of insect cells in helical ribbon impeller bioreactor

An 11-L helical ribbon impeller (HRI) bioreactor was tested for the culture of Spodoptera frugiperda (Sf-9) cells. This impeller and surface baffling ensured homogeneous mixing and high oxygen transfer through surface aeration and surface-induced babble generation. Serum-supplemented and serum-free cultures, using TNMFH and IPL/41 media, respectively, grew a similar specific growth rates(0.031 and 0.028 h(-1)) to maximum cell densities of 5.5 x 10(6)-6.0 x 10(6) cells. mL(-1) with viability exceeding 98% during exponential growth phase. Growth limitation coincided with glucose and glutamine depletion and production of significant amounts of alanine. The bioreactor was further tested under more stringent conditions by infecting a serum-free medium culture with a recombinant baculovirus. Heterologous protein production of approximately 35 mug per 10(6) cells was comparable to yields obtained in serum-free cultures grown in spinner flasks and petri dishes. Average specific oxygen up-take and carbon dioxide production rates of the serum-free culture prior to infection as measured by on-line mass spectroscopy were 0.20 mumol O(2)mu.(10(6) cells)(-1) h(-1) and 0.22 mumol CO(2) . (10(6) cells)(-1)h(-1) and increased by 30-40% during infection. Therefore, the mixing and oxygenation conditions of this bioreactor were suitable for insect cell culture and recombinant protein production, with limitation being mainly attributed to nutrient depletion and toxic by-product generation.     

Isolation and characterization of microorganisms able to produce 1,3-propanediol under aerobic conditions

Microbial fermentation under strictly anaerobic conditions has been conventionally used for the production of 1,3-propanediol, a key raw material required for the synthesis of polytrimethylene terephthalate (PTT) and other polyester fibers. In the current study, we have identified eight strains of microorganism which are able to produce 1,3-propanediol under aerobic condition. Those strains were isolated from garden soil, which were enriched by culturing in LB medium with glycerol added under aerobic condition. The identities of those strains were established based on their 16S rRNA sequences and physiological characteristics. Results indicated 6 strains are Citrobacter freundii and 2 strains are Klebsiella pneumoniae subsp Penumoniae. One of Klebsiella pneumoniae subsp Penumoniae strains, designated as TUAC01, demonstrated comparable levels of 1,3-propanediol oxidoreductase, glycerol dehydratase and glycerol dehydrogenase activity to the anaerobic microorganisms described in the literature. Accordingly, in larger scales (5 l) fed-batch culture the TUAC01 strain showed a remarkable 1,3-propanediol producing potency under aerobic conditions. 60.1 g/l of 1,3-propanediol was yield after 42 h incubation in an agitating bioreactor; and in air-lift bioreactor 66.3 g/l of 1,3-propanediol was yield after 58.5 h incubation. The aerobic ferment process, reduced the product cost and made the biological method of 1,3-propanediol production more attractive.     

Microaerophilic degradation of hexahydro‐1,3,5‐trinitro‐1,3,5‐triazine (RDX) by three Rhodococcus strains

Aim: The goal of this study was to compare the degradation of hexahydro‐1,3,5‐trinitro‐1,3,5‐triazine (RDX) by three Rhodococcus strains under anaerobic, microaerophilic (<0·04 mg l^?1 dissolved oxygen) and aerobic (dissolved oxygen (DO) maintained at 8 mg l^?1) conditions. Methods and Results: Three Rhodococcus strains were incubated with no, low and ambient concentrations of oxygen in minimal media with succinate as the carbon source and RDX as the sole nitrogen source. RDX and RDX metabolite concentrations were measured over time. Under microaerophilic conditions, the bacteria degraded RDX, albeit about 60‐fold slower than under fully aerobic conditions. Only the breakdown product, 4‐nitro‐2,4‐diazabutanal (NDAB) accumulated to measurable concentrations under microaerophilic conditions. RDX degraded quickly under both aerated and static aerobic conditions (DO allowed to drop below 1 mg l^?1) with the accumulation of both NDAB and methylenedinitramine (MEDINA). No RDX degradation was observed under strict anaerobic conditions. Conclusions: The Rhodococcus strains did not degrade RDX under strict anaerobic conditions, while slow degradation was observed under microaerophilic conditions. The RDX metabolite NDAB was detected under both microaerophilic and aerobic conditions, while MEDINA was detected only under aerobic conditions. Impact and Significance of the Study: This work confirmed the production of MEDINA under aerobic conditions, which has not been previously associated with aerobic RDX degradation by these organisms. More importantly, it demonstrated that aerobic rhodococci are able to degrade RDX under a broader range of oxygen concentrations than previously reported.     

Microaerophilic–aerobic sequential decolourization/biodegradation of textile azo dyes by a facultative Klebsiella sp. strain VN-31

Four different azo dyes were decolourized and biodegraded in a sequential microaerophilic–aerobic treatment by a facultative Klebsiella sp. strain VN-31, a bacterium isolated from activated sludge process of the textile industry. Dye decolourization was performed under microaerophilic conditions until no colour was observed (decolourization percentage >94%). The medium was then aerated to promote the biodegradation of the amines produced. The presence of aromatic amine in the microaerophilic stage and its absence in the aerobic stage demonstrate azo bond reduction and an oxidative biodegradation process, respectively. Total Organic Carbon (TOC) reduction for the growth medium plus dyes was ～50% in the microaerophilic stage and ～80% in the aerobic stage. The degradation products were also characterized by FT-IR and UV–vis techniques and their toxicity measured using Daphnia magna. The results provide evidence that the successive microaerophilic/aerobic stages, using a single Klebsiella sp. strain VN-31 in the same bioreactor, were able to form aromatic amines by the reductive break down of the azo bond and to oxidize them into non-toxic metabolites.     

A mutant of Halobacterium halobium with constitutive production of bacteriorhodopsin

Abstract A purple mutant of Halobacterium halobium was isolated in a previous study. The 'in vitro' absorption spectra of the cells gave a broad shoulder around 570 nm. The amounts of bacteriorhodopsin were high under any growth condition (including aerobic) inhibitory for the wild-type strain. The mutant grew faster under illuminated microaerophilic conditions and showed faster proton extrusion than the wild-type strain. This evidence shows that the mutant has a constitutive bacteriorhodopsin production not influenced by the oxygen concentration in the medium. However, some stimulation by light was found.     

Simple method to grow enteric campylobacters in unsupplemented liquid medium without the need for microaerophilic kits

Campylobacter strains (100 human, animal and environmental isolates) were grown in untreated brain heart infusion broth medium (10 ml in tightly capped 20 ml capacity universal tubes) without using microaerophilic kits. Cells grown in these conditions did not differ in their growth rates, protein profiles or substrate utilisation even after 40 passages compared to cells grown under microaerophilic conditions. Growth in such conditions provides a cost effective, convenient and simple system for growing pure culture of campylobacters and obviates the generation of microaerobic conditions using specialised kits.     

Oxygen transfer and consumption in a thiosulfate oxidizing bioreactor with sulfur production

AIMS: To evaluate the contribution of oxygen transfer and consumption in a sulfoxidizing system to increase the elemental sulfur yield from thiosulfate oxidation. METHODS AND RESULTS: A 10 l thiosulfate oxidizing bioreactor with suspended cells operating under microaerophilic conditions and a separated aerator with a variable volume of 0.8--1.7 l were operated with a consortium containing mainly Thiobacillus sp. that oxidizes several sulfide species to elemental sulfur and sulfate. From the gas-liquid oxygen balance, the k(L)a was estimated under different operation conditions. A k(L)a of around 200 h(-1) favoured elemental sulfur production and can serve as scale-up criterion. It was further shown that more than 50% of the oxygen fed to the system was consumed in the aerator. CONCLUSIONS: The performance of the sulfoxidizing system can be improved by controlling oxygen transfer. SIGNIFICANCE AND IMPACT OF THE STUDY: The proposed method for the k(L)a determination was based on the oxygen balance, which incorporates the oxygen concentrations measured in the liquid in steady state, reducing the interference of the response time in the traditional non-steady state methods. This approach can be used to optimize reactors where microaerophilic conditions are desirable.     

Multiple and flexible roles of facultative anaerobic bacteria in microaerophilic oleate degradation

Anaerobic degradation of long-chain fatty acids (LCFA) involves syntrophic bacteria and methanogens, but facultative anaerobic bacteria (FAB) might have a relevant role as well. Here we investigated oleate degradation by a syntrophic synthetic co-culture of Syntrophomonas zehnderi (Sz) and Methanobacterium formicicum (Mf) and FAB (two oleate-degrading Pseudomonas spp. I1 + I2). Sz + Mf were first cultivated in a continuous bioreactor under strict anaerobic conditions. Thereafter, I1 + I2 were inoculated and microaerophilic conditions were provided. Methane and acetate were the main degradation products by Sz + Mf in anaerobiosis and by Sz + Mf + I1 + I2 in microaerophilic conditions. However, acetate production from oleate was higher in microaerophilic conditions (5% O₂) with the four microorganisms together (0.41 ± 0.07 mmol day⁻¹) than in anaerobiosis with Sz + Mf (0.23 ± 0.05 mmol day⁻¹). Oleate degradation in batch assays was faster by Sz + Mf + I1 + I2 (under microaerophilic conditions) than by Sz + Mf alone (under strict anaerobic conditions). I1 + I2 were able to grow with oleate and with intermediates of oleate degradation (hydrogen, acetate and formate). This work highlights the importance of FAB, particularly Pseudomonas sp., in anaerobic reactors treating oleate-based wastewater, because they accelerate oleate conversion to methane, by protecting strict anaerobes from oxygen toxicity and also by acting as alternative hydrogen/formate and acetate scavengers for LCFA-degrading anaerobes.     

Sodium-Dependent Azotobacter chroococcum Strains Are Aeroadaptive, Microaerophilic, Nitrogen-Fixing Bacteria

Na⁺ -dependent strains of Azotobacter chroococcum were observed to have very low reactivities with the H₂O₂ spot test for catalase. The cell extract of the representative Na⁺ -dependent strain 184 contained a catalase specific activity that was 10-to 600-fold lower than those found in Na⁺ -independent strains of A. chroococcum. Peroxidase and superoxide dismutase activities existed in all strains, although only certain Na⁺ -dependent strains contained a peroxidase reactive with p-phenylenediamine. The activities of catalase and peroxidase in the Na⁺ -dependent strain 184 were dependent on iron availability, which helped to explain the iron-dependent growth characteristic of this strain. The activities of these enzymes were not increased by subjecting the cells to increased aeration, nitrogen-fixing conditions, or paraquat. Strain 184 was found to be very sensitive to H₂O₂ or paraquat, even under iron-sufficient conditions, and was difficult to recover quantitatively on solid plating media. Strain 184 was more susceptible to H₂O₂ when grown under low-aeration, nitrogen-fixing conditions than when it was grown in the presence of NH₄⁺. Low population densities of strain 184 grew in nitrogen-free medium under microaerophilic conditions, while more dense populations were able to fix nitrogen under aerobic conditions. Therefore, these bacteria appeared to be aeroadaptive, microaerophilic, nitrogen-fixing bacteria.     

Tolerance to challenges miming gastrointestinal transit by spores and vegetative cells of Bacillus clausii

AIMS: To study Bacillus clausii from a pharmaceutical product (Enterogermina O/C, N/R, SIN, T) and reference strains (B. clausii and Bacillus subtilis) for eco-physiological aspects regarding the gut environment. METHODS AND RESULTS: Spores and vegetative cells were challenged in vitro miming the injury of gastrointestinal transit: pH variations, exposure to conjugated and free bile salts, microaerophilic and anaerobic growth. No relevant differences were found studying the growth at pH 8 and 10, whereas at pH 7 the yields obtained for O/C and SIN were higher than those obtained for N/R and T strains. The spores were able to germinate and grow in the presence of conjugated bile salts (up to 1%, w/v) or free bile salts (0.2%) and also exhibited tolerance for the combined acid-bile challenge. As evidenced by lag-time, growth rate and cell yield the tolerance of Enterogermina isolates for conjugated salts was comparable with that of B. clausii type strain (DSM 8716(T)), and resulted higher than that observed for B. subtilis (ATCC 6051(T)). All the considered B. clausii strains demonstrated microaerophilic growth, but only some grew anaerobically in a nitrate medium. CONCLUSIONS: The ability of B. clausii spores to germinate after an acid challenge and grow as vegetative cells both in the presence of bile and under limited oxygen availability is consistent with the beneficial health effects evidenced for spore-forming probiotics in recent clinical studies. SIGNIFICANCE AND IMPACT OF THE STUDY: The experimental evidence from this study emphasizes some functional properties of B. clausii strains regarding their use as probiotics.     

Energy--dependent bacterivory in Ochromonas minima--a strategy promoting the use of substitutable resources and survival at insufficient light supply

Phagotrophy and competitive ability of the mixotrophic Ochromonas minima were investigated in a three-factorial experiment where light intensity (low: 1.0 micromol m(-2)s(-1) and high: 60 micromol m(-2)s(-1) PPFD), nutrient concentration (ambient: 7.0 micromolNl(-1), 0.11 micromol P l(-1) and enriched: 88 micromol N l(-1), 6.3 micro mol P l(-1)) and DOC supply (without and with enrichment, 250 micromol C l(-1)) were manipulated. Ochromonas minima and bacterial abundance were monitored for 12 days. We found significant and interacting effects of light and nutrients on Ochromonas minima growth rate and abundance. At high light intensity, nutrient enrichment resulted in increased growth rates and population sizes. In contrast, reduced growth rates and population sizes were observed for nutrient enrichment when light intensity was low. Although, Ochromonas minima was able to ingest bacteria under both high and low light conditions, it grew only when light intensity was high. At high light intensity, Ochromonas minima grew exponentially under nutrient conditions that would have been limiting for photoautotrophic microalgae. In non-enriched low light treatments, Ochromonas minima populations survived, probably by using background DOC as an energy source, indicating that this ability can be of relevance for natural systems even when DOC concentrations are relatively low. When competing with photoautotrophic microalgae, the ability to grow under severe nutrient limitation and to survive under light limitation should be advantageous for Ochromonas minima.     

Anaerobic oxidation of 2-chloroethanol under denitrifying conditions by<Emphasis Type="Italic"> Pseudomonas stutzeri</Emphasis> strain JJ

A bacterium that uses 2-chloroethanol as sole energy and carbon source coupled to denitrification was isolated from 1,2-dichloroethane-contaminated soil. Its 16 S rDNA sequence showed 98% similarity with the type strain of Pseudomonas stutzeri (DSM 5190) and the isolate was tentatively identified as Pseudomonas stutzeri strain JJ. Strain JJ oxidized 2-chloroethanol completely to CO(2) with NO(3)(- )or O(2) as electron acceptor, with a preference for O(2) if supplied in combination. Optimum growth on 2-chloroethanol with nitrate occurred at 30 degrees C with a mu(max) of 0.14 h(-1) and a yield of 4.4 g protein per mol 2-chloroethanol metabolized. Under aerobic conditions, the mu(max) was 0.31 h(-1). NO(2)(-) also served as electron acceptor, but reduction of Fe(OH)(3), MnO(2), SO(4)(2-), fumarate or ClO(3)(-) was not observed. Another chlorinated compound used as sole energy and carbon source under aerobic and denitrifying conditions was chloroacetate. Various different bacterial strains, including some closely related Pseudomonas stutzeri strains, were tested for their ability to grow on 2-chloroethanol as sole energy and carbon source under aerobic and denitrifying conditions, respectively. Only three strains, Pseudomonas stutzeri strain LMD 76.42, Pseudomonas putida US2 and Xanthobacter autotrophicus GJ10, grew aerobically on 2-chloroethanol. This is the first report of oxidation of 2-chloroethanol under denitrifying conditions by a pure bacterial culture.     

Growth Properties of Rhodospirillum rubrum Mutants and Fermentation of Pyruvate in Anaerobic, Dark Conditions

Mutant C and G1 were obtained earlier from Rhodospirillum rubrum S(1) during growth in the dark under strict anaerobic conditions in medium containing sodium pyruvate. Mutant C and mutant G1 grew in the dark with generation times of 5.8 h and 4.6 h, respectively. Mutant C cells grew equally well when switched between anaerobic (dark or light) or aerobic, dark conditions. Mutant G1 cells grew only in the dark (anaerobic or aerobic conditions), but a fraction of cells in anaerobic, dark cultures grew when placed in light. This number increased about 3,000-fold when G1 cells were incubated aerobically in the dark. During anaerobic, dark growth, C and G1 organisms incorporated similar amounts of [2-(14)C]sodium pyruvate. About 34% of the incorporated radioactivity was found in lipid fractions from C cells that developed chromatophores during dark growth. Similar results were obtained using G1 cells, which formed only trace amounts of photosynthetic structures. Both mutants fermented sodium pyruvate and produced acetate, formate, carbon dioxide, and hydrogen gas. Molar growth yield coefficients indicated that the cells obtained about 1 mol of adenosine triphosphate per mol of sodium pyruvate fermented. Results suggested that pyruvate fermentation during dark growth occurred via a pyruvate formate-lyase or the pyruvate ferredoxin-oxidoreductase pathway, or both.