Survival of sulfate-reducing bacteria after oxygen stress, and growth in sulfate-free oxygen-sulfide gradients

Abstract The survival after oxygen stress was studied with eight species of sulfate-reducing bacteria. In the absence of sulfide all species tolerated 6 min of aeration without loss of viability. Even after 3 h of aeration the viability of four species ( Desulfovibrio vulgaris, D. desulfuricans, D. salexigens and Desulfobacter postgatei ) was not impaired. Four other species were sensitive to 3 h of aeration: the surviving fractions of Desulfotomaculum ruminis, D. nigrificans and Desulfococcus multivorans were about 1%, that of Desulfotomaculum orientis about 0.01%. Formation of spores resulted in oxygen resistance of D. orientis . Reducing agents did not protect the vegetative cells of this strain against oxygen toxicity. In contrast, sulfhydryl group-containing agents increased the oxygen sensitivity considerably.
Growth of sulfate- and sulfur-reducing bacteria in oxygen-sulfide gradients in agar tubes was studied. In the gradients these strictly anaerobic bacteria revealed oxygen-dependent growth in sulfate- and sulfur-free medium. Three sulfate-reducing bacteria that could not use thiosulfate or sulfur as electron acceptor failed to grow in oxygen-sulfide gradients. Obviously, not directly molecular oxygen, but oxidation products of sulfide, such as thiosulfate or sulfur, were used as electron acceptors and were continuously regenerated in a cycling process from sulfide by autoxidation. The conceivable ecological significance of a short sulfur cycle driven by autoxidation of sulfide is discussed.     

Sulfate reduction and possible aerobic metabolism of the sulfate-reducing bacterium Desulfovibrio oxyclinae in a chemostat coculture with Marinobacter sp. Strain MB under exposure to increasing oxygen concentrations

A chemostat coculture of the sulfate-reducing bacterium Desulfovibrio oxyclinae together with a facultative aerobe heterotroph tentatively identified as Marinobacter sp. strain MB was grown under anaerobic conditions and then exposed to a stepwise-increasing oxygen influx (0 to 20% O(2) in the incoming gas phase). The coculture consumed oxygen efficiently, and no residual oxygen was detected with an oxygen supply of up to 5%. Sulfate reduction persisted at all levels of oxygen input, even at the maximal level, when residual oxygen in the growth vessel was 87 microM. The portion of D. oxyclinae cells in the coculture decreased gradually from 92% under anaerobic conditions to 27% under aeration. Both absolute cell numbers and viable cell counts of the organism were the same as or even higher than those observed in the absence of oxygen input. The patterns of consumption of electron donors and acceptors suggest that aerobic incomplete oxidation of lactate to acetate is performed by D. oxyclinae under high oxygen input. Both organisms were isolated from the same oxic zone of a cyanobacterial mat where they have to adapt to daily shifts from oxic to anoxic conditions. This type of syntrophic association may occur in natural habitats, enabling sulfate-reducing bacteria to cope with periodic exposure to oxygen.     

Transition from anaerobic to aerobic growth conditions for the sulfate-reducing bacterium Desulfovibrio oxyclinae results in flocculation

A chemostat culture of the sulfate-reducing bacterium Desulfovibrio oxyclinae isolated from the oxic layer of a hypersaline cyanobacterial mat was grown anaerobically and then subjected to gassing with 1% oxygen, both at a dilution rate of 0.05 h(-1). The sulfate reduction rate under anaerobic conditions was 370 nmol of SO(4)(2-) mg of protein(-1) min(-1). At the onset of aerobic gassing, sulfate reduction decreased by 40%, although viable cell numbers did not decrease. After 42 h, the sulfate reduction rate returned to the level observed in the anaerobic culture. At this stage the growth yield increased by 180% compared to the anaerobic culture to 4.4 g of protein per mol of sulfate reduced. Protein content per cell increased at the same time by 40%. The oxygen consumption rate per milligram of protein measured in washed cell suspensions increased by 80%, and the thiosulfate reduction rate of the same samples increased by 29% with lactate as the electron donor. These findings indicated possible oxygen-dependent enhancement of growth. After 140 h of growth under oxygen flux, formation of cell aggregates 0.1 to 3 mm in diameter was observed. Micrometer-sized aggregates were found to form earlier, during the first hours of exposure to oxygen. The respiration rate of D. oxyclinae was sufficient to create anoxia inside clumps larger than 3 microm, while the levels of dissolved oxygen in the growth vessel were 0.7 +/- 0.5 microM. Aggregation of sulfate-reducing bacteria was observed within a Microcoleus chthonoplastes-dominated layer of a cyanobacterial mat under daily exposure to oxygen concentrations of up to 900 microM. Desulfonema-like sulfate-reducing bacteria were also common in this environment along with other nonaggregated sulfate-reducing bacteria. Two-dimensional mapping of sulfate reduction showed heterogeneity of sulfate reduction activity in this oxic zone.     

Examination of Lactobacillus plantarum lactate metabolism side effects in relation to the modulation of aeration parameters

AIMS: The characterization of global aerobic metabolism of Lactobacillus plantarum LP652 under different aeration levels, in order to optimize acetate production kinetics and to suppress H2O2 toxicity. METHODS AND RESULTS: Cultures of L. plantarum were grown on different aeration conditions. After sugar exhaustion and in the presence of oxygen, lactate was converted to acetate, H2O2 and carbon dioxide with concomitant ATP production. Physiological assays were performed at selected intervals in order to assess enzyme activity and vitality of the strain during lactic acid conversion. The maximal aerated condition led to fast lactate-to-acetate conversion kinetics between 8 and 12 h, but H2O2 immediately accumulated, thus affecting cell metabolism. Pyruvate oxidase activity was highly enhanced by oxygen tension and was responsible for H2O2 production after 12 h of culture, whereas lactate oxidase and NADH-dependent lactate dehydrogenase activities were not correlated to metabolite production. Limited NADH oxidase (NOX) and NADH peroxidase (NPR) activities were probably responsible for toxic H2O2 levels in over-aerated cultures. CONCLUSION: Modulating initial airflow led to the maximal specific activity of NOX and NPR observed after 24 h of culture, thus promoting H2O2 destruction and strain vitality at the end of the process. SIGNIFICANCE AND IMPACT OF THE STUDY: Optimal aeration conditions were determined to minimize H2O2 concentration level during growth on lactate.     

A device for aeration and mixing of cell and organelle suspensions during nuclear magnetic resonance studies

A device is described which maintains homogeneous aerobic or anaerobic cell and organelle suspensions within an NMR sample tube. Line broadening due to magnetic field inhomogeneity is reduced by elimination of gas bubbles from the area of the probe receiver coils. The linewidth of the extracellular orthophosphate resonance of a yeast suspension in 31P NMR was 0.21 ppm compared with 0.4-0.7 ppm in conventionally aerated suspensions. Recirculation of the sample results in complete mixing within 90 s of addition of aliquots of acid or alkali. The maximum rate of oxygen transfer from the gaseous to the liquid phase was approximately 600 microM min-1 when aerated with 95% oxygen/5% carbon dioxide. A 60% wet weight suspension of yeast cells was recirculated for 20 h without settling of cells occurring. A method for estimating oxygen transfer rate is described.     

Desulfovibrio aerotolerans sp. nov., an oxygen tolerant sulphate-reducing bacterium isolated from activated sludge

A new mesophilic sulphate-reducing bacterium, designated strain DvO5(T) (T=type strain), was isolated from the outermost sulphate reduction-positive most-probable-number tube (10(-6) dilution) of an activated sludge sample, which had been oxygenated at 100% air saturation for 120 h. The motile, Gram-negative, curved 1 by 2-5 microm and non-spore-forming cells of strain DvO5(T) existed singly or in chains. Strain DvO5(T) grew optimally at 29 degrees C, pH 6.9 and 0.05% (w/v) NaCl in a medium containing lactate, sulphate and yeast extract. Sulphite, thiosulphate and elemental sulphur also served as electron acceptors whereas nitrate, nitrite or ferric iron were not reduced. Lactate, pyruvate, H(2) (with acetate as carbon source), ethanol and glycerol efficiently supported growth as electron donors. Pyruvate and malate were fermented. Strain DvO5(T) reduced oxygen by oxidising endogenous polyglucose at rates ranging from 0.4 to 6.0 nmol O(2)/mg protein min depending on the oxygen concentration, the highest rates being observed at atmospheric oxygen saturation. The G+C content of the DNA was 57.2 mol%. Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain DvO5(T) was a member of the genus Desulfovibrio with D. magneticus (98.2% 16S rRNA gene sequence similarity) and D. burkinensis (97.5% 16S rRNA gene sequence similarity) being its closest relatives among validly described species. A similar phylogenetic affiliation was obtained by sequence analyses of the genes encoding the alpha and the beta subunit of dissimilatory sulphite reductase (dsrAB) as well as the alpha subunit of adenosine-5'-phosphosulphate reductase (apsA) of strain DvO5(T). On the basis of genotypic and phenotypic characteristics, strain DvO5(T) (DSM 16695(T), JCM 12613(T)) is proposed as the type strain of a new species, Desulfovibrio aerotolerans sp. nov.     

Effects of Environmental Parameters on the Formation and Turnover of Acetate by Forest Soils

The capacity to form acetate from endogenous matter was a common property of diverse forest soils when incubated under anaerobic conditions. At 15 to 20(deg)C, acetate synthesis occurred without appreciable delay when forest soils were incubated as buffered suspensions or in microcosms at various percentages of their maximum water holding capacity. Rates for acetate formation with soil suspensions ranged from 35 to 220 (mu)g of acetate per g (dry weight) of soil per 24 h, and maximal acetate concentrations obtained in soil suspensions were two- to threefold greater than those obtained with soil microcosms at the average water holding capacity of the soil. Cellobiose degradation in soil suspensions yielded H(inf2) as a transient product. Under anaerobic conditions, supplemental H(inf2) and CO(inf2) were directed towards the acetogenic synthesis of acetate, and enrichments yielded a syringate-H(inf2)-consuming acetogenic consortium. At in situ temperatures, acetate was a relatively stable anaerobic end product; however, extended incubation periods induced acetoclastic methanogenesis and sulfate reduction. Higher mesophilic and thermophilic temperatures greatly enhanced the capacity of soils to form methane. Although methanogenic and sulfate-reducing activities under in situ-relevant conditions were negligible, these findings nonetheless demonstrated the occurrence of methanogens and sulfate-reducing bacteria in these aerated terrestrial soils. In contrast to the protracted stability of acetate under anaerobic conditions at 15 to 20(deg)C with unsupplemented soils, acetate formed by forest soils was rapidly consumed in the presence of oxygen and nitrate, and substrate-product stoichiometries indicated that acetate turnover was coupled to oxygen-dependent respiration and denitrification. The collective results suggest that acetate formed under anaerobic conditions might constitute a trophic link between anaerobic and aerobic processes in forest soils.     

A sulfate-reducing bacterium from the oxic layer of a microbial mat from Solar Lake (Sinai), Desulfovibrio oxyclinae sp. nov.

In an investigation on the oxygen tolerance of sulfate-reducing bacteria, a strain was isolated from a 10⁷-fold dilution of the upper 3-mm layer of a hypersaline cyanobacterial mat (transferred from Solar Lake, Sinai). The isolate, designated P1B, appeared to be well-adapted to the varying concentrations of oxygen and sulfide that occur in this environment. In the presence of oxygen strain P1B respired aerobically with the highest rates [260 nmol O₂ min^–1 (mg protein)^–1] found so far among marine sulfate-reducing bacteria. Besides H₂ and lactate, even sulfide or sulfite could be oxidized with oxygen. The sulfur compounds were completely oxidized to sulfate. Under anoxic conditions, it grew with sulfate, sulfite, or thiosulfate as the electron acceptor using H₂, lactate, pyruvate, ethanol, propanol, or butanol as the electron donor. Furthermore, in the absence of electron donors the isolate grew by disproportionation of sulfite or thiosulfate to sulfate and sulfide. The highest respiration rates with oxygen were obtained with H₂ at low oxygen concentrations. Aerobic growth of homogeneous suspensions was not obtained. Additions of 1% oxygen to the gas phase of a continuous culture resulted in the formation of cell clumps wherein the cells remained viable for at least 200 h. It is concluded that strain P1B is oxygen-tolerant but does not carry out sulfate reduction in the presence of oxygen under the conditions tested. Analysis of the 16S rDNA sequence indicated that strain P1B belongs to the genus Desulfovibrio, with Desulfovibrio halophilus as its closest relative. Based on physiological properties strain P1B could not be assigned to this species. Therefore, a new species, Desulfovibrio oxyclinae, is proposed. Received: 7 August 1996 / Accepted: 29 January 1997     

Sulfate Reduction and Possible Aerobic Metabolism of the Sulfate-Reducing Bacterium Desulfovibrio oxyclinae in a Chemostat Coculture with Marinobacter sp. Strain MB under Exposure to Increasing Oxygen Concentrations

A chemostat coculture of the sulfate-reducing bacterium Desulfovibrio oxyclinae together with a facultative aerobe heterotroph tentatively identified as Marinobacter sp. strain MB was grown under anaerobic conditions and then exposed to a stepwise-increasing oxygen influx (0 to 20% O₂ in the incoming gas phase). The coculture consumed oxygen efficiently, and no residual oxygen was detected with an oxygen supply of up to 5%. Sulfate reduction persisted at all levels of oxygen input, even at the maximal level, when residual oxygen in the growth vessel was 87 μM. The portion of D. oxyclinae cells in the coculture decreased gradually from 92% under anaerobic conditions to 27% under aeration. Both absolute cell numbers and viable cell counts of the organism were the same as or even higher than those observed in the absence of oxygen input. The patterns of consumption of electron donors and acceptors suggest that aerobic incomplete oxidation of lactate to acetate is performed by D. oxyclinae under high oxygen input. Both organisms were isolated from the same oxic zone of a cyanobacterial mat where they have to adapt to daily shifts from oxic to anoxic conditions. This type of syntrophic association may occur in natural habitats, enabling sulfate-reducing bacteria to cope with periodic exposure to oxygen.     

Oxygen-dependent growth of the sulfate-reducing bacterium Desulfovibrio oxyclinae in coculture with Marinobacter sp. Strain MB in an aerated sulfate-depleted chemostat

A chemostat coculture of the sulfate-reducing bacterium Desulfovibrio oxyclinae and the facultatively aerobic heterotroph Marinobacter sp. strain MB was grown for 1 week under anaerobic conditions at a dilution rate of 0.05 h(-1). It was then exposed to an oxygen flux of 223 micromol min(-1) by gassing the growth vessel with 5% O(2). Sulfate reduction persisted under these conditions, though the amount of sulfate reduced decreased by 45% compared to the amount reduced during the initial anaerobic mode. After 1 week of growth under these conditions, sulfate was excluded from the incoming medium. The sulfate concentration in the growth vessel decreased exponentially from 4.1 mM to 2.5 microM. The coculture consumed oxygen effectively, and no residual oxygen was detected during either growth mode in which oxygen was supplied. The proportion of D. oxyclinae cells in the coculture as determined by in situ hybridization decreased from 86% under anaerobic conditions to 70% in the microaerobic sulfate-reducing mode and 34% in the microaerobic sulfate-depleted mode. As determined by the most-probable-number (MPN) method, the numbers of viable D. oxyclinae cells during the two microaerobic growth modes decreased compared to the numbers during the anaerobic growth mode. However, there was no significant difference between the MPN values for the two modes when oxygen was supplied. The patterns of consumption of electron donors and acceptors suggested that when oxygen was supplied in the absence of sulfate and thiosulfate, D. oxyclinae performed incomplete aerobic oxidation of lactate to acetate. This is the first observation of oxygen-dependent growth of a sulfate-reducing bacterium in the absence of either sulfate or thiosulfate. Cells harvested during the microaerobic sulfate-depleted stage and exposed to sulfate and thiosulfate in a respiration chamber were capable of anaerobic sulfate and thiosulfate reduction.     

Complete anaerobic oxidation of hydroquinone by Desulfococcus sp. strain Hy5: indications of hydroquinone carboxylation to gentisate

The sulfate-reducing strain Hy5 was able to grow with hydroquinone as sole source of carbon and energy. In experiments with dense cell suspensions, several indications were found that gentisate was the first intermediate in anaerobic degradation of hydroquinone: (1) degradation of hydroquinone was accelerated by addition of bicarbonate; (2) cell suspensions grown with hydroquinone oxidized gentisate at a rate similar to that of suspensions grown with gentisate, whereas the latter were not able to degrade hydroquinone in the presence of chloramphenicol; (3) in SDS-PAGE analysis of cell-free extracts of strain Hy5, two additional protein bands were found after growth with hydroquinone that were not detected in cells grown with gentisate, probably representing a hydroquinone carboxylating enzyme. A corresponding enzyme activity could not be detected. In cell-free extracts of hydroquinone-grown strain Hy5, the specific acyl-CoA ligase activity with gentisate as substrate was detected at 70 nmol x mg^-1 x min^-1. Gentisyl-CoA was enzymatically reduced to several unidentified nonaromatic products in the presence of dithionite-reduced methyl viologen.     

Studies on in vivo inactivation of gramicidin S synthetase and its retardation

The oxygen-dependent in vivo inactivation of gramicidin S synthetase was investigated in Bacillus brevis ATCC 9999. Inhibitors of energy metabolism and of protein synthesis added to aerated cell suspensions did not provide any protection against inactivation, thus indicating that the process does not depend on energy-yielding metabolism nor on de novo protein synthesis. Organic thiols added to anaerobic long-term incubations retarded synthetase inactivation for several hours, whereas in short-term incubations of previously air-exposed cells they resulted in partial restoration of activity. The in vivo inactivation of the enzyme was found to be accompanied by a parallel drop in intracellular thiols. These results implicate enzyme SH oxidation as a mechanism of in vivo inactivation. Retardation of inactivation was achieved upon addition of utilizable carbon sources (glycerol, fructose, inositol) to aerated cell suspensions in buffer, the degree of stabilization being proportional to the ease of uptake and to the concentration of C source. This effect involves actual consumption of the exogenous C source and is accompanied by lower dissolved oxygen levels in the cell suspension. Pulsed additions of C source could retard inactivation but could not restore partly or fully lost activity. The C-source effect was blocked by the uncoupler dinitrophenol, while dissolved oxygen levels in the suspension remained low. C-source-supplemented cell suspensions incubated under air had a decreased intracellular redox state, as revealed by intracellular SH concentration.     

Denitrification under various aeration conditions in Comamonas sp., strain SGLY2

Abstract A number of bacteria were isolated from different anoxic reactors. Those having denitrifying potential were tested for their ability to denitrify under aerobic conditions. The activity of their denitrifying enzymes varied from partial inactivation by oxygen (strains NO2B9 and TCET1) to oxygen-independent activity in a strain named SGLY2 which was tentatively identified as Comamonas sp. The effect of different aeration conditions on growth and on denitrification of SGLY2 was studied more extensively. This strain was able to consume oxygen and nitrate simultaneously with the production of nitrogen and without build-up of nitrite. The dissimilatory nitrate-reductase of nitrate-adapted cells was found to be more active in the presence of oxygen than in micro-aerobic or strictly anaerobic conditions.     

Isolation of a sulfate-reducing bacterium growing with methanol

Abstract A sulfate-reducing bacterium capable of growth with methanol as sole source of energy was isolated from an anaerobic waste-water treatment plant via enrichment in an ethanol/sulfate medium. The cells were curved, Gram-negative, nonmotile rods. No spore formation was observed. Glycerol could be fermented to 1,3-propanediol and 3-hydroxypropionate. In the presence of sulfate, glycerol was stoichiometrically converted to 3-hydroxypropionate. The cells contained stacked intracellular membranes during growth with ethanol or lactate but not with methanol. The doubling time during growth on methanol was approx. 75 h. Methanol was metabolized according to the following equation:
The possible role of sulfate-reducing bacteria in the anaerobic mineralization of methanol is discussed.     

Oxygen dependent lactate utilization by Lactobacillus plantarum

Lactobacillus plantarum P5 grew aerobically in rich media at the expense of lactate; no growth was observed in the absence of aeration. The oxygen-dependent growth was accompanied by the conversion of lactate to acetate which accumulated in the growth medium. Utilization of oxygen with lactate as substrate was observed in buffered suspensions of washed whole cells and in cell-free extracts. A pathway which accounts for the generation of adenosine triphosphate during aerobic metabolism of lactate to acetate via pyruvate and acetyl phosphate is proposed. Each of the enzyme activities involved, nicotinamide adenine dinucleotide independent lactic dehydrogenase, nicotinamide adenine dinucleotide dependent lactic dehydrogenase, pyruvate oxidase, acetate kinase and NADH oxidase were demonstrated in cell-free extracts. The production of pyruvate, acetyl phosphate and acetate was demonstrated using cell-free extracts and cofactors for the enzymes of the proposed pathway.Abbreviations MRS Man, Rogosa and Sharpe (1960) medium modified as in Materials and methods - TY Tryptone Yeast Extract broth - OUL Oxygen uptake with lactate as substrate - DCPIP 2,6-Dichlorophenolindophenol - LDH Lactic dehydrogenase     

Effects of Oxygen and Heme on the Development of a Microbial Respiratory System

The effect of adding hemin to anaerobically grown cells of a strain of Staphylococcus epidermidis, which was heme-deficient due to anaerobic growth, has been examined. Cells grown anaerobically in media containing hemin exhibited a marked increase in several oxidative activities as compared with cells grown anaerobically without hemin. The respiratory activity of whole cells and a cyamide-sensitive reduced nicotinamide adenine dinucleotide oxidase activity of cell-free extracts were increased fourfold. The content of enzymatically reducible pigments which exhibit difference spectra similar to cytochromes b(1) and o was also markedly increased. These pigments are mostly sedimented at 100,000 x g (1 hr). Hemin also caused a marked increase in respiratory activity when added directly to the anaerobic culture after the period of growth, but did not cause a similar increase in respiration when added to washed, resting-cell suspensions. Under the latter conditions, heme pigments were formed which exhibited difference spectra similar to, but not identical with, the spectra of pigments found in anaerobic cells grown in the presence of hemin. When resting suspensions of cells grown anaerobically without hemin were exposed to air, a rapid fourfold increase in respiratory activity and a limited increase in cytochrome-like pigments occurred. The presence of the heme precursor Delta-aminolevulinic acid during this aeration resulted in a rapid and marked increase in heme pigments, but only a slight stimulation of respiratory activity. The possible implications of these results for the roles which heme and oxygen play in the development of the respiratory system of this organism are discussed.     

Transition from Anaerobic to Aerobic Growth Conditions for the Sulfate-Reducing Bacterium Desulfovibrio oxyclinae Results in Flocculation

A chemostat culture of the sulfate-reducing bacterium Desulfovibrio oxyclinae isolated from the oxic layer of a hypersaline cyanobacterial mat was grown anaerobically and then subjected to gassing with 1% oxygen, both at a dilution rate of 0.05 h⁻¹. The sulfate reduction rate under anaerobic conditions was 370 nmol of SO₄²⁻ mg of protein⁻¹ min⁻¹. At the onset of aerobic gassing, sulfate reduction decreased by 40%, although viable cell numbers did not decrease. After 42 h, the sulfate reduction rate returned to the level observed in the anaerobic culture. At this stage the growth yield increased by 180% compared to the anaerobic culture to 4.4 g of protein per mol of sulfate reduced. Protein content per cell increased at the same time by 40%. The oxygen consumption rate per milligram of protein measured in washed cell suspensions increased by 80%, and the thiosulfate reduction rate of the same samples increased by 29% with lactate as the electron donor. These findings indicated possible oxygen-dependent enhancement of growth. After 140 h of growth under oxygen flux, formation of cell aggregates 0.1 to 3 mm in diameter was observed. Micrometer-sized aggregates were found to form earlier, during the first hours of exposure to oxygen. The respiration rate of D. oxyclinae was sufficient to create anoxia inside clumps larger than 3 μm, while the levels of dissolved oxygen in the growth vessel were 0.7 ± 0.5 μM. Aggregation of sulfate-reducing bacteria was observed within a Microcoleus chthonoplastes-dominated layer of a cyanobacterial mat under daily exposure to oxygen concentrations of up to 900 μM. Desulfonema-like sulfate-reducing bacteria were also common in this environment along with other nonaggregated sulfate-reducing bacteria. Two-dimensional mapping of sulfate reduction showed heterogeneity of sulfate reduction activity in this oxic zone.     

<Emphasis Type="Italic">Sporotalea propionica</Emphasis> gen. nov. sp. nov., a hydrogen-oxidizing,oxygen-reducing,propionigenic firmicute from the intestinal tract of a soil-feeding termite

An unusual propionigenic bacterium was isolated from the intestinal tract of the soil-feeding termite Thoracotermes macrothorax. Strain TmPN3 is a motile, long rod that stains gram-positive, but reacts gram-negative in the KOH test. It forms terminal endospores and ferments lactate, glucose, lactose, fructose, and pyruvate to propionate and acetate via the methyl-malonyl-CoA pathway. Propionate and acetate are formed at a ratio of 2:1, typical of most propionigenic bacteria. Under a H₂/CO₂ atmosphere, the fermentation product pattern of glucose, fructose, and pyruvate shifts towards propionate formation at the expense of acetate. Cell suspensions reduce oxygen with lactate, glucose, glycerol, or hydrogen as electron donor. In the presence of oxygen, the product pattern of lactate fermentation shifts from propionate to acetate production. 16S rRNA gene sequence analysis showed that strain TmPN3 is a firmicute that clusters among the Acidaminococcaceae, a subgroup of the Clostridiales comprising obligately anaerobic, often endospore-forming bacteria that possess an outer membrane. Based on phenotypic differences and less than 92% sequence similarity to the 16S rRNA gene sequence of its closest relative, the termite hindgut isolate Acetonema longum, strain TmPN3^T is proposed as the type species of a new genus, Sporotalea propionica gen. nov. sp. nov. (DSM 13327^T, ATCC BAA-626^T).     

β-Galactosidase of Propionibacterium shermanii

Ten strains of Propionibacterium shermanii were tested for beta-galactosidase (beta-gal) activity. Of these ten strains, five yielded enhanced enzyme activity when cell suspensions were treated with toluene-acetone; on solvent treatment, the remaining five lost a considerable portion of the activity found in whole-cell suspensions. By using a strain yielding decreased activity upon solvent treatment, explanations for the loss in activity were sought through assays for possible alternative beta-galactoside utilization mechanisms. When this strain was assayed for beta-D-phosphogalactoside galactohydrolase by using orthonitrophenyl-beta-D-galactopyranoside-6-P04 as a substrate, the activity was wither lower or indiffernt as compared with beta-gal activity determined simultaneously. Cell suspensions of P. shermanii 7 and 22 (strains chosen for further work) grown separately on the individual substrates (lactose, glucose, galactose, and sodium lactate) did not show significant differences in beta-gal activity. Optimal temperature for beta-gal activity in untreated and toluene-acetone-treated cell suspensions of strain 7 was 52 C. With strain 22, of the temperatures tested, maximal activity in untreated cell suspensions was noted at 58 C and with solvent-treated cells at 32 C. In the cell-free extract (CFE) system, both strains exhibited maximal activity at 52 C. Optimal pH for untreated and solvent-treated cell suspensions of both strains was around 7.5. In the P. shermanii 22 CFE system, maximal activity occurred at pH 7.0; pH had very little effect on enzyme activity in P. shermanii 7 CFE. Sodium or potassium phosphate buffers in the assay system yielded the best activity. In the CFE system of these two strains, Mn2+ was definitely stimulatory, but in untreated and solvent-treated cell systems of these strains presence or absence of Mn2+ in the assay system had variable effects on enzyme activity. Maximal beta-gal activity was noted in P. shermanii 7 cells harvested after 28 h of growth at 32 C in sodium lactate broth. Sulfhydryl-group blocking agents inhibited enzyme activity in P. shermanii 22 CFE; the inhibition was partly reversed by dithiothreitol.