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PubMed2006

Secondary mitochondrial dysfunction in propionic aciduria: a pathogenic role for endogenous mitochondrial toxins.

Mitochondrial dysfunction during acute metabolic crises is considered an important pathomechanism in inherited disorders of propionate metabolism, i.e. propionic and methylmalonic acidurias. Biochemically, these disorders are characterized by accumulation of propionyl-CoA and metabolites of alternative propionate oxidation. In the present study, we demonstrate uncompetitive inhibition of PDHc (pyruvate dehydrogenase complex) by propionyl-CoA in purified porcine enzyme and in submitochondrial particles from bovine heart being in the same range as the inhibition induced by acetyl-CoA, the physiological product and known inhibitor of PDHc. Evaluation of similar monocarboxylic CoA esters showed a chain-length specificity for PDHc inhibition. In contrast with CoA esters, non-esterified fatty acids did not inhibit PDHc activity. In addition to PDHc inhibition, analysis of respiratory chain and tricarboxylic acid cycle enzymes also revealed an inhibition by propionyl-CoA on respiratory chain complex III and alpha-ketoglutarate dehydrogenase complex. To test whether impairment of mitochondrial energy metabolism is involved in the pathogenesis of propionic aciduria, we performed a thorough bioenergetic analysis in muscle biopsy specimens of two patients. In line with the in vitro results, oxidative phosphorylation was severely compromised in both patients. Furthermore, expression of respiratory chain complexes I-IV and the amount of mitochondrial DNA were strongly decreased, and ultrastructural mitochondrial abnormalities were found, highlighting severe mitochondrial dysfunction. In conclusion, our results favour the hypothesis that toxic metabolites, in particular propionyl-CoA, are involved in the pathogenesis of inherited disorders of propionate metabolism, sharing mechanistic similarities with propionate toxicity in micro-organisms.

Acetyl Coenzyme A↗
PubMed1994

Polarized Na+/H+ exchange function is pliable in response to transepithelial gradients of propionate.

Short-chain fatty acids are produced at high concentration in the colonic lumen and stimulate electroneutral Na+ absorption by activating apical Na+/H+ exchange in colonocytes. We used an epithelial cell line derived from a human colon carcinoma (HT29-18-C1) to study activation of apical and basolateral Na+/H+ exchange by a short-chain fatty acid, propionate. Confluent cell monolayers on membrane filters were loaded with 2',7'-bis(2-carboxyethyl)-5 (and 6)-carboxyfluorescein (a fluorescent pH indicator) and intracellular pH was monitored with a digital fluorescence imaging microscope. Cells acidified by transient exposure to NH4Cl demonstrated both apical and basolateral Na+/H+ exchange. In this condition, apical Na+/H+ exchange was 50% of the total Na+/H+ exchange activity. Similar results were obtained when cells were bilaterally perfused with apical and basolateral propionate in an isosmotic medium (130 mM propionate at each membrane surface). However, apical Na+/H+ exchange was a significantly larger fraction (76%) of the total Na+/H+ exchange activity when cells were acidified by exposure to apical propionate alone. Conversely, in cells acidified by basolateral propionate alone, apical Na+/H+ exchange was 21% of the total Na+/H+ exchange activity. The change in relative activity was observed in individual cells which expressed both apical and basolateral Na+/H+ exchange and occurred rapidly (within 7 min). In the presence of transepithelial propionate gradients, all Na(+)-dependent alkalinization was sensitive to 3 microM 5-(N-ethyl-N-isopropyl)amiloride, a potent Na+/H+ exchange inhibitor. These results suggest that transepithelial gradients of short-chain fatty acids, which occur in vivo, can cause preferential activation of apical Na+/H+ exchange.

Ammonium Chloride↗
PubMed1975

A relationship between the molar proportion of propionic acid and the clearance rate of the liquid phase in the rumen of the sheep.

1. Four rumen-cannulated sheep were given a forage mixture (F) of chopped hay-ground, pelleted, dried grass (92:8, w/w) and two concentrate mixtures (C and S) of ground barley-ground hay-flaked maize (46:24:30 and 56:24:20, by wt respectively) in twenty-four hourly meals each day. Each of the diets was offered in successive periods of 16 d to give a feeding sequence F-S-C-S for one pair of sheep and C-S-F-S for the other pair. 2. The average composition (mol/100 mol) of the mixture of short-chain fatty acids, acetic, propionic and butyric, in the rumen was respectively 70-1, 18-5 and 7-5 with diet F, and 55-8, 24-8 and 13-6 with diet C. With diet S, the pattern of fermentation varied both between animals and in the same animal for different periods having either 'high' (28-39 mol/100 mol) or 'low' (16-21 mol/100 mol) proportions of propionic acid. On average when diet S followed diet F there was less propionic acid in the fermentation mixture than when diet S followed diet C (59-3 acetic, 22-2 propionic and 14-1 butyric as compared with 52-7, 29-4 and 13-1 respectively) but this trend was not significant and there was evidence of interactions between the feeding sequences and the individual sheep. 3. The mean concentrations of ammonia, sodium, potassium and chloride were similar for all diets but the pH and concentrations of calcium, magnesium and phosphorus tended to be higher and the buffering capacity lower for diet F than for diets C or S. In animals receiving diet S there was no relationship between the concentrations of minerals, the pH or buffering capacity and the pattern of fermentation except for ammonia, the concentration of which was high when the molar proportion of propionic acid was low. 4. Rumen volume, outflow rate and clearance rate, determined using polyethylene glycol, were higher for diet F than for diets C and S but within each diet, particularly for diet S, values varied considerably between sheep and between periods. 5. There was evidence of an interrelationship between the molar proportion of propionic acid in the fermentation products and the clearance rate, which indicated that the clearance rate may be an important factor influencing the pattern of fermentation in the rumen.

Acetates↗
PubMed1987

Effects on plasma insulin of intermittent infusions of propionic acid, glucose or casein into the alimentary tract of non-lactating cows maintained on a liquid diet.

1. An experiment was conducted using three non-lactating cows completely maintained by infusions of volatile fatty acids into the rumen, and casein into the abomasum. Plasma insulin responses to propionic acid, glucose or casein were recorded. Further information was obtained using protein-free infusions. 2. When part of the propionic acid was infused into the rumen in a twice-daily 3 h dose and the remainder infused continuously with acetic and butyric acids and casein, there were large increases in the concentrations of propionic acid and insulin in the jugular blood. When glucose, corresponding in energy to that supplied by the intermittent propionic acid infusions was similarly infused, the plasma levels of glucose and insulin were increased. Glucose appeared to stimulate a greater increase in insulin than did propionic acid. Casein infused into the abomasum in intermittent doses produced a rise in plasma insulin, but smaller than that observed with propionic acid or with glucose. 3. The protein-free infusion was characterized by a lower concentration of insulin in the blood plasma, a reduction in plasma urea and free amino nitrogen and unchanged plasma glucose.

Abomasum↗
PubMed1993

The role of propionate and acetate in the control of food intake in sheep.

Sheep fed ad lib. on a good-quality pelleted diet (g/kg; hay 500, barley 300, molasses, fish meal and minerals) were infused via the hepatic portal vein with mixtures of the sodium salts of volatile fatty acids, acetate and propionate, and a variety of equivalent osmotic loads. Propionate infused at rates between 0.6 and 2.5 mmol/min consistently reduced food intake in a linear, dose-related manner. Propionate infusions resulted in consistent dose-related increases in peripheral venous plasma glucose concentration but variable changes in insulin concentration. Infusion of osmotically balanced mixtures of propionate, acetate, mannitol or saline (9 g NaCl/l) indicated that at constant osmotic loading propionate caused a greater reduction in intake than other infusions. Acetate infusions only depressed food intake when administered as a 1 M solution. Lower concentrations had little effect, similar to that of equivalent osmotic loads of mannitol or saline. It is concluded that portal propionate flow has a potential role in the control of food intake in ruminants which is independent of osmotic effects or changes in plasma insulin concentration.

Acetates↗
PubMed2003

Intraruminal propionate supplementation modifies hindlimb energy metabolism without changing the splanchnic release of glucose in growing lambs.

The influence of propionate supplementation on the splanchnic metabolism of energy-yielding nutrients and the supply of glucose to the hindlimb was investigated in growing lambs. Six rumen-cannulated and multicatheterized lambs (32.2 kg), fed frozen rye grass at 690 kJ metabolizable energy intake/d per kg body weight0.75, were infused intraruminally with a salt solution (control) or with propionate solutions at 0.55 mol/d (P1) or 0.98 mol/d (P2) according to a replicated Latin square design. In the rumen fluid, supplementation decreased the acetate:propionate molar ratio from 2.36:1 to 1.37:1, without modifying the ruminal concentrations of acetate and NH4. As a result, the portal appearance of propionate increased by 51 and 72 % with P1 and P2, respectively, and that of l-lactate doubled. Across the liver, net extraction of propionate increased by 47 and 67 % with P1 and P2, respectively. However, the net hepatic production of glucose remained unchanged, probably as the result of a substantial rise in insulin secretion and its hepatic extraction. Overall, the net splanchnic release of acetate, glucose and butyrate was not modified while that of l-lactate increased. Despite this, the net uptake of acetate, glucose, l-lactate and non-esterified fatty acids by the hindlimb increased. Propionate probably enhanced the storage of energy-yielding nutrients in the hindlimb, despite their unchanged release by the splanchnic tissues and the unmodified insulinaemia. Regulatory mechanisms are not clear.

Animal Nutritional Physiological Phenomena↗
PubMed1991

Propionate lowers blood glucose and alters lipid metabolism in healthy subjects.

Incorporation of 3.3 g sodium propionate per 50 g available carbohydrate portion of bread reduced acutely the blood glucose response area in six healthy volunteers by 47.6 +/- 12.1% Similarly, in vitro digestibility was reduced by 47.4 +/- 1.1% (P less than 0.01). One week of dietary supplementation with 9.9 g sodium propionate in bread/d reduced the blood glucose area in comparison with standard propionate-free bread by 38.0 +/- 8.7% (P less than 0.05), but increased fecal bulk by 28.3 +/- 8.7% (P less than 0.05) and anaerobic microflora by 0.564 +/- 0.165 X 10(6)/g feces (P less than 0.05), specifically as bifidobacteria. Day-long breath hydrogen concentrations did not increase after 1 wk on propionate bread but methane production increased in the three methane producers. Although lipid changes were not significant, five subjects showed reduced high-density-lipoprotein and increased triglyceride concentrations, both of which correlated with increased fecal weight (P less than 0.05). Because propionate reduces the rate of starch digestion, studies using oral propionate must take into account its action as an enzyme inhibitor.

Adult↗
PubMed1995

Propionate inhibits incorporation of colonic [1,2-13C]acetate into plasma lipids in humans.

Acetate and propionate, produced during colonic fermentation of unabsorbed carbohydrate, may influence systemic lipid metabolism. As a preliminary study to see whether colonic acetate is incorporated into plasma lipids and whether propionate inhibits this process, 5 healthy males were studied after fasting overnight. They were given, in random order, 12.5 mmol (1.05 g) [1,2-13C]sodium acetate by intravenous or rectal infusion, and the rectal infusion was given with or without 6 mmol (0.58 g) sodium propionate. Two hours after rectal acetate, 13C recoveries in plasma cholesterol (0.59 +/- 0.22%) and triglycerides (1.24 +/- 0.69%) were significantly greater than after intravenous acetate (0.09 +/- 0.12% and 0.29 +/- 0.18%, respectively). Addition of propionate reduced 13C recovery in triglycerides (0.19 +/- 0.06%, P = 0.024) compared with rectal acetate alone, but the effect on cholesterol (0.26 +/- 0.05%) was not significant. These data suggest that incorporation of colonic acetate into plasma triglycerides is inhibited by propionate. Further studies are required to quantify the effects of colonic acetate and propionate on lipid synthesis.

Acetates↗
PubMed2001

Measurements of the gastric emptying rate by use of ultrasonography: studies in humans using bread with added sodium propionate.

BACKGROUND: Foods with a low glycemic index are increasingly being acknowledged as beneficial for individuals with disorders related to the insulin resistance syndrome. The presence of certain salts of organic acids has been shown to lower the glycemic index of bread products and one of the suggested mechanisms is a lowered gastric emptying rate (GER). One obvious pitfall with many of the common techniques for GER measurement is that the food structure, and hence the gastric release of nutrients, may be affected by enclosure of the marker for gastric emptying, eg, paracetamol. Ultrasonography is a noninvasive method for which the above pitfall is to a large extent avoided. OBJECTIVE: The main objective was to evaluate the use of ultrasonography to determine whether the lowered glycemic and insulinemic responses to bread ingestion after the addition of sodium propionate are explained by a specific effect of propionate on the GER. DESIGN: The effect of sodium propionate in bread was evaluated in 9 healthy volunteers. Barley bread products, with or without added sodium propionate, were ingested as breakfast after an overnight fast. The GER was monitored for 2 h by ultrasonography; during this period, capillary blood was withdrawn repeatedly for measurement of blood glucose and insulin. RESULTS: The GER of the barley bread decreased markedly after the addition of sodium propionate and was accompanied by lowered glycemic and insulinemic responses. CONCLUSION: The lowered glycemic response to ingestion of bread with added sodium propionate appears to be related to a lowered GER.

Adolescent↗
PubMed1983

Influence of vitamin B-12 status on hepatic propionic acid uptake in sheep.

Pregnant ewes were fed a depletion diet low in cobalt (0.06 ppm) for 3 1/2 months. Chronic catheters were implanted 8 weeks postpartum and 7 experiments were performed on these nonlactating vitamin B-12-depleted sheep (de-B12: 340 +/- 30 ng vitamin B-12 per gram wet liver) prior to repletion by intramuscular injection of hydroxocobalamin. Six experiments were then repeated after vitamin B-12 repletion (re-B12: 2220 +/- 50 ng vitamin B-12 per gram wet liver). The hepatic extraction ratios (HER) in continuously fed sheep were 0.81 and 0.77 for de-B12 and re-B12 corresponding to net hepatic uptakes of 460 +/- 50 and 440 +/- 40 mumol propionate per minute, respectively. Continuous infusion of unlabeled propionate into a mesenteric vein at 1 mmol/minute reduced the HER, yet this depression was greatest for re-B12 (0.74 vs. 0.63 for de-B12 and re-B12, respectively). Net hepatic uptake of propionate was increased (1145 +/- 100 vs. 985 +/- 95 mumol/minute, respectively), although vitamin B-12 status was without effect. It is concluded that the ability of liver to extract propionate is not affected at vitamin B-12 concentrations greater than 250 ng/g wet liver. However, when propionate entry rate was enhanced by intramesenteric infusion, the livers of de-B12 sheep had a greater capacity to remove propionate suggesting that alternate routes of metabolism may occur.

Animals↗
PubMed1983

Effects of avoparcin on ruminal propionate production and amino acid degradation in sheep fed high and low fiber diets.

The effect of the antibiotic, avoparcin, on ruminal propionate production and amino acid degradation was investigated by using four rumen fistulated wethers. Wethers were fed each of four diets during 28-day periods in a 4 X 4 Latin square design. Diets were high fiber (HF), high fiber plus 50 ppm avoparcin (HF-AVO), low fiber (LF) and low fiber plus 50 ppm avoparcin (LF-AVO). Total digestible energy intake (1.25 X maintenance) and crude protein intake for each animal were similar for all diets. Propionate production was determined each period from all animals by using a single injection of [1-14C]propionate and polyethylene glycol. Avoparcin decreased total volatile fatty acid concentration, increased ruminal pH and molar proportions of propionate on both HF and LF diets. Daily propionate production and pool turnover rate increased in sheep fed LF-AVO. Similar changes were noted in HF-AVO animals, although they were not statistically significant. Avoparcin decreased ruminal ammonia (50 vs. 45 mg/100 ml) and increased alpha amino nitrogen (8.6 vs. 14.3 mM) in LF-fed sheep. In vitro studies demonstrated a decreased degradation of crystalline amino acids by mixed populations of rumen microorganisms from sheep receiving LF diets containing avoparcin. Avoparcin appears to modify rumen fermentation by increasing propionate production and inhibiting protein or amino acid degradation.

Amino Acids↗
PubMed1999

Characterization of the anaerobic propionate-degrading syntrophs Smithella propionica gen. nov., sp. nov. and Syntrophobacter wolinii.

A strain of anaerobic, syntrophic, propionate-oxidizing bacteria, strain LYPT (= OCM 661T; T = type strain), was isolated and proposed as representative of a new genus and new species, Smithella propionica gen. nov., sp. nov. The strain was enriched from an anaerobic digestor and isolated. Initial isolation was as a monoxenic propionate-degrading co-culture containing Methanospirillum hungateii JF-1T as an H2- and formate-using partner. Later, an axenic culture was obtained by using crotonate as the catabolic substrate. The previously described propionate-degrading syntrophs of the genus Syntrophobacter also grow in co-culture with methanogens such as Methanospirillum hungateii, forming acetate, CO2 and methane from propionate. However, Smithella propionica differs by producing less methane and more acetate; in addition, it forms small amounts of butyrate. Smithella propionica and Syntrophobacter wolinii grew within similar ranges of pH, temperature and salinity, but they differed significantly in substrate ranges and catabolic products. Unlike Syntrophobacter wolinii, Smithella propionica grew axenically on crotonate, although very slowly. Co-cultures of Smithella propionica grew on propionate, and grew slowly on crotonate or butyrate. Syntrophobacter wolinii and Syntrophobacter pfennigii grow on propionate plus sulfate, whereas Smithella propionica did not. Comparisons of 16S rDNA genes indicated that Smithella propionica is most closely related to Syntrophus, and is more distantly related to Syntrophobacter.

Acetates↗
PubMed2005

Syntrophobacter sulfatireducens sp. nov., a novel syntrophic, propionate-oxidizing bacterium isolated from UASB reactors.

Two obligate anaerobes, TB8106(T) and WZH410, which degraded propionate in syntrophic association with methanogens, were isolated from two upflow anaerobic sludge blanket reactors, one treating brewery wastewater and the other bean curd wastewater. The strains were Gram-negative, non-spore-forming and non-motile. Cells were egg-shaped, with a size of 1.0-1.3 x 1.8-2.2 microm. Growth was observed at 20-48 degrees C and pH 6.2-8.8. Both strains converted propionate to acetate and methane in co-culture with methanogens, and grew on propionate plus sulfate in pure culture, with a doubling time of 52-55 h at 37 degrees C. Sulfate and thiosulfate both served as electron acceptors for propionate degradation. The DNA G + C contents of the two strains were 58.5 and 58.7 mol%, respectively. Phylogenetic analysis based on 16S rRNA gene sequences revealed that the strains were closely related to a propionate-oxidizing syntrophic bacterium, Syntrophobacter fumaroxidans DSM 10017(T) (94.7 % similarity). However, the novel strains could not ferment fumarate, and grew at a more alkaline pH range than Syntrophobacter fumaroxidans. Moreover, the novel isolates had obviously higher growth rates on propionate plus sulfate (0.12 day(-1)) than Syntrophobacter fumaroxidans DSM 10017(T) (0.024 day(-1)). Therefore, a novel species, Syntrophobacter sulfatireducens sp. nov., is proposed, with strain TB8106(T) (=AS 1.5016(T) = DSM 16706(T)) as the type strain.

Acetic Acid↗
PubMed2004

Cloning, expression, purification, crystallization and preliminary X-ray diffraction analysis of propionate kinase (TdcD) from Salmonella typhimurium.

In the cell, propionate is mainly formed during beta-oxidation of odd-numbered carbon-chain fatty acids, fermentation of carbohydrates and degradation of the amino acids threonine, valine, isoleucine and methionine. Recently, it has been shown that L-threonine is non-oxidatively cleaved to propionate via 2-ketobutyrate. The last step in this process, conversion of propionyl phosphate and ADP to propionate and ATP, is catalysed by propionate kinase (EC 2.7.1.-). Here, the cloning of propionate kinase (molecular weight 44 kDa) from Salmonella typhimurium with an N-terminal hexahistidine affinity tag and its overexpression in Escherichia coli are reported. Purified propionate kinase was found to cocrystallize with ADP in the hanging-drop vapour-diffusion and microbatch methods. Crystals belong to space group P3(1)21 or P3(2)21, with unit-cell parameters a = b = 111.47, c = 66.52 A. A complete data set to 2.2 A resolution has been collected using an image-plate detector system mounted on a rotating-anode X-ray generator.

Bacterial Proteins↗
PubMed2006

Evaluation and management of patients with propionic acidemia undergoing liver transplantation: a comprehensive review.

Propionic acidemia is a rare metabolic disorder that often results in episodic hyperammonemia, basal ganglia infarction, mental retardation, and cardiomyopathy. OLT has been used as a treatment for propionic acidemia, but its benefit in patients with this disease is unclear. The current study was undertaken to clarify the role of OLT in the management of this disease. The medical literature, a national registry of US OLT recipients, and a single institution liver transplant experience were reviewed for cases of OLT for propionic acidemia. Accumulated cases demonstrate that OLT has resulted in clear evidence of clinical improvement in several patients, often obviating the need for dietary restriction or other forms of medical management. OLT appears to halt the decline in neurocognitive function often associated with propionic acidemia. In total, 12 patients with propionic acidemia have undergone a total of 14 OLTs. A quantitative analysis of outcomes shows an overall patient survival rate of 72.2% at one year after OLT. In conclusion, OLT should be considered a treatment option for patients with propionic acidemia who continue to experience episodes of hyperammonemia in spite of maximal medical therapy. Early OLT may limit the development of mental retardation and/or cardiomyopathy.

Adult↗
PubMed1988

The roles of hepatic nerves in the reduction of food intake as a consequence of intraportal sodium propionate administration in sheep.

Like other mammals, ruminants can also control their food intake. One of the potential sites where feed-back signals arise is the liver. We have already shown in our previous study that intraportal administration of propionate depresses intake and that this action is dependent on an intact nerve supply to the liver because sectioning the hepatic plexus abolishes the effect of propionate. In the following work the relative importance and the possible roles of different hepatic nerves were investigated. Three-hour continuous infusions of sodium propionate or saline into the hepatic portal vein of sheep were carried out following applications of different surgical procedures and food intakes were measured. In experiment 1 bilateral splanchnotomies were sufficient to prevent the effect of propionate on food intake. The subsequent total hepatic denervations also resulted in the removal of the depressing effect of propionate. Experiment 2 was designed to compare the involvement of the splanchnic afferents from the liver and the hepatic vagal afferents. Intraportal propionate was demonstrated to depress feeding in the intact animal whereas splanchnic nerve blockade with local anaesthetic removed this effect. Paradoxically selective hepatic vagotomy also abolished this effect. It was concluded that there may be more than one pathway involved in transmitting information from the liver to the central nervous system. Possible implications of the results are discussed and attempts are made to explain the mechanism of action and compare different theories by other workers.

Animals↗
PubMed1993

Effects of hydrogen and formate on the degradation of propionate and butyrate in thermophilic granules from an upflow anaerobic sludge blanket reactor.

Degradation of propionate and butyrate in whole and disintegrated granules from a thermophilic (55 degrees C) upflow anaerobic sludge blanket reactor fed with acetate, propionate, and butyrate as substrates was examined. The propionate and butyrate degradation rates in whole granules were 1.16 and 4.0 mumol/min/g of volatile solids, respectively, and the rates decreased 35 and 25%, respectively, after disintegration of the granules. The effect of adding different hydrogen-oxidizing bacteria (both sulfate reducers and methanogens), some of which used formate in addition to hydrogen, to disintegrated granules was tested. Addition of either Methanobacterium thermoautotrophicum delta H, a hydrogen-utilizing methanogen that does not use formate, or Methanobacterium sp. strain CB12, a hydrogen- and formate-utilizing methanogen, to disintegrated granules increased the degradation rate of both propionate and butyrate. Furthermore, addition of a thermophilic sulfate-reducing bacterium (a Desulfotomaculum sp. isolated in our laboratory) to disintegrated granules improved the degradation of both substrates even more than the addition of methanogens. By monitoring the hydrogen partial pressure in the cultures, a correlation between the hydrogen partial pressure and the degradation rate of propionate and butyrate was observed, showing a decrease in the degradation rate with increased hydrogen partial pressure. No significant differences in the stimulation of the degradation rates were observed when the disintegrated granules were supplied with methanogens that utilized hydrogen only or hydrogen and formate. This indicated that interspecies formate transfer was not important for stimulation of propionate and butyrate degradation.

Anaerobiosis↗
PubMed2000

Cultivation and in situ detection of a thermophilic bacterium capable of oxidizing propionate in syntrophic association with hydrogenotrophic methanogens in a thermophilic methanogenic granular sludge.

The thermophilic, anaerobic, propionate-oxidizing bacterial populations present in the methanogenic granular sludge in a thermophilic (55 degrees C) upflow anaerobic sludge blanket reactor were studied by cultivation and in situ hybridization analysis. For isolation of propionate-degrading microbes, primary enrichment was made with propionate as the sole energy source at 55 degrees C. After several attempts to purify the microbes, a thermophilic, syntrophic, propionate-oxidizing bacterium, designated strain SI, was isolated in both pure culture and coculture with Methanobacterium thermoautotrophicum. Under thermophilic (55 degrees C) conditions, strain SI oxidized propionate, ethanol, and lactate in coculture with M. thermoautotrophicum. In pure culture, the isolate was found to ferment pyruvate. 16S ribosomal DNA sequence analysis revealed that the strain was relatively close to members of the genus Desulfotomaculum, but it was only distantly related to any known species. To elucidate the abundance and spatial distribution of organisms of the strain SI type within the sludge granules, a 16S rRNA-targeted oligonucleotide probe specific for strain SI was developed and applied to thin sections of the granules. Fluorescence in situ hybridization combined with confocal laser scanning microscopy revealed that a number of rod-shaped cells were present in the middle and inner layers of the thermophilic granule sections and that they formed close associations with hydrogenotrophic methanogens. They accounted for approximately 1.1% of the total cells in the sludge. These results demonstrated that strain SI was one of the significant populations in the granular sludge and that it was responsible for propionate oxidation in the methanogenic granular sludge in the reactor.

Bacteria↗