Biomedical subjects
Publications and source records attributed to R A Mah.
The effects of fermentation products and media supplements on a colorimetric assay for formate were evaluated. Formate was detected at concentrations as low as 0.5 mM in fermentation media.
A new anaerobic, mesophilic, spore-forming cellulolytic bacterium is described. Cellulose is cleared within 24 to 48 h around colonies formed in cellulose agar roll tubes. Cells stain gram negative and are nonmotile rods which form oblong spores either centrally or subterminally in a clostridial swelling. Colonies are irregular with an opaque edge and a center devoid of both vegetative cells and spores. Cellulose, xylan, pectin, cellobiose, glucose, maltose, galactose, sucrose, lactose, and mannose serve as substrates for growth. H(2), CO(2), acetate, butyrate, formate, and lactate are produced during fermentation of cellulose or cellobiose. The temperature and pH for optimum growth are 37 degrees C and 7.0, respectively. The DNA composition is 26 to 27 mol% guanine plus cytosine. This bacterium resembles "Clostridium lochheadii" in morphological and some biochemical characteristics but is not identical to it. The name Clostridium cellulovorans sp. nov. is proposed. The type strain is 743B (ATCC 35296).
A thermophilic methanogen was isolated from enrichment cultures originally inoculated with sludge from an anaerobic kelp digester (55 degrees C). This isolate exhibited a temperature optimum of 55 to 60 degrees C and a maximum near 70 degrees C. Growth occurred throughout the pH range of 5.5 to 9.0, with optimal growth near pH 7.2. Although 4% salt was present in the isolation medium, salt was not required for optimal growth. The thermophile utilized formate or H(2)-CO(2) but not acetate, methanol, or methylamines for growth and methanogenesis. Growth in complex medium was very rapid, and a minimum doubling time of 1.8 h was recorded in media supplemented with rumen fluid. Growth in defined media required the addition of acetate and an unknown factor(s) from digester supernatant, rumen fluid, or Trypticase. Cells in liquid culture were oval to coccoid, 0.7 to 1.8 mum in diameter, often occurring in pairs. The cells were easily lysed upon exposure to oxygen or 0.08 mg of sodium dodecyl sulfate per ml. The isolate was sensitive to tetracycline and chloramphenicol but not penicillin G or cycloserine. The DNA base composition was 59.69 mol% guanine plus cytosine.
Growth of Methanosarcina sp. strain 227 and Methanosarcina mazei on H(2)-CO(2) and mixtures of H(2)-CO(2) and acetate or methanol was examined. The growth yield of strain 227 on H(2)-CO(2) in complex medium was 8.4 mg/mmol of methane produced. Growth in defined medium was characteristically slower, and cell yields were proportionately lower. Labeling studies confirmed that CO(2) was rapidly reduced to CH(4) in the presence of H(2), and little acetate was used for methanogenesis until H(2) was exhausted. This resulted in a biphasic pattern of growth similar to that reported for strain 227 grown on methanol-acetate mixtures. Biphasic growth was not observed in cultures on mixtures of H(2)-CO(2) and methanol, and less methanol oxidation occurred in the presence of H(2). In M. mazei the aceticlastic reaction was also inhibited by the added H(2), but since the cultures did not immediately metabolize H(2), the duration of the inhibition was much longer.
Explore the source record for details and available documents.
A method is presented for determining methyl mercury that combines thin layer chromatography and atomic absorption spectrophotometry. The procedure is based on a previously reported method and entails isolating methyl mercury on a thin layer chromatogram, scraping adsorbent into an ignition tube, heating the tube, and moving the mercury vapor under vacuum into an absorption cell where it absorbs light at 253.7 nm. The procedure is sensitive to 0.7 ng Hg. Methyl mercury is extracted from biological samples by distilling, and then extracting with dithizone-chloroform. The extract is concentrated and applied to a thin layer chromatographic plate. The overall sensitivity of the method varies with sample size. For 100 g samples, the method is sensitive to about 8 ng Hg (as methyl mercury)/g (9 ppb).
Methanosarcina strain 227 grew rapidly and produced methane on a mineral medium containing acetate as the sole added organic substrate. Cell yields but not doubling times were affected by the presence or absence of yeast extract. Greater cell yields occurred in yeast extract medium than in mineral medium. Radioactive labeling studies showed that acetate was decarboxylated in mineral medium, as was shown previously in complex medium. The specific radioactivity of methane produced per specific acitvity of acetate added was not significantly different in yeast extract medium compared with mineral medium. Unequivocal evidence indicates that the cleavage of acetate to methane and carbon dioxide provided the energy for growth in the presence or absence of other organic compounds; these latter compounds do not serve as energy sources, electron donors, or significant sources of methane during this aceticlastic reaction.
A thermophilic strain of Methanosarcina, designated Methanosarcina strain TM-1, was isolated from a laboratory-scale 55 degrees C anaerobic sludge digestor by the Hungate roll-tube technique. Penicillin and d-cycloserine, inhibitors of peptidoglycan synthesis, were used as selective agents to eliminate contaminating non-methanogens. Methanosarcina strain TM-1 had a temperature optimum for methanogenesis near 50 degrees C and grew at 55 degrees C but not at 60 degrees C. Substrates used for methanogenesis and growth by Methanosarcina strain TM-1 were acetate (12-h doubling time), methanol (7- to 10-h doubling time), methanol-acetate mixtures (5-h doubling time), methylamine, and trimethylamine. When radioactively labeled acetate was the sole methanogenic substrate added to the growth medium, it was predominantly split to methane and carbon dioxide. When methanol was also present in the medium, the metabolism of acetate shifted to its oxidation and incorporation into cell material. Electrons derived from acetate oxidation apparently were used to reduce methanol. H(2)-CO(2) was not used for growth and methanogenesis by Methanosarcina strain TM-1. When presented with both H(2)-CO(2) and methanol, Methanosarcina strain TM-1 was capable of limited hydrogen metabolism during growth on methanol, but hydrogen metabolism ceased once the methanol was depleted. Methanosarcina strain TM-1 required a growth factor (or growth factors) present in the supernatant of anaerobic digestor sludge. Growth factor requirements and the inability to use H(2)-CO(2) are characteristics not found in other described Methanosarcina strains. The high numbers of Methanosarcina-like clumps in sludges from thermophilic digestors and the fast generation times reported here for Methanosarcina TM-1 indicate that Methanosarcina may play an important role in thermophilic methanogenesis.
An acetate-fermenting strain of Methanosarcina was isolated from an acetate enrichment culture inoculated with anaerobic sludge from a waste treatment digestor. In pure culture, this organism fermented acetate in the absence of added hydrogen at rates comparable in magnitude to those found in digestor systems. This rate was significantly higher than previously obtained for pure cultures of this genus. Mineral components of yeast extract were highly stimulatory for cultures growing on methanol. Comparable stimulation was not observed for cultures growing on acetate. Labeling studies indicated that acetate was converted to methane and CO2 as predicted by previous studies on mixed cultures. Total oxidation or reduction of acetate was not the mechanism of conversion of acetate to methane by the pure culture. The ability of this strain to form colonies or to produce methane from acetate was apparently influenced by the choice of substrate and conditions used for growing the inoculum.
A methanogenic acetate enrichment was initiated by inoculation of an acetate-mineral salts medium with domestic anaerobic digestor sludge and maintained by weekly transfer for 2 years. The enrichment culture contained a Methanosarcina and several obligately anaerobic nonmethanogenic bacteria. These latter organisms formed varying degrees of association with the Methanosarcina, ranging from the nutritionally fastidious gram-negative rod called the satellite bacterium to the nutritionally nonfastidious Eubacterium limosum. The satellite bacterium had growth requirements for amino acids, a peptide, a purine base, vitamin B12, and other B vitamins. Glucose, mannitol, starch, pyruvate, cysteine, lysine, leucine, isoleucine, arginine, and asparagine stimulated growth and hydrogen production. Acetate was neither incorporated nor metabolized by the satellite organism. Since acetate was the sole organic carbon source in the enrichment culture, organism(s) which metabolize acetate (such as the Methanosarcina) must produce substrates and growth factors for associated organisms which do not metabolize acetate.
An acetate enrichment culture was initiated by inoculating anaerobic sludge from a mesophilic methane digestor into a mineral salts medium with calcium acetate as the sole carbon and energy source. This enrichment was maintained indefinitely by weekly transfer into medium of the same composition. A study of this enrichment disclosed an unexpected age-dependent inhibition of methanogenesis by H2 and formate which apparently differed from the inhibition by chloroform and benzyl viologen. This age-dependent inhibition indicated that microbial interactions of the mixed enrichment population may play a regulatory role in methane formation. Futhermore, stimulation of methanogenesis in the acetate enrichment by addition of yeast extract showed a nutrient limitation which indicated that syntrophic interactions leading to formation of growth factors may also occur. A model is presented to illustrate the possible interrelationships between methanogenic and nonmethanogenic bacteria in their growth and formation of methane and carbon dioxide from acetate.
Methanosarcina strain 227 exhibited exponential growth on sodium acetate in the absence of added H(2). Under these conditions, rates of methanogenesis were limited by concentrations of acetate below 0.05 M. One mole of methane was formed per mole of acetate consumed. Additional evidence from radioactive labeling studies indicated that sufficient energy for growth was obtained by the decarboxylation of acetate. Diauxic growth and sequential methanogenesis from methanol followed by acetate occurred in the presence of mixtures of methanol and acetate. Detailed studies showed that methanol-grown cells did not metabolize acetate in the presence of methanol, although acetate-grown cells did metabolize methanol and acetate simultaneously before shifting to methanol. Acetate catabolism appeared to be regulated in response to the presence of better metabolizable substrates such as methanol or H(2)-CO(2) by a mechanism resembling catabolite repression. Inhibition of methanogenesis from acetate by 2-bromoethanesulfonate, an analog of coenzyme M, was reversed by addition of coenzyme M. Labeling studies also showed that methanol may lie on the acetate pathway. These results suggested that methanogenesis from acetate, methanol, and H(2)-CO(2) may have some steps in common, as originally proposed by Barker. Studies with various inhibitors, together with molar growth yield data, suggest a role for electron transport mechanisms in energy metabolism during methanogenesis from methanol, acetate, and H(2)-CO(2).
Stream and lake water from the Mammoth Lakes region of California was sampled for Yersinia enterocolitica. From 10 of the 34 sites examined, organisms were isolated that were biochemically identified as Y. enterocolitica. Only one of the ten strains could be serologically confirmed. This strain was identified as Y. enterocolitica serotype 16. Although an outbreak of enteritis in the area prompted this study, no correlation with gastrointestinal disease could be established since the majority of the strains were untypeable.
The metabolism of glucose during enterotoxin B synthesis in Staphylococcus aureus S-6 was examined under anaerobic conditions in the presence and absence of nitrate. The repression of enterotoxin synthesis which occurs during the oxidative metabolism of glucose was relieved after a shift to anaerobic conditions; glucose was then converted primarily to lactic acid and was metabolized more rapidly, presumably to obtain the equivalent amount of energy available aerobically. A greater proportion of oxidized end products and evidently more energy per glucose molecule was produced in the presence of oxygen. Thus, available energy as judged by a change in the type and proportion of end products appears to be related to the degree of toxin repression. As expected, the addition of nitrate during anaerobic glucose metabolism prevented derepression of toxin synthesis.
Explore the source record for details and available documents.
Explore the source record for details and available documents.
Several factors influenced the formation of enterotoxin B by Staphylococcus aureus strain S-6. In the standard casein hydrolysate medium, toxin was not produced in detectable quantities during exponential growth; it was produced during the post-exponential phase when total protein synthesis was arithmetic. The rate of toxin synthesis was much greater than the rate of total protein synthesis. The appearance of enterotoxin was inhibited by chloramphenicol; thus, the presence of toxin was dependent on de novo protein synthesis. When low concentrations of glucose (<0.30%) were added to the casein hydrolysate medium, growth was diauxic; glucose was completely metabolized during the first growth period. During the second growth period, enterotoxin was synthesized. In unbuffered casein hydrolysate medium containing excess glucose, toxin synthesis was completely repressed. The absence of toxin production under such conditions might be explained by the low (4.6) pH resulting from the acid end products of glucose metabolism. At pH <5.0, little or no toxin was produced. Toxin synthesis was initiated in the presence of glucose when the medium were buffered at any pH above 5.6. In such media, the differential rates of toxin synthesis, with respect to the rates of total protein synthesis, were lower than the differential rates in casein hydrolysate medium alone. Addition of glucose to a culture synthesizing toxin resulted in an immediate decrease in the differential rate without any change in pH. Thus, toxin synthesis appeared to be regulated by catabolite repression.