Effects of carbohydrate pulses and pH on population shifts within oral microbial communities in vitro.

A mixed culture chemostat system was used to distinguish between the effects of carbohydrate availability per se and the low pH generated from carbohydrate metabolism on the proportions of bacteria within microbial communities. Nine oral bacteria were grown at pH 7 and pulsed with glucose on ten consecutive days. In one chemostat, the pH was maintained automatically at 7 throughout the experimental period, while in the other, pH control was discontinued for six hours after each pulse. Glucose pulses at neutral pH had little effect on the composition of the microflora. Only the proportions of A. viscosus and V. dispar increased; L. casei and S. mutans remained at low levels (0.2% and 1.0%, respectively). Acetate and propionate were low. In contrast, when pH was allowed to fall after each glucose pulse, the composition of the microflora altered dramatically. The amounts of L. casei and S. mutans increased both as a proportion of the total count and in absolute numbers, as did V. dispar, whereas the amounts of the other Gram-negative organisms (B. intermedius, F. nucleatum, and N. subflava) and S. sanguis were considerably reduced. Lactate formed a major portion of the metabolic end-products. Successive glucose pulses resulted in both amplified changes in the microflora and a steadily greater rate and final extent of acid production. This is in agreement with the reported shifts in the oral microflora in vivo in response to frequent carbohydrate intake. Analysis of the data strongly suggests that the pH generated from carbohydrate metabolism, rather than carbohydrate availability per se, is responsible for the widely reported shifts in composition and metabolism of the oral microflora in vivo.

Factors affecting the in vitro production of volatile fatty acids by mixed bacterial populations from the bovine rumen.

Factors affecting in vitro ruminal bacterial VFA production were examined. Treatments consisted of high and low initial pH (6.7, 5.7), osmolality (600, 400 mOsm) and concentrations of acetic (40, 0 mM) and propionic acids (20, 0 mM). Response variables measured included the production of acetic, propionic and total VFA, total gas and methane. Initial pH affected (P less than .05) most variables either independently or in combination with one or more of the other factors. Acetic acid production was reduced 40% (P = .03) when initial acetic acid concentrations were 40 mM compared with 0 mM. Also, acetic acid production was less (P less than .01) at low initial pH (5.7) than at high initial pH (6.7). Propionic acid production was greater (P = .05) at high vs low initial acetic acid concentrations. Propionic acid production was greater in response to low vs high initial osmolality, although the magnitude of this difference depended on initial pH (interaction P = .02). Total production of VFA was greater (P less than .01) at high than at low initial pH; however, at low initial pH, no difference (P greater than .05) was observed due to initial osmolality, whereas at high pH, production was greater (interaction P = .04) for low than for high initial osmolality. The diminished production of total VFA at pH 5.7 occurred primarily due to reduced acetic acid production, although increased production of propionic and butyric acids was noted.(ABSTRACT TRUNCATED AT 250 WORDS)

Short-chain fatty acid production from mono- and disaccharides in a fecal incubation system: implications for colonic fermentation of dietary fiber in humans.

An in vitro fecal incubation system was used to demonstrate how lactose, lactulose and monosaccharides (mainly constituents of dietary fiber) influence short-chain fatty acid production in colon. Short-chain fatty acids were formed from all mono- and disaccharides tested (except L-glucose): D-glucose, D-galactose, D-fructose, D-mannose, L-rhamnose, D-sorbitol, D-arabinose, D-xylose, D-ribose, D-galacturonate, D-glucuronate, lactose and lactulose. All saccharides increased acetate formation; propionate production was increased from rhamnose, arabinose, xylose, ribose, galacturonic and glucuronic acid, whereas the synthesis of butyrate was elevated in assays incubated with sorbitol, galacturonic and glucuronic acid, and to a lesser degree ribose. Isobutyrate, valerate, isovalerate and hexanoate were produced in increased amounts in assays incubated with albumin, but in fact decreased in many incubations with saccharides. It is speculated that saccharide fermentation always results in formation of acetate, and that the relative production of acetate, propionate and butyrate is related to the monosaccharide composition of dietary fiber available for colonic bacteria. However, the production of isobutyrate, valerate, isovalerate and hexanoate is probably not due to saccharide fermentation, but is rather of polypeptide origin.

Forced synthesis of trace amounts of juvenile hormone II from propionate by corpora allata of a juvenile hormone III-producing insect.

Corpora allata of the cockroach Diploptera punctata normally synthesize only the isoprenoid juvenile hormone III (JH III). Only under extreme in vitro conditions (absence of carbon sources other than propionate) do they produce trace amounts of the homoisoprenoid JH II in addition to JH III. The specificity of the in vitro synthesis of JH III by D. punctata is thus consistent with the observed lack of homoisoprenoid JHs in this insect.

[Search for propionic acid bacteria in the human intestine]. / Poisk propionovokislykh bakterii v kishechnike cheloveka.

The possibility of detecting propionic acid-producing bacteria in the intestine of healthy humans with a view to obtaining a strain which is physiologically most suitable for therapeutic purposes has been studied. The selective conditions for the isolation of propionic acid-producing bacteria from the large intestine have been experimentally established. Analysis of 70 puncture biopsy specimens of parietal mucus and luminal contents from different sections of the intestine has not shown the presence of the representatives of propionic acid-producing bacteria. The strains isolated under the conditions selective for such bacteria have been found to belong to the family Enterobacteriaceae. These strains have proved capable of synthesizing vitamin B12, but the synthesis of propionic acid has not been observed.

Biochemical changes during the aerobic-anaerobic transition in Ascaris suum larvae.

Ascaris suum L3 larvae isolated from rabbit lungs undergo the third ecdysis to L4 larvae after 3 days in culture under a gas phase of 85% N2/10% CO2/5% O2. The L3 larvae contain substantial malic enzyme activity and are capable of producing small amounts of the reduced organic acids characteristic of the fermentative pathways which operate in the adult. However, only a small portion of the total carbon utilized is accounted for by these reduced acids and their motility is cyanide-sensitive, suggesting that their energy-generating pathways are predominantly aerobic. In contrast, after ecdysis, the L4 larvae begin to utilize glucose at a greater rate and the proportion of total carbon utilized which is accounted for as propionate, 2-methylbutyrate and 2-methylvalerate also increases. In addition, motility becomes increasingly cyanide-insensitive, suggesting that these L4 larvae are able to utilize the anaerobic energy-generating pathways of the adult. Surprisingly, on day 10 in culture, these L4 larvae, although capable of producing reduced volatile acids, still retain substantial cyanide-sensitive cytochrome oxidase activity.

Characterization of Flavobacterium species by analysis of volatile fatty acid production.

Seventy-four Flavobacterium strains were characterized by gas-liquid chromatographic analysis of volatile fatty acids produced in the culture medium. Principal components analysis permitted the graphic representation of the relative positions of the different strains, and aggregation according to the variance enabled a hierarchical classification to be established. The study revealed three subgroups each for F. meningosepticum and F. odoratum. Our F. breve, Flavobacterium sp. group IIb and F. multivorum strains appeared to be homogeneous. These results tallied with those of previous studies on DNA base composition and reassociation, electrophoretic protein profiles and cellular fatty acid composition.

Cytotoxic activity of Propionibacterium acnes and other skin organisms.

Culture supernatants from four species of skin micro-organisms were tested against VERO (monkey kidney cells) and skin fibroblasts for cytotoxic activity. Cytotoxic activity was produced by the three species of Propionibacterium tested (P. acnes, P. avidum and P. granulosum), and this activity was highest when cultures were grown in the presence of glucose. In contrast, Staphylococcus epidermidis was devoid of cytotoxic activity whether grown in the presence of glucose or not. The agent responsible for the cytotoxic activity was heat stable, of low molecular weight and removable from supernatants by ether extraction. These properties, coupled with the finding that the levels of cytotoxicity are directly proportional to the concentrations of propionate measured in samples, suggests that propionate is the agent responsible for the cytotoxicity of the culture supernatants. Pure propionate and the salts of other carboxylic acids (CI to C5) were tested at the same concentrations and showed different degrees of cytotoxicity depending on their chain length. Propionate may have an important role in the aetiology of the disease acne vulgaris.

Isolation, characterization and ecological distribution of recently recognized propionic acid-negative Bacteroides species from human sources.

140 strains of anaerobic, Gram-negative, propionate-negative, non-pigmented rods (Bacteroides oralis group) were isolated from normal flora and from clinical specimens. They were characterized by physiological tests, gas chromatography of the acid end products, and determination of the G + C content of their DNA. In addition, their distribution in the various biotopes of the human body (oropharyngeal tract, genitourinary tract, and gut) and in clinical specimens was investigated. The strains could be assigned to the recently described species Bacteroides denticola, B. buccae, B. buccalis, and B. oris, or to B. bivius, B. disiens and B. oralis. In the investigated group, especially B. bivius exhibits an antibiotic resistance--particularly against the beta-lactam antibiotics--comparable to that of B. fragilis.

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.

Organic acids and anaerobic microorganisms in the contents of the cholesteatoma sac.

Organic acids in the contents of the cholesteatoma sac from 28 cases were studied by gas chromatographic technique. Five volatile fatty acids (acetate, propionate, isobutyrate, butyrate and isovalerate) and lactate were detected in large amounts, which may lower the pH of the cholesteatoma content. These acids were considered to be derived from products of anaerobic microorganisms. Therefore, the contents from 12 cases were cultured anaerobically in a glove box. Obligate microorganisms were identified in 92% of the cases and Peptococcus, Bacteroides, and Clostridium species were frequently isolated. In vitro, such obligate anaerobes produced various organic acids from the cholesteatoma content. Facultatives such as Staphylococcus aureus and Proteus mirabilis produced acetate in the content under aerobic and anaerobic conditions, whereas no organic acid was produced by Pseudomonas aeruginosa. Organic acids in the cholesteatoma content could be fermentative products made by the microorganisms, anaerobes and facultatives, which use the content as a substrate for acid production.

Presence of diaminopimelic acid in propionate-negative Bacteroides species and in some butyric acid-producing strains.

AThe presence of diaminopimelic acid (m-DAP) in strains of Bacteroides melaninogenicus, B. bivius and other species as well as in unidentified strains of Bacteroides was investigated by thin-layer chromatography. Strains of B. bivius and B. disiens all contained m-DAP as did the subspecies intermedius and melaninogenicus of B. melaninogenicus. Strains of B. asaccharolyticus and similar black pigment-producing butyrate-positive isolates showed heterogeneity. Asaccharolytic strains were DAP negative, whereas two strains fermenting glucose were positive. Some of the non-pigmented propionate-negative and butyrate-negative unidentified strains also contained DAP. The consistent finding of m-DAP in strains of B. bivius, B. disiens, and B. melaninogenicus indicates that DAP detection might be of value in the identification of these species.

Influence of heme and vitamin B12 on growth and fermentations of Bacteroides species.

We examined the effects of heme on the growth and fermentations of Bacteroides species. Bacteroides fragilis ATCC 25285 required heme for growth and produced malate and lactate as major products of glucose fermentation when the concentration of heme was 1 ng/ml. With 1 microgram of heme per ml, malate was not formed, lactate production decreased, and succinate and acetate were the major fermentation products. B. eggerthii ATCC 27754 grew without heme, with the production of mainly malate and lactate from glucose. Its fermentation with 1 microgram of heme per ml was similar to that of B. fragilis grown with the same concentration of heme. B. splanchicus VPI 6842 grew without heme, with the production of mainly malate, acetate, and H2 from glucose. With 1 microgram of heme per ml, malate disappeared, H2 decreased significantly, and succinate, acetate, and butyrate were the major products. The addition of vitamin B12 to media containing 1 microgram of heme per ml caused all species to produce propionate at the expense of succinate and, with B. splanchnicus, also at the expense of butyrate. Thus, the concentration of heme and the presence of vitamin B12 significantly influenced the course of glucose fermentation by these bacteria.

Spontaneous variability of Streptomyces glomeratus, a producer of the amthracycline antibiotics beromycins.

Spontaneous variants of the beromycin-producing strain Streptomyces glomeratus 3980 were divided into five groups (A-E) according to increasing antibiotic activity. The most active variants (group E) differed from the other types and the wild strain by a suppressed ability to produce aerial mycelium and melanoid piogment and by an increased production of propionic acid. Strains with a 12-fold higher antibiotic production capacity (with respect to strain 3980) were obtained by selection of superior segregants from submerged cultures of the E type.

[Changes in the physiologic properties of a chemostat culture of Propionibacterium shermanii during growth limitation by a deficiency of sodium lactate]. / Izmenenie fiziologicheskikh svoistv khemostatnoi kul'tury Propionibacterium shermanii pri limitatsii rosta nedostatkom laktata natriia.

A chemostat curve has been constructed for changes in the Propionibacterium shermanii population density with an increase in the flow rate of the medium from 0.04 to 0.55 h-1 upon limitation of the growth with sodium lactate deficiency. The curve differs from the classic chemostat curve described with the Monod equation, thus indicating changes in metabolism at different growth rates. Fermentation of lactate to acetic and propionic acids by the culture is most complete at low flow rates of the medium (D = 0.04 h-1). The rates of oxidation processes increase at average flow rates (D = 0.1--0.33 h-1). Lactate is both fermented to yield volatile acids, and oxidized. The growth rate D = mu = 0.55 h-1 approaches the maximal one. The rate of fermentation by the culture at this growth rate decreases while the rate of oxidation increases. The ratio between the rates of these two processes shifts towards oxidation. The cells display now a very high synthetic activity: constructive metabolism of the cells is realized practically only at the account of the nitrogen from ammonium rather than the nitrogen of amino acids. The chemical composition of the cells along the chemostat curve is relatively constant. The content of protein and DNA does not change; the content of RNA slightly rises with an increase in the growth rate, and the protein synthesizing activity of RNA is directly proportional to D = mu.

[Comparison of the metabolism of isolated streptococci and dental plaque]. / Der Stoffwechsel isolierter Streptokokken und Plaque im Verleich.

1. Isolated cultures of Streptococcus mutans form lactic acid as well as small amounts of formic acid and acetic acid during the anaerobic break down of glucose. The composition of the acids in terms of percent is independent of time. 2. Plaque specimens incubated in vitro show a complicated spectrum of substances; propionic and butyric acids are found in addition to lactic, formic, and acetic acids. The composition in terms of percent depends on the incubation period. The percentage of lactic acid drops as the incubation period is extended. 3. Just as the plaque, the product spectrum of isolated streptococci in terms of quantitative composition depends on the buffer capacity.