Biomedical subjects

James B Russell

Publications and source records attributed to James B Russell.

At least 19 recordsLinked to original sources

PubMed2007

Effect of acidic pH on the ability of Clostridium sporogenes MD1 to take up and retain intracellular potassium.

At pH values >5.5, Clostridium sporogenes MD1 accumulated potassium even though it had little protonmotive force, but an ATPase inhibitor (N, N'- dicyclohexylcarbodiimide) prevented this uptake. The results suggested that potassium transport was ATP-driven, and a protonophore (3, 3', 4', 5 - tetrachlorosalicylanilide) did not eliminate uptake. However, potassium uptake could also be driven by an artificial pH gradient, and in this case the protonophore acted as an inhibitor. These latter results indicated that the cells also had a protonmotive force-driven transporter. When the pH <5.1, the cells could not retain potassium, rapid efflux was observed, and intracellular volume collapsed.

Adenosine Triphosphatases↗

PubMed2006

The effect of calcium and magnesium on the activity of bovicin HC5 and nisin.

Some Gram-positive bacteria produce small peptides (bacteriocins) that have antimicrobial activity, but many bacteria can become bacteriocin resistant. Bovicin HC5, a lantibiotic produced by Streptococcus bovis HC5, has the ability to inhibit nisin-resistant bacteria. Because nisin resistance has in many cases been correlated with an alteration of lipoteichoic acids or the polar head groups of membrane phospholipids, we decided to examine the effect of divalent cations on nisin and bovicin HC5 activity. Both bacteriocins catalyzed potassium efflux from S. bovis JB1, a non-bacteriocin-producing strain. The addition of large amounts (100 mM) of calcium or magnesium increased the ability of S. bovis JB1 to bind Congo red (an anionic dye) and counteracted bacteriocin-mediated potassium loss. Calcium was more effective than magnesium in decreasing nisin activity, but the reverse was observed with bovicin HC5. Nisin-resistant S. bovis JB1 cells bound three times as much Congo red as nisin-sensitive cells, and this result is consistent with the idea that changes in cell surface charge can be a mechanism of bacteriocin resistance. The nisin-resistant cells were less susceptible to bovicin HC5, but bovicin HC5 still caused a 50% depletion of intracellular potassium. These results indicate that nisin and bovicin HC5 react differently with the cell surfaces of Gram-positive bacteria.

Bacteriocins↗

PubMed2006

Fermentation acids inhibit amino acid deamination by Clostridium sporogenes MD1 via a mechanism involving a decline in intracellular glutamate rather than protonmotive force.

Fermentation acids inhibited the growth and ammonia production of the amino-acid-fermenting bacterium Clostridium sporogenes MD1, but only when the pH was acidic. Such inhibition was traditionally explained by the ability of fermentation acids to act as uncouplers and decrease protonmotive force (Deltap), but C. sporogenes MD1 grows even if the Deltap is very low. Cell suspensions incubated with additional sodium chloride produced ammonia as rapidly at pH 5.0 as at pH 7.0, but cells incubated with additional sodium lactate were sensitive to even small decreases in extracellular pH. Similar results were obtained if the sodium lactate was replaced by sodium acetate or propionate. When extracellular pH declined, DeltapH increased even if sodium lactate was present. The cells accumulated intracellular lactate anion when the pH was acidic, and intracellular glutamate declined. Because amino acid deamination is linked to a transamination reaction involving glutamate dehydrogenase, the decrease in ammonia production could be explained by the decrease in intracellular glutamate. This latter hypothesis was consistent with the observation that extracellular glutamate addition restored amino acid deamination even though glutamate alone did not allow for the generation of ammonia.

Adenosine Triphosphate↗

PubMed2006

Factors affecting lysine degradation by ruminal fusobacteria.

Fusobacterium necrophorum can readily be enriched from the rumen with lysine, and its deamination rate is very rapid. The addition of F. necrophorum JB2 to mixed ruminal bacteria significantly increased lysine degradation, but only if the ratio of ruminal fluid to basal medium was less than 25%. If more ruminal fluid (pH 6.1) was added, ammonia production decreased by as much as 80%. Clarified, autoclaved ruminal fluid was also inhibitory. When F. necrophorum JB2 was grown in a lysine-limited continuous culture (0.1 h(-1) dilution rate) and pH was decreased using HCl, optical density decreased linearly, and the culture washed out at pH 5.6. Batch cultures of F. necrophorum JB2 deaminated as much lysine at pH 6.1 as at pH 6.6, but only if fermentation acids were not present. Sodium acetate (100 mM) had little effect at pH 6.6, but the same concentration inhibited ammonia production by 80% at pH 6.1. The idea that fermentation acids could prevent the enrichment of fusobacteria in vivo was supported by the observation that dietary lysine supplementation did not enhance the lysine deamination rate of the mixed ruminal bacteria.

Ammonia↗

PubMed2006

Bacterial competition between a bacteriocin-producing and a bacteriocin-negative strain of Streptococcus bovis in batch and continuous culture.

A bacteriocin-producing Streptococcus bovis strain (HC5) outcompeted a sensitive strain (JB1) before it reached stationary phase (pH 6.4), even though it grew 10% slower and cell-free bovicin HC5 could not yet be detected. The success of bacteriocin-negative S. bovis isolates was enhanced by the presence of another sensitive bacterium (Clostridium sticklandii SR). PCR based on repetitive DNA sequences indicated that S. bovis HC5 was not simply transferring bacteriocin genes to S. bovis JB1. When the two S. bovis strains were coinoculated into minimal medium, bacteriocin-negative isolates predominated, and this effect could be explained by the longer lag time (0.5 vs. 1.5 h) of S. bovis HC5. If the glucose concentration of the minimal medium was increased from 2 to 7 mg mL(-1), the effect of lag time was diminished and bacteriocin-producing isolates once again dominated the coculture. When the competition was examined in continuous culture, it became apparent that batch culture inocula were never able to displace a strain that had already reached steady state, even if the inoculum was large. This result indicated that bacterial selection for substrate affinity was even more important than bacteriocin production.

Antibiosis↗

PubMed2005

The ability of acidic pH, growth inhibitors, and glucose to increase the proton motive force and energy spilling of amino acid-fermenting Clostridium sporogenes MD1 cultures.

Clostridium sporogenes MD1 grew rapidly with peptides and amino acids as an energy source at pH 6.7. However, the proton motive force (Deltap) was only -25 mV, and protonophores did not inhibit growth. When extracellular pH was decreased with HCl, the chemical gradient of protons (ZDeltapH) and the electrical membrane potential (DeltaPsi) increased. The Deltap was -125 mV at pH 4.7, even though growth was not observed. At pH 6.7, glucose addition did not cause an increase in growth rate, but DeltaPsi increased to -70 mV. Protein synthesis inhibitors also significantly increased DeltaPsi. Non-growing, arginine-energized cells had a DeltaPsi of -80 mV at pH 6.7 or pH 4.7, but DeltaPsi was not detected if the F1F0 ATPase was inhibited. Arginine-energized cells initiated growth if other amino acids were added at pH 6.7, and DeltaPsi and ATP declined. At pH 4.7, ATP production remained high. However, growth could not be initiated, and neither DeltaPsi nor the intracellular ATP concentration declined. Based on these results, it appears that C. sporogenes MD1 does not need a large Deltap to grow, and Deltap appears to serve as a mechanism of ATP dissipation or energy spilling.

Adenosine Triphosphate↗

PubMed2005

Transcriptional regulation of beta-glucanase activity in the ruminal bacterium, Prevotella bryantii B14.

The ruminal bacterium, Prevotella bryantii B(1)4, grew more rapidly with glucose as an energy source than mannose (0.73 versus 0.47 h(-1)) and had 8-fold less beta-glucanase activity (50 versus 400 nmol reducing sugar mg protein(-1) min(-1)). Cultures that were provided with glucose and mannose had little beta-glucanase activity even though both sugars were utilized simultaneously. The observation that glucose and mannose were utilized simultaneously indicated that beta-glucanase expression was not merely a simple induction or inducer exclusion. When glucose was added to cultures growing on mannose, hexose flux through the glucomannokinase increased 1.5-fold, and this increase was associated with an almost immediate decrease in beta-glucanase mRNA. After only three generation (doubling) times, the amount of beta-glucanase mRNA was comparable to that observed in cells growing only with glucose. These results indicate that beta-glucanase activity is transcriptionally regulated. However, further work will be needed to define more precisely the nature of this regulation and to identify the intermediate in this response.

Blotting, Northern↗

PubMed2005

Enrichment of fusobacteria from the rumen that can utilize lysine as an energy source for growth.

Ruminal lysine degradation is a wasteful process that deprives the animal of an essential amino acid. Mixed ruminal bacteria did not deaminate lysine (50 mM) at a rapid rate, but lysine degrading bacteria could be enriched if Trypticase (5 mg/mL) was also added. Lysine degrading isolates produced acetate, butyrate and ammonia, were non-motile, stained Gram-negative and could also utilize lactate, glucose, maltose or galactose as an energy source for growth. Lactate was converted to acetate and propionate, and 16S rDNA indicated that their closest relatives were Fusobacterium necrophorum. Growing cultures produced ammonia at rates as high as 2400 nmol/mg protein/mL/min. Washed cell suspensions took up (14)C lysine (3 microM) at an initial rate of 6 nmol/mg protein/min, and glucose addition did not affect the transport. Cells washed aerobically had the same transport rate as those handled anaerobically, but only if the transport buffer contained sodium. The affinity constant for sodium was 8 mM, and sodium could not be replaced by lithium. Cells treated with the sodium/proton antiporter, monensin (5 microM), did not take up lysine, but a protonophore that inhibited growth (tetrachlorosalicylanilide, 10 microM) had no effect. An artificial membrane potential created by potassium diffusion did not increase the rate of lysine transport, and an Eadie-Hofstee plot indicated the transport rate was directly proportional to the lysine concentration. Decreasing the pH from 6.7 to 5.5 caused an 85% decrease in the rate of lysine transport. The addition of F. necrophorum JB2 (130 microg protein/mL) to mixed ruminal bacteria increased lysine degradation 10-fold, but only if the pH was 6.7 and monensin was not present. Further work will be needed to see if dietary lysine enriches fusobacteria in vivo.

Journal Article↗

PubMed2004

Effect of pH on the activity of bovicin HC5, a bacteriocin from Streptococcus bovis HC5.

The bacteriocin, bovicin HC5, catalyzed potassium efflux from Streptococcus bovis JB1, and this activity was highly pH dependent. When the pH was near neutral, glucose-energized cells were not affected by bovicin HC5, but the intracellular steady-state concentration of potassium decreased at acidic pH values. The idea that pH was affecting bovicin HC5 binding was supported by the observation that acidic pH also enhanced the efflux of potassium from non-energized cells that had been loaded with potassium. The relationship between bovicin HC5 concentration and potassium depletion was a saturation function, but cooperativity plots indicated that the binding of one bovicin molecule to the cell membrane facilitated the binding of another.

Bacteriocins↗

PubMed2004

The effect of cellobiose, glucose, and cellulose on the survival of Fibrobacter succinogenes A3C cultures grown under ammonia limitation.

The ruminal, cellulolytic bacterium, Fibrobacter succinogenes A3C, grew rapidly on cellulose, cellobiose, or glucose, but it could not withstand long periods of energy source starvation. If ammonia was limiting and either cellobiose or glucose was in excess, the viability declined even faster. The carbohydrate-excess, ammonia-limited cultures did not spill energy, but they accumulated large amounts of cellular polysaccharide. Cultures that were carbohydrate-limited had approximately 4 nmol ATP mg cell protein(-1), but ATP could not be detected in cultures that had an excess of soluble carbohydrates. However, if F. succinogenes A3C was provided with excess cellulose and ammonia was limiting, ATP did not decline, and the cultures digested the cellulose soon after additional nitrogen sources were added. From these results, it appears that excess soluble carbohydrates can promote the death of F. succinogenes, but cellulose does not.

Acids↗

PubMed2004

Nutritional requirements of Allisonella histaminiformans, a ruminal bacterium that decarboxylates histidine and produces histamine.

Histamine is an inflammatory agent that contributes to bovine laminitis. Cattle fed silage-containing rations often have large populations of Allisonella histaminiformans, but this obligate histidine-decarboxylating bacterium could not be isolated from cattle fed timothy hay. The growth of A. histaminiformans was stimulated by yeast extract, protein hydrolysates, and water-soluble extracts of alfalfa or corn silage. Extracts of alfalfa were more potent than corn silage. Because growth and histamine production were not stimulated by Casamino Acids or a mixture of purified amino acids, it appeared that A. histaminiformans requires peptides. The idea that A. histaminiformans requires peptides is consistent with the observation that alfalfa silages often have a large amount of peptide nitrogen.

Animal Feed↗

PubMed2003

Inhibition of Listeria monocytogenes by bovicin HC5, a bacteriocin produced by Streptococcus bovis HC5.

Cattle can be infected with Listeria monocytogenes by consuming contaminated plant materials, soil or silage, and farmers have sought ways of preventing this contamination. Recent work indicated that Streptococcus bovis HC5 produced a bacteriocin (bovicin HC5) that could inhibit a variety of gram-positive bacteria, and we examined the ability of bovicin HC5 to inhibit 10 strains of L. monocytogenes that had been isolated from plant materials, soil, silage and infected cattle. Growth experiments indicated that all of the L. monocytogenes strains were inhibited by 100 activity units (AU) of bovicin HC5 ml(-1). L. monocytogenes cultures that were transferred with sublethal doses (12.5 AU ml(-1)) could be adapted in stepwise fashion to higher doses of bovicin HC5. However, even 'adapted' cultures did not grow if 400 AU ml(-1) was added. The effect of bovicin HC5 on L. monocytogenes was bactericidal, and viability decreased 5-7 logs after only 2 h of exposure. Bovicin HC5 caused a nearly complete efflux of intracellular potassium in 15 min but only if the pH was less than 6.0. When the pH was greater than 6.0, the cells maintained their potassium pool. L. monocytogenes cells that were acid-adapted (final pH of 4.6) were as sensitive to bovicin HC5 as those that were not acid-adapted (final pH of 6.3). These results support the idea that bovicin HC5 could be effective in controlling listeria in contaminated silages.

Adaptation, Physiological↗

PubMed2003

The susceptibility of ionophore-resistant Clostridium aminophilum F to other antibiotics.

OBJECTIVE: To determine if ionophore-resistant ruminal bacteria are cross-resistant to other classes of antibiotics. Clostridium aminophilum was used as a model organism because this Gram-positive ruminal bacterium can adapt to ionophores (monensin and lasalocid). Non-adapted cultures lagged for at least 12 h with 1 microM monensin or lasalocid, but initiated no growth if the concentration was 10 microM. Adapted cultures did not lag with 1 microM monensin or lasalocid, grew well even if the ionophore concentration was 10 microM and contained cells at least 100,000-fold more resistant than those in non-adapted cultures. METHODS: Ionophore-adapted and non-adapted cultures were assayed for their susceptibility to other classes of antibiotics (penicillin G, ampicillin, cephalosporin C, vancomycin, carbenicillin, tetracycline, chloramphenicol, erythromycin, streptomycin, linocomycin, rifampicin, trimethoprim, novobiocin, polymyxin B and bacitracin) using a broth microdilution method. RESULTS: Adapted cultures retained their resistance phenotype for at least 28 generations even if ionophore was no longer present. Monensin-adapted cultures were as resistant to lasalocid as those adapted to lasalocid, but lasalocid-adapted cultures lagged with 1 microM monensin. Monensin- and lasalocid-resistant C. aminophilum F cultures were as susceptible to most antibiotics as non-adapted cultures. The only antibiotic that seemed to have a common mechanism of resistance was bacitracin, and the ionophore-adapted cultures had a 32-fold greater MIC. CONCLUSION: The use of ionophores in cattle feed and the selection of ionophore-resistant ruminal bacteria does not necessarily lead to other types of antibiotic resistance.

Anti-Bacterial Agents↗

PubMed2003

Factors affecting the antibacterial activity of the ruminal bacterium, Streptococcus bovis HC5.

Streptococcus bovis HC5 inhibits a variety of S. bovis strains and other Gram-positive bacteria, but factors affecting this activity had not been defined. Batch culture studies indicated that S. bovis HC5 did not inhibit S. bovis JB1 (a non-bacteriocin-producing strain) until glucose was depleted and cells were entering stationary phase, but slow-dilution-rate, continuous cultures (0.2 h(-1)) had as much antibacterial activity as stationary-phase batch cultures. Because the activity of continuous cultures (0.2-1.2 h(-1)) was inversely related to the glucose consumption rate, it appeared that the antibacterial activity was being catabolite repressed by glucose. When the pH of continuous cultures (0.2 h(-1)) was decreased from 6.7 to 5.4, antibacterial activity doubled, but this activity declined at pH values less than 5.0. Continuous cultures (0.2 h(-1)) that had only ammonia as a nitrogen source had antibacterial activity, and large amounts of Trypticase (10 mg ml(-1)) caused only a 2.0-fold decline in the amount of HC5 cell-associated protein that was needed to prevent S. bovis JB1 growth. Because S. bovis HC5 was able to produce antibacterial activity over a wide range of culture conditions, there is an increased likelihood that this activity could have commercial application.

Animals↗

PubMed2003

Ionophore resistance of ruminal bacteria and its potential impact on human health.

In recent years, there has been a debate concerning the causes of antibiotic resistance and the steps that should be taken. Beef cattle in feedlots are routinely fed a class of antibiotics known as ionophores, and these compounds increase feed efficiency by as much as 10%. Some groups have argued that ionophore resistance poses the same public health threat as conventional antibiotics, but humans are not given ionophores to combat bacterial infection. Many ruminal bacteria are ionophore-resistant, but until recently the mechanism of this resistance was not well defined. Ionophores are highly lipophilic polyethers that accumulate in cell membranes and catalyze rapid ion movement. When sensitive bacteria counteract futile ion flux with membrane ATPases and transporters, they are eventually de-energized. Aerobic bacteria and mammalian enzymes can degrade ionophores, but these pathways are oxygen-dependent and not functional in anaerobic environments like the rumen or lower GI tract. Gram-positive ruminal bacteria are in many cases more sensitive to ionophores than Gram-negative species, but this model of resistance is not always clear-cut. Some Gram-negative ruminal bacteria are initially ionophore-sensitive, and even Gram-positive bacteria can adapt. Ionophore resistance appears to be mediated by extracellular polysaccharides (glycocalyx) that exclude ionophores from the cell membrane. Because cattle not receiving ionophores have large populations of resistant bacteria, it appears that this trait is due to a physiological selection rather than a mutation per se. Genes responsible for ionophore resistance in ruminal bacteria have not been identified, but there is little evidence that ionophore resistance can be spread from one bacterium to another. Given these observations, use of ionophores in animal feed is not likely to have a significant impact on the transfer of antibiotic resistance from animals to man.

Animal Feed↗

PubMed2003

Ability of lysozyme and 2-deoxyglucose to differentiate human and bovine Streptococcus bovis strains.

Human and bovine Streptococcus bovis strains had the same 16S ribosomal DNA restriction fragment length polymorphism and often had the same patterns of starch, mannitol, lactose, and raffinose utilization. PCRs of BOX sequences differed, but numerical analyses indicated that some human strains clustered with bovine strains. However, human and bovine strains had distinctly different sensitivities to lysozyme and 2-deoxyglucose.

Animals↗

PubMed2002

The effect of bovicin HC5, a bacteriocin from Streptococcus bovis HC5, on ruminal methane production in vitro.

Methane represents a loss of feed energy to ruminant animals, and nutritionists have sought methods of inhibiting ruminal methane production. When mixed ruminal bacteria (approximately 400 mg protein ml(-1)) from a cow fed timothy hay were incubated in vitro with carbon dioxide and hydrogen (0.5 atm) for less than 8 h, the first-order rate of methane production was 17 micromol ml(-1). Semi-purified bacteriocin from Streptococcus bovis HC5 (bovicin HC5) inhibited methane production, by as much as 50%, and even a low concentration of bovicin HC5 (128 activity units (AU) ml(-1)) caused a significant decrease. Mixed ruminal bacteria that were transferred successively retained their ability to produce methane from carbon dioxide and hydrogen, and the first-order rate of methane production did not decrease. Cultures that were treated with bovicin HC5 (128 AU ml(-1)) gradually lost their ability to produce methane, and methane was not detected after four transfers. These latter results indicated that ruminal methanogens could not adapt and become resistant to bovicin HC5. When the chromosomal DNA was amplified with 16S rDNA primers specific to archaea, digested with restriction enzymes (HhaI and HaeIII) and separated on agarose gels, approximately 12 fragments were observed. DNA from control and treated cultures (third transfer) had the same fragment pattern indicating bovicin HC5 was not selective. Given the perception that the routine use of antibiotics in animal feeds should be avoided, bacteriocins may provide an alternative strategy for decreasing ruminal methane production.

Animals↗

PubMed2002

The adaptation and resistance of Clostridium aminophilum F to the butyrivibriocin-like substance of Butyrivibrio fibrisolvens JL5 and monensin.

When the amino acid-fermenting bacterium Clostridium aminophilum F was inoculated into media containing 1 microM monensin or a bacteriocin-like inhibitory substance (BLIS) from Butyrivibrio fibrisolvens JL5, the cultures lagged and growth was not observed for more than 12 h. The monensin- and BLIS-treated cultures eventually grew rapidly and did not lag a second time. Because cross-resistance could not be demonstrated, it appeared that the adaptation was specific. Non-adapted cells that were incubated with monensin lost their ability to produce ammonia from amino acids, and ATP, intracellular potassium, and electrical potential (DeltaPsi) were lower than untreated cells. Monensin-adapted cells regained their ability to produce ammonia, and intracellular potassium and DeltaPsi increased, but ATP was still 40% lower than untreated cells. When non-adapted cells were treated with the BLIS, ammonia production did not decline. Non-adapted cells were agglutinated by lysozyme, but in each case, adapted cells were not agglutinated. Adapted cells had more cellular polysaccharide and bound less of either inhibitor. Based on these results, it appears that the adapted cells had altered cell wall characteristics that prevented the binding of either monensin or the B. fibrisolvens JL5 BLIS.

Adenosine Triphosphate↗