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PubMedDate not supplied

The crucial question of standardisation when measuring drug consumption.

The reliability of drug consumption studies will depend on agreement on an international unit of measurement and the provision of accurate descriptions of patterns of use. This measurement unit should permit comparisons between countries and periods of time. Different units have been proposed and published. Consumption may be expressed in terms of pharmaceutical firm turnover, therapeutic costs, weight (total weight or dose equivalent), treatment doses such as defined daily dose and prescribed daily dose, or as number of items or packages sold. The advantages and disadvantages of the different evaluation units used in veterinary medicine are reviewed.

Animals↗
PubMed1988

Olaquindox resistance in the coliform flora of pigs and their environment: an ecological study.

Faecal samples were taken weekly from a chosen pen on each of four commercial pig farms, two of which used olaquindox as a feed additive. Coliform bacteria were isolated from the samples and the incidence and level of resistance to olaquindox and to four therapeutic antibiotics determined. The coliforms isolated were biotyped to follow the emergence of drug-resistant sub-populations. The results showed that the coliform flora was complex and that a turnover of biotypes was associated with changes in the occupancy of the pen and, possibly, diet. This turnover led to large fluctuations in both incidence and level of resistance to olaquindox and the antibiotics. Olaquindox resistance was not specifically linked with resistance to any therapeutic antibiotic, and the possible origin of olaquindox-resistant strains is discussed in relation to the biotypes.

Ampicillin↗
PubMed1974

Streptomycin-induced synthesis of abnormal protein in an Escherichia coli mutant.

To determine directly the effects of streptomycin on translational fidelity in intact cells, we studied the synthesis of beta-galactosidase and of the coat protein of bacteriophage R17 in an Escherichia coli mutant in which the bactericidal effects of streptomycin are delayed. After the addition of streptomycin to exponentially growing mutant cells, protein synthesis continues at an undiminished rate for approximately an hour; however, as measured by enzyme assays, little functional protein is produced. Serological assays designed to detect beta-galactosidase and bacteriophage R17 coat protein show that substantial amounts of the protein synthesized can react with antisera prepared against active beta-galactosidase and phage R17, indicating the aberrance of the protein produced in the presence of the antibiotic. The polypeptides synthesized in the presence of streptomycin are degraded in the cell to a much greater extent than protein synthesized in the absence of the antibiotic. The proteolytic attack on this protein is not affected by inhibitors of serine proteases, suggesting that enzymes other than those involved in "normal turnover" of cellular protein are responsible. In this strain, certain of the multiple effects of streptomycin are separated in time and the production of abnormal protein (enzymatically inactive and susceptible to proteolytic attack) could be studied in the absence of the lethal effect of the drug.

Arginine↗
PubMed1990

Penicillin-binding proteins and ampicillin resistance in Haemophilus influenzae.

Ampicillin-resistant, non-beta-lactamase-producing isolates of Haemophilus influenzae contain a variety of penicillin-binding protein (PBP) patterns that differ from the single pattern of eight PBPs characteristic of susceptible strains. During genetic transformation of resistance, only some of the anomalies in PBP pattern were transformed, specifically those relating to the penicillin-binding capacities of PBPs 4 (Mr of 62,000) and 5 (Mr of 59,000) and, in some transformations, PBP 3 (Mr of 71,000). Comparison of the binding of penicillin by PBPs 4 and 5 of three resistant transformants (derived with DNA from different donors) revealed a decrease in the rate of PBP acylation and no appreciable change in the rate of deacylation as compared to the susceptible recipient. Thus, rapid turnover of these PBPs does not play a role. Retransformation studies confirm that altered PBPs 3, 4, and 5 are associated with resistance and suggest that these PBPs are major targets for the beta-lactam antibiotics in H. influenzae.

Acylation↗
PubMed1979

Collagenolytic activity of crevicular fluid and of adjacent gingival tissue.

The fluid in and gingival tissue lining periodontal pockets were collected from male patients undergoing periodontal therapy. The collagenolytic activity of the crevicular fluid, and the ability of the gingiva in culture to degrade (a) an exogenous collagen substrate, and (b) endogenous collagen newly synthesized and labeled with H3-hydroxyproline, were related to the severity of gingival inflammation. Although inflammation appeared to have only a slight effect on gingival collagenolytic activity and on the turnover of collagen newly synthesized in culture, a marked effect was observed on the collagenase activity in the crevicular fluid. This study suggests that the collagen destructive activity of the periodontal lesion can be assessed by monitoring crevicular fluid collagenolytic activity.

Adult↗
PubMed1992

Taxol blocks processes essential for prostate tumor cell (PC-3 ML) invasion and metastases.

We have examined the antimetastatic effects of taxol on a PC-3 human prostatic tumor variant (PC-3 ML) which metastasizes to the lumbar vertebrae in severe combined immunodeficiency-carrying (SCID) mice. Immunofluorescence labeling indicated that taxol (0.5 to 1.0 microM for 6 h) produced an abnormal bundling of microtubules in a dosage-dependent manner. Slot blotting and gelatinase assays revealed that taxol inhibited secretion of the M(r) 72,000 and M(r) 92,000 type IV collagenases plus a M(r) 57,000 gelatinase. Radioimmunoprecipitation measurements confirmed that the drug inhibited both the secretion and the synthesis of the M(r) 72,000 collagenase. Taxol also blocked total protein secretion but did not influence total protein synthesis or turnover. Boyden chamber chemotactic studies further showed that taxol (0.5 to 1.0 microM) inhibited invasion of Matrigel. More importantly, studies in SCID mice demonstrated that taxol (50 to 250 mg/m2/day) blocked the establishment, growth, and long-term survival of PC-3 ML cells.

Alkaloids↗
PubMed2006

Modeling the reactions of superoxide and myeloperoxidase in the neutrophil phagosome: implications for microbial killing.

Neutrophils kill bacteria by ingesting them into phagosomes where superoxide and cytoplasmic granule constituents, including myeloperoxidase, are released. Myeloperoxidase converts chloride and hydrogen peroxide to hypochlorous acid (HOCl), which is strongly microbicidal. However, the role of oxidants in killing and the species responsible are poorly understood and the subject of current debate. To assess what oxidative mechanisms are likely to operate in the narrow confines of the phagosome, we have used a kinetic model to examine the fate of superoxide and its interactions with myeloperoxidase. Known rate constants for reactions of myeloperoxidase have been used and substrate concentrations estimated from neutrophil morphology. In the model, superoxide is generated at several mm/s. Most react with myeloperoxidase, which is present at millimolar concentrations, and rapidly convert the enzyme to compound III. Compound III turnover by superoxide is essential to maintain enzyme activity. Superoxide stabilizes at approximately 25 microM and hydrogen peroxide in the low micromolar range. HOCl production is efficient if there is adequate chloride supply, but further knowledge on chloride concentrations and transport mechanisms is needed to assess whether this is the case. Low myeloperoxidase concentrations also limit HOCl production by allowing more hydrogen peroxide to escape from the phagosome. In the absence of myeloperoxidase, superoxide increases to >100 microM but hydrogen peroxide to only approximately 30 microM. Most of the HOCl reacts with released granule proteins before reaching the bacterium, and chloramine products may be effectors of its antimicrobial activity. Hydroxyl radicals should form only after all susceptible protein targets are consumed.

Blood Bactericidal Activity↗
PubMed1991

Evidence that estramustine binds MAP-1A to inhibit type IV collagenase secretion.

Estramustine is a novel anti-microtubule drug shown to bind MAP-1 and MAP-2 (microtubule-associated proteins) in vitro. In this paper we have shown that estramustine specifically binds MAP-1A in Du 145a cells, resulting in disruption of MAP-1A microtubules and inhibition of type IV collagenase secretion. Immunofluorescence studies revealed that at 30 microM levels estramustine blocked type IV collagenase secretion by partial disruption of the MAP-1A microtubule networks. Immunoprecipitation studies with polyclonal antibodies provided quantitative evidence that 30-60 microM estramustine blocked secretion of a 105 x 10(3) Mr type IV collagenase. Pulse-labeling experiments confirmed that the effect was not a result of inhibition of either protein synthesis or altered rates of type IV collagenase turnover. Finally, drug uptake studies with [3H]estramustine, scintillation counting and fluorography demonstrated that the principal target of the drug was MAP-1A. For the first time we have shown that the drug blocks secretion by binding MAP-1A and causing incomplete disruption of the microtubule networks.

Cell Line↗
PubMed1987

Studies of rumen function in an in vitro continuous culture system.

An in vitro continuous culture system to simulate processes in the rumen is described. This comprises a culture vessel (Fig. 2) fed continuously with pelleted solid feed (Fig. 1); artificial saliva also enters the vessel while effluent leaves it continuously in two streams, one is filtered and the other simply overflows (Fig. 1). In this way liquid and solid turnover times may be manipulated independently; in a trial experiment the former and latter were 6.3% and 3.4% per hour respectively. In four replicated experiments a steady-state was achieved in 5-6 days with a feed of barley, tapioca, fishmeal, urea and straw and maintained for a further 7 to 14 days. During a steady-state period, rumen characteristics in terms of protozoal count, pH, redox potential, total volatile fatty acid concentration and ammonia concentration were stable and similar to those found in the rumens of animals given similar diets. Flows of microbial N compounds were assessed by infused Na2H32PO4 with the artificial saliva and estimating the 32P label incorporated in bacterial fractions. The system demonstrably provided a satisfactory means for studying many aspects of rumen function.

Animal Feed↗
PubMed2002

Mutant TEM beta-lactamase producing resistance to ceftazidime, ampicillins, and beta-lactamase inhibitors.

A derivative of the TEM-1 beta-lactamase producing clinically significant levels of resistance to ceftazidime and beta-lactamase inhibitors in the presence of penicillins was generated following five rounds of DNA shuffling and selection. This complex mutant enzyme contained three amino acid substitutions including those of residues 104 and 276 that are known to produce extended-spectrum resistance and, correspondingly, resistance to beta-lactamase inhibitors. Although the Glu104Lys substitution by itself produced low levels of ceftazidime resistance, additional amino acid replacements in the enzyme with the triple mutation resulted in further enhancement of resistance to ceftazidime. Kinetic studies of the purified beta-lactamase enzyme with the triple mutation indicated enhancement of the catalytic efficiency for turnover (kcat/Km) of ceftazidime. The increases in the Ki values of both clavulanic acid and tazobactam for the enzyme with the triple mutation were consistent with the observed bacterial resistance to the reversibility of beta-lactam resistance with these inhibitors.

Amino Acid Substitution↗
PubMed2006

Characterization of the Staphylococcus aureus heat shock, cold shock, stringent, and SOS responses and their effects on log-phase mRNA turnover.

Despite its being a leading cause of nosocomal and community-acquired infections, surprisingly little is known about Staphylococcus aureus stress responses. In the current study, Affymetrix S. aureus GeneChips were used to define transcriptome changes in response to cold shock, heat shock, stringent, and SOS response-inducing conditions. Additionally, the RNA turnover properties of each response were measured. Each stress response induced distinct biological processes, subsets of virulence factors, and antibiotic determinants. The results were validated by real-time PCR and stress-mediated changes in antimicrobial agent susceptibility. Collectively, many S. aureus stress-responsive functions are conserved across bacteria, whereas others are unique to the organism. Sets of small stable RNA molecules with no open reading frames were also components of each response. Induction of the stringent, cold shock, and heat shock responses dramatically stabilized most mRNA species. Correlations between mRNA turnover properties and transcript titers suggest that S. aureus stress response-dependent alterations in transcript abundances can, in part, be attributed to alterations in RNA stability. This phenomenon was not observed within SOS-responsive cells.

Adaptation, Physiological↗
PubMed2005

Nitric oxide dioxygenase function and mechanism of flavohemoglobin, hemoglobin, myoglobin and their associated reductases.

Microbial flavohemoglobins (flavoHbs) and hemoglobins (Hbs) show large *NO dioxygenation rate constants ranging from 745 to 2900 microM(-1) s(-1) suggesting a primal *NO dioxygenase (NOD) (EC 1.14.12.17) function for the ancient Hb superfamily. Indeed, modern O2-transporting and storing mammalian red blood cell Hb and related muscle myoglobin (Mb) show vestigial *NO dioxygenation activity with rate constants of 34-89 microM(-1) s(-1). In support of a NOD function, microbial flavoHbs and Hbs catalyze O2-dependent cellular *NO metabolism, protect cells from *NO poisoning, and are induced by *NO exposures. Red blood cell Hb, myocyte Mb, and flavoHb-like activities metabolize *NO in the vascular lumen, muscle, and other mammalian cells, respectively, decreasing *NO signalling and toxicity. HbFe(III)-OO*, HbFe(III)-OONO and protein-caged [HbFe(III)-O**NO2] are proposed intermediates in a reaction mechanism that combines both O-atoms of O2 with *NO to form nitrate and HbFe(III). A conserved Hb heme pocket structure facilitates the dioxygenation reaction and efficient turnover is achieved through the univalent reduction of HbFe(III) by associated reductases. High affinity flavoHb and Hb heme ligands, and other inhibitors, may find application as antibiotics and antitumor agents that enhance the toxicity of immune cell-derived *NO or as vasorelaxants that increase *NO signalling.

Amino Acid Sequence↗
PubMed1990

Stereoselective Reduction of Ethyl 4-Chloro-3-Oxobutanoate by a Microbial Aldehyde Reductase in an Organic Solvent-Water Diphasic System.

Enzyme-catalyzed asymmetric reduction of ethyl 4-chloro-3-oxobutanoate in an organic solvent-water diphasic system was studied. NADPH-dependent aldehyde reductase isolated from Sporobolomyces salmonicolor AKU4429 and glucose dehydrogenase were used as catalysts for reduction of ethyl 4-chloro-3-oxobutanoate and recycling of NADPH, respectively, in this system. In an aqueous system, the substrate was unstable. Inhibition of the reaction and inactivation of the enzymes by the substrate and the product were also observed. An n-butyl acetate-water diphasic system very efficiently overcame these limitations. In a 1,600-ml-1,600-ml scale diphasic reaction, ethyl (R)-4-chloro-3-hydroxybutanoate (0.80 mol; 86% enantiomeric excess) was produced from the corresponding oxoester in a molar yield of 95.4% with an NADPH turnover of 5,500 mol/mol.

Journal Article↗
PubMed1992

In vitro induction of cecropin genes--an immune response in a Drosophila blood cell line.

The Drosophila melanogaster cell line mbn-2 was explored as a model system to study insect immune responses in vitro. This cell line is of blood cell origin, derived from larval hemocytes of the mutant lethal (2) malignant blood neoplasm (1(2)mbn). The mbn-2 cells respond to microbial substances by the activation of cecropin genes, coding for bactericidal peptides. The response is stronger than that previously described for SL2 cells, and four other tested Drosophila cell lines were totally unresponsive. Bacterial lipopolysaccharide, algal laminarin (a beta-1,3-glucan), and bacterial flagellin were strong inducers, bacterial peptidoglycan fragments gave a weaker response, whereas a formyl-methionine-containing peptide had no effect. Experiments with different drugs indicate that the response may be mediated by a G protein, but not by protein kinase C or eicosanoids, and that it requires a protein factor with a high rate of turnover.

Animals↗
PubMed2026

Transcriptomic changes in the gut mucosa of fasting northern elephant seal pups reveal immune modulation during early microbiome establishment.

Fasting is an integral component of the life-history of many species. Following abrupt weaning, northern elephant seal pups (Mirounga angustirostris) undergo an extended post-weaning fast of approximately 60 days. During this period, enteric bacterial diversity increases, suggesting that host immune regulation may facilitate the establishment of microbial communities. However, the molecular processes occurring within the intestinal mucosa during this transition remain poorly understood. To investigate these mechanisms, we characterized transcriptional changes in the enteric mucosa of male and female northern elephant seal pups sampled at weaning and after one month of fasting. Total RNA isolated from rectal swabs was sequenced and aligned to the Mirounga angustirostris reference genome. Differential gene expression and gene set enrichment analyses were used to identify genes and pathways associated with fasting and sex-specific responses. Fasting was accompanied primarily by transcriptional downregulation, including genes involved in antimicrobial defense, inflammation, protein turnover, and epithelial remodeling. In contrast, several genes associated with B-cell activity and immune recognition were upregulated. Gene Set Enrichment Analysis revealed coordinated activation of immune-regulatory pathways indicating dynamic modulation of intestinal immunity rather than generalized immune suppression. Pronounced sex-specific differences were also observed. Male pups exhibited transcriptional patterns consistent with enhanced immune tolerance, whereas females showed broader immune-pathway activation, including enrichment of pro-inflammatory and stress-response pathways. Several non-coding RNAs also displayed sex-specific changes in expression. Together, these findings suggest that fasting induces transcriptional remodeling of the gut and may contribute to immune regulation during a critical period of microbiome establishment in northern elephant seal pups.

Animals↗
PubMed1986

Bacterial organomercurial lyase: overproduction, isolation, and characterization.

Organomercurial lyase mediates the first of two steps in the microbial detoxification of organomercurial salts. This enzyme encoded on the plasmid R831 obtained from Escherichia coli J53-1 has been overproduced to the level of 3% of the soluble cell protein in E. coli by a construction using the T7 promoter. The enzyme has been purified to homogeneity in quantity in three steps. It is a monomer of Mr 22,400 with no detectable cofactors or metal ions. It catalyzes the protonolysis of the C-Hg bond in a wide range of organomercurial salts (primary, secondary, tertiary, alkyl, vinyl, allyl, and aryl) to the hydrocarbon and mercuric ion with turnover rates in the range of 1-240 min-1.

Amino Acid Sequence↗
PubMed2005

Compound-specific delta13C analysis of individual amino sugars--a tool to quantify timing and amount of soil microbial residue stabilization.

There is strong scientific evidence that microbial residues such as amino sugars may be stabilized in soil. However, up to now, no investigation has been carried out to quantify both the amount and timing of such stabilization. This is primarily due to methodological constraints, because it is not possible to differentiate between stabilized (old) and recently produced (new) amino sugars when these biomarkers are conventionally analyzed, e.g. by means of gas chromatography and flame ionization detection. Therefore, the aim of the present study was to test whether compound-specific isotope analysis (delta13C) of amino sugars extracted from soil could be used to differentiate between old and new microbial residues. For this aim a method for the delta13C analysis of individual amino sugars was developed and optimized. First results of delta13C values of glucosamine, galactosamine, mannosamine, and muramic acid in soil samples from two different ecological studies are presented, clearly indicating that discrimination between soil inherent and newly formed amino sugars is possible in stable isotope labeling experiments. Our results further showed that, in the short term (within 1 month), only few amino sugars were built, thus making highly 13C-enriched substrates necessary for the quantification of new amino sugar production and for the determination of amino sugar turnover rates.

Amino Sugars↗
PubMed2003

Disposal and degradation of pesticide waste.

Generation of pesticide waste is inevitable during every agricultural operation from storage to use and equipment cleanup. Large-scale pesticide manufacturers can afford sophisticated recovery, treatment, and cleanup techniques. Small-scale pesticide users, for example, single farms or small application businesses, struggle with both past waste problems, including contaminated soils, and disposal of unused product and equipment rinsewater. Many of these problems have arisen as a result of inability to properly handle spills during, equipment loading and rinsewater generated after application. Small-scale facilities also face continued problems of wastewater handling. Old, obsolete pesticide stocks are a vexing problem in numerous developing countries. Pesticide waste is characterized by high concentrations of a diversity of chemicals and associated adjuvants. Dissipation of chemicals at elevated concentrations is much slower than at lower concentrations, in part because of microbial toxicity and mass transfer limitations. High concentrations of pesticides may also move faster to lower soil depths, especially when pore water becomes saturated wish a compound. Thus, if pesticide waste is not properly disposed of, groundwater and surface water contamination become probable. The Waste Management Hierarchy developed as an Australian Code of Practice can serve as a guide for development of a sound waste management plan. In order of desirability, the course of actions include waste avoidance, waste reduction, waste recycling, waste treatment, and waste disposal. Proper management of pesticide stocks, including adequate storage conditions, good inventory practices, and regular turnover of products,. will contribute to waste avoidance and reduction over the long-term. Farmers can also choose to use registered materials that have the lowest recommended application rates or are applied in the least volume of water. Wastewater that is generated during equipment rinsing can be recycled by spraying it onto cropland, thus avoiding a soil contamination problem. If it is not feasible to spray out rinsates, then water treatment becomes necessary. However, for small waste generators, practical technology is still too experimental and not easily implemented on an individual farm or at a small application business. Nevertheless, research has been quite active in application of advanced oxidation processes (UV/ozonation: photoassisted Fenton reaction: photocatalysis using TiO2). Obsolete pesticide stocks in developing countries are being packaged and shipped to developed countries for incineration. Contaminated soil can also be incinerated, but this is not practical nor affordable for small waste generators. Chemical degradation of chlorinated hydrocarbon pesticides may be amenable to dechlorination by alkali polyethylene glycol treatment, but further study is needed to make the technique practical for small waste generators. Contaminated soils may be amenable to cleanup by one of several biological treatment methods, including composting, landfarming, and bioaugmentation/ biostimulation. Composting and landfarming (which may be used in combination with biostimulation) may be the most practical of the biological methods that is immediately ready for implementation by small-scale pesticide waste generators.

Agriculture↗