Antibiotic-tolerant mutants of Streptococcus pneumoniae that are not deficient in autolytic activity.

Several mutants of Streptococcus pneumoniae were isolated that appeared tolerant, to varying extents, to the lytic and bactericidal effects of some antibiotics that inhibit peptidoglycan synthesis, but were not deficient in autolytic activity. The method used to select the mutants was based on the survival of tolerant mutants during treatment with either bacitracin, benzylpenicillin, D-cycloserine plus beta-chloro-D-alanine, or vancomycin. Most (60 to 80%) of the surviving isolates were found to be deficient in autolytic activity, and these were rejected. The smaller proportion that had wild-type sensitivity to deoxycholate-induced lysis was studied further with respect to tolerance to the other antibiotics used in the selection procedures. Two of these mutants (selected by treatment with benzylpenicillin) were tolerant to either benzylpenicillin or D-cycloserine plus beta-chloro-D-alanine, but were supersusceptible, in terms of initiation of lysis, to either bacitracin or vancomycin. The minimal inhibitory concentration values of several antibiotics for these two mutants were identical to those for the wild-type strain. Moreover, the interaction of radioactive benzylpenicillin with the penicillin-binding proteins, examined in whole organisms, also appeared the same as previously found for either wild-type or autolytic-deficient strains of S. pneumoniae.     

Sensitivity to antibiotics of pneumococci isolated from patients with chronic nonspecific pneumonia and bronchitis

Four hundred and twenty two pneumococcal strains isolated from 300 patients with chronic nonspecific pneumonia and bronchitis were studied with respect to their sensitivity to 18 antibiotics within a period from 1982 to 1985. It was shown with the method of serial dilutions on solid media that 91.7, 87.8, 85 and 81 per cent of the isolates were sensitive to benzylpenicillin, ampicillin, lincomycin and cefuroxime, respectively. A significant percentage of the pneumococcal strains had decreased sensitivity to benzylpenicillin (MIC close to the therapeutic concentration). On this basis it was recommended to use lower concentrations of benzylpenicillin (less than 0.25 units/ml) in assay of sensitivity in clinical strains of Pneumococcus.     

The sensitivity to 12 antibiotics of 125 strains of Streptococcus group B isolated in a maternity home

Sensitivity of 125 strains of group B streptococci isolated from newborns, their mothers and personnel in a maternity home was studied with respect to 12 antibiotics: benzylpenicillin, ampicillin, methicillin, cephalotin, erythromycin, lincomycin, levomycetin (chloramphenicol), oxacillin, tetracycline, streptomycin, gentamicin and ristomycin. The method of serial dilutions in a solid medium was applied. All the strains were sensitive to ristomycin and erythromycin. The predominating number of the strains were sensitive to lincomycin, levomycetin and the beta-lactam antibiotics. Strains resistant or moderately resistant to benzylpenicillin, ampicillin, oxacillin, methicillin and cephalotin were detected. The majority of the strains were resistant to streptomycin, tetracycline and gentamicin. Multiple antibiotic resistance with 2-7 determinants was revealed in 11.2 per cent of the strains. The antibiotic sensitivity of the strains isolated from the newborns, their mothers and the personnel in the maternity home was on the whole similar or insignificantly differed.     

两种抗生素对龙须菜的光合生理效应

探讨了不同浓度的两种抗生素(氯霉素和青霉素G钠)对龙须菜(Gracilaria lemaneiformis)生长、光合作用、呼吸作用、色素含量以及可溶性蛋白含量等生理特性的影响。结果表明:龙须菜的生长受到两种抗生素的影响,但是氯霉素的影响要比青霉素G钠的影响大的多。在氯霉素处理的过程中,光合作用、有效光化学效率(Yield)、藻红藻蓝蛋白以及可溶性蛋白含量都随着氯霉素浓度的升高而显著下降,但是呼吸作用速率由于氯霉素的处理而升高;此外光合色素含量不受氯霉素的影响。在青霉素G钠的处理中,光合作用、有效光化学效率随着青霉素G钠的升高而下降,龙须菜叶绿素a与类胡萝卜素含量随着青霉素G钠浓度的升高而具有升高的趋势,但藻红蛋白、藻蓝蛋白以及可溶性蛋白在各处理组之间均没有表现出一定趋势。这些结果说明,氯霉素对生长的影响主要是光合作用速率的下降,以及有关蛋白合成下降引起,而青霉素G钠对生长的影响可能原因是呼吸作用速率的增加引起。由于龙须菜对氯霉素的敏感性比对青霉素G钠的敏感性更强,氯霉素在基因工程的育种中可能更适合作为选择压力。     

Demonstration of the presence of an inducibleβ-lactamase inAzospirillum lipoferum

Azospirillum lipoferum is a soil microorganism that has been shown to be resistant to penicillins. It has been suggested that resistance is due to the presence of a -lactamase activity, but no conclusive evidence has been reported. The incubation of benzylpenicillin, or nitrocephin with either wholeAzospirillum cells or cell-free extracts was accompanied, by hydrolysis of the -lactam ring of the antibiotics. Such hydrolytic activity exhibited Michaelis and Menten-like kinetics. The enzyme was produced at a low, basal level that was increased approximately 50 times by the addition of benzylpenicillin, an increase that was completely blocked by chloramphenicol or rifampicin.     

Benzylpenicillin efficacy for neutropenic infection prophylaxis in patients with cancer and postcytostatic neutropenia]

Evaluation of benzylpenicillin (penicillin G) effect for infection prophylaxis at the oncological patients with severe postcytostatic neutropenia was performed. All the patients with neutrophils levels lower than 0.5 x 10(9)/L were recommended to use antibiotics for infection prophylaxis. Test-group (n = 40) used ciprofloxacin (0.5 g twice daily, per os) combined with benzylpenicillin (1.0 g four times daily, i/v); control group was treated by ciprofloxacin in the same dose only. Combination with benzylpenicillin resulted in statistically significant reduction of infections frequency among oncological patients.     

Assessment of the sensitivity of Neisseria meningitidis strains to benzylpenicillin using the disc diffusion method]

The sensitivity of 235 N. meningitidis strains to 5 antibiotics was estimated by the diameter of growth inhibition zones according to the criteria recommended by a Laboratory (Marseilles, France) collaborating with the WHO. All the strains proved to be sensitive to benzylpenicillin when disks containing 10 and 2 IU of the antibiotic were used. The strains were also shown to be sensitive to chloramphenicol and tetracycline. 95.7 and 7.7 per cent of the strains were sensitive to rifampicin and oleandomycin, respectively. When the strain sensitivity was assayed with the disks containing 10 and 2 IU of benzylpenicillin by the more severe criteria recommended by J. Saez-Nieto et al., significant changes were detected: meningococci with relative resistance to benzylpenicillin were detected in various regions of this country and the number of such strains was found to have a tendency to slightly increase.     

Effect of hydroacridine derivatives on the sensitivity of polyresistant strains of Staphylococcus aureus and Escherichia coli to antibiotics]

Sensitivity of 4 clinical strains of Staph. aureus and E. coli to 13 hydroacridine derivatives and their combinations with antibiotics, such as benzylpenicillin, ampicillin, semi-synthetic penicillins, streptomycin, chloramphenicol, tetracycline, chlortetracycline, monomycin, oleandomycin and erythromycin was studied. The highest bacteriostatic effect was observed on the use of perhydroactidine derivatives with benzylpenicillin or ampicillin with respect to polyresistant penicillinase-producing strains of Staph. aureus, resistance of which to these antibiotics was decreased 250--1000 times. Under the effect of the above compounds the staphylococcal resistance to chloramphenicol, tetracycline, chlortetracycline, oleandomycine and erythromycin decreased 2--66 times. The combinations of hydroacridine with the antibiotics, except 10-amino-trans-syn-trans-perhydroacridine had no effect on the resistance of the E. coli strains. The results of the combined effect of the above substances were associated with their chemical nature, the bacterial type and possibly the character of the strain resistance.     

Purification and characterization of the penicillin-binding protein that is the lethal target of penicillin in Bacillus megaterium and Bacillus licheniformis. Protein exchange and complex stability.

The penicillin-binding protein that is thought to be the lethal target of penicillin in Bacillus megaterium (protein 1) has been purified to greater than 95% homogeneity. The membrane-bound penicillin-binding proteins were solubilized with a non-ionic detergent and partially separated from each other by ion-exchange chromatography on DEAE-Sepharose CL-6B. Protein 1 was subsequently purified by covalent affinity chromatography on ampicillin-affinose. Bacillus licheniformis contains an equivalent penicillin-binding protein (protein 1) that can be more readily purified to virtual homogeneity in a one-step procedure. It was separated from the other penicillin-binding proteins by utilizing the observation that in this organism, this particular protein is the only one whose covalent complex with benzylpenicillin subsequently breaks down. Membranes were treated with saturating concentrations of benzylpenicillin followed by the removal of free penicillin and further incubation to allow the complex between benzylpenicillin and protein 1 to break down. The penicillin-binding proteins were then solubilized and applied to a column of ampicillin-affinose to which only protein 1 was bound as the other penicillin-binding proteins still had benzylpenicillin bound to them. Pure protein 1 was eluted from the affinity resin with hydroxylamine. The interaction of benzylpenicillin with purified protein 1 has been studied by separating unbound antibiotic from the benzylpenicillin . protein complex by paper electrophoresis. Benzylpenicillin reacts with the protein rapidly to form a covalent complex and the fully saturated complex has a molar ratio of bound [14C] benzylpenicillin: protein of 0.7:1. The complex breaks down, obeying first-order kinetics, with a half-life of 16 min at 35 degrees C, a value identical to that obtained with the membrane-bound protein. The concentration of benzylpenicillin that results in the formation of 50% of the maximum amount of benzylpenicillin . protein complex is that at which the molar amount of benzylpenicillin present is equal to 50% of the molar amount of penicillin-binding protein, rather than being a measure of any of the kinetic parameters of the binding reaction. This observation may be significant in the interpretation of previous results where the amounts of penicillins needed to kill cells or to inhibit penicillin-sensitive reactions have been expressed as concentrations. The possible importance of the breakdown of beta-lactam . protein complexes in the clinical use of these antibiotics is discussed.     

Effect of the functional state of the internal organs on the pharmacokinetics of benzylpenicillin in angina]

The levels of benzylpenicillin on its intramuscular administration in doses of 100000 or 200000 units, as well as the renal and hepatic functions and the antibiotic binding by the host protein components were studied in 44 angina patients and 29 healthy persons. The benzylpenicillin levels were always markedly higher in the angina cases than those in the healthy persons. The ratio of the free and protein bound fractions of the antibiotic in the above groups was practically the same. At the same time the amount of benzylpenicillin excreted with the urine was higher in the angina patients than in the healthy persons approximately by 2 times. The study of the function of the internal organs revealed a marked decrease in the daily diuresis in all the angina patients and microhematuria in the 1/4 of the cases. The impairment of the renal function was determined by the laboratory methods practically in all the patients. On the basis of the data obtained it was concluded that the increased benzylpenicillin levels in the angina cases were connected with a decreased capacity for inactivating this substance by the liver enzymatic systems due to the changed function of this organ. The fact of the increased antibiotics levels in the host is considered as a positive phenomenon providing inhibition of the disease causative agents resistant to usual concentrations of the drug.     

Side effects of antibiotics in patients with thermal burns]

The possibilities of antibacterial therapy in the clinics of burn diseases has at present decreased because of increasing microflora resistance to antibiotics. This phenomenon is one of the most often causes of antibacterial drug side effects in burn patients. For control of infections complications in burn patients which are most often lethal it is necessary to use biologically active subtance, such as prodigiozan and lysozime in addition to the directed antibiotic therapy. The use of specific antitoxic antistaphylococcal drugs, such as antistaphylococcal plasma and antistaphylococcal gamma-globulin in combination with the antibiotics of the direct action provided effective control of infectious complications and sepsis of staphylococcal genesis in burn patients. Decamine proved to be effective along with the usual use of nystatin in cases with dysbacteriosis as a result of the antibiotic side effects. In the patients treated with decamine the sings of candidosis disappeared by the 5th--7th day. Therefore, for decreasing the side effects of antibiotics in the clinics of burn patients it is expedient to use antibiotics in combination with the biologically active and immune preparations which increases the efficacy of antibiotic therapy, impfoves the treatment results and decreases the antibiotic side effects.     

Incidence of antibodies to antibiotics in subjects with chronic staphylococcal diseases

Antibody response to antibiotics in patients with chronic staphylococcal diseases was studied and the results were compared with the production of specific antibodies. Increased titres of antibodies to antibiotics were found in cases with inadequate antibody response. The possibility of an immunosuppressive effect of antibiotics on the formation of specific antistaphylococcal antibodies is discussed.     

Purification and partial characterization of a penicillin-binding protein from Mycobacterium smegmatis.

Penicillin-binding proteins (PBPs), although characterized from several organisms, have so far not been studied in mycobacteria. The present study is the first characterization of a PBP from Mycobacterium smegmatis. The PBP was purified by solubilization of the membranes with Triton X-100 and successive chromatography of the solubilized proteins on ampicillin-linked CH Sepharose 4B and DE-52. The purified PBP (M(r), 49,500) catalyzed a model transpeptidase reaction with the tripeptide acetyl2-L-Lys-D-Ala-D-Ala as the substrate and Gly-Gly as the acceptor. The transpeptidase activity was inhibited by 50% at a benzylpenicillin concentration of 1.8 x 10(-7) M, which was similar to the concentration (1.1 x 10(-7) M) of benzylpenicillin required to saturate to 50% this PBP. Of several antibiotics tested, the concentration of antibiotic required to inhibit [35S]penicillin binding by 90% was found to be the lowest for cefoxitin and Sch 34343.     

Effect of benzylpenicillin on murein biosynthesis, turnover and structure in Listeria monocytogenes cells

The action of beta-lactam antibiotics such as ampicillin and benzylpenicillin on cells of Listeria monocytogenes appears to be bacteriostatic. However, after approximately two hours in the presence of 10 x MIC of benzylpenicillin the cells begin to rapidly lose viability without undergoing lysis. In this report we present the results of studies on the biosynthesis of murein in L. monocytogenes cells during the first 120 min of their exposure to benzylpenicillin as measured by the continuous and pulse incorporation of the precursor N-acetyl-D-[1-3H]glucosamine. The turnover of the murein sacculus in the presence of penicillin as well as the lack of discernible changes in the molecular structure of the murein synthesised in the presence of benzylpenicillin is also discussed.     

Crystallographic mapping of beta-lactams bound to a D-alanyl-D-alanine peptidase target enzyme

X-ray crystallography has been used to examine the binding of three members of the beta-lactam family of antibiotics to the D-alanyl-D-alanine peptidase from Streptomyces R61, a target of penicillins. Cephalosporin C, the monobactam analog of penicillin G and (2,3)-alpha-methylene benzylpenicillin have been mapped at 2.3 A resolution in the form of acyl-enzyme complexes bound to serine 62. On the basis of the positions of these inhibitors, the binding of a tripeptide substrate for the enzyme, L-lysyl-D-alanyl-D-alanine, has been modeled in the active site. The binding of both inhibitors and substrate is facilitated by hydrogen-bonding interactions with a conserved beta-strand (297-303), which is antiparallel to the beta-lactam's acylamide linkage or the substrate's peptide bond. The active site is similar to that in beta-lactamases.     

Antibiotic Sensitivities of Organisms Isolated from Mastitic and Nonmastitic Mammary Secretions

Antibiotic sensitivity tests to determine the minimal inhibitory concentrations (MIC) and the minimal lethal concentrations (MLC) for six antibiotics were conducted against mastitic isolates of staphylococci in skim milk and broth. The MIC and the MLC for all the antibiotics except benzylpenicillin were considerably higher in skim milk than in broth. Benzylpenicillin was the most effective antibiotic tested in either medium, and dihydrostreptomycin was the least effective against the organisms tested.     

The activity of the dinuclear cobalt-beta-lactamase from Bacillus cereus in catalysing the hydrolysis of beta-lactams

Metallo-beta-lactamases are native zinc enzymes that catalyse the hydrolysis of beta-lactam antibiotics, but are also able to function with cobalt(II) and require one or two metal-ions for catalytic activity. The hydrolysis of cefoxitin, cephaloridine and benzylpenicillin catalysed by CoBcII (cobalt-substituted beta-lactamase from Bacillus cereus) has been studied at different pHs and metal-ion concentrations. An enzyme group of pK(a) 6.52+/-0.1 is found to be required in its deprotonated form for metal-ion binding and catalysis. The species that results from the loss of one cobalt ion from the enzyme has no significant catalytic activity and is thought to be the mononuclear CoBcII. It appears that dinuclear CoBcII is the active form of the enzyme necessary for turnover, while the mononuclear CoBcII is only involved in substrate binding. The cobalt-substituted enzyme is a more efficient catalyst than the native enzyme for the hydrolysis of some beta-lactam antibiotics suggesting that the role of the metal-ion is predominantly to provide the nucleophilic hydroxide, rather than to act as a Lewis acid to polarize the carbonyl group and stabilize the oxyanion tetrahedral intermediate.     

Use of paper indicator discs for determining the minimal inhibitory concentration of antibiotics]

Sensitivity of clinical strains of Staphylococcus and some Enterobacteriaceae to a number of widely used antibiotics was compared simultaneously with the use of two methods, i. e. the agar diffusion method and the method of serial dilutions. Regularities in distribution of the staphylococcal strains according to their sensitivity to antibiotics, such as erythromycin, benzylpenicillin, levomycetin and others were also studied with respect to every year using indicator paper discs. Interrelation observed during the comparison of the microbial sensitivity with the use of the two assay methods provided elaboration of the criteria for classification of the strains as "resistant" or "sensitive". The differentiation boarder for these two groups was determined according to the principle of the assay error minimization. A necessity of using standard dry media for specification of individual characteristics of various drugs in estimation of the microbial sensitivity to them by the agar diffusion method is emphasized.     

Influence of antibiotics (benzylpenicillin,pharmazin, and nystatin) on the number of microorganisms in ordinary chernozem

In model experiments, the influence of pharmaceutical antibiotics (benzylpenicillin, pharmazin, and nystatin) in different doses (100, 300, 450, and 600 mg/kg) on a number of microorganisms of the chernozem region has been studied. All the studied doses of antibiotics had reliable and overwhelming impacts on the amount of soil microorganisms. The time between a dose of antibiotics and a change in the number of microorganisms in the soil is shown to have a linear dependence. The studied groups of soil microorganisms in relation to pharmaceutical antibiotics showed a hierarchy of resistance (high concentration): Azotobacter > amylolytic bacteria > ammonifying bacteria > micromycetes.     

Procedures for the Assay of Carbenicillin in Body Fluids

The assay of carbenicillin in clinical specimens is complicated by the fact that carbenicillin also contains a small amount of benzylpenicillin, thereby precluding the use of conventional penicillin assay organisms. This report gives details of a microbiological assay method involving the use of a strain of Pseudomonas aeruginosa which is very sensitive to carbenicillin but insensitive to benzylpenicillin. The outline of a microassay method with this organism is presented, and a method for the assay of specimens containing mixtures of carbenicillin and other antibiotics is described.