Dependence of equilibrium and kinetic parameters of erythromycin: a sorption on the structural characteristics of the biosorbent

The use of special synthetic sorbents with surface carboxyl groups allowed us to increase fourfold the effective diffusion coefficient for erythromycin. The maximum sorption capacity for the antibiotic exceeded twofold that observed in experiments with a macroporous sulfocation exchanger MN-500. The sorption of the antibiotic was completely reversible upon treatment with a combined eluant that competitively interacts with the sorbent and does not impair the structural characteristics of erythromycin.     

Molecularly imprinted hydrophilic polymer sorbents for selective sorption of erythromycin

New hydrophilic polymer sorbents comprising reactionary sites which are complementary to a molecule of antibiotic erythromycin were synthesized by the method of molecular imprinting. A series of similar sorbents without reactionary sites was used for comparison of sorption characteristics. Sorption of erythromycin on both types of polymer sorbents synthesized was studied in a wide range of pH and ionic strength. Selectivity of erythromycin sorption on molecularly imprinted cross-linked polymers was shown to depend on the specific interaction of target molecule with polymer matrix. This type of sorbent is perspective for the development of antibiotic purification directly from a culture medium Saccharopolyspora erythreus.     

Enhancement of the efficacy of erythromycin in multiple antibiotic-resistant gram-negative bacterial pathogens

Aims:  To improve the efficacy of erythromycin, a hydrophobic antibiotic, against multiple antibiotic-resistant gram-negative bacterial pathogens by enhancing their outer membrane permeability.
Methods and Results:  Fifty-one nonrepeat gram-negative bacterial pathogens of various genera, resistant to multiple antibiotics, including erythromycin, were selected by disc agar diffusion tests. The amphiphilic cationic steroid antibiotic, Ceragenin CSA-13, a potent permeabilizer of bacterial outer membranes, reduced the minimum inhibitory concentration of erythromycin in 92% of the bacterial pathogens selected for the test, when supplemented with erythromycin. A synergistic effect of Ceragenin CSA-13 and erythromycin in combination was also observed. Spectrofluorimetric study confirmed that Ceragenin CSA-13 acts by depolarizing the bacterial outer membrane. The toxicity of Ceragenin CSA-13 was evaluated to be insignificant by measuring 'median lethal dose' (LD₅₀) on mouse model.
Conclusions:  Ceragenin CSA-13 may be useful as an agent to make erythromycin effective against infections caused by multiple antibiotic resistant gram-negative bacteria.
Significance and Impact of the Study:  The outcome of the study suggests erythromycin–Ceragenin combination as a new approach to overcome the problem associated with the rapid emergence of multi-drug-resistant pathogens. The insignificant toxicity of Ceragenin CSA-13, as found, supports the possibility of the application of this compound for human therapeutics.     

Identification of a Saccharopolyspora erythraea gene required for the final hydroxylation step in erythromycin biosynthesis.

In analyzing the region of the Saccharopolyspora erythraea chromosome responsible for the biosynthesis of the macrolide antibiotic erythromycin, we identified a gene, designated eryK, located about 50 kb downstream of the erythromycin resistance gene, ermE. eryK encodes a 44-kDa protein which, on the basis of comparative analysis, belongs to the P450 monooxygenase family. An S. erythraea strain disrupted in eryK no longer produced erythromycin A but accumulated the B and D forms of the antibiotic, indicating that eryK is responsible for the C-12 hydroxylation of the macrolactone ring, one of the last steps in erythromycin biosynthesis.     

Comparative pharmacokinetics of erythromycin target cell-associated transport and intravenous administration in patients with pneumonia]

The method of cell-associated antibiotic therapy consists of extracorporal exposure of the autoblood formed elements to antibiotic solution followed by their reinfusion. Pharmacokinetics of erythromycin after its intravenous and cell-associated administration in patients with community-acquired pneumonia and the clinical efficacy of the method were evaluated. HPLC of the erythromycin pharmacokinetic pattern in 20 patients showed that after the antibiotic target transport the pharmacokinetic model changed from one-compartment to two-compartment one and the antibiotic maximum concentration and elimination rate were lower vs. the intravenous administration. It was also shown that the clinical efficacies of the erythromycin intravenous administration and target transport did not significantly differ, whereas after the cell-associated transport of the antibiotic the therapeutic effect was observed earlier and the side effects were less frequent.     

Accumulation in gram-postive and gram-negative bacteria as a mechanism of resistance to erythromycin

Erythromycin was recovered in high yield after incubation with gram-negative bacteria. The cell-free protein-synthesizing preparation from gram-negative bacteria is equally as susceptible to the antibiotic as is that from gram-positive bacteria. Thus, neither destruction of erythromycin nor the absence of the step susceptible to the antibiotic plays an important role in the resistance mechanism of gram-negative bacteria. A 100-fold difference in accumulation of erythromycin between gram-positive and gram-negative bacteria was observed. This alone explains the resistance of gram-negative bacteria to erythromycin. Furthermore, data showed that the inhibition of growth is closely related to the accumulation of erythromycin. The concentration of intracellular erythromycin in gram-positive bacteria was found to be 44- to 90-fold greater than that of the extracellular medium. However, the antibiotic did not accumulate on the cell walls, nor was the accumulation energy-dependent. It is proposed that it takes place by the binding of erythromycin to the bacterial ribosomes, forming a very stable complex. The dissociation constants of erythromycin-Staphylococcus aureus complex and erythromycin-Bacillus subtilis complex were determined to be 1.1 x 10(-7) and 3.4 x 11(-7)m, respectively.     

Erythromycin penetration into the cerebrospinal fluid of patients]

Permeability of erythromycin through the barrier of blood-cerebrospinal fluid in neurosurgical patients after its oral administration in a dose of 300-500 mg and intravenous administration in a dose of 200 mg was studied. The erythromycin was determined after the antibiotic single administration at intervals of 40 minutes to 6 hours. A total of 31 observations were performed. Low penetration of erythromycin into the cerebrospinal fluid of the patients was shown. The administration route (oral or intravenous) practically had no effect on the antibiotic penetration level into the subarachnoidal spaces. The highest liquor levels were observed within the period of 3 to 6 hours after the drug administration. The maximum index of penetration from the blood into the cerebrospinal fluid was about 10 per cent. The erythromycin penetration increased in cases with inflammatory changes in the meninges.     

Mathematical model for the control and standardization of discs]

Analysis of tetracycline and erythromycin diagnostic discs manufactured in the USSR showed their conformity with the requirements of the USA Federal Register. Comparison of the antibiotic amounts extracting and diffusing from the discs showed that as an average 92 and 90 per cent of the extracted amounts of erythromycin and tetracycline respectively diffused into the agar. Subsequently, it is desirable that on control testing of the disc quality both the similarity of the antibiotic content in the discs and the real amount of the antibiotic diffusing into the agar should be considered. A statistically reliable correlation between the values of the growth inhibition zones around the discs with definite and constant amounts of erythromycin, tetracycline or oxacillin and different resistance levels for every staphylococcal strain was found. On the basis of such a control system it is possible to divide the staphyloccal strains into the groups with high, low and intermediate resistance levels to the above antibiotics. However, it is not possible to use such data for accurate calculation of the value of the minimum inhibitory concentration of unknown strains because of a high value of the main error.     

Antibiotic Resistance Among Cultured Bacterial Isolates from Bioethanol Fermentation Facilities Across the United States

Bacterial contamination of fuel ethanol fermentations by lactic acid bacteria (LAB) can have crippling effects on bioethanol production. Producers have had success controlling bacterial growth through prophylactic addition of antibiotics to fermentors, yet concerns have arisen about antibiotic resistance among the LAB. Here, we report on mechanisms used by 32 LAB isolates from eight different US bioethanol facilities to persist under conditions of antibiotic stress. Minimum inhibitory concentration assays with penicillin, erythromycin, and virginiamycin revealed broad resistance to each of the antibiotics as well as high levels of resistance to individual antibiotics. Phenotypic assays revealed that antibiotic inactivation mechanisms contributed to the high levels of individual resistances among the isolates, especially to erythromycin and virginiamycin, yet none of the isolates appeared to use a β-lactamase. Biofilm formation was noted among the majority of the isolates and may contribute to persistence under low levels of antibiotics. Nearly all of the isolates carried at least one canonical antibiotic resistance gene and many carried more than one. The erythromycin ribosomal methyltransferase (erm) gene class was found in 19 of 32 isolates, yet a number of these isolates exhibit little to no resistance to erythromycin. The erm genes were present in 15 isolates that encoded more than one antibiotic resistance mechanism, suggestive of potential genetic linkages.     

On the Origin of Mitochondrial Mutants: Evidence for Intracellular Selection of Mitochondria in the Origin of Antibiotic-Resistant Cells in Yeast

In wild-type Saccharomyces cerevisiae, erythromycin and certain other antibacterial antibiotics inhibit the formation of respiratory enzymes in mitochondria by inhibiting translation on mitochondrial ribosomes. This paper is concerned with the origin of mutant cells, resistant to erythromycin by virtue of having a homogeneous population of mutant mitochondrial DNA molecules. Such mutant cells are obtained by plating wild-type (sensitive) cells on a nonfermentable substrate plus the antibiotic. Colonies of mutant cells appear first about four days after the time of appearance of established mutant cells; new colonies continue to appear, often at a constant rate, for many days. Application of the Newcombe respreading experiment demonstrates that most or all of the mutant cells which form the resistant colonies on selective medium arise only after exposure of the population to erythromycin. It is suggested that this result is most probably due to intracellular selection for mitochondrial genomes. Resistant mitochondria arising from spontaneous mutation are postulated to be at a selective disadvantage in the absence of erythromycin; reproducing more slowly than wild-type sensitive mitochondria, they cannot easily accumulate in sufficient numbers in a cell to render it resistant as a whole. In the presence of erythromycin, resistant mitochondria can continue to reproduce while sensitive mitochondria cannot, until there is a sufficient number to make the cell resistant, i.e. to permit normal cell growth. The same phenomenon is seen with respect to chloramphenicol resistance. Intracellular selection is considered more likely than direct induction of mutation by the antibiotic, since mutant cells do not accumulate in the presence of erythromycin if the mitochondrial genome is rendered non-essential by growth on glucose or nontranslatable by chloramphenicol. Intra-cellular selection provides a mechanism for direct adaptation at the cell level, compatible with currently acceptable ideas of spontaneous mutation and selection at the organelle level.     

Dependence of erythromycin biosynthesis on the active acidity of the medium]

Dependence of erythromycin biosynthesis on the medium active acidity was studied by the following methods: by changing pH of the initial medium, by changing the concentration of the medium components determining the active acidity of the culture, by using buffer mixtures by automatic control of pH. It was found that pH of the initial medium within 5.7-8.1 had no effect on the culture growth. Biosynthesis of erythromycin markedly decreased at pH 6.3 or lower. The values of pH within 6.6-7.5 (optimal values 6.7-6.9) were favourable for the antibiotic biosynthesis. At pH 6.2-6.3 the antibiotic accumulation was equal to 5-10 per cent of the control.     

In vitro activity of kaempferol isolated from the Impatiens balsamina alone and in combination with erythromycin or clindamycin against Propionibacterium acnes

The in vitro antibacterial activity against antibiotic-resistant Propionibacterium acnes of kaempferol isolated from the Impatiens balsamina alone and in combination with erythromycin or clindamycin antibiotics was investigated. The antibiotic combination effect against antibiotic-resistant P. acnes was studied by checkerboard test. Kaempferol and quercetin demonstrated antibacterial activities against P. acnes. Minimum inhibitory concentrations (MICs) for both compounds were < or =32 ug/ml and < or =64 ug/ml for clindamycin-sensitive and-resistant P. acnes, respectively. The four combination formulations (kaempferol and either erythromycin or clindamycin; quercetin and either erythromycin or clindamycin) exhibited a synergic inhibition of P. acnes growth. The combination of kaempferol with quercetin showed an indifferent effect. The combination of clindamycin with kaempferol or quercetin showed a greater synergic effect than that of erythromycin with kaempferol or quercetin. Thus, these combinations demonstrated the potential to treat acne.     

The interaction between carbamazepine and erythromycin

Erythromycin has been reported to interact with the anticonvulsant, carbamazepine, in both children and adults. Toxic serum levels of carbamazepine are observed within 24 h of antibiotic administration, suggesting a mechanism not previously described for other erythromycin-based drug interactions. In rats erythromycin significantly depressed the elimination of carbamazepine in animals induced with carbamazepine for 4 days but had no effect on carbamazepine elimination in noninduced animals. Although the in vitro metabolism of carbamazepine to its epoxide by hepatic microsomes prepared from noninduced rats was significantly inhibited by erythromycin, the inhibition of carbamazepine epoxidation was greatly enhanced in carbamazepine-induced rats. In the pig the sensitivity of carbamazepine metabolism to erythromycin was much greater than in the rat, indicating the existence of a large species difference in this particular drug interaction. It is concluded that the interaction between erythromycin and carbamazepine is caused by a direct inhibition of carbamazepine oxidation by the antibiotic.     

Erythromycin block of the HERG K+ channel: accessibility to F656 and Y652

The HERG potassium channel might have a non-canonical drug binding site, distinct from the channel's inner cavity, that could be responsible for elements of closed-state pharmacological inhibition of the channel. The macrolide antibiotic erythromycin is a drug that may block unconventionally because of its size. Here we used whole-cell patch-clamp recording at 37 degrees C from heterologously expressed HERG channels in a mammalian cell line to show that erythromycin either produces a rapid open-state-dependent HERG channel inhibition, or components of both open-state-dependent and closed-state-dependent inhibition. Alanine-substitution of HERG's canonical determinants of blockade revealed that Y652 was not important as a molecular determinant of blockade, and that mutation of F656 resulted in only weak attenuation of inhibition. In computer models of the channel, erythromycin could make several direct contacts with F656, but not with Y652, in the open-state model, and erythromycin was unable to fit into a closed-state channel model.     

Genetic analysis of erythromycin production in Streptomyces erythreus

Streptomyces erythreus produces the 14-membered macrolide antibiotic erythromycin A. The properties of erythromycin A nonproducing mutants and their genetic linkage to chromosomal markers were used to establish the rudiments of genetic organization of antibiotic production. Thirty-three Ery- mutants, produced by mutagenesis of S. erythreus NRRL 2338 and affecting the formation of the macrolactone and deoxysugar intermediates of erythromycin A biosynthesis, were classified into four phenotypically different groups based on their cosynthesis behavior, the type of biosynthetic intermediate accumulated, and their ability to biotransform known biochemical intermediates of erythromycin A. Demonstration of the occurrence of natural genetic recombination during conjugal mating in S. erythreus enabled comparison of the genetic linkage relationships of three different ery mutations with seven other markers on a simple chromosome map. This established a chromosomal location for the ery mutations, which appear to be located in at least two positions within one interval of the map.     

Transduction transmission of the extrachromosomal markers of antibiotic resistance in staphylococcal populations]

Transduction of extrachromosomal markers of resistance to penicillin and erythromycin in staphylococcal strains isolated from patients was studied. Two transduction methods were compared, i. e. transduction with a phage filtrate of the donor culture resistant to erythromycin and transduction on mixed cultivation of the donor and recipient. A higher transduction rate was observed with the latter method. Mixed cultivation of the donor cultures resistant to penicillin or erythromycin and the recipient strains of the wild type sensitive to these antibiotic resulted in transduction of the respective markers. Transductants which acquired prophage 6 simultaneously with marker Egg became donors of erythromycin resistance.     

Infectivity and sensitivity to antibiotics of Rickettsia prowazekii mutants of the low-pathogenic strain E cultured in chick embryos

Selection of mutants of a low pathogenic strain E of R. prowazekii is a trend in genetic investigation of this Rickettsia species and one of the approaches to stabilizing the strain avirulent properties with a purpose of using in vaccine prophylaxis of typhus. The mutants of R. prowazekii, strain E selected by the authors earlier were characterized with respect to their infective capacity for chick embryos (CE) and antibiotic sensitivity. It was found that the infective capacity for CE of the erythromycin resistant mutant induced by nitroso guanidine (EErrI) was by ID50 2-3 logarithms lower than that of the initial strain E. The infective capacity for CE of the rifampicin resistant mutant induced by nitroso guanidine (ERifrI) and the spontaneous erythromycin resistant mutant was similar to that of strain E. The ERifrI strain differed from the initial strain E by higher sensitivity to tetracycline and erythromycin and the EErrI strain differed from the initial strain E by higher sensitivity to tetracycline and rifampicin. It was shown that the biological properties of the nitroso guanidine-induced mutants resistant to rifampicin and erythromycin differed from those of the initial strain E and the properties of the spontaneous erythromycin resistant mutant were similar to those of the initial strain E.     

Pharmacological availability of erythromycin granules for children's use

Pharmaceutical availability of erythromycin granules with polymeric coating of different composition+ was studied. With an account of the ++anatomo-physiological features of a child organism and the properties of the antibiotic, acetylphthalyl cellulose in combination with hydroxypropyl methylcellulose or methyl cellulose was used as a film forming agent. The coated granules were estimated by such parameters as the time of disintegration and the rate of dissolution in various media. The results of the study showed that coating of the erythromycin granules with the film composed of acetylphthalyl cellulose and hydroxypropyl methylcellulose in the ratio of 8 to 2 provided the required protection of the antibiotic in acid media and high pharmaceutical availability of the drug.     

Biosynthesis of the anti-parasitic agent megalomicin: transformation of erythromycin to megalomicin in Saccharopolyspora erythraea

Megalomicin is a therapeutically diverse compound which possesses antiparasitic, antiviral and antibacterial properties. It is produced by Micromonospora megalomicea and differs from the well-known macrolide antibiotic erythromycin by the addition of a unique deoxyamino sugar, megosamine, to the C-6 hydroxyl. We have cloned and sequenced a 48 kb segment of the megalomicin (meg) biosynthetic gene cluster which contains the modular polyketide synthase (PKS) and the complete pathway for megosamine biosynthesis. The similarities and distinctions between the related megalomicin and erythromycin gene clusters are discussed. Heterologous expression of the megalomicin PKS in Streptomyces lividans led to production of 6-deoxyerythronolide B, the same macrolactone intermediate for erythromycin. A 12 kb fragment harbouring the putative megosamine pathway was expressed in Saccharopolyspora erythraea, resulting in the conversion of erythromycin to megalomicin. Considering the extensive knowledge surrounding the genetic engineering of the erythromycin PKS and the familiarity with genetic manipulation and fermentation of S. erythraea, the ability to produce megalomicin in this strain should allow the engineering of novel megalomicin analogues with potentially improved therapeutic activities.