Electrooptical parameters of kanamycin-treated E. coli cell suspensions

The effect of kanamycin on the electrophysical parameters of cell suspensions of Escherichia coli K-12 and pMMB33 was investigated. Incubation of the sensitive K-12 strain with kanamycin resulted in significant changes in the orientation spectra (OS) of the cell suspensions; these changes were not revealed in the case of the resistant pMMB33 strain. In the case of the sensitive K-12 strain incubated with different kanamycin concentrations, changes in the OS of the cell suspensions occurred within the 10-1000 kHz frequency range of the orienting electrical field. The most pronounced change in the electrooptical signal was observed at 10 microg/ml of kanamycin. Control experiments were carried out by standard plating on nutrient media. Thus, the OS changes of suspensions in the presence of antibiotics may be used as a test for microbial resistance to such antibiotics.     

Changes in electrophysical properties of Escherichia coli K-12 cells under the action of kanamycin and tetracycline

Essential changes in the orientation spectra (OS) of cell suspensions incubated at different concentrations of kanamycin, were found to occur only in the first 5 frequencies of the orienting electrical field (10-1,000 kHz)). The maximum change in the intensity of the electrooptical signal occurred at a concentration of kanamycin of 10 microg/ml. Antibiotic concentration of 5 microg/ml caused no changes in OS. During the incubation of the cells with tetracycline (1.7, 2.5, 5.0 microg/ml) no changes in OS of the cell suspension were registered. Considerable changes in the intensity of the electrooptical signal occurred during the incubation of the cells with kanamycin (5 microg/ml) and tetracycline (1.7 microg/ml) simultaneously, which was due to the synergic action of these two antibiotics. Thus, as found with the use of the electrooptical analysis, the joint action of kanamycin and tetracycline could increase their antibacterial effect. The results demonstrated the effectiveness of using electrophysical methods for the study of the mechanism of action of antibiotics on bacterial cells and for the control of such action.     

The Electro-Optical Investigation of Suspensions of Escherichia coli K-12 Cells Metabolizing Glucose,Lactose, and Galactose

The electro-optical characteristics of suspensions of Escherichia coli K-12 cells metabolizing glucose, lactose, and galactose were studied by measuring the suspension turbidity as a function of cell alignment in an orienting electric field whose frequency was varied from 10 kHz to 10 MHz. In a frequency range of 10 kHz to 1 MHz, the orientational spectra of E. coli K-12 cells grown on glucose and lactose considerably changed after their incubation in the presence of the sugars. These changes likely reflect alterations in the polarizability of the cells induced by sugar metabolism.     

Electrooptic analysis of Escherichia coli cell suspension for assessment of antibacterial activity of levomycetin

Influence of chloramphenicol on electrophysiologic charateristics of Escherichia coli strains susceptible (K-12 strain) and resistant (pBR-325 strain) to it has been studied. It has been shown that incubation of susceptible bacteria with chloramphenicol leads to significant change of magnitute of electrooptic (EO) signal. Significant changes in orientantional spectra of suspensions of susceptible to chloramphenicol cells incubated with different concentrations of antibiotic were observed only on first five frequencies of orienting electric field (10 - 1000 kHz). Maximal change of EO signal occurred at chloramphenicol concentration 35 mg/ml and it didn't depend on the time of antibiotic exposure. Incubation of resistant strain pBR-325 with chloramphenicol did not lead to change of EO parameters of cell suspension. Potential for use of electrophysical analytic methods for assessment of antibacterial activity of chloramphenicol to control effect of antibiotics on microorganisms has been proposed.     

Exchange of colicin receptor capacity between strains of Escherichia coli sensitive or resistant to colicin K-K235

Phage and colicin-resistant mutants were derived from Escherichia coli K-12P678. Two classes of phage T6 and colicin K-resistant mutants (genotype tsx) were isolated. Tsx-2 mutants, which demonstrated mucoid growth and increased sensitivities to many antibiotics, became sensitive to colicin K when pretreated with ethylenediaminetetraacetate (EDTA), whereas Tsx-1 mutants did not. Reassociation of EDTA-released material partially restored resistance to colicin K for Tsx-2 mutants. When EDTA-released material from strain P678 was associated with either class of K-resistant mutant, an increase in colicin K sensitivity resulted. Observations suggest that colicin K can act on its target site once it penetrates the cell surface. In addition, results suggest that functional colicin K receptors can be transferred from sensitive to resistant strains, thus conferring colicin sensitivity.Non-standard Abbreviations SDS sodium dodecyl sulfate     

The electrooptical parameters of suspensions of <Emphasis Type="Italic">Escherichia coli</Emphasis> XL-1 cells interacting with helper phage M13K07

The electrophysical properties of Escherichia coli XL-1 cells interacting with helper phage M13K07 were studied as a function of the phage-to-cell ratio and the contact time. The electro-optical signal of bacterial cells changed considerably as soon as 10 min after the onset of their incubation with phage particles, presumably due to phage adsorption on the cell surface. The maximum changes in the orientational spectra of cell suspensions were observed when the phage-to-cell ratio was 20. Selectivity studies showed that E. coli XL-1 cells interacting with the helper phage M13K07 in the presence of foreign microflora, such as E. coli K-12 or Azospirillum brasilense Sp7, can be identified by using their electrophysical properties. Changes in the orientational spectra of cell suspensions are interpreted with the stage of phage-bacterium interaction taken into account. The results obtained can probably be used to devise a new rapid method for identification of microorganisms and to study the particular stages of cell infection by bacteriophages.__________Translated from Mikrobiologiya, Vol. 74, No. 2, 2005, pp. 198–203.Original Russian Text Copyright © 2005 by Bunin, Ignatov, Guliy, Zaitseva, ONeil, Ivnitskii     

The electro-optical investigation of suspensions of Escherichia coli K-12 cells metabolizing glucose,lactose, and galactose

The electro-optical characteristics of suspensions of Escherichia coli K-12 cells metabolizing glucose, lactose, and galactose were studied my measuring the suspension turbidity as a function of cell alignment in an orienting electric field whose frequency was varied from 10 kHz to 10 MHz. In a frequency range of 10 kHz to 1 MHz, the orientational spectra of E. coli K-12 cells grown on glucose and lactose considerably changed after their incubation in the presence of the sugars. These changes likely reflect alterations in the polarizability of the cells induced by sugar metabolism.     

Involvement of O8-antigen in altering β-lactam antibiotic susceptibilities in Escherichia coli

In spite of being dispensable, O-antigens are believed to facilitate various cellular processes and alter antibiotic sensitivities. Escherichia coli K-12 (CS109) strains are lacking in O-antigens and are reported to be sensitive to antibiotics. To our surprise, E. coli 2443 (expressing O8-antigen) manifested two- to fourfold higher sensitivities toward penicillin and its derivatives than strain CS109. However, sensitivities toward other structurally unrelated antibiotics remained unchanged. To understand the rationale behind such observations, we replaced the rfb locus of strain 2443 with that of E. coli K-12. The β-lactam sensitivities of 2443 cells with replaced rfb locus appeared to be identical to those for CS109. Therefore, it is quite reasonable to hypothesize the possible involvement of O8-antigen in β-lactam sensitization.     

Coresistance to Neomycin and Kanamycin by Mutations in an Escherichia coli Locus that Affects Ribosomes

Mutant strains resistant to neomycin or to kanamycin sulfate were isolated from Escherichia coli K-12. Nine mutants were analyzed; all were resistant to both antibiotics (about 150 and 100 mug/ml, respectively), and were designated nek. In the mutant strains, the ribosomes are changed from those of the parental strain; for when they were used in assays for polypeptide formation directed by polyadenylic acid or polycytidylic acid, coding fidelity in presence of the drugs was increased and inhibition of synthesis by the drugs was lessened. Mating experiments and transduction tests showed that all of the nine nek mutants are either closely linked or allelic, and the nek locus is closely linked to two genes-str (streptomycin) and spc (spectinomycin)-known to affect the 30S ribosome. The two nek mutants tested were recessive to the sensitive, wild-type allele. When the nek mutants were compared to the parental strain, pleiotropic effects of the nek mutations were observed. Resistance to low levels of streptomycin and spectinomycin was increased, whereas resistance to chloramphenicol was decreased. Also, the mutants were less able to adapt to high concentrations of lincomycin, and could no longer show phenotypic suppression of an arginine requirement by neomycin or kanamycin. Such pleiotropic effects are suggested to be the rule for mutations in genes that participate in the biosynthesis of a cellular organelle.     

In vitro selection of a glyphosate-tolerant sugarcane cellular line

Cell suspensions derived fromSaccharum spp. varieties (PR62258, V64-10, V71-51) were used to study the effects of glyphosate on cell suspensions of commercial cultivars of sugarcane in order to select and develop a glyphosate-tolerant cellular line. A liquid medium with 0.8 mM/L glyphosate was used for selection. The results showed growth inhibition of approximately 50% for cell suspensions in mediums with 0.8 mM/L glyphosate. The results obtained in vitro were consistent with the results reported from in vivo tests. Plants were regenerated from sensitive cell suspensions and tolerant cell suspensions from cultivar V71-51. Tolerant cell suspensions tolerated a concentration that was 12.5-fold higher than that of the sensitive cell suspensions. Plants regenerated from tolerant cell suspensions tolerated a glyphosate concentration that was 6-fold higher than that of the plants regenerated from sensitive cell suspensions. Cell suspensions derived from tolerant regenerated plants tolerated a glyphosate concentration that was 5-fold higher than that of those derived from sensitive regenerated plants. The RAPD patterns obtained with OPA-07 revealed a 564-bp band that can be used to characterize the tolerant cellular line. This band is not present in the sensitive cellular line. Consequently, the tolerance persisted throughout the differentiation into plants and dedifferentiation into cell suspensions. This performance suggests that an in vitro selection of pre-existing variability has taken place.     

<Emphasis Type="Italic">Escherichia coli</Emphasis> Membrane-Associated Energy-Dependent Processes and Sensitivity Toward Antibiotics Changes as Responses to Low-Intensity Electromagnetic Irradiation of 70.6 and 73 GHz Frequencies

Escherichia coli K-12(λ) was sensitive toward low-intensity (non-thermal, flux capacity 0.06 mW cm⁻²) electromagnetic irradiation (EMI) of extremely high frequency—70.6 and 73 GHz. 1 h exposure to EMI markedly depressed growth and cell viability of bacteria. Membrane-associated processes—total H⁺ efflux and H₂ evaluation by whole cells during glucose fermentation were shown to be lowered as well. At the same time, the F₀F₁-ATPase activity of membrane vesicles was little depressed with 70.6 GHz irradiation only. This finding was in conformity with non-changed N,N′-dicyclohexylcarbodiimide-sensitive H⁺ efflux. Furthermore, for understanding the different frequencies action mechanisms, the effects of antibiotics (chloramphenicol, ceftriaxone, kanamycin, and tetracycline) on irradiated cells growth and survival were determined. EMI with the frequencies of 70.6 and 73 GHz as with 51.8 and 53.0 GHz enhanced the sensitivity of bacteria toward antibiotics, but comparison revealed that each frequency had a different portion. Probably, EMI of specific frequency triggered changes in biological processes and afterward in growth and viability of bacteria, creating conditions when the action of antibiotics became facilitated.     

Dependence of Escherichia coli hyperbaric oxygen toxicity on the lipid acyl chain composition.

This study examines certain membrane-related aspects of oxygen poisoning in Escherichia coli K1060 (fabB fadE lacI) and its parent strain, K-12 Ymel. Cells were grown to exponential or stationary phase in a minimal medium and exposed to air plus 300 lb/in2 of O2 as a suspension in minimal salts. After an initial lag, both strains lost viability with apparent first-order kinetics. Hypebaric oxygen was more toxic to cells harvested during the exponential phase of growth than to cells harvested from the stationary phase of growth for both strains K-12 Ymel and K1060. Control suspensions exposed to air plus 300 lb/in2 of N2 did not lose viability during a 96-h exposure. The sensitivity of the unsaturated fatty acid auxotroph, strain K1060, to hyperbaric oxygen increased as the degree of unsaturation of the fatty acid supplement increased. Cells grown with a cyclopropane fatty acid (9,10=methylenoctadecanoate) were the most resistant; cells grown with a monounsaturated fatty acid (oleate) were intermediate; and those grown with polyunsaturated fatty acids (linoleate and linolenate) were most sensitive to hyperbaric oxygen. The parent strain, K-12 Ymel, lost viability in hyperbaric oxygen most similarly to strain K1060 supplemented with oleate. To determine the relative effect of hyperbaric oxygen on the survival of E. coli with saturated membranes, substrains of K1060 were selected for growth on 12-methyltetrade-canoate or on 9 or 10-monobromostearate. Substrains grown with a saturated fatty acid supplement were equally or more sensitive to hyperbaric oxygen than when the same substrains were grown with a cyclopropane fatty acid supplement. The lipid acyl chain composition was determined in E. coli K1060 before and after exposure to hyperbaric oxygen or hyperbaric nitrogen. The proportion of nonsaturated acyl chain lipid of either the oleate- or the 9,10-methyleneoctade-canoate-supplemented K1060 remained unchanged after hyperbaric gas exposure. In strain K1060 supplemented with linoleate and grown to stationary phase, however, the relative unsaturated acyl chain content after hyperbaric exposure decreased in both gases. This finding prompted an investigation of the role of lipid oxidation in hyperbaric oxygen toxicity. Assays of potential lipid oxidation products were performed with linoleate-grown cells. The lipid hydroperoxide and peroxide content of the lipid extract increased by 6.9 times after 48 h of air plus 300 lb/in2 of O2; malondialdehyde and fluorescent complex lipid oxidation products showed much smaller or no changes. Lipid extracts from hyperbaric oxygen-exposed cells were not toxic to viable E. coli K1060, nor did they increase the rate of loss of viability in cells simultaneously exposed to hyperbaric oxygen. Linoleic acid hydroperoxide at 1.0 mM had no effect on the viability of E. coli K-12 Ymel and only marginally decreased the viability of E. coli K1060 supplemented with linoleate. We conclude that the kinetics of oxygen toxicity in E...     

On the interaction between heparin and some aminoglycoside antibiotics

An interaction between the aminoglycoside antibiotics and heparin wherein charge transfer complexes are formed has been investigated to determine the degree of inhibition of antibacterial function of the antibiotic in the complexed form.Minimum inhibitory concentration (MIC) values have been obtained for the action of the aminoclycoside antibiotics tobramycin, gentamicin, amikacin, kanamycin, and streptomycin, on a sensitive strain ofE. coli. Growth curves ofE. coli determined at concentrations of these antibiotics just below the MIC demonstrated significant lengthening of the lag phase relative to control growth curves generated in the absence of antibiotic. Heparin (1 U ml^–1 and 10 U ml^–1) had no effect on control growth curves; however, particularly at the higher concentration, it reduced the effect on the lag phase produced by the aminoglycoside antibiotics. Thus kanamycin, gentamicin, and tobramycin were most affected, while amikacin and streptomycin were least affected. The rank order of inhibition of antibiotic activity by interaction with heparin was in qualitative agreement with previously published figures for the degree of complexation between antibiotics and heparin.     

Comparative study of Streptomyces--producer of aminoglycoside antibiotics, monomycin and kanamycin, and the strain 344 obtained by fusion of their protoplasts by the method of restrictive total DNA fingerprinting]

The method of total DNA restriction finger prints was applied to the study of Streptomyces monomycini INA 1465 producing monomycin, Streptomyces kanamyceticus INA K-13 producing kanamycin and strain 344 isolated after fusion of the protoplasts of strain 1465 and K-13, which produced albofungin and chloralbofungin, aminoglycoside antibiotics. For preparing the finger prints of the strains splitting by endonucleases BamHI, PstI, PvuII, and BgIII was used. The finger prints showed that strain 344 was related to the strain of S. monomycini and markedly differed from the strain of S. kanamyceticus. Strain 344 was likely to result from reconstruction (probably 20-kb deletion) of the genome of S. monomycini INA 1465 induced by the preparation and regeneration of its protoplasts. The reconstruction could affect the genome area with localization of the genes involved in monomycin biosynthesis and monomycin resistance genes.     

The effect of ampicillin on the electrophysical properties of Escherichia coli cells

The study of the effect of ampicillin on the electrophysical properties of Escherichia coli cells showed that this antibiotic influences the orientational spectra (OS) of the ampicillin-susceptible E. coli strains K-12 and XL-1 within the frequency range 10-1000 kHz of the orienting electric field and does not affect the OS of the ampicillin-resistant strains K-12(pUC-18) and XL-1(pHEN1). The change in the electrooptical signal of the ampicillin-susceptible cells was maximum at an ampicillin concentration of 50 microg/ml and did not depend on the exposure time. The conclusion is drawn that changes in the OS of cells can be used to evaluate their resistance to ampicillin.     

Antibiotic Control of Mycoplasma in Tissue Culture

Seven of eight strains of Mycoplasma (PPLO) were found to be sensitive to the deoxystreptamines, certain macrolides, and the tetracyclines. These antibiotics are relative noncytotoxic. Kanamycin and tetracycline were useful in eliminating PPLO (pleuropneumonia-like organisms) strain Squibb no. 1 from a HeLa cell line which was deliberately contaminated with PPLO. Repeated exposure of M. laidlawii type B cells to neomycin resulted in a 50-fold increase in resistance, and the resistant strain was also resistant to gentamicin, kanamycin, neomycin, and paromomycin. A tetracycline-resistant strain of this culture was found to be resistant to 7-chlortetracycline, 7-chlor-6-demethyltetracycline, and 5-hydroxytetracycline. One PPLO strain, Squibb no. 2, derived from a contaminated HeLa cell culture, was resistant to all antibiotics studied.     

长江江豚杀鲑气单胞菌的分离鉴定及生物学特性分析

本文通过细菌的分离培养,首次从临床症状表现为溃疡、腐烂的患病江豚表皮分离到一株杀鲑气单胞菌XJ-JT株。通过细菌理化性质鉴定、遗传进化分析、药敏试验、致病性试验对其生物学特性进行分析,结果表明：菌株XJ-JT为革兰氏阴性短杆菌,两端钝圆,无芽孢;细菌分离鉴定的结果显示分离菌株为杀鲑气单胞菌;系统进化分析揭示其基因序列与银鲫源性杀鲑气单胞菌分离株高度同源;20种抗生素的药敏试验结果表明分离菌株对阿米卡星、克拉霉素、克林霉素等8种药物敏感,对头孢噻肟、头孢曲松、卡那霉素中介,对阿莫西林、氨苄西林、四环素等9种药物耐药;人工感染试验,结果显示分离菌对鲫鱼有较强的致病性。     

Exocellular Cyclic Dipeptides from a <Emphasis Type="Italic">Ruegeria</Emphasis> Strain Associated with Cell Cultures of <Emphasis Type="Italic">Suberites domuncula</Emphasis>

From cell cultures of Suberites domuncula was isolated a bacterial strain, SDC-1, which was identified by 16S ribosomal RNA sequence analysis as an -Proteobacterium of the genus Ruegeria. The occurrence of the strain in sponge cell culture could be explained by its resistance to the antibiotics used in the isolation of sponge cell cultures or by the preservation of SDC-1 by host sponge cells. The fatty acid composition of SDC-1 is characterized by branched C-12 methyl fatty acids. Two new and 8 known cyclic dipeptides were isolated and characterized from the fermentation broth of SDC-1. Cyclodipeptides are one of the families of cell-cell signaling compounds and may have some role to play in sponge-bacteria interactions.     

Involvement of O8-antigen in altering beta-lactam antibiotic susceptibilities in Escherichia coli.

In spite of being dispensable, O-antigens are believed to facilitate various cellular processes and alter antibiotic sensitivities. Escherichia coli K-12 (CS109) strains are lacking in O-antigens and are reported to be sensitive to antibiotics. To our surprise, E. coli 2443 (expressing O8-antigen) manifested two- to fourfold higher sensitivities toward penicillin and its derivatives than strain CS109. However, sensitivities toward other structurally unrelated antibiotics remained unchanged. To understand the rationale behind such observations, we replaced the rfb locus of strain 2443 with that of E. coli K-12. The beta-lactam sensitivities of 2443 cells with replaced rfb locus appeared to be identical to those for CS109. Therefore, it is quite reasonable to hypothesize the possible involvement of O8-antigen in beta-lactam sensitization.     

High and selective resistance to mecillinam in adenylate cyclase-deficient or cyclic adenosine 3'',5''-monophosphate receptor protein-deficient mutants of Escherichia coli.

Adenylate cyclase-deficient (cya) mutants of Escherichia coli K-12 were selectively and highly resistant to mecillinam (FL1060) among several beta-lactam antibiotics in the absence of cyclic adenosine 3',5'-monophosphate (cAMP). They became sensitive to the drug in the presence of cAMP. Also, cAMP receptor protein-negative (crp) mutants, with the exception of strain 5333, were highly resistant to mecillinam in the presence and in the absence of cAMP. Mecillinam exerted two distinct and sequential effects in both cya+ strains and cya strains supplemented with cAMP: (i) rounding of cells and (ii) cessation of cell division. The first effect was accompanied by a decrease in growth rate, whereas the second effect was accompanied by enlargement and lysis of the rounded cells. The second effect of mecillinam was dependent on inoculum size and cAMP. When the cell density was above about 10(6) cells per ml, the rounded cells stopped dividing but did not lyse. In the absence of cAMP, cya strains neither stopped dividing nor lysed; they were resistant to the second, lethal effect of mecillinam.