We studied the impact of a sublethal concentration of erythromycin on the fitness and proteome of a continuously cultivated population of Escherichia coli. The development of resistance to erythromycin in the population was followed over time by the gradient plate method and minimum inhibitory concentration (MIC) measurements. We measured the growth rate, standardized efficiency of synthesis of radiolabeled proteins, and translation accuracy of the system. The proteome changes were followed over time in two parallel experiments that differed in the presence or absence of erythromycin. A comparison of the proteomes at each time point (43, 68, and 103 h) revealed a group of unique proteins differing in expression. From all 35 proteins differing throughout the cultivation, only three were common to more than one time point. In the final population, a significant proportion of upregulated proteins was localized to the outer or inner cytoplasmic membranes or to the periplasmic space. In a population growing for more than 100 generations in the presence of antibiotic, erythromycin-resistant bacterial clones with improved fitness in comparison to early resistant culture predominated. This phenomenon was accompanied by distinct changes in protein expression during a stepwise, population-based development of erythromycin resistance.

• Klíčová slova
• Continuous cultivation system, Escherichia coli, erythromycin, fitness, proteome, resistance,
• MeSH
• antibakteriální látky farmakologie   MeSH
• bakteriální léková rezistence genetika   MeSH
• časové faktory MeSH
• erythromycin farmakologie   MeSH
• Escherichia coli účinky léků   genetika   metabolismus   MeSH
• genetická zdatnost účinky léků   MeSH
• kultivační média MeSH
• mikrobiální testy citlivosti MeSH
• proteiny z Escherichia coli genetika   metabolismus   MeSH
• proteom genetika   metabolismus   MeSH
• regulace genové exprese u bakterií * MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• antibakteriální látky MeSH
• erythromycin MeSH
• kultivační média MeSH
• proteiny z Escherichia coli MeSH
• proteom MeSH

Thirty-five Staphylococcus aureus strains from auricular infections were isolated. The identification of strains was confirmed by Api ID 32 Staph strips, the antibiotic susceptibility test was performed using ATB Staph kit. PCR assay was used to detect the oxacillin resistance gene (mecA) and the erythromycin genes (ermA, ermB, ermC, msrA and mef). The susceptibility profile of all strains revealed a low resistance level to oxacillin and erythromycin. The PCR results show that 60 % of the strains are mecA positive. The frequency of erythromycin genes was: ermA (+) 22.8 %, ermB (+) 45.7, ermC (+) 17.1, msrA (+) 28.6. The mef gene was not detected in any strain. No correlations between genotypic and phenotypic methods for the determination of oxacillin and erythromycin resistance was found. However, multiplex PCR technique was shown to be a fast, practical and economic technique for the detection of methicillin-and erythromycin-resistant staphylococci.

• MeSH
• antibakteriální látky farmakologie   MeSH
• bakteriální léková rezistence * MeSH
• bakteriální proteiny genetika   metabolismus   MeSH
• erythromycin farmakologie   MeSH
• lidé MeSH
• membránové proteiny genetika   metabolismus   MeSH
• methyltransferasy genetika   metabolismus   MeSH
• oxacilin farmakologie   MeSH
• polymerázová řetězová reakce metody   MeSH
• proteiny vázající penicilin MeSH
• stafylokokové infekce mikrobiologie   MeSH
• Staphylococcus aureus účinky léků   genetika   izolace a purifikace   MeSH
• ušní boltec mikrobiologie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• hodnotící studie MeSH
• Názvy látek
• antibakteriální látky MeSH
• bakteriální proteiny MeSH
• ErmA protein, Bacteria MeSH Prohlížeč
• erythromycin MeSH
• mecA protein, Staphylococcus aureus MeSH Prohlížeč
• MefA protein, Streptococcus MeSH Prohlížeč
• membránové proteiny MeSH
• methyltransferasy MeSH
• oxacilin MeSH
• proteiny vázající penicilin MeSH

We found a new variant of the streptogramin A resistance gene, vga(A)LC, in clinical isolates of Staphylococcus haemolyticus resistant to lincomycin and clindamycin but susceptible to erythromycin and in which no relevant lincosamide resistance gene was detected. The gene vga(A)LC, differing from the gene vga(A) at the protein level by seven amino acid substitutions, was present exclusively in S. haemolyticus strains resistant to both lincosamides and streptogramin A (LS(A) phenotype). Antibiotic resistance profiles of the ATP-binding cassette (ABC) proteins Vga(A)(LC) and Vga(A) in the antibiotic-susceptible host S. aureus RN4220 were compared. It was shown that Vga(A)LC conferred resistance to both lincosamides and streptogramin A, while Vga(A) conferred significant resistance to streptogramin A only. Detailed analysis of the seven amino acid substitutions, distinguishing the two related ABC proteins with different substrate specificities, identified the substrate-recognizing site: four clustered substitutions (L212S, G219V, A220T, and G226S) in the spacer between the two ATP-binding cassettes altered the substrate specificity and constituted the lincosamide-streptogramin A resistance phenotype. A transport experiment with radiolabeled lincomycin demonstrated that the mechanism of lincosamide resistance in S. haemolyticus was identical to that of the reported macrolide-streptogramin B resistance conferred by Msr(A).

• MeSH
• bakteriální geny MeSH
• bakteriální proteiny chemie   genetika   metabolismus   fyziologie   MeSH
• DNA bakterií genetika   izolace a purifikace   MeSH
• genetická variace * MeSH
• lidé MeSH
• linkosamidy MeSH
• makrolidy farmakologie   MeSH
• mikrobiální testy citlivosti MeSH
• mnohočetná bakteriální léková rezistence MeSH
• molekulární evoluce * MeSH
• sekvence aminokyselin MeSH
• sekvence nukleotidů MeSH
• sekvenční analýza DNA MeSH
• Staphylococcus aureus účinky léků   genetika   MeSH
• Staphylococcus haemolyticus účinky léků   genetika   izolace a purifikace   MeSH
• streptogramin A farmakologie   MeSH
• substituce aminokyselin MeSH
• substrátová specifita MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH
• srovnávací studie MeSH
• Názvy látek
• bakteriální proteiny MeSH
• DNA bakterií MeSH
• linkosamidy MeSH
• makrolidy MeSH
• streptogramin A MeSH

A rapid and simple microdilution technique on 96-well microplate based on turbidimetry was optimized and validated for screening of antimicrobial activity against erythromycin-resistant bacterial strains of Streptococcus pyogenes and Staphylococcus simulans isolated from Finnish patients. Using S. pyogenes ATCC 12351 as reference strain the developed method was evaluated by reproducibility measurements and using parameters typically employed for screening methods, i.e. signal-to-background, signal-to-noise and a screening-window coefficient, the Z' factor. The method was further used for screening a group of natural compounds and their synthetic derivatives against resistant bacterial strains. Of these, octyl and dodecyl gallates, and usnic and ursolic acids were the most active. The described method is a rapid, homogeneous, cost-effective and easy-to-perform system for screening of new potential antimicrobial agents in drug discovery.

• MeSH
• antibakteriální látky chemie   farmakologie   MeSH
• bakteriální léková rezistence * MeSH
• časové faktory MeSH
• erythromycin farmakologie   MeSH
• flavonoidy farmakologie   MeSH
• hydroxybenzoáty farmakologie   MeSH
• kumariny farmakologie   MeSH
• kyselina gallová analogy a deriváty   farmakologie   MeSH
• lidé MeSH
• mikrobiální testy citlivosti metody   normy   MeSH
• nefelometrie a turbidimetrie MeSH
• penicilin G farmakologie   MeSH
• Staphylococcus účinky léků   růst a vývoj   MeSH
• Streptococcus pyogenes účinky léků   růst a vývoj   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• hodnotící studie MeSH
• práce podpořená grantem MeSH
• Geografické názvy
• Finsko MeSH
• Názvy látek
• antibakteriální látky MeSH
• erythromycin MeSH
• flavonoidy MeSH
• hydroxybenzoáty MeSH
• kumariny MeSH
• kyselina gallová MeSH
• lauryl gallate MeSH Prohlížeč
• octyl gallate MeSH Prohlížeč
• penicilin G MeSH
• phenolic acid MeSH Prohlížeč

Erm methyltransferases mediate the resistance to the macrolide-lincosamide-streptogramin B antibiotics via dimethylation of a specific adenine residue in 23S rRNA. The role of positively charged N-terminal residues of the ErmC' methyltransferase in RNA binding and/or catalysis was determined. Mutational analysis of amino acids K4 and K7 was performed and the mutants were characterized in in vivo and in vitro experiments. The K4 and K7 residues were suggested not to be essential for the enzyme activity but to provide a considerable support for the catalytic step of the reaction, probably by maintaining the optimum conformation of the transition state through interactions with the phosphate backbone of RNA.

• MeSH
• aminokyseliny bazické genetika   MeSH
• antibakteriální látky farmakologie   MeSH
• Bacillus subtilis enzymologie   genetika   MeSH
• bakteriální léková rezistence genetika   MeSH
• bakteriální RNA metabolismus   MeSH
• DNA bakterií genetika   fyziologie   MeSH
• erythromycin farmakologie   MeSH
• konzervovaná sekvence MeSH
• methyltransferasy genetika   izolace a purifikace   metabolismus   fyziologie   MeSH
• mikrobiální testy citlivosti MeSH
• molekulární sekvence - údaje MeSH
• mutageneze cílená MeSH
• oligoribonukleotidy metabolismus   MeSH
• posunová mutace MeSH
• RNA ribozomální 23S metabolismus   MeSH
• S-adenosylmethionin metabolismus   MeSH
• sekundární struktura proteinů MeSH
• sekvence aminokyselin MeSH
• sekvenční seřazení MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• aminokyseliny bazické MeSH
• antibakteriální látky MeSH
• bakteriální RNA MeSH
• DNA bakterií MeSH
• erythromycin MeSH
• methyltransferasy MeSH
• oligoribonukleotidy MeSH
• RNA ribozomální 23S MeSH
• rRNA (adenosine-O-2'-)methyltransferase MeSH Prohlížeč
• S-adenosylmethionin MeSH