BACKGROUND: Aminopenicillins are recommended agents for non-invasive Haemophilus influenzae infections. One of the mechanisms of resistance to β-lactams is the alteration of the transpeptidase region of penicillin binding protein 3 (PBP3) which is caused by mutations in the ftsI gene. It was shown that exposure to beta-lactams has a stimulating effect on increase of prevalence of H. influenzae strains with the non-enzymatic mechanism of resistance. OBJECTIVES: The aim of our study was to compare the mutational potential of ampicillin and cefuroxime in H. influenzae strains, determination of minimum inhibitory concentration and the evolution of mutations over time, focusing on amino acid substitutions in PBP3. METHODS: 30 days of serial passaging of strains in liquid broth containing increasing concentrations of ampicillin or cefuroxime was followed by whole-genome sequencing. RESULTS: On average, cefuroxime increased the minimum inhibitory concentration more than ampicillin. The minimum inhibitory concentration was increased by a maximum of 32 fold. Substitutions in the PBP3 started to appear after 15 days of passaging. In PBP3, cefuroxime caused different substitutions than ampicillin. CONCLUSIONS: Our experiment observed differences in mutation selection by ampicillin and cefuroxime. Selection pressure of antibiotics in vitro generated substitutions that do not occur in clinical strains in the Czech Republic.
- MeSH
- ampicilin * farmakologie MeSH
- antibakteriální látky * farmakologie MeSH
- bakteriální proteiny genetika metabolismus MeSH
- cefuroxim * farmakologie MeSH
- Haemophilus influenzae * genetika účinky léků MeSH
- hemofilové infekce mikrobiologie MeSH
- lidé MeSH
- mikrobiální testy citlivosti * MeSH
- molekulární evoluce MeSH
- mutace * MeSH
- proteiny vázající penicilin * genetika metabolismus MeSH
- sekvenování celého genomu MeSH
- selekce (genetika) MeSH
- sériové pasážování MeSH
- substituce aminokyselin * MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Equine herpesvirus 1 (EHV-1), like other members of the Alphaherpesvirinae subfamily, is a neurotropic virus causing latent infections in the nervous system of the natural host. In the present study, we have investigated EHV-1 replication (wild-type Jan-E strain and Rac-H laboratory strain) during long-term infection and during the passages of the virus in cultured neurons. The studies were performed on primary murine neurons, which are an excellent in vitro model for studying neurotropism and neurovirulence of EHV-1. Using real-time cell growth analysis, we have demonstrated for the first time that primary murine neurons are able to survive long-term EHV-1 infection. Positive results of real-time PCR test indicated a high level of virus DNA in cultured neurons, and during long-term infection, these neurons were still able to transmit the virus to the other cells. We also compared the neurovirulence of Rac-H and Jan-E EHV-1 strains after multiple passages of these strains in neuron cell culture. The results showed that multiple passages of EHV-1 in neurons lead to the inhibition of viral replication as early as in the third passage. Interestingly, the inhibition of the EHV-1 replication occurred exclusively in neurons, because the equine dermal (ED) cells co-cultivated with neuroculture medium from the third passage showed the presence of large amount of viral DNA. In conclusion, our results showed that certain balance between EHV-1 and neurons has been established during in vitro infection allowing neurons to survive long-term infection.
- MeSH
- herpesvirus 1 koní genetika růst a vývoj patogenita fyziologie MeSH
- herpetické infekce veterinární virologie MeSH
- hostitelská specificita MeSH
- koně MeSH
- kultivované buňky MeSH
- myši inbrední BALB C MeSH
- myši MeSH
- nemoci koní virologie MeSH
- neurony virologie MeSH
- replikace viru MeSH
- sériové pasážování MeSH
- virulence MeSH
- zvířata MeSH
- Check Tag
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
Colonies of Saccharomyces cerevisiae laboratory strains pass through specific developmental phases when growing on solid respiratory medium. During entry into the so-called alkali phase, in which ammonia signaling is initiated, 2 prominent cell types are formed within the colonies: U cells in upper colony regions, which have a longevity phenotype and activate the expression of a large number of metabolic genes, and L cells in lower regions, which die more quickly and exhibit a starvation phenotype. Here, we performed a detailed analysis of the activities of enzymes of central carbon metabolism in lysates of both cell types and determined several fermentation end products, showing that previously reported expression differences are reflected in the different enzymatic capabilities of each cell type. Hence, U cells, despite being grown on respiratory medium, behave as fermenting cells, whereas L cells rely on respiratory metabolism and possess active gluconeogenesis. Using a spectrum of different inhibitors, we showed that glycolysis is essential for the formation, and particularly, the survival of U cells. We also showed that β-1,3-glucans that are released from the cell walls of L cells are the most likely source of carbohydrates for U cells.
- MeSH
- beta-glukany metabolismus MeSH
- buněčná stěna metabolismus MeSH
- časové faktory MeSH
- fenotyp MeSH
- fermentace * účinky léků MeSH
- genotyp MeSH
- glykolýza * účinky léků MeSH
- inhibitory enzymů farmakologie MeSH
- kultivační média chemie metabolismus MeSH
- mikrobiální viabilita MeSH
- mikrobiologické techniky metody MeSH
- počet mikrobiálních kolonií MeSH
- Saccharomyces cerevisiae - proteiny genetika metabolismus MeSH
- Saccharomyces cerevisiae účinky léků enzymologie genetika růst a vývoj MeSH
- sériové pasážování MeSH
- substrátová specifita MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
The growth adaptability to increasing concentration of the biocide 2-phenoxyethanol (PE) was determined in Pseudomonas aeruginosa PAO1 (P.a.) as part of efforts to understand and control the biocide tolerance and its effect on cross-resistance to other biocides and resistance to antibiotics. After repeated subculturing in media containing increasing sub-minimum-inhibitory PE concentration, P.a. exhibited an adaptive resistance indicated by two-fold increase in MIC at the 10th passage. The resistance was stable and remained after passaging the strain in further 7 successive passages in PE-free growth media. The strain showed cross-resistance towards dissimilar biocides and displayed increased susceptibility to antibiotics, which was not influenced by the presence of the efflux inhibitor 'carbonyl cyanide m-chlorophenyl hydrazone'. Outer membranes of adapted strain showed altered protein profile when examined by SDS-PAGE.
- MeSH
- antibakteriální látky farmakologie MeSH
- biologická adaptace MeSH
- dezinficiencia metabolismus toxicita MeSH
- elektroforéza v polyakrylamidovém gelu MeSH
- ethylenglykoly metabolismus toxicita MeSH
- financování organizované MeSH
- mikrobiální testy citlivosti MeSH
- proteiny vnější bakteriální membrány analýza MeSH
- proteom analýza MeSH
- Pseudomonas aeruginosa fyziologie MeSH
- sériové pasážování MeSH
- tolerance léku MeSH
An attenuated strain (263) of the tick-borne encephalitis virus, isolated from field ticks, was either serially subcultured, 5 times in mice, or at 40 degrees C in PS cells, producing 2 independent strains, 263-m5 and 263-TR with identical genomes; both strains exhibited increased plaque size, neuroinvasiveness and temperature-resistance. Sequencing revealed two unique amino acid substitutions, one mapping close to the catalytic site of the viral protease. These observations imply that virus adaptation from ticks to mammals occurs by selection of pre-existing virulent variants from the quasispecies population rather than by the emergence of new random mutations. The significance of these observations is discussed.
- MeSH
- financování organizované MeSH
- genetika MeSH
- klíšťata virologie MeSH
- klíšťová encefalitida mortalita patofyziologie virologie MeSH
- kojená zvířata MeSH
- molekulární modely MeSH
- mozek virologie MeSH
- mutace MeSH
- myši inbrední ICR MeSH
- myši MeSH
- RNA-helikasy genetika chemie MeSH
- sekvence aminokyselin MeSH
- sekvence nukleotidů MeSH
- sekvenční analýza DNA MeSH
- selekce (genetika) MeSH
- serinové endopeptidasy genetika chemie MeSH
- sériové pasážování MeSH
- substituce aminokyselin MeSH
- virové nestrukturální proteiny genetika chemie MeSH
- virulence MeSH
- viry klíšťové encefalitidy genetika izolace a purifikace patogenita MeSH
- zvířata MeSH
- Check Tag
- myši MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Geografické názvy
- Evropa MeSH
