RESUMO
Siderophores selectively bind ferric iron and are involved in receptor-specific iron transport into bacteria. Several types of siderophores were synthesized, and growth-promoting or inhibitory activities when they were conjugated to carbacephalosporin, erythromycylamine, or nalidixic acid were investigated. Overall, 11 types of siderophores and 21 drug conjugates were tested against seven different bacterial species: Escherichia coli, Bordetella bronchiseptica, Pasteurella multocida, Pasteurella haemolytica, Streptococcus suis, Staphylococcus aureus, and Staphylococcus epidermidis. In some species, the inhibitory activities of the drug conjugates were associated with the ability of the bacteria to use the siderophore portion of the molecules for growth promotion in disc diffusion tests (0.04 mumol of conjugate or siderophore per disc). E. coli used catechol-based siderophore portions as well as hydroxamate-based tri-delta-OH-N-OH-delta-N-acetyl-L-ornithine ferric iron ligands for growth under iron-restricted conditions achieved by supplemental ethylenediamine di (O-hydroxyphenylacetic acid) (100 micrograms/ml) and was sensitive to carbacephalosporin conjugated to these siderophore types (up to a 34-mm-diameter inhibition zone). B. bronchiseptica used desferrioxamine B and an isocyanurate-based or trihydroxamate in addition to catechol-based siderophore portions for promotion but was not inhibited by beta-lactam conjugates partly because of the presence of beta-lactamase. P. multocida and P. haemolytica did not use any of the synthetic siderophores for growth promotion, and the inhibitory activities of some conjugates seemed partly linked to their ability to withhold iron from these bacteria, since individual siderophore portions showed some antibacterial effects. Individual siderophores did not promote S. suis growth in restrictive conditions, but the type of ferric iron ligands attached to beta-lactams affected inhibitory activities. The antibacterial activities of the intracellular-acting agents erythromycylamine and nalidixic acid were reduced or lost, even against S. aureus and S. epidermidis, when the agents were conjugated to siderophores. Conjugate-resistant E. coli mutants showed the absence of some iron-regulated outer membrane proteins in gel electrophoresis profiles and in specific phage or colicin sensitivity tests, implying that the drugs used outer membrane receptors of ferric complexes to get into cells.
Assuntos
Antibacterianos/metabolismo , Bactérias/metabolismo , Ferro/metabolismo , Sideróforos/metabolismo , Doenças dos Suínos/microbiologia , Animais , Antibacterianos/química , Bactérias/genética , Proteínas da Membrana Bacteriana Externa/metabolismo , Bordetella/genética , Bordetella/crescimento & desenvolvimento , Bordetella/metabolismo , Colicinas/farmacologia , Resistência Microbiana a Medicamentos , Escherichia coli/efeitos dos fármacos , Escherichia coli/genética , Ácidos Hidroxâmicos/farmacologia , Testes de Sensibilidade Microbiana , Mutação , Pasteurella/genética , Pasteurella/crescimento & desenvolvimento , Pasteurella/metabolismo , Sideróforos/química , Streptococcus suis/genética , Streptococcus suis/crescimento & desenvolvimento , Streptococcus suis/metabolismo , SuínosRESUMO
Bacteria from the subcutaneous abscesses which appeared in a laboratory colony of DS mouse since October of 1977 were identified as Pasteurella pneumotropica by various biological examinations. The abscess formation was limited to multiparous female mice over 100 day-age, but virgin females were free from the disease. The MIC of various antibacterial substances showed that potassium tellurite, kanamycin and bacitracin were effective to isolate the organism selectively from various infection sites harboring many other species of bacteria. A novel NKBT medium was prepared by adding these antibacterial substances to the heart infusion agar (HIA) supplemented by 10% Fildes digested blood. A fluid culture medium, TGN broth was prepared for multiplication of the organism by adding 10% Fildes digested blood and potassium tellurite to GN broth. To isolate the organism from the pharyngo-larynx a direct application of mucus wiped off the infection site onto the culture medium was sufficient, but pre-multiplication in the TGN broth was required for isolation of the organism from gut contents before inoculation onto the NKBT medium. The pre-cultivation in the TGN broth vastly improved the recovery of the organism especially from feces. Thereby we could easily detect the latent infection of this bacterium without sacrificing animals.
Assuntos
Meios de Cultura , Pasteurella/crescimento & desenvolvimento , Abscesso/patologia , Animais , Anticorpos Antibacterianos/análise , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Testes de Sensibilidade Microbiana , Pasteurella/efeitos dos fármacos , Pasteurella/isolamento & purificação , Infecções por Pasteurella/patologiaRESUMO
Sterile, concentrated culture supernatant from Pasteurella haemolytica (biotype A, serotype 1) strain 630 was subjected to physical, chemical, and immunologic treatments to determine their influence on leukotoxin (cytotoxin) activity contained in the supernatant. Each treated sample contained approximately 8 chemiluminescence inhibitory units of leukotoxin. Treatment effects were evaluated for their ability to inactivate leukotoxin activity. Leukotoxin activity in treated samples was determined by inhibition of the luminol-dependent chemiluminescence response of bovine neutrophils. Optimal leukotoxin synthesis by P haemolytica occurred when the bacteria were at the logarithmic growth phase, whereas stationary phase cultures contained minimal amounts of leukotoxin activity in their culture supernatant. Leukotoxin activity was heat labile; activity was substantially decreased when concentrated culture supernatant samples containing leukotoxin activity were incubated at 37 C for several hours. When concentrated culture supernatant was incubated at progressively decreasing temperatures, there was a progressive increase in the length of time that the leukotoxin retained its biologic activity. Samples stored at -70 C retained activity for at least 2 months. Leukotoxin activity was nondialyzable and was able to withstand considerable extremes in hydrogen ion concentration. Leukotoxin activity could not be pelleted when subjected to forces of 100,000 X g for 1 hour. Chemical and enzymatic studies suggested that P haemolytica leukotoxin contained carbohydrate and protein moieties. Chemical treatment with 0.2% sodium lauryl sulfate, 0.5% sodium deoxycholate, 7.5 mM EDTA and 8M urea with 8 mM 2-mercaptoethanol and enzymatic treatment with lipase, ribonuclease, and deoxyribonuclease had no discernible effect on leukotoxin activity.(ABSTRACT TRUNCATED AT 250 WORDS)
Assuntos
Doenças dos Bovinos/microbiologia , Exotoxinas/toxicidade , Hidrolases/farmacologia , Neutrófilos/fisiologia , Infecções por Pasteurella/veterinária , Pasteurella/crescimento & desenvolvimento , Animais , Bovinos , Exotoxinas/biossíntese , Exotoxinas/isolamento & purificação , Hemólise/efeitos dos fármacos , Concentração de Íons de Hidrogênio , Cinética , Medições Luminescentes , Luminol , Neutrófilos/efeitos dos fármacos , Pasteurella/isolamento & purificação , TemperaturaRESUMO
A previous investigation demonstrated that infusion of an antipyretic drug into the preoptic anterior hypothalamus (PO/AH) of rabbits reduced the fever usually seen during the initial stages of infection. This was followed by an increased fever and an increased mortality rate [32]. The work reported here investigated the hypothesis that the increased mortality was the result of decreased killing and/or increased multiplication of bacteria during the initial, attenuated phase of the febrile course in the antipyretic-treated rabbits. Rabbits were injected intravenously with Pasteurella multocida and either sodium salicylate or a control solution was infused directly into the PO/AH. Infusion of sodium salicylate reduced the mean fever 4 hours after injection of bacteria from 2.07 +/- 0.28 degrees C (S.E.M.) to 0.62 +/- 0.43 degrees C. Rabbits with reduced fevers had decreased blood leucocyte counts and greater numbers of bacteria in lung and liver samples. No differences were seen in reticuloendothelial clearance of carbon, hematocrit, or intracellular viability of bacteria when antipyretics were administered. This increase in bacterial numbers corresponds well to the increased mortality found in previous studies in animals with reduced fevers.