Alteration in the electrophoretic mobility of OmpC due to variations in the ammonium persulfate concentration in sodium dodecyl sulfate-polyacrylamide gel electrophoresis.

Studies on Salmonella typhi and Salmonella typhimurium outer membrane proteins have shown that the relative position of OmpC porin in sodium dodecyl sulfate.polyacrylamide gel electrophoresis undergoes an important shift when the concentration of ammonium persulfate in the running gel is increased from 6 to 12 mM. The apparent molecular mass at these concentrations was estimated to be 34 and 40 kDa, respectively. Under similar electrophoretic conditions the apparent molecular mass estimated for OmpF was 37.6 and 38.2 kDa. Therefore, OmpC moves from a leading position to a position behind OmpF. For Escherichia coli OmpC the shift observed is less pronounced than that occurring in Salmonellae.

Epidemias por salmonella y por shigella, importancia de la vigilancia epidemiologíca y control de la resistencia a los antibióticos / Epidemics by salmonella and shigella, importance of epidemiologic vigilance and control of the antibiotics resistence

El descubrimiento de los antibióticos y su aplicación clínica ha permitido el tratamiento y control de padecimientos infecciosos. Por otra parte, se ha descubierto que algunas enterobacterias desarrollan resistencia a las drogas antimocrobianas causada, principalmente por la presencia de elementos extracromosómicos denominados plasmidos. De entre estos microorganismos se han estudiado la salmonella y shigella con la finalidad de determinar las características de las mismas, los antibióticos a que se resisten y con cuales pueden ser tratados los padecimientos provocados por las mismas. La presencia de epidemias y las endemias en regiones de México así como en otras regiones del mundo, y consecuentemente el alto índice de morbilidad han inducido a los especialistas a investigar los procesos infecciosos provocados por shigella y la salmonella, se ha logrado establecer una clasificación de las mismas así como la sintomatología que producen las mismas. Para establecer un tratamiento determinado, se aisló al agente infeccioso, se le sometió a estudios para poder establecer a que drogas eran resistentes y a cuales no. De este modo se ha podido descrubir que algunos tipos de estas bacterias son resistentes, por ejemplo, al clorafenicol, la tetraciclina, la estreptomicina y a la ampicilina, en tanto que otras son suceptibles a los mismos, así mismo se ha observado que la región guarda cierta relación con el desarrollo de la resistencia

Heterogeneity of lipopolysaccharide phenotype among Salmonella typhi strains.

Comparison of the Vi antigen, lipopolysaccharide, and protein components of the cell surface of three strains of Salmonella typhi showed that differences in lipopolysaccharide contributed most to distinctions in serum survival, whereas differences in Vi antigen content had no apparent effect.

[Elaboration of an immunosorbent for the purification of porins from Salmonella typhi 9, 12, Vi:d]. / Elaboración de un inmunoadsorbente para la purificación de porinas de Salmonella typhi 9, 12, Vi: d.

The current work was undertaken to purify porins of Salmonella typhi, which are outer membrane proteins (OMPs) that induce protection in mice against challenge with the bacteria in mucin. OMPs, isolated with a non-ionic detergent, had a 4% contamination with LPS (endotoxin) and molecular sizes ranging from 17 to 70 KDa. Porins (Mw 38-41 KDa) were isolated from OMPs preparative SDS-PAGE. Anti-porins antisera were raised in rabbits and specific IgG was purified, which was coupled to Sepharose-CNBr. This immunosorbent was used to purify LPS-free porins.

Immunoblot analysis of Salmonella typhi lipopolysaccharide (LPS) using typhoid sera.

Lipopolysaccharide (LPS) of Salmonella typhi has been analyzed by immunoblotting with pooled sera from typhoid patients. Pooled typhoid sera have recognized all the antigenic determinants of S. typhi LPS, giving a strong reaction with the repeating units on the O-side chains as well as with the core region. Cross-reacting antigens have been observed with the LPS of S. typhimurium and S. enteritidis, while no heterologous reactions were seen with the LPS of E. coli strains.

Native and chemically modified porin channels from Salmonella typhi Ty2 in planar lipid bilayers.

Native porins, from Salmonella typhi Ty2 outer membrane, and porins alkylated with pyridoxal phosphate (Plp) were studied in planar lipid bilayers. The conductance of bilayers exposed to native or chemically modified porins increases in discrete jumps. Conductance histograms for native porins displayed two major peaks at 1.7 and 6.7 nS (in 0.5 M KCl). On the other hand, Plp-treated porins exhibited a single major peak at 1 nS. The relation between bilayer conductance and native porin concentration was linear. However, this relation became logarithmic in the presence of modified porins. The results support the notion that alkaline reduction of S. typhi Ty2 porins with Plp dissociates porin channel trimers in a reversible fashion.

Effect of bacterial endotoxin on the transmembrane electrical potential and plasma membrane fluidity of human monocytes.

In order to gain insight into the physical interaction between bacterial endotoxins and the surface of human monocytes, we investigated the effects of Salmonella typhi endotoxin and lipid A on two functional properties of the plasma membrane of these cells: (1) the transmembrane electrical potential and (2) the fluidity of the lipid bilayer. Using the fluorescent lipophilic cationic probe 3,3'-dipropylthiodicarbocyanine (di-S-C3(5] to monitor the transmembrane electrical potential, we found that neither endotoxin nor lipid A induced depolarization of the monocyte's plasma membrane or impeded its ability to undergo depolarization in response to phorbol myristate acetate. When the resting transmembrane potential of the monocyte was analyzed by exposing di-S-C3(5)-labeled cells suspended in media containing incremental concentrations of potassium ion (K+) to valinomycin, no difference between the response of control cells and cells pretreated with endotoxin was noted. We next examined the effect of endotoxin and lipid A on the fluidity of the monocyte's plasma membrane by monitoring the intensity of the fluorescence of 1,6-diphenyl-1,3,5-hexatriene. By quantifying the intensity of parallel and perpendicular polarized light emitted by this membrane-embedded probe between 8 and 56 degrees C, measurements of molecular anisotropy were used to identify temperature-dependent phase transitions within the hydrocarbon region of the plasma membrane and to estimate the relative microviscosity of the lipid bilayer before and after exposing the cells to endotoxin or lipid A. Although the temperature at which phase transitions occurred was the same in all experimental groups of cells, preincubation of monocytes with either endotoxin or lipid A appeared to increase both the apparent microviscosity of the cell membrane and the order of the lipid bilayer as reflected by a decrease in its flow-activation energy. Our data indicate that when endotoxin molecules contact the surface of the monocyte, the lipid A moiety appears to become incorporated into the plasma membrane, increasing the microviscosity of the lipid bilayer without significantly altering its ionic permeability. We therefore conclude that the metabolic activation of monocytes by endotoxin is not coupled to, or initiated by, membrane depolarization.

The hemolytic effect of Salmonella typhi Ty 2 porins.

Two outer membrane proteins of Salmonella typhi Ty 2 were extensively co-purified. According to their migration in dodecylsulfate/polyacrylamide gel electrophoresis and solubility characteristics, these proteins are homologous to the 35-kDa and 36-kDa porins found in Salmonella typhimurium. A porin homologous to the 34-kDa one has not been found in S. typhi Ty 2. A critical step in the purification of porins is heating at 100 degrees C in 2% sodium dodecyl sulfate before Sephadex gel filtration. The absence of detergent in aqueous suspensions enhances porin aggregation, these aggregations inducing human red cell lysis. Porins obtained by an alternative procedure consisting of heating at 60 degrees C instead of 100 degrees C were also hemolytic. Using nanomolar concentration of porins a strong influence of temperature on the hemolytic effect was observed. Porin-induced hemolysis was inhibited with anti-porin serum, as well as by a treatment with phenylglyoxal, which reacts with the arginine residues of proteins. The membrane-disrupting ability of porins aggregates might explain some pathogenic characteristics of gram-negative bacterial infections.

Surveillance for typhoid fever in Matsuyama city during 1974-1981 and detection of Salmonella typhi in sewage and river waters.

Eighty-two cases of typhoid fever were found in Matsuyama city in the period from 1974 to 1981. Seventy-six cases were found to be infected with Salmonella typhi other three with Salmonella paratyphi A, and the remaining three were diagnosed only clinically. The strains of S. typhi isolated from these patients showed such a variety of Vi-phage types as D1, D2, E1, M1, 53 and degraded Vi-positive strain (DVS). The concurrent survey of the city sewage and river waters for typhoid bacilli was conducted with total 578 samples taken therefrom. S. typhi was isolated from 120 of those samples. The Vi-phage types of the isolates were closely related with those of the isolates from the patients. The periodical examinations of the city sewage and the draining river may serve as a useful means for the controlling typhoid fever epidemics.

[Potential use of infrared spectroscopy for the quantitative characterization of the chemical composition of Salmonella typhi preparations]. / Vozmozhnost' primeneniia infrakrasnoi spektroskopii dlia kolichestvennoi kharakteristiki khimicheskogo sostava preparatov Salmonella typhi.

The possibility of using infrared spectroscopy for the evaluation of the chemical composition of S. typhi preparations is shown. The existence of a clear-cut correlation between the optical density of adsorption bands in the region between 1,665 and 1,100 cm-1 and the content of protein, polysaccharide and nucleic acids in the preparations subjected to analysis has been revealed. The character of changes in the infrared spectra of the preparations at different stages of deep cultivation has been studied.

Diversity of Vi-related antigens in the microcapsule of Salmonella typhi.

Two new antigenic acidic polysaccharides, Vi-P and Vi-C, were isolated from the microcapsule of Salmonella typhi by a very mild procedure. The antigens were purified to serological homogeneity, and it was found that each of them shares with Vi antigen one of its two serological determinants (a different one in each case). One of the antigens Vi-C, was also isolated from Escherichia coli B, which is not a Vi-producing strain.

[Effect of bacterial polysaccharide on the primary and secondary immunologic response following immunization with sheep erythrocytes]. / Deistvie bakterial'nogo polisakharida na pervichnyi i vtorichnyi immunologicheskii otvet pri immunizatsi éritrotsitami barana.

The effect of typhoid bacterial polysaccharide on the primary and secondary immune response to SRBS was studied. The polysaccharide was shown to have both stimulating and depressive effect on the population of antibody-producing cells. This effect depended on the time and the number of polysaccharide injections. Thus, a single polysaccharide injection made on the day preceding immunization resulted in the maximum stimulation in the system of IgM- and IgG-producing cells, while the maximum depression of these cells could be observed after 2 polysaccharide injections: on the day preceding immunization and on the day of immunization. In the secondary immune response considerable stimulation of the populations of antibody-producing cells was observed after polysaccharide injections made on days 2 and 3 after reimmunization.

Studies on the lipid components of endotoxin. II. Chemical analyses and biological properties of neutral, polar-I and polar-II subfractions.

Lipid components obtained from Salmonella typhosa O-901 endotoxin by acid hydrolysis were separated into neutral, polar-I and polar-II lipid fractions by silica gel column chromatography. These lipids were further separated by silica gel column and/or thin-layer chromatography. The subfractions were analyzed by thin-layer chromatography, gas chromatography and infrared spectrophotometry. Seven subfractions obtained from the neutral lipid fraction contained lauric, myristic, palmitic, 3-OH-myristic acid, artificial products of 3-OH-myristic acid, or a small amount of two unidentified fatty acids. These fatty acids and glucosamine were commonly detected in six subfractions obtained from the polar-I lipid fraction. Fatty acids, glucosamine, and O-phosphorylethanolamine were detected in all of the 13 subfractions obtained from the polar-II lipid fraction. Chick embryo lethal activity, rabbit pyrogenicity and in vitro interferon inducing activity were found in three polar-I lipid subfractions and five polar-II lipid subfractions, but not in neutral lipids. The activities were highest in a polar-II lipid subfraction which contained smaller amounts of O-phosphorylethanolamine and glucosamine than the other subfractions. However, no particular chemical constituent(s) related to the biological activities could be found. Prolonged acid hydrolysis of the polar-II lipids gave rise to neutral and polar-I lipids. Chemical and biological aspects of the lipid constituents of endotoxin are discussed.

[Trace element content in comparison with the physiological and immunogenic properties of S. typhi obtained by various continuous cultivation regimens]. / Soderzhanie mikroélementov v sopostavlenii s fiziologicheskimi i immunogennymi svoistvami S. typhi, poluchennykh pri razlichnykh rezhimakh nepreryvnogo kul'tivirovaniia.

The content of trace elements in the S. typhi cells was studied during continuous cultivation in the synthetic nutrient medium. Growing under conditions of complete or incomplete glucose utilization failed to influence the content of trace elements in these cells. The same tendencies were revealed in the changes of manganese, iron, magnesium, calcium, and strontium content with increase of the growth rate. The content of aluminium in the cells altered in parallel with the changes in the immunogenic properties with increase of the growth rate from 0.17 to 0.44 hour-1.

Tolerance and other biological properties of Vibrio parahaemolyticus endotoxins.

Biological properties of endotoxins prepared from three strains of Vibrio parahaemolyticus were compared with reference to commercially prepared Salmonella typhi endotoxin. Endotoxin assays performed in rabbits included dermal Shwartzman reactivity, pyrogenicity, heat stability, and ability to induce tolerance as well as cross-tolerance. Mice were used for endotoxin LD50 determinations. Results showed V. parahaemolyticus endotoxins were similar to that of S. typhi strain O901. Induction of tolerance to V. parahaemolyticus strain 11590 endotoxin resulted in complete cross-tolerance to S. typhi endotoxin, and vice versa. Partial cross-tolerance to S. typhi endotoxin was demonstrated with rabbits rendered tolerant to endotoxin from V. parahaemolyticus strains Sak-3 and FC1011. Absorption spectra, nitrogen, phosphorus and carbohydrate analyses revealed additional similarities between endotoxins from V. parahaemolyticus and endotoxin from a member of the Enterbacteriaceae.

[Polyamines in microorganisms sensitive and resistant to antibiotics]. / Poliaminy u mikroorganizmov, chuvstvitel'nykh i ustoichivykh k antibiotikam

Intracellular contents of polyamine dancyl derivatives were determined by the method of thin-layer chromatography in Coli and typhoid fever bacteria sensitive to antibiotics and their variants resistant to streptomycin, levomycetin, chlortetracyline and neomycin. Increased levels of spermidine and putrescin in the antibiotic resistant cultures were found. The increased levels of the polyamines correlated with changes in the content of nucleic acids. It is supposed that the increase in the content of the polyamines in the antibiotic resistant bacteria was the result of accumulation of nucleic acids by them.

The molecular relatedness of R factors in enterobacteria of human and animal origin.

The molecular length and DNA homology of R factors isolated from enterobacteria of human and animal origin have been examined. DNA from plasmids of the same compatibility group, whether of human or animal origin is indistinguishable, after allowance has been made for the regions coding for different antibiotic resistances. These results indicate that there is a common pool of R factors in man and ainmals.