Natural antibodies in Pseudomonas aeruginosa infections

Studying 347 sera of 49 patients suffering from Pseudomonas aeruginosa infection the mean titre value (MTV) calculated from the exponent of the binary logarithm of natural antibody (NA) titres, was found suitable to characterize the humoral immune status. As long as the organism is in equilibrium with the infection, the NA level rises. In septic shock, before death or at the development of a massive infection, the NA titre decreases rapidly. The decrease may be due partly to the permeability increasing effect of endotoxin and antigen-antibody reactions exerted on the capillaries. Consequently, in the most severe phase of sepsis, when bacteria enter the circulation less NA is available to fight against them. This might be a cause of the still very high lethality of septic shock due to Gram-negative bacteria. Finally, when applying the MTV one always has to consider that despite its advantages it gives less information than the Backhausz immunogram.     

Potentiation of vasopressin analgesia in rats treated neonatally with monosodium glutamate

The analgesic response elicited by central administration of arginine vasopressin (AVP) appears to be dependent upon the integrity of the hypothalamic paraventricular nucleus (PVN), since lesions placed in the PVN eliminate AVP analgesia. A projection to the zona externa of the median eminence constitutes one of the VP-containing efferents of the PVN. Neonatal treatment with monosodium glutamate (MSG) destroys perikarya of the arcuate nucleus and median eminence. The present study examined whether AVP analgesia was affected in the MSG-treated rat and whether these alterations were accompanied by specific changes in VP immunoreactivity in the zona externa of the median eminence. Female rats, neonatally treated with either MSG or a saline control, were tested as adults on the tail-flick test following intracerebroventricular injections of 0, 75, 150 and 500 ng doses of AVP. After testing, selected animals were prepared for AVP and oxytocin immunocytochemistry of the median eminence. Significant potentiations in the magnitude of AVP analgesia were observed in MSG-treated rats. AVP and oxytocin immunoreactivity in the zona interna and oxytocin immunoreactivity in the zona externa of the median eminence were similar in MSG-treated and control rats. In contrast, AVP immunoreactivity in the zona externa of the median eminence was markedly reduced in the MSG-treated rat. These data suggest that VP analgesia may normally be inhibited by those medial-basal hypothalamic neurons affected by neonatal MSG treatment.     

Discrimination between the tastes of sucrose and monosodium glutamate in rats

Conditioned taste aversion studies have demonstrated that rats conditioned to avoid monosodium glutamate (MSG) with amiloride added to reduce the intensity of the sodium component of MSG taste, will generalize an aversion for MSG to sucrose and vice versa. This suggests that taste transduction for sodium, sucrose and MSG may intersect at some point. Generalization of conditioned taste aversion indicates that two substances share similar taste features, but it does not reveal the extent of their differences. In this study, we tested how well rats can discriminate sucrose and MSG under a variety of conditions. Water-deprived rats were trained on a combination of water reinforcement and shock avoidance to discriminate between MSG and sucrose, both with and without amiloride, and with and without equimolar NaCl in all solutions. In the absence of amiloride, rats reliably distinguished between MSG and sucrose down to 10 mM solutions. However, they could correctly identify solutions only above 50 mM in the presence of amiloride, equimolar sodium chloride, or both. These results suggest that gustatory stimulation by MSG and sucrose interact somewhere in taste transduction, perhaps within taste receptor cells or gustatory afferent pathways.     

Glutamate taste: Discrimination between the tastes of glutamate agonists and monosodium glutamate in rats

Taste aversion studies have demonstrated that rats conditioned to avoid monosodium glutamate (MSG) with amiloride added to reduce the intensity of the sodium component of MSG taste, generalize this aversion to aspartic acid and to L-AP4, but not to ionotropic glutamate receptor agonists. That is, MSG, L-AP4 and aspartate have similar tastes to rats. However, conditioned taste aversion methods are unable to show to what extent the tastes of two substances are different. If two substances activate the same afferent processes (e.g. taste receptors), they are likely to produce the same tastes, but if they activate different afferent processes, the subject may detect differences between the tastes of the substances. In this study, rats were tested to determine if they could discriminate between the tastes of these agonists and MSG. We also established the detection thresholds for NMDA, aspartic acid and L-AP4, with and without amiloride (a sodium channel antagonist). Taste threshold values were 1-4 mM for NMDA and aspartic acid and 0.5-2.5 microM for L-AP4. None were affected by 30 micro M amiloride. Rats could readily distinguish between the tastes of MSG and NMDA but they had difficulty discriminating between the tastes of aspartic acid and MSG. Rats could also easily distinguish between 10-100 mM MSG and 0.01-5 mM L-AP4. However, in two separate experiments error rates increased significantly when L-AP4 concentrations were between 10-100 mM, indicating that the tastes of L-AP4 and MSG were similar at these concentrations.     

Bacteriophages for the treatment of Pseudomonas aeruginosa infections

Bacteriophages were first identified in 1915 and were used as antimicrobial agents from 1919 onwards. Despite apparent successes and widespread application, early users did not understand the nature of these agents and their efficacy remained controversial. As a result, they were replaced in the west by chemical antibiotics once these became available. However, bacteriophages remained a common therapeutic approach in parts of Eastern Europe where they are still in use. Increasing levels of antibiotic-resistant bacterial infections are now driving demand for novel therapeutic approaches. In cases where antibiotic options are limited or nonexistent, the pressure for new agents is greatest. One of the most prominent areas of concern is multidrug-resistant Gram-negative bacteria. Pseudomonas aeruginosa is a prominent member of this class and is the cause of damaging infections that can be resistant to successful treatment with conventional antibiotics. At the same time, it exhibits a number of properties that make it a suitable target for bacteriophage-based approaches, including growth in biofilms that can hydrolyse following phage infection. Pseudomonas aeruginosa provides a striking example of an infection where clinical need and the availability of a practical therapy coincide.     

Mutations affecting the synthesis of NADP-dependent glutamate dehydrogenase in Pseudomonas aeruginosa

NADP-dependent glutamate dehydrogenase (NADP-GDH) was purified to homogeneity from Pseudomonas aeruginosa strain 8602 (PAC 1). The Mr determined by Sephadex gel filtration was 280,000; the subunit Mr determined by SDS-PAGE was 45,000. Mutant strains lacking NADP-GDH and glutamate synthase (Gdh-Glt-) required glutamate for growth. Transductants that lacked only NADP-GDH were indistinguishable from the wild-type strain in growth properties. It was concluded that NADP-GDH is not essential for growth of the wild-type organism and that glutamate formation via NAD-dependent glutamate dehydrogenase does not occur to a significant extent. A mutant strain, 39, producing high NADP-GDH activity, synthesized normal NADP-GDH and had the same intracellular glutamate concentrations as its parent. The mutation responsible for the synthesis of high levels of NADP-GDH was shown, by transduction, to be closely linked to the NADP-GDH structural gene (gdhA).     

Cooperation and virulence in acute Pseudomonas aeruginosa infections

Background  

Efficient host exploitation by parasites is frequently likely to depend on cooperative behaviour. Under these conditions, mixed-strain infections are predicted to show lower virulence (host mortality) than are single-clone infections, due to competition favouring non-contributing social 'cheats' whose presence will reduce within-host growth. We tested this hypothesis using the cooperative production of iron-scavenging siderophores by the pathogenic bacterium Pseudomonas aeruginosa in an insect host.     

Behavioral comparisons of the tastes of L-alanine and monosodium glutamate in rats

Recent research has implicated T1R1/T1R3 as the primary taste receptor in mammals for detecting L-amino acids, including L-monosodium glutamate (MSG) and L-alanine. Previous behavioral studies with rodents found only minimal evidence that these two substances share perceptual qualities, but those studies did not control for the taste of sodium associated with MSG. This study used several behavioral methods to compare the perceptual qualities of MSG and L-alanine in rats, using amiloride (a sodium channel blocker) to reduce the sodium component of MSG taste. Detection thresholds of L-alanine in rats ranged between 0.4 and 2.5 mM, with or without amiloride added, which are similar to threshold estimates for MSG. Conditioned taste aversion (CTA) found that rats showed strong cross-generalization of CTA between MSG and L-alanine when mixed with amiloride, indicating the two substances have similar perceptual qualities. Discrimination methods showed that rats easily discriminated between L-alanine and MSG unless the cue function of sodium was reduced. The discrimination became significantly more difficult at concentrations < 100 mM when amiloride was added to all stimuli and became even more difficult when NaCl was also added to L-alanine solutions to match the sodium concentrations of MSG. These results indicate that, perceptually, MSG and L-alanine have quite similar taste qualities and support the hypothesis that these two L-amino acids activate a common taste receptor. The differences in perceptual qualities also suggest separate afferent processing of one or both substances may also be involved.     

Loss of morphine hyperphagia following neonatal monosodium glutamate treatment in rats

Morphine stimulates food intake in mildly-deprived and nondeprived rats. Neonatal administration of monosodium glutamate (MSG) destroys the medial-basal hypothalamus and other circumventricular organs, including cells containing beta-endorphin that project to other hypothalamic nuclei proposed in the modulation of morphine hyperphagia. Food intake of MSG-treated and control rats were assessed following vehicle and morphine (1.0-5.0 mg/kg, sc) treatment in a mild (5h) food deprivation paradigm. Morphine hyperphagia was found to be absent in MSG-treated rats, although they responded normally to mild deprivation following vehicle treatment. These results add to the types of ingestive deficits observed in the MSG-treated rat, and suggest that the circumventricular system in general, and opioid medial-basal hypothalamic cells in particular may be implicated in morphine hyperphagia.     

Quantitative observations of heterogeneities in Pseudomonas aeruginosa biofilms.

Heterogeneity in a Pseudomonas aeruginosa biofilm was quantified by measuring distributions of thickness in biofilm samples and a distribution of particle sizes in effluent samples. The mean steady-state thickness was approximately 33 microns, but individual measurements ranged from 13.3 to 60.0 microns. Particles exceeding 100 microns3 were observed in the reactor effluent. The results reveal a rough biofilm surface and indicate that most biomass detaches in the form of multicellular particles.     

Pseudomonas aeruginosa infections associated with use of contaminated medicaments.

Pseudomonas aeruginosa was recovered from three hospital-prepared medicaments being used on the wards. Sixty-six patients were studied to observe the effect of using these contaminated medicaments. Psaeruginosa was recovered from 29 patients; in five the strains recovered bore a close resemblance to strains previously isolated from the contaminated medicaments.     

Effects of monosodium glutamate on food acceptance and toxicity of selenium in rats

Food acceptance and toxic effects of feeding sodium selenite (Se) alone and in combination with monosodium glutamate (MSG), a taste enhancer were studied in the laboratory rat. Dose-dependent stimulation of daily food intake was observed with MSG offered in no-choice or bi-choice with the plain food. Consumption of pellets containing 0.05, 0.5 and 1.0% Se was significantly low than the plain or MSG containing pellets but their active ingredient was sufficient to cause mortality of rats. Food pellets containing both MSG and Se in no-choice feeding trial were not preferred by the rats, as their consumption remained low as compared to pellets containing only MSG. However, prior feeding on MSG containing pellets for two days increased the amount of intake of Se-containing pellets. No mortality of rats feeding on pellets containing different concentrations of MSG was recorded. Feeding on Se-containing pellets caused dose-dependent mortality on the third day of the trial. As compared to rats feeding on Se-containing pellets, the mortality rate was reduced in those provided Se in combination with MSG but the intake of active ingredient of Se in both these trials did not differ significantly. Decrease in death rate of rats feeding on Se in combination with MSG containing pellets suggested that addition of MSG to seleniferous food probably provide protection to some extent from the toxic effects of selenium. However, combination of excess doses of MSG and Se in food pellets caused mortality of all experimental animals.     

Pseudomonas aeruginosa infections are prevented in cystic fibrosis patients by avian antibodies binding Pseudomonas aeruginosa flagellin

Pseudomonas aeruginosa (PA) is the main cause of morbidity and mortality in cystic fibrosis (CF) patients. CF patients with chronic PA infections have a more rapid deterioration of their lung function and the bacteria become impossible to eradicate from the lungs. Antibiotic resistance among PA strains in CF patients is steadily increasing. Specific chicken (IgY) antibodies against PA have been shown to have potential to prevent PA infections in CF. Anti-Pseudomonas IgY reduces PA adhesion to epithelia, but the mechanism has not been fully elucidated. To gain further insight into the prophylactic effect of these antibodies, the immunoreactivity was investigated by 2D electrophoresis of PA strains, immunoblotting and MALDI-TOF-MS. To confirm the identity of the proteins, the tryptic peptides were analyzed by MALDI-TOF-MS to accurately measure their monoisotopic masses as well as determine their amino acid sequences. In order to facilitate fragmentation of the peptides they were N-terminally or C-terminally labeled. Several strains were investigated and anti-Pseudomonas IgY was immunoreactive against all of these strains, which strengthens its potential as a prophylactic treatment against PA. Flagellin was identified as the major antigen. Flagellin is the main protein of the flagella and is crucial for establishing infections in hosts as well as being involved in PA chemotaxis, motility, adhesion and inflammation. Furthermore, secreted flagellin elicits an inflammatory response. In conclusion, anti-Pseudomonas IgY binds flagellin, which may prevent PA infections in CF patients by hindering host invasion.     

Effect of perinatal monosodium glutamate treatment on endocrine functions of rats in maturity

Rats were injected subcutaneously with monosodium glutamate (MSG); either a single injection of 2 mg/g b.w. was given on the 2nd day of life or repeated daily injections of the same dose were given from the 2nd to the 10th day. Controls received saline. The growth of repeatedly treated females was slightly retarded from day 80. Repeated treatment caused a slight reduction in endocrine organ weights in maturity; however, endocrine functions tested between 120-150 days of life were similar in control and treated animals, e.g., AM-PM difference in plasma corticosterone levels, adrenocortical stress responsiveness and glucose tolerance test (in males) and normal vaginal cyclicity, compensatory hypertrophy of ovaries and presence of pituitary castration cells after spaying (in females). None of these treatments influenced the sexual activity and fertility of either sex. These data indicate that perinatal MSG treatment does not cause hypopituitary syndrome in the mature rat.     

Effect of adrenalectomy on the activity of small intestine enzymes in monosodium glutamate obese rats

It is well known that adrenalectomy (ADX) reverses the eating and energy balance disturbances in a variety of models of obesity associated with elevated food intake. We have previously demonstrated enhanced functional activity in the small intestine of neonatally monosodium glutamate-treated (MSG) obese rats despite the absence of overeating and we concluded that these changes might also contribute to the development of MSG obesity. The objective of the present experiments was to investigate whether ADX would affect the small intestinal functions and whether their changes would counteract attenuation or prevention of obesity development in MSG rats. Therefore the investigation was carried out in MSG-obese Wistar male rats and untreated intact rats adrenalectomized on day 40, as well as in lean littermates of MSG rats and intact rats subjected to Sham-ADX surgery. All animals had free access to a standard pellet diet after weaning. At the age of 80 days, body mass, body fat content and food consumption as well as changes of the brush-border-bound duodenal and jejunal alkaline phosphatase (AP), the dipeptidyl(amino)peptidase IV (DPP IV) and maltase activity were measured. During the postoperative period, ADX resulted in a significant decrease of mass gain in both MSG and control rats (P<0.05). ADX fully prevented the development of obesity in MSG rats (significantly decreased epididymal+retroperitoneal fat pad mass, P<0.05) and increased mean daily food intake (P<0.001). These effects were only minimal in the ADX controls suggesting that enhanced adrenal secretion is involved in the expression of MSG obesity and its complications. The AP activity in obese MSG rats was increased by about 21 % (P<0.01) in both intestinal segments when compared to the lean controls, whereas no parallel variations in the activities of DPP IV and maltase were observed in the intestinal parts mentioned. In MSG rats, ADX significantly reduced the AP activity in the duodenum and jejunum (P<0.01). A similar tendency was also seen in the DPP IV activity of adrenalectomized MSG rats as well as in lean control rats. Nevertheless, no significant effect of adrenal withdrawal on maltase activity was found. These results indicate that the decrease of enzyme activities in the small intestine and the different effectiveness of nutrient absorption might be a significant factor preventing the development of excess adiposity in glutamate-treated rats. This information contributes to a better understanding of the importance of small intestinal function for the development of obesity and its maintenance in later life.     

Pyocyanin reactions with Pseudomonas aeruginosa cells and their fragments

Pyocyanin added to suspensions of Pseudomonas aeruginosa P changes the structural organization of the cells depending on their physiological characteristics, in particular, the capability to liberate pyocyanin into the cultural broth. Exogenous pyocyanin does not interact with the cells of the parent strain producing the pigment. However, the structure of the isolated cell walls deformed after the fractionation becomes similar, upon contact with pyocyanin, to the structure of the freshly isolated cell walls. The fraction of the cytoplasmic membranes of the parent strain also slightly changes its spatial organization after addition of pyocyanin. The cells of the mutant which do not produce pyocyanin display the ability for structural interaction with exogenous pyocyanin. In contrast, the fraction of their cell walls does not react to addition of the pigment. The cytoplasmic membranes of the mutant interact with pyocyanin in the same manner as the whole cells. These changes caused by pyocyanin always involve certain parts of the molecules. As a result, the following features are observed in their spectra: the absorption at 1475 cm-1 becomes more intensive; the "knee" at the band Amide II is more pronounced in the range of 1500 cm-1; the absorption at 1520 cm-1 decreases as well as the intensity of the band Amide II as a whole. The interaction of pyocyanin with biological objects stems, apparently, from the presence of certain proteins in them since peroxidase was found to respond to pyocyanin in a manner similar to that described for the cells and their fractions.