Effect of Zuccagnia punctata Cav. (Fabaceae) extract on pro-inflammatory enzymes and on planktonic cells and biofilm from Staphylococcus aureus. Toxicity studies.

Zuccagnia punctata Cav. (Fabaceae), a native plant from Argentina has been used traditionally as medicinal species. The aim of the study was to validate the antibiotic and anti-inflammatory potential of Z. punctata organic extract (ZpE) and the major compounds; 2',4'-dihydroxy-3'-methoxychalcone (DHMC), 2',4'-dihydroxychalcone (DHC), 7-hydroxyflavanone (7-HF) and 3,7-dihydroxyflavone (DHF); using an in vitro model. The antibiotic activity was determined using a broth microdilution method and the minimum inhibitory concentration (MIC) was determined. The extract and the isolation compounds affect the normal growth of all assayed Staphylococcus aureus strains. The MIC values for ZpE and isolated compounds were between 125 and 500 µg/mL and between 25 and 400 µg/mL, respectively, against all assayed strains. The inhibitory effect of extract and isolated compounds on biofilm formation and on pro-inflammatory enzymes (sPLA2, COX-2, LOX) was analyzed. The compound DHC was the most active on sPLA2 while DHF and DHMC showed the highest activity on LOX. Both the extract and pure compounds except DHMC were active against COX-2. It can be concluded that the phytocomplex and the pure compounds possessed antibiotic and anti-inflammatory activities under the conditions tested, and could be a good alternative therapy for infective and inflammatory processes.

Comparison of seven structurally related coumarins on the inhibition of Quorum sensing of Pseudomonas aeruginosa and Chromobacterium violaceum.

Quorum sensing (QS) is a cell-to-cell signaling communication system that controls the virulence behavior of a broad spectrum of bacterial pathogens, participating also in the development of biofilms, responsible of the antibiotic ineffectiveness in many infections. Therefore, QS system is an attractive target for antimicrobial therapy. In this study, we compare the effect of seven structurally related coumarins against bacterial growth, biofilm formation and elastase activity of Pseudomonas aeruginosa. In addition, the anti-pathogenic capacity of the seven coumarins was evaluated on the wild type and the biosensor strain of Chromobacterium violaceum. The comparative study of coumarins showed that molecules with hydroxyl groups on the aromatic ring displayed higher activity on the inhibition of biofilm formation of P. aeruginosa over coumarins with substituents in positions 3 and 4 or without the double 3,4-bond. These 3 or 4-hydroxylated positions caused a decrease in the anti-biofilm activity obtained for coumarin. However, the hydroxyl group in position 3 of the pyrone ring was important for the inhibition of C. violaceum QS and elastolytic activity of P. aeruginosa. The effects observed were active independently of any effect on growth. According to our results, coumarin and its hydroxylated derivatives represent an interesting group of compounds to use as anti-virulence agents against the human pathogen P. aeruginosa.

Probiotic administration modifies the milk fatty acid profile, intestinal morphology, and intestinal fatty acid profile of goats.

The effect of a mixture of potentially probiotic bacteria (MPPB; Lactobacillus reuteri DDL 19, Lactobacillus alimentarius DDL 48, Enterococcus faecium DDE 39, and Bifidobacterium bifidum strains) on the milk fatty acid (FA) profile, with emphasis on cis-9,trans-11 conjugated linoleic acid (CLA) in the middle stage of goat lactation, was determined. In addition, the effects of MPPB feeding on the FA profile in intestinal content and intestinal morphology in weaned goats were analyzed. The probiotic supplement was able to modify FA composition of milk and intestinal content. The unsaturated FA concentrations in milk (g of FA/L of milk) increased from 4.49 to 7.86 for oleic (18:1), from 0.70 to 1.39 for linoleic (18:2), from 0.063 to 0.187 for linolenic (18:3) acid, and from 0.093 to 0.232 for CLA. The atherogenicity index diminished 2-fold after MPPB ingestion. In the intestinal content of the weaned goats, no significant difference in saturated FA concentration compared with the control was observed. However, oleic acid, linolenic acid, CLA, and docosahexaenoic acid concentrations increased by 81, 23, 344, and 74%, respectively, after probiotic consumption. The ruminal production of CLA was increased by the MPPB. However, bacterial strains of MPPB were unable to produce CLA in culture media. By histological techniques, it was observed that the treated group had intestinally more conserved morphological structures than the control group. The results obtained in this study indicate that the MPPB administration in lactating and weaned goats allows for the production of milk with improved concentrations of beneficial compounds, and also produces a protective effect in the goat intestine. The results obtained in this study reinforce the strategy of probiotics application to enhance goat health with the production of milk with higher concentrations of polyunsaturated FA.

Inhibition of mycotoxin-producing Aspergillus nomius vsc 23 by lactic acid bacteria and Saccharomyces cerevisiae

The effect of different fermenting microorganisms on growth of a mycotoxin- producing Aspergillus nomius was assayed. Two lactic acid bacteria, Lactobacillus fermentum and Lactobacillus rhamnosus, and Saccharomyces cerevisiae, all of which are widely used in fermentation and preservation of food, were assayed on their fungus inhibitory properties. Assays were carried out by simultaneous inoculation of one of the possible inhibiting microorganisms and the fungus or subsequent inoculation of one of the microorganisms followed by the fungus. All three microorganisms assayed showed growth inhibition of the mycotoxin-producing Aspergillus strain. L. rhamnosus O236, isolated from sheep milk and selected for its technological properties, showed highest fungal inhibition of the microorganisms assayed. The use of antifungal LAB with excellent technological properties rather than chemical preservatives would enable the food industry to produce organic food without addition of chemical substances.

Inhibition of mycotoxin-producing Aspergillus nomius vsc 23 by lactic acid bacteria and Saccharomyces cerevisiae.

The effect of different fermenting microorganisms on growth of a mycotoxin- producing Aspergillus nomius was assayed. Two lactic acid bacteria, Lactobacillus fermentum and Lactobacillus rhamnosus, and Saccharomyces cerevisiae, all of which are widely used in fermentation and preservation of food, were assayed on their fungus inhibitory properties. Assays were carried out by simultaneous inoculation of one of the possible inhibiting microorganisms and the fungus or subsequent inoculation of one of the microorganisms followed by the fungus. All three microorganisms assayed showed growth inhibition of the mycotoxin-producing Aspergillus strain. L. rhamnosus O236, isolated from sheep milk and selected for its technological properties, showed highest fungal inhibition of the microorganisms assayed. The use of antifungal LAB with excellent technological properties rather than chemical preservatives would enable the food industry to produce organic food without addition of chemical substances.

Inhibition of mycotoxin-producing Aspergillus nomius vsc 23 by lactic acid bacteria and Saccharomyces cerevisiae

The effect of different fermenting microorganisms on growth of a mycotoxin- producing Aspergillus nomius was assayed. Two lactic acid bacteria, Lactobacillus fermentum and Lactobacillus rhamnosus, and Saccharomyces cerevisiae, all of which are widely used in fermentation and preservation of food, were assayed on their fungus inhibitory properties. Assays were carried out by simultaneous inoculation of one of the possible inhibiting microorganisms and the fungus or subsequent inoculation of one of the microorganisms followed by the fungus. All three microorganisms assayed showed growth inhibition of the mycotoxin-producing Aspergillus strain. L. rhamnosus O236, isolated from sheep milk and selected for its technological properties, showed highest fungal inhibition of the microorganisms assayed. The use of antifungal LAB with excellent technological properties rather than chemical preservatives would enable the food industry to produce organic food without addition of chemical substances.

Characterization of a Lactobacillus plantarum strain able to produce tyramine and partial cloning of a putative tyrosine decarboxylase gene.

The aim of this article was to analyze the ability of wine Lactobacillus plantarum strains to form tyramine. Preliminary identification of L. plantarum strains was performed by amplification of the recA gene. Primers pREV and PlanF, ParaF and PentF were used respectively as reverse and forward primers in the polymerase chain reaction tests as previously reported. Furthermore, the gene encoding for the tyrosine decarboxylase (TDC) was partially cloned from one strain identified as L. plantarum. The strain was further analyzed by 16S rDNA sequence and confirmed as belonging to L. plantarum species. The tyrosine decarboxylase activity was investigated and tyramine was determined by the high-performance liquid chromatography method. Moreover, a negative effect of sugars such as glucose and fructose and L: -malic acid on tyrosine decarboxylase activity was observed. The results suggest that, occasionally, L. plantarum is able to produce tyramine in wine and this ability is apparently confined only to L. plantarum strains harboring the tdc gene.

Biogenic amine production by Lactobacillus.

AIMS: The aim of this work was to demonstrate that strains of Lactobacillus may be able to produce putrescine and agmatine from one of the major amino acids present in fruit juices and wine, arginine, and from amino acid-derived ornithine. METHODS AND RESULTS: Biogenic amines were determined by HPLC. Their production in the culture medium was similar under both microaerophilic and anaerobic conditions. The presence of Mn2+ had a minimal influence on the results, whereas the addition of pyridoxal phosphate increased amine production 10-fold. Lactobacillus hilgardii X1B, isolated from wine, was able to degrade arginine by two pathways: arginine deiminase and arginine decarboxylase. The isolate was able to produce putrescine from ornithine and from agmatine. Lactobacillus plantarum strains N4 and N8, isolated from orange, utilized arginine via the arginine deiminase system. Only the N4 strain was able to produce putrescine from ornithine. CONCLUSION: It has been demonstrated that Lact. hilgardii X1B is able to produce the most important biogenic amine found in wine, putrescine, and also agmatine from arginine and ornithine, and that Lactobacillus plantarum, considered to be an innocuous spoilage micro-organism in fruit juices, is able to produce amines. SIGNIFICANCE AND IMPACT OF THE STUDY: The results have significance in relation to food poisoning caused by beverages that have been contaminated with biogenic amines.

Calcium and boron for in vitro rooting of Nothofagus nervosa.

Several mineral compounds influence adventitious rooting, and those containing boron and calcium, play an important role. They are associated to the enzyme regulation, cell division, cellular metabolism (induction stage) and roots enlargement (expression stage). The aims of this work were to study the variation of the endogenous calcium and boron concentrations along the rooting, and determine their optimal concentrations in the rooting basal medium of Nothofagus nervosa. Significant variation in endogenous calcium concentrations was detected along the rooting phase, strongly correlated with rhizogenesis phenomenons and linked to total peroxidase variation as cited elsewhere. Calcium and boron concentrations in the basal medium significantly affected the rooting. Treatment containing 60 mg.L-1 Ca and 1 mg.L-1 B produced root formation in fewer days than other combinations. The variation of these nutrient concentrations in the medium is a useful tool to improve microcutting rooting. Roots are obtained earlier and in a shorter period of time, finally resulting in more homogeneous microplants with a better developed rooting system.

In vitro propagation of Berberis buxifolia Lam.

Berberis buxifolia is a native shrub of Patagonia with a great importance due to its crop production as soon its medicinal and tinctorial applications. The aim of this work was to develop a protocol for in vitro propagation of B. buxifolia, with special emphasys on the rooting stage. The culture of the explants on Murashige and Skoog (1962) medium added with 0.55 microM BA allowed to attain a multiplication rate of 1:4.7 at day 63. Rooted shoots were obtained on Murashige and Skoog medium with half strength of macronutrient salts. The culture of the shoots with a period of 4 days of darkness at the beginning of the rooting, on a medium with 1.25 microM IBA for 7 days, followed by a IBA free medium until day 28, allowed to attain 80% rooting. These results show that B. buxifolia can be in vitro propagated.

Calcium and boron for in vitro rooting of Nothofagus nervosa.

Several mineral compounds influence adventitious rooting, and those containing boron and calcium, play an important role. They are associated to the enzyme regulation, cell division, cellular metabolism (induction stage) and roots enlargement (expression stage). The aims of this work were to study the variation of the endogenous calcium and boron concentrations along the rooting, and determine their optimal concentrations in the rooting basal medium of Nothofagus nervosa. Significant variation in endogenous calcium concentrations was detected along the rooting phase, strongly correlated with rhizogenesis phenomenons and linked to total peroxidase variation as cited elsewhere. Calcium and boron concentrations in the basal medium significantly affected the rooting. Treatment containing 60 mg.L-1 Ca and 1 mg.L-1 B produced root formation in fewer days than other combinations. The variation of these nutrient concentrations in the medium is a useful tool to improve microcutting rooting. Roots are obtained earlier and in a shorter period of time, finally resulting in more homogeneous microplants with a better developed rooting system.

In vitro propagation of Berberis buxifolia Lam.

Berberis buxifolia is a native shrub of Patagonia with a great importance due to its crop production as soon its medicinal and tinctorial applications. The aim of this work was to develop a protocol for in vitro propagation of B. buxifolia, with special emphasys on the rooting stage. The culture of the explants on Murashige and Skoog (1962) medium added with 0.55 microM BA allowed to attain a multiplication rate of 1:4.7 at day 63. Rooted shoots were obtained on Murashige and Skoog medium with half strength of macronutrient salts. The culture of the shoots with a period of 4 days of darkness at the beginning of the rooting, on a medium with 1.25 microM IBA for 7 days, followed by a IBA free medium until day 28, allowed to attain 80

rooting. These results show that B. buxifolia can be in vitro propagated.

[Inhibitory and bactericidal activity and selection capacity of mutants resistant and tolerant to vancomycin in Staphylococcus aureus strains]. / Actividad inhibitoria, bactericida y capacidad de selección de mutantes resistentes y tolerantes de vancomicina en cepas de Staphylococcus aureus.

BACKGROUND: The aim of the present was to study the inhibitory and bactericide activity and risk of selection of mutants resistant or tolerant to vancomycin in strains of Staphylococcus aureus resistant to methicillin from out patient and hospitalary patients. MATERIALS AND METHODS: The minimum inhibitory concentration was obtained by the macrodilution method in liquid medium and by the Etest system. The minimum bactericide concentration was obtained by the macrodilution method in liquid medium. The selection of resistant or tolerant mutants was performed inoculating more than 5 x 10(6) UFC/ml in brain-heart agar plates and in tripteinsoya brought with different concentrations of vancomycin. RESULTS: Vancomycin showed greater inhibitory and bactericide activity in strains sensitive to methicillin of out patient and hospitalary origin with respect to strains resistant to methicillin with a minimum inhibitory concentration > 1 mg/l in 23.3, 20.0 and 70.0%, respectively and a minimum bactericide concentration of 90%, 8.16 and 64 mg/l. The incidence of tolerance was greater in the strains resistant to methicillin in comparison with the out patient and hospitalary strains sensitive to methicillin, since a minimum bactericide/inhibitory concentration greater than 4 was obtained in 63.3 in comparison with 20.0%. Only mutants with diminished sensitivity (minimum inhibitory concentration of 3 and 6 mg/l) or tolerant in strains resistant to methicillin could be selected. The increases in the minimum inhibitory concentrations were not stable, while the tolerant mutants preserved this characteristic after 20 subcultures. CONCLUSIONS: The in vitro selection of mutants with decreased sensitivity or tolerant was only obtained in strains resistant to methicillin. The changes in the minimum inhibitory concentrations were not stable in contrast to the increase in the minimum bactericide concentrations which remained unchanged. The differences in the selection of mutants with diminished sensitivity or tolerant among strains resistant and sensitive to methicillin found in this study were correlated with the differences in minimum inhibitory and bactericide concentrations and the relation between both in the clinical isolates.

Arginine dihydrolase pathway in Lactobacillus plantarum from orange.

Lactobacillus plantarum N8 and N4 strains isolated from orange degraded L-arginine to citrulline, ornithine and ammonia. Citrulline and ornithine were consumed. Lactobacillus plantarum N4 utilized arginine and ornithine to a higher extent than Lactobacillus plantarum N8. Urea was not detected during arginine degradation, indicating that the amino acid degradation was carried out only by the arginine dihydrolase pathway. Citrulline increased the growth of the two strains, arginine only increased the growth of Lactobacillus plantarum N4. Ornithine did not modify the growth of the strains studied. With different behavior, Lactobacillus plantarum N8 and N4 strains were able to derive energy and ammonia from arginine or citrulline catabolism. This is interesting for microorganisms developing in a stressful environment.

Arginine, citrulline and ornithine metabolism by lactic acid bacteria from wine.

The catabolism of arginine, an amino acid found in grape juice and wine, citrulline and ornithine was investigated in four lactic acid bacteria. Only Lactobacillus hilgardii X1B catabolized arginine and excreted citrulline into the medium. The recovery of arginine as ornithine was lower than the expected theoretical value. The arginase-urease pathway was not detected indicating that the amino acid degradation was carried out only by the arginine dihydrolase pathway. Oenococcus oeni m, a strain not able to utilize arginine, degraded citrulline that was completely recovered as ornithine, ammonia and CO2. Lactobacillus hilgardii X1B catabolized citrulline but it was only 44% recovered as ornithine. The citrulline utilization by Oenococcus oeni m may be important for two reasons: it can gain extra energy for growth from citrulline metabolism, and the amino-acid diminution could avoid the possibility of ethyl carbamate formation from the citrulline naturally present in wine.

Physiological variability arising from in vitro propagation of Nothofagus obliqua

As with cuttings, the microcuttings produced from in vitro generated shoots could be affected by topophysis. Very little research has been done about such in vitro effect. This paper reports the influence of the topophysis on the regeneration response (shoot multiplication, microcutting rooting and plantlet acclimation) of explants on the in vitro culture of juvenile N. obliqua trees. The results showed that the main responsible of the physiological variability arising from all the in vitro propagation stages is topophysis. This influence was found in the first culture and can be extended to the next subculture (multiplication stage). Growth of the obtained plants was also affected (rooting of microcuttings and plantlet acclimation). The multiplication rate was lower in apical portion (1:3.66) than basal portion (1:5.8) while rooting was 48.6 and 27.0 respectively. By the way, length of acclimated explants was 52.4 mm in the apical rooted explants and 28.7 mm in the basal ones at the 30th day. In order to reduce such variability, it is necessary to take into account the initial position of the explants during the establishment and multiplication stages of micropropagation of forest trees

Physiological variability arising from in vitro propagation of Nothofagus obliqua

As with cuttings, the microcuttings produced from in vitro generated shoots could be affected by topophysis. Very little research has been done about such in vitro effect. This paper reports the influence of the topophysis on the regeneration response (shoot multiplication, microcutting rooting and plantlet acclimation) of explants on the in vitro culture of juvenile N. obliqua trees. The results showed that the main responsible of the physiological variability arising from all the in vitro propagation stages is topophysis. This influence was found in the first culture and can be extended to the next subculture (multiplication stage). Growth of the obtained plants was also affected (rooting of microcuttings and plantlet acclimation). The multiplication rate was lower in apical portion (1:3.66) than basal portion (1:5.8) while rooting was 48.6 and 27.0 respectively. By the way, length of acclimated explants was 52.4 mm in the apical rooted explants and 28.7 mm in the basal ones at the 30th day. In order to reduce such variability, it is necessary to take into account the initial position of the explants during the establishment and multiplication stages of micropropagation of forest trees

[Bactericidal activity and capacity for selection of mutants resistant to imipenmen and meropenem in strains of Proteus, Morganella and Providencia]. / Actividad bactericida y capacidad de selección de mutantes resistentes a imipenem y meropenem en cepas de Proteus, Morganella y Providencia.

BACKGROUND: To compare the bactericidal activity and frequency of mutation for meropenem and imipenem against Proteus mirabilis, Morganella morganii and Providencia rettgeri. MATERIAL AND METHODS: Minimum inhibitory concentrations (MIC) were determined by agar dilution method. Bactericidal activities were evaluated by killing curves method employing 4 and 16x MIC. One-step resistant mutant selection was performed by spreading more than 5 x 10(8) UFC/ml on cystine-lactose-electrolyte deficient agar containing 4 times the MIC of meropenem or imipenem. RESULTS: MIC were 8 to 16 times lower for meropenem. After 24 h, bactericidal activity was observed for meropenem at 4 and 16 x MIC against 76.7 and 100% of the strains in comparison to 26.7 and 83.3% with imipenem. After 24 h incubation with imipenem, re-growth occurred in 80 and 90% of P. mirabilis and M. morganii strains, respectively. Imipenem resistant mutants were selected from 3 strains of P. mirabilis. One of them was stable and MIC of meropenem and imipenem were 8 to 16-fold higher. CONCLUSIONS: From the laboratory point of view we consider that meropenem is more active against Proteeae because it was more potent in terms of inhibitory and bactericidal activity. In addition the risk to select for resistant mutants was significant with imipenem and P. mirabilis.

Alteraciones histológicas relacionadas al Helicobacter pylori en niños / Helicobacter pylori associated with histological changes in children

El helicobacter pylori(HP) es descripto como causante de inflamación crónica primaria de la mucosa gástrica,tanto en adultos como en niños. Estudios de la histología de la mucosa gástrica relacionados con HP en el niño, incluyen en general adolescentes y adultos jóvenes mostrando una casuística poco numerosa. El objetivo fue evaluar alteraciones histólogicas de la mucosa gástrica infectada por HP, en una población exclusivamente pediátrica con molestias abdominales

Alteraciones histológicas relacionadas al Helicobacter pylori en niños / Helicobacter pylori associated with histological changes in children

El helicobacter pylori(HP) es descripto como causante de inflamación crónica primaria de la mucosa gástrica,tanto en adultos como en niños. Estudios de la histología de la mucosa gástrica relacionados con HP en el niño, incluyen en general adolescentes y adultos jóvenes mostrando una casuística poco numerosa. El objetivo fue evaluar alteraciones histólogicas de la mucosa gástrica infectada por HP, en una población exclusivamente pediátrica con molestias abdominales