Contribución al estudio de la resistencia a diversos agentes antimicrobianos en cepas salvajes de Pseudomonas aeruginosa aisladas de aguas y suelos de Cataluña

Tesi de Llicenciatura per a la obtenció del Grau de Farmàcia. Facultat de Farmàcia. Universitat de Barcelona. Director: Ma. Dolores Simón Pujol. 1977

Aislamiento y caracterización de Pseudomonas aeruginosa procedentes de distintos habitats: aspectos ecológicos y sanitarios

[spa] Pseudomonas aeruginosa es un microoganismo Gram-negativo, potencialmente patógeno, agente causal de serias infecciones en el hombre que cursan con gran variedad de síntomas (HOMMA, 1971). Entre sus principales características destacan su elevada resistencia a los agentes antimicrobianos y su potencial metabólico, siendo capaz de utilizar un número muy elevado de compuestos (LIU, 1976; STANIER et al., 1977). La existencia en el medio ambiente de bacterias patógenas multirresistentes, como P. aeruginosa, puede significar un insospechado peligro para el hombre. Las implicaciones de la presencia y origen de estos microorganismos son cuestiones importantes que deben seírcontestadas (COLWELL y SIZEMORE, 1974). Para entender el comportamiento de un microorganismo patógeno oportunista como P, aeruginosa y poderlo combatir de forma efectiva, es necesario estudiar su ciclo y las relaciones que presenta en el medio ambiente, pudiendo llegar de esta forma a conocer sus vías de transmisión (HOADLEY, 1977, YOUNG, 1977). El fin de esta tesis es eliminar algunas barreras existentes en el campo de la Microbiología, presentando una información conjunta esencial para el entendimiento de la P. aeruginosa. En esta memoria se estudian algunos aspectos de su biología, sobre todo los que hacen referencia a su resistencia ante agentes antimicrobianos. Su incidencia y nivel de resistencia puede significar un verdadero reto para la salud pública

Multilayer emulsions stabilized by vegetable proteins

There is great interest in the food, cosmetic and pharmaceutical industry in the use of proteins and polysaccharides as natural hydrocolloids to create novel emulsion systems with improved stability and functionality. For example, the electrostatic interaction between proteins and polysaccharides may be used to form oil-in-water (O/W) emulsions with multilayered interfacial membranes around oil droplets or multilayer emulsions. This type of emulsions have been developed using the layer-by-layer (LbL) technique, which consists of direct adsorption of an oppositely charged polyelectrolyte layer (e.g. polysaccharides) on a primary layer of ionic emulsifiers (e.g. proteins). The polymeric structure and electrical charge of proteins make them a special class of compounds very suitable for its utilization in the LbL technique. In recent years, the utilization of proteins as emulsifier in the food and pharmaceutical industry has been turning towards plants as a preferred alternative to animal-based sources. This article reviews the current understanding of the utilization of different vegetable proteins as emulsifier in order to stabilize O/W multilayer emulsions systems. Additionally, it highlights some potential applications of the multilayer emulsion technology in the industry for improving the stability of emulsions to environmental stresses or and for developing controlled or triggered release systems

Unveiling the Membrane and Cell Wall Action of Antimicrobial Cyclic Lipopeptides: Modulation of the Spectrum of Activity

Antibiotic resistance is a major public health challenge, and Gram-negative multidrug-resistant bacteria are particularly dangerous. The threat of running out of active molecules is accelerated by the extensive use of antibiotics in the context of the COVID-19 pandemic, and new antibiotics are urgently needed. Colistin and polymyxin B are natural antibiotics considered as last resort drugs for multi-resistant infections, but their use is limited because of neuro- and nephrotoxicity. We previously reported a series of synthetic analogues inspired in natural polymyxins with a flexible scaffold that allows multiple modifications to improve activity and reduce toxicity. In this work, we focus on modifications in the hydrophobic domains, describing analogues that broaden or narrow the spectrum of activity including both Gram-positive and Gram-negative bacteria, with MICs in the low µM range and low hemolytic activity. Using biophysical methods, we explore the interaction of the new molecules with model membranes that mimic the bacterial inner and outer membranes, finding a selective effect on anionic membranes and a mechanism of action based on the alteration of membrane function. Transmission electron microscopy observation confirms that polymyxin analogues kill microbial cells primarily by damaging membrane integrity. Redistribution of the hydrophobicity within the polymyxin molecule seems a plausible approach for the design and development of safer and more selective antibiotics

Optimizing the production of the biosurfactant lichenysin and its application in biofilm control

Aims: Apply response surface methodology (RSM) to develop and optimize an economical medium for lichenysin production, which is a surfactant produced by Bacillus licheniformis and evaluate the application of lichenysin in the prevention and disruption of pathogenic micro-organism biofilm that creates health problems in the food industry and hospitals. Results: An economical medium containing molasses was optimized to enhance lichenysin production by RSM. A production of 3.2 g l 1 of lichenysin was achieved with an optimum medium containing 107.82 g l 1 of molasses, 6.47 g l 1 of NaNO3 and 9.7 g l 1 of K2HPO4/KH2PO4, in which molasses and phosphate salts had a significant effect on biosurfactant production. Lichenysin was effectively applied in a surface pre-treatment to avoid microbial biofilm development of methicillin-resistant Staphylococcus aureus (MRSA) (68.73%) and Candida albicans (74.35%), with ED50 values of 8.3 and 17.2 lg ml 1 respectively. It was also very efficient in a surface posttreatment to remove biofilms of MRSA (55.74%) and Yersinia enterocolitica (51.51%), with an ED50 of 2.79 and 4.09 lg ml 1 respectively. Conclusions: Lichenysin was found to have notable anti-adhesion activity, being able to prevent and eliminate the biofilm formation by pathogenic strains associated with foodborne illness. This new medium resulted in a fourfold increase in production compared with the nonoptimized medium. Significance and Impact of the Study: Molasses can be regarded as a useful resource for biotechnological applications, such as the production of lichenysin. The use of agro-industrial substrates has an important role in the sustainable and competitive development of several industrial sectors, as well as in industrial residues management. Additionally, lichenysin is particularly effective in preventing biofilm formation by strains problematic for the food industry and in the hospital environment. Lichenysin also efficiently disrupts biofilm

Green Catanionic Gemini Surfactant-Lichenysin Mixture: Improved Surface, Antimicrobial, and Physiological Properties

Catanionic surfactant mixtures form a wide variety of organized assemblies and aggregates with improved physicochemical and biological properties. The green catanionic mixture (NN omega)-N-alpha-Bis(N(alpha)caproylarginine) alpha,omega-propyldiamide (C-3(CA)(2)):Lichenysin (molar ratio 8:2) showed antimicrobial synergies against Yersinia enterocolitica, Bacillus subtilis, Escherichia coli O157:H7, and Candida albicans. Flow cytometry and viability studies indicated that this catanionic mixture increases the probability of Y. enterocolitica (38.2%) and B. subtilis (17.1%) cells entering a viable but nonculturable state. Zeta potential showed that one of the cationic charges of C-3(CA)(2) is neutralized by Lichenysin. An isotherm study demonstrated the formation of a stable aggregate between the two surfactants. The catanionic aggregate was able to interact with bacterial phospholipids. The lowest hemolysis (22.1 mu M) was obtained with the catanionic mixture, although an irritant potential (0.70) was characterized. According to the therapeutic index, the C-3(CA)(2):Lichenysin mixture was the formulation least toxic to eukatyotic cells. Partial neutralization of C-3(CA)(2) by Lichenysin modified the mode of action that enhances the transition of bacterial cells into a viable but nonculturable state (VBNC) and improved the cell selectivity. KEYWORDS:antimicrobial activity catanionic mixtures biosurfactant Lichenysin gemini surfactants flow cytometry VBNC Langmuir balanc

Lichenysin production and application in the pharmaceutical field

Podeu consultar el llibre complet a: http://hdl.handle.net/2445/103042Lipopeptides such as lichenysin are biosurfactants of great interest, due to the demand for natural surface-active agents with low toxicity. Bacillus licheniformis AL 1.1 produces a lipopeptide characterized as lichenysin (LchAL1.1), which acts as a powerful surfactant, able to reduce surface tension to 28.5 mN m-1 and with a critical micelle concentration of 15 mg L-1. LchAL1.1 is particularly effective in preventing biofilm formation by pathogenic strains, has an emulsifying capacity and permeabilizes membranes by a colloid-osmotic process. The production of lipopeptides from agro-industrial residues, particularly molasses, is a sustainable process of great potential for the development of economic bioprocesses

Biocompatible Catanionic Vesicles from Arginine-Based Surfactants: A New Strategy to Tune the Antimicrobial Activity and Cytotoxicity of Vesicular Systems

Their stability and low cost make catanionic vesicles suitable for application as drug delivery systems. In this work we prepared catanionic vesicles using biocompatible surfactants: two cationic arginine-based surfactants (the monocatenary Nα-lauroyl-arginine methyl ester LAM and the gemini Nα,Nϖ-bis(Nα-lauroylarginine) α, ϖ-propylendiamide C3(CA)2) and three anionic amphiphiles (the single chain sodium dodecanoate, sodium myristate, and the double chain 8-SH). The critical aggregation concentration, colloidal stability, size, and charge density of these systems were comprehensively studied for the first time. These catanionic vesicles, which form spontaneously after mixing two aqueous solutions of oppositely charged surfactants, exhibited a monodisperse population of medium-size aggregates and good stability. The antimicrobial and hemolytic activity of the vesicles can be modulated by changing the cationic/anionic surfactant ratio. Vesicles with a positive charge efficiently killed Gram-negative and Gram-positive bacteria as well as yeasts; the antibacterial activity declined with the decrease of the cationic charge density. The catanionic systems also effectively eradicated MRSA (Methicillin-resistant Staphylococcus Aureus) and Pseudomonas aeruginosa biofilms. Interestingly, the incorporation of cholesterol in the catanionic mixtures improved the stability of these colloidal systems and considerably reduced their cytotoxicity without affecting their antimicrobial activity. Additionally, these catanionic vesicles showed good DNA affinity. Their antimicrobial efficiency and low hemolytic activity render these catanionic vesicles very promising candidates for biomedical application

Viability qPCR, a new tool for Legionella risk management

Background Viability quantitative Polymerase Chain Reaction (v-qPCR) is a recent analytical approach for only detecting live microorganisms by DNA amplification-based methods This approach is based on the use of a reagent that irreversibly fixes dead cells DNA. In this study, we evaluate the utility of v-qPCR versus culture method for Legionellosis risk management. Methods The present study was performed using 116 real samples. Water samples were simultaneously analysed by culture, v-qPCR and qPCR methods. Results were compared by means of a non-parametric test. Results In 11.6% of samples using both methods (culture method and v-qPCR) results were positive, in 50.0% of samples both methods gave rise to negative results. As expected, equivalence between methods was not observed in all cases, as in 32.1% of samples positive results were obtained by v-qPCR and all of them gave rise to negative results by culture. Only in 6.3% of samples, with very low Legionella levels, was culture positive and v-qPCR negative. In 3.5% of samples, overgrowth of other bacteria did not allow performing the culture. When comparing both methods, significant differences between culture and v-qPCR were in the samples belonging to the cooling towers-evaporative condensers group. The v-qPCR method detected greater presence and obtained higher concentrations of Legionella spp. (p < 0.001). Otherwise, no significant differences between methods were found in the rest of the groups. Conclusions The v-qPCR method can be used as a quick tool to evaluate Legionellosis risk, especially in cooling towers-evaporative condensers, where this technique can detect higher levels than culture. The combined interpretation of PCR results along with the ratio of live cells is proposed as a tool for understanding the sample context and estimating the Legionellosis risk potential according to 4 levels of hierarchy