RESUMEN: La aparición de cepas de Pseudomonas aeruginosa con resistencias a diversos antibióticos es un problema clínico de gran relevancia. Los métodos de detección de dichas resistencias son laboriosos y lentos, lo que genera una complicación al momento de tratar a los pacientes oportunamente. En este trabajo desarrollamos un método simple de detección simultánea de varios genes de resistencia a carbapenem, mediante un sistema de PCR múltiple. El ensayo de PCR desarrollado, seguido de una separación electroforética de los amplicones, permite distinguir simultáneamente la presencia de 6 genes de resistencia a antibióticos: bla-VIM (261 pb), bla-IMP (587 pb), bla-SPM (648 pb), bla-GIM-1 (753 pb), bla-NDM-1 (813 pb) y bla-KPC (882 pb). Analizamos 7 aislados clínicos obtenidos en Chile, encontrando en 5 de ellos los genes de resistencia bla-VIM, bla-IMP, bla-SPM, bla-GIM y bla-NDM. Encontramos una perfecta correlación entre la detección de diversos genes de resistencia y los resultados obtenidos mediante antibiogramas. Interesantemente, 2 de las cepas mostraron poseer simultáneamente 3 genes de resistencia distintos. Por último, en este trabajo encontramos la presencia de 3 genes nunca antes descritos en aislados clínicos de P. aeruginosa en Chile (bla-IMP, bla-SPM y bla-GIM-1). Hemos desarrollado un test rápido de PCR múltiple, para la detección simultánea de hasta 6 genes de resistencia a antibióticos de la familia.a de las metallo-b-lactamases en P. aeruginosa.

SUMMARY: The appearance of Pseudomonas aeruginosa strains with multi-resistance to antibiotics is a clinical problem of great relevance. The methods for detecting these resistances are laborious and slow, which is a complication when treating patients promptly. In this work, we developed a simple method for simultaneous detection of several carbapenem resistance genes using a multiplex PCR assay. The PCR assay developed, followed by electrophoretic separation of fragments, allows to simultaneously identify the presence of 6 antibiotic resistance genes: bla-VIM (261 bp), bla-IMP (587 bp), bla-SPM (648 bp), bla-GIM-1 (753 bp), bla-NDM-1 (813 bp) and bla-KPC (882 bp). We analyzed 7 clinical isolates of P. aeruginosa obtained in Chile, finding the resistance genes bla-VIM, bla-IMP, bla-SPM, bla-GIM, and bla-NDM in 5 of them. We found a perfect correlation between the detection of various resistance genes by PCR and the results obtained by antibiograms. Interestingly, 2 of the strains possessed 3 different resistance genes simultaneously. Finally, in this work, we found the presence of 3 genes never described before in clinical isolates of P. aeruginosa in Chile (bla-IMP, bla-SPM, and bla-GIM-1). We developed a rapid multiplex PCR test for the simultaneous detection of up to 6 antibiotic resistance genes of the metallo-β-lactamase family in P. aeruginosa.

SUMMARY: Cancer is the second leading cause of death in the world and colorectal cancer is the only cancer that has shown a sustained increase in mortality in the last decade. In the search for new chemotherapeutic agents against cancer, extremophilic microorganisms have shown to be a potential source to obtain molecules of natural origin and with selective cytotoxic action towards cancer cells. In this work we analyzed the ability of a collection of Antarctic soil bacteria, isolated on Collins Glacier from the rhizosphere of Deschampsia antarctica Desv plant, to secrete molecules capable of inhibiting cell proliferation of a colorectal cancer tumor line. Our results demonstrated that culture supernatants from the Antarctic bacteria K2I17 and MI12 decreased the viability of LoVo cells, a colorectal adenocarcinoma cell line. Phenotypic and genotypic characterization of the Antarctic bacteria showed that they were taxonomically related and nucleotide identity analysis based on the 16S rRNA gene sequence identified the bacterium K2I17 as a species belonging to the genus Bacillus.

RESUMEN: El cáncer es la segunda causa de muerte en el mundo y el cáncer colorrectal es el único que presenta un aumento sostenido de la mortalidad en la última década. En la búsqueda de nuevos agentes quimioterapeúticos contra el cáncer, se ha propuesto a los microorganismos extremófilos como una fuente potencial para obtener moléculas de origen natural y con acción citotóxica selectiva hacia las células cancerígenas. En este trabajo analizamos la capacidad de una colección de bacterias de suelo antártico, aisladas en el glaciar Collins desde rizosfera de la planta de Deschampsia antarctica Desv, de secretar moléculas capaces de inhibir la proliferación celular de una línea tumoral de cáncer colorrectal. Nuestros resultados demostraron que los sobrenadantes de cultivo de las bacterias antárticas K2I17 y MI12 disminuyeron la viabilidad de la línea celular de adenocarcinoma colorrectal LoVo, en un ensayo de reducción metabólica de MTT. La caracterización fenotípica y genotípica de las bacterias antárticas, demostró que estaban relacionadas taxonómicamente y el análisis de la identidad nucleotídica en base a la secuencia del gen ARNr 16S identificó a la bacteria K2I17 como una especie perteneciente al género Bacillus.

ABSTRACT: The sustained decrease in the availability of new antimicrobial molecules for thera peutic use in recent decades and the large increase in microorganisms resistant to these compounds makes it necessary to search for new substances with antimicrobial properties. Extreme environments, where there is little availability of nutrients, are favorable scenarios to search for this type of compounds, since bacteria compete with other bacteria or other organisms to position themselves in the ecological niche and secrete, associated with their secondary metabolism, a series of molecules that inhibit or kill other microorganisms thus preventing their proliferation. Our group has a collection of rhizosphere bacteria from soil of the Antarctic continent that has ability to grow under conditions of food deprivation and at 4°C. Due to the extreme habitat where they develop, Antarctic soil bacteria are in constant competition for nutritional resources and therefore develop different strategies to colonize their ecological niche and compete with other microorganisms of the Antarctic microbiota. The objective of this work was to identify bacteria isolated from the rhizosphere of Deschampsia antarctica Desv. that inhibit the growth of bacteria and fungi that are pathogenic for humans. From a collection of 55 rhizosphere isolates of the Deschampsia antarctica Desv. we identified 11 isolates with the capacity to eliminate human pathogenic bacteria, but no isolate from the collection eliminated the Candida albicans fungus. The 11 Antarctic bacteria with antibacterial capacity are strict aerobic Gram-negative bacilli, psychro phile to the genus Pseudomonas.

RESUMEN: La disminución sostenida de la disponibilidad de nuevas moléculas antimicrobianas para el uso terapéutico en las últimas décadas y el gran aumento de microorganismos resistentes a estos compuestos hace necesario buscar nuevas sustancias con propiedades antimicrobianas. Los ambientes extremos, donde hay poca disponibilidad de nutrientes, son escenarios propicios para buscar este tipo de compuestos, ya que las bacterias compiten entre sí y con otros organismos por posicionarse en el nicho ecológico y secretan, asociado a su metabolismo secundario, una serie de moléculas que inhiben o matan a otros microorga nismos, evitando su multiplicación. Nuestro grupo posee una colección de bacterias aisladas de la rizósfera de suelo del con tinente antártico, que logran crecer en condiciones de deprivación de nutrientes y a 4°C. Debido al hábitat extremo donde se desarrollan, las bacterias de los suelos de la Antártica están en una constante competencia por los recursos nutricionales, desarrollando distintas estrategias para colonizar su nicho ecológico y competir con otros microorganismos de la microbiota antártica. El objetivo de este trabajo consistió en identificar bacterias aisladas de la rizósfera de Deschampsia antarctica Desv. que inhiben el crecimiento de bacterias y hongos patógenos para humanos. A partir de una colección de 55 aislados de la rizósfera de la planta D. antarctica Desv. se identificaron 11 aislados con capacidad de matar o inhibir bacterias patógenas humanas, pero ningún aislado inhibió el crecimiento del hongo Candida albicans. Los resultados mos traron que las 11 bacterias antárticas con actividad antibacteriana corresponden a bacilos aerobios estrictos, Gram negativo, psicrófilos que pertenecen al género Pseudomonas.

SUMMARY: Bacteriocins are antimicrobial peptides of ribosomal synthesis secreted by bacteria. Among these, nisin stands out, which has potential uses in antibiotic therapies, as a food bio preservative and probiotics. Nisin has also been reported to have cytotoxicity on neoplastic cell lines, but there is little information on its effect on blood tumor cells. Due to the potential use that nisin presents, it is relevant to determine the toxicity it presents on tumor cell lines of the blood type. For this, hemolytic activity tests were carried out on human erythrocytes and toxicity on human peripheral blood mononuclear cells, determining that nisin does not have a toxic effect on this type of normal human blood cells. Cytotoxicity tests were also carried out with tumor cell lines (K562 and U937), to determine dose, exposure time and selectivity in the toxic effect of nisin on human tumor cells. These tests show that nisin shows cytotoxic activity on K562 and U937 cell lines at 72 h of exposure, at a concentration of 40 µg / mL, which corresponds to 100 times the minimum inhibitory concentration (MIC) used for its action on bacteria. When comparing the effect of nisin on human peripheral blood mononuclear cells with lymphoid and myeloid tumor lines (K562 and U937 respectively), a selective effect of nisin on blood tumor cells is observed.

RESUMEN: Las bacteriocinas son péptidos antimicrobianos de síntesis ribosomal secretadas por bacterias. Dentro de estas destaca nisina que posee potenciales usos en terapias antibióticas, como biopreservante de alimentos y probióticos. También se ha descrito que nisina posee citotoxicidad sobre líneas celulares neoplásicas, pero existe poca información de su efecto sobre células tumorales sanguíneas. Debido al potencial uso que presenta nisina, es relevante determinar la toxicidad que presenta sobre líneas celulares tumorales del tipo sanguíneo. Para esto, se realizaron ensayos de actividad hemolítica sobre eritrocitos humanos y de toxicidad sobre células mononucleares de sangre periférica humanas, determinándose que nisina no posee efecto citotóxico sobre este tipo de células normales humanas sanguíneas. Se realizaron también, ensayos de citotoxicidad con líneas celulares tumorales (K562 y U937), con el fin de determinar dosis, tiempo de exposición y selectividad en el efecto tóxico de nisina sobre las células tumorales humanas. Estos ensayos muestran que nisina presenta actividad citotóxica sobre líneas celulares K562 y U937 a las 72 h de exposición, a una concentración de 40 µg/mL, que corresponde a 100 veces la concentración mínima inhibitoria (MIC) usada para su acción sobre bacterias. Al comparar el efecto de nisina sobre células mononucleares de sangre periférica humanas con las líneas tumorales linfoides y mieloides (K562 y U937 respectivamente), se observa un efecto selectivo de nisina sobre las células tumorales sanguíneas.

Abstract: In this study, 50 bacterial isolates from pristine Antarctic soil samples from King George and Greenwich Islands (South Shetland Islands) and 25 bacterial isolates from the rhizosphere of Deschampsia antarctica Desv, were characterized for their antimicrobial activity and response to 21 antibiotics. Antibiotic susceptibility was assayed following the disc diffusion method using diffe rent groups of antibiotics: penicillins, cephalosporins, carbapenems, aminoglycosides, quinolones, tetracycline, phenicols, macrolides, sulfonamides, and trimethoprim. Anti microbial compounds production by the Antarctic bacterial isolates were determined using the agar diffusion method, using a set of human pathogenic bacteria. The bacterial isolates studied showed resistance to 14 (67%) out of the 21 antibiotics tested. Three bacterial isolates (4%) were resistant to at least one antibiotic and 47 (63%) were multidrug-resistant. Additionally, 26 (35%) bacterial isolates do not show antimicrobial activity and were susceptible to all antibiotics studied. Finally, 27 (36%) bacterial isolates combining antimicrobial activity and multiple resistance to antibiotics. These bacterial isolates on average presented resistance to 10 antibiotics. The bacterial isolates combining antimicrobial activity and multiple resistance to anti biotics are especially interesting. These two capabilities probably provide a competitive advantage to Antarctic bacteria to enable them to survive in a harsh environment. These microbial isolates are potential new sources of active compounds for the control of pathogenic microorganisms.

Resumen: En este estudio 50 aislados bacterianos obtenidos de muestras de suelo antártico prístino, de las islas Rey George y Greenwich (Islas Shetland del Sur), y 25 aislados bacterianos de la rizósfera de Deschampsia antarctica Desv, se estudiaron para determinar si presentan actividad antimicrobiana y para determinar su respuesta a 21 antibióticos. La susceptibilidad a los antibióticos se evaluó siguiendo el método de difusión en disco utilizando diferentes grupos de antibióticos: penicilinas, cefalosporinas, carbapenémicos, aminoglicósidos, qui- nolonas, tetraciclina, fenicoles, macrólidos, sulfonamidas y trimetoprim. La producción de compuestos antimicrobianos por los aislados bacterianos antarticos se determinó mediante el método de difusión en agar, utilizando como indicadores un panel de bacterias patógenas humanas Gram positivo y Gram negativo. Los aislados bacterianos estudiados mostraron resistencia a 14 (67%) de los 21 antibióticos probados. Tres aislados bacterianos (4%) fueron resistentes a al menos un antibiótico y 47 (63%) fueron multirresistentes. Además, 26 aislados bacterianos (35%) no muestran acti vidad antimicrobiana y fueron susceptibles a todos los antibióticos estudiados. Finalmente, 27 aislados bacterianos (36%) combinaron actividad antimicrobiana y resistencia múltiple a antibióticos y en promedio presentaron resistencia a 10 antibióticos. Los aislados bacterianos que combinan actividad antimicrobiana y resistencia múltiple a los antibióticos son especialmente interesantes ya que es probable que estas dos capacidades proporcionen una ventaja competitiva a las bacterias antárticas para permitirles sobrevivir en un entorno hostil. Adicionalmente, estos aislados bacterianos son nuevas fuentes po tenciales de compuestos activos para el control de microorganismos patógenos.

ABSTRACT Thraustochytrids are unicellular protists belonging to the Labyrinthulomycetes class, which are characterized by the presence of a high lipid content that could replace conventional fatty acids. They show a wide geographic distribution, however their diversity in the Antarctic Region is rather scarce. The analysis based on the complete sequence of 18S rRNA gene showed that strain 34-2 belongs to the species Thraustochytrium kinnei, with 99% identity. The total lipid profile shows a wide range of saturated fatty acids with abundance of palmitic acid (16:0), showing a range of 16.1-19.7%. On the other hand, long-chain polyunsaturated fatty acids, mainly docosahexaenoic acid and eicosapentaenoic acid are present in a range of 24-48% and 6.1-9.3%, respectively. All factors analyzed in cells (biomass, carbon consumption and lipid content) changed with variations of culture temperature (10 °C and 25 °C). The growth in glucose at a temperature of 10 °C presented the most favorable conditions to produce omega-3fatty acid. This research provides the identification and characterization of a Thraustochytrids strain, with a total lipid content that presents potential applications in the production of nutritional supplements and as well biofuels.

ABSTRACT The mononuclear cobalt (III) complex derived from 1,10-phenanthroline with lactose [Co(phen)2lactose]Cl2·3H2O (1) has been prepared and its properties have been compared with the sucrose complex [Co(Phen)2sucrose]Cl2·3H2O(2) and the complex without carbohydrate [Co(Phen)2Cl2]Cl·3H2O(3). The chemical structure of (1) was assigned by 1H-NMR, IR, CD and UV-Vis spectral data. The antibacterial activity of (1) - (3) was evaluated by disc-diffusion assays, using Gram-negative and positive bacteria. The minimum inhibitory concentration of the three complexes on the studied bacteria and their cytotoxicity on HEK293 human cells was determined. A colorimetric plate assay was used to distinguish bacteriostatic from bactericidal effect. Finally, the complexes uptake mechanism was evaluated using bacteria with mutated genes that encode for carbohydrate and siderophore receptors. The results indicate that complex (1) has an antibacterial activity similar to (3), while (2) presents a more restricted one. Moreover, all three complexes act by a bacteriostatic effect against bacterial cells and both (1) and (3) use a siderophore uptake mechanism to enter on bacterial cytoplasm. Cytotoxicity assays show that carbohydrate complexes are not cytotoxic to human cells, in contrast with complex (3), which is highly toxic. These results suggest that the use of the lactose ligand would maintain the antibacterial activity and uptake mechanism of the complex at reasonable levels, and would also reduce its toxicity against human cells. Thus, its strategic use would allow a decrease in toxicity of complexes used in eventual studies on eukaryotic systems.

RESUMEN. Introducción: el objetivo del presente estudio consistió en implementar la técnica de qPCR en tiempo real para identificar y cuantificar la especie Streptococcus mutans en muestras de saliva y biopelícula dentaria. Métodos: se seleccionaron al azar 27 niños de 8 años de edad, de nivel socio-económico bajo del área norte de la región metropolitana de Santiago de Chile, que se citaron en ayunas y sin cepillado durante al menos 12 horas, para colectar saliva no estimulada y un pool de biopelícula dentaria supragingival de todas las caras mesio-vestibulares de dientes anteriores y posteriores. Se cuantificó la cantidad de S. mutans en las muestras mediante qPCR empleando partidores que amplifican un fragmento del gen gtfB de S. mutans. Resultados: la amplificación presentó 98% de eficiencia con delta de fluorescencia de 3,36 ciclos. La curva de fusión (melting) presentó un solo máximo a una misma temperatura para todas las muestras. Conclusión: la metodología permite la identificación y cuantificación específica del gen gtfB de S. mutans en muestras de saliva y biopelícula dentaria, de forma rápida y exacta, aportando a la determinación del riesgo cariogénico individual.

ABSTRACT Introduction: the objective of this study was to use real-time qPCR to identify and quantify the Streptococcus mutans species in samples of saliva and dental biofilm. Methods: 27 children were randomly chosen with the following criteria: 8 years of age, low socio-economic levels, residing in the northern metropolitan area of Santiago de Chile; they were asked to attend an appointment while fasting with no teeth brushing for at least 12 hours, in order to collect non-stimulated saliva and a pool of supragingival dental biofilm of all the mesio-vestibular sides of anterior and posterior teeth. The amount of S. mutans in the samples was quantified by qPCR using primers that amplify a fragment of the gtfB gene of S. mutans. Results: the amplification showed 98% efficiency with a fluorescence of 3.36 cycles. The melting curve presented a single maximum at the same temperature for all samples. Conclusion: the methodology allows the specific identification and quantification of gene gtfB of S. mutans in saliva and dental biofilm in a quick and reliable manner, contributing to the identification of individual cariogenic risk.

The mononuclear cobalt(III) complex derived from 1,10-phenanthroline and maltose ([Co(phen)2maltose]Cl2∙3H2O) (1) has been synthesized and characterized in aqueous solution. Its characterization was based on its optical and spectroscopic properties. The antimicrobial activity of this complex was screened in vitro against the microorganisms Escherichia coli DH5a, Salmonella enterica sv Enteritidis ISP/953, Klebsiella pneumoniae RYC492, Pseudomonas aeruginosa PAO1, Enterococcus faecalis ATCC 29212, Bacillus cereus GCA234, Micrococcus sp. Staphylococcus aureus ATCC25923. Complex (1) showed antibacterial activity with a bacteriostatic effect over Gram positive and negative bacteria. The cytotoxicity of complex (1) was tested in vitro on human embryonic kidney cells.

Obesity is a risk factor for cancer. Epidemiological evidences associate ovarian cancer with obesity. Epithelial ovarian cancer (EOC) is the most common type of ovarian cancer and accounts for a high rate of mortality. The association between ovarian cancer and obesity could be explained by molecular factors secreted by adipose tissue such as leptin. In EOC, leptin increases cell proliferation and inhibits apoptosis. Additionally, adipose tissue synthesizes endogenous estrogens, which increase cell proliferation of epithelial ovarian cells. Also, obesity associated hyperinsulinism could increase ovarian estrogen secretion.

The complexes Diaqua-bis(5-nitroimidazole)-copper(II)-dinitrate (1); Tetrakis(4- phenylimidazole)-copper(II)-dinitrate, ethanol solvate (2); Bis(4-phenylimidazole)-bis(acetate)-copper(II) (3); Hexakis(4-phenylimidazole)-cobalt(II)-acetate (4) and Bis(2- phenylimidazole)-bis(acetate)-cobalt(II) (5), have been synthesized. Their structures were determined by X-ray analysis. The antimicrobial activities of these complexes have been screened in vitro against the microorganisms Escherichia coli, Klebsiella pneumoniae, Shigella flexneri, Salmonella typhi, Salmonella enteritidis, Salmonella entérica, Staphylococcus aureus, Listeria monocytogenes. Complexes (1) and (3) showed antimicrobial activity over Gram positive and negative bacteria by a bacteriostatic effect. On the other hand, complexes (4) and (5) showed antibacterial activity with a bacteriolytic effect over Gram positive and negative bacteria. The cytotoxicity of complexes (3) and (4) was tested in vitro. These complexes presented cytotoxic effect to values near the MIC.

The preparation, magnetic characterization and antimicrobial properties of the complexes [Fe(3)0(CH3COO)6(CH3COOH)(H(2)0)]Cl(MeImid)-(H(2)0) (1) and [Fe(4)0(2)(CH3COO)(7)0(2)(BPA)2(H(2)0)]- CH.25(CH3CH2OH)(H(2)0) (2), where Melmid is 2-methyl-imidazolium and BPA is 2,2'-bipyridine, are described. The variable temperature susceptibilities of these complexes were investigated in the temperature range 2-300K. The data have been modeled on the assumption of "butterfly" arranged for (2) and equilateral triangle of ferric ions for (1). The antimicrobial activities of these complexes have been screened in vitro against different bacterial strains. The complexes (1) and (2) display activity over Gram negative bacteria and these compounds not present activity over Gram positive strains. These complexes have bacteriostatic effect over bacterial target. The toxicity analyses of iron complexes showed these have cytotoxicity effect to values around the MIC on human cell.