Pichia membranifaciens CYC 1086 secretes a killer toxin (PMKT2) that is inhibitory to a variety of spoilage yeasts and fungi of agronomical interest. The killer toxin in the culture supernatant was concentrated by ultrafiltration and purified to homogeneity by two successive steps, including native electrophoresis and HPLC gel filtration. Biochemical characterization of the toxin showed it to be a protein with an apparent molecular mass of 30 kDa and an isoelectric point of 3.7. At pH 4.5, optimal killer activity was observed at temperatures up to 20 6C. Above approximately this pH, activity decreased sharply and was barely noticeable at pH 6. The toxin concentrations present in the supernatant during optimal production conditions exerted a fungicidal effect on a variety of fungal and yeast strains. The results obtained suggest that PMKT2 has different physico-chemical properties from PMKT as well as different potential uses in the biocontrol of spoilage yeasts. PMKT2 was able to inhibit Brettanomyces bruxellensis while Saccharomyces cerevisiae was fully resistant, indicating that PMKT2 could be used in wine fermentations to avoid the development of the spoilage yeast without deleterious effects on the fermentative strain. In small-scale fermentations, PMKT2, as well as P. membranifaciens CYC 1086, was able to inhibit B. bruxellensis, verifying the biocontrol activity of PMKT2 in simulated winemaking conditions. INTRODUCTIONWorldwide, microbial growth destroys large amounts of various products, causing yield losses in the agronomical and biotechnological industries. Traditionally, biocides have been used to deal with these problems but different disadvantages such as establishment of resistant strains and suppression of natural competitors have made alternatives such as biological control necessary (Beever et al., 1989;Raposo et al., 2000). Biological control strategies include natural plant-and animal-derived compounds, as well as antagonistic micro-organisms (Ciani & Fatichenti, 2001).During recent decades, microbiological control of spoilage micro-organisms has evolved as a possibility. Many yeast strains and other micro-organisms inhibiting plant pathogens have been reported, especially within the fruit-and vegetable-producing sector, and several new products have reached the commercial market (Janisiewicz & Korsten, 2002). The suggested modes of action of biocontrol yeasts are not likely to constitute any hazard for the consumer (Janisiewicz et al., 2001;Masih & Paul, 2002;Comitini et al., 2004;Santos & Marquina, 2004a).The food and beverage industries were among the first to explore the application of killer-toxin-producing yeasts to kill spoilage micro-organisms (Lowes et al., 2000). Yeast strains often achieve competitive advantage by producing killer toxins, which kill off competing sensitive cells belonging to either the same or a different species (Young, 1987;Ciani & Fatichenti, 2001). The most thoroughly studied examples are the Saccharomyces cerevisiae toxins K1, K2 and K28; producers of these toxi...
Fungal cells trigger adaptive mechanisms to survive in situations that compromise cell wall integrity. We show here that the global transcriptional response elicited by inhibition of the synthesis of β-1,3-glucan by caspofungin, encompasses a set of genes that are dependent on Slt2, the MAPK of the Cell Wall Integrity (CWI) pathway, and a broad group of genes regulated independently of Slt2. Genes negatively regulated by the cyclic AMP/Protein Kinase A (PKA) signaling pathway were overrepresented in the latter group. Moreover, cell wall stress mediated by inhibition of β-1,3-glucan synthesis, but not by other cell wall interfering compounds, negatively regulated PKA signaling as indicated by the nuclear localisation of Msn2, cellular glycogen accumulation, a decrease of intracellular cAMP levels and a severe decrease in both the activation of the small GTPase Ras2 and the phosphorylation of known substrates of PKA. All these effects relied on the plasma membrane-spanning sensor of the CWI pathway Wsc1. In addition, caspofungin induced a reduction in the cytosolic pH, which was dependent on the extracellular region of Wsc1. Therefore, alterations of the β-1,3-glucan network in the fungal cell wall, induce, through Wsc1, the activation of the CWI pathway and parallel inhibition of PKA signaling.Fungi cause a large number of infections worldwide and the incidence of fungi-related infections, primarily in hosts with impaired immunity, has risen sharply over the last few decades. The most significant fungal infections, accounting for approximately 90% of human mortality cases, are caused by species of Aspergillus, Candida, Cryptococcus, and Pneumocystis 1 . The most common classes of antifungals that are currently in use are polyenes, azoles, and echinocandins 2 . Echinocandins are synthetically modified lipopeptides that specifically target the fungal cell wall, a structure that is essential in fungi but absent in animal cells, and they represent the most recent group of antifungal agents introduced into the clinical practice. Echinocandins include micafungin, anidulafungin, and caspofungin (CAS), of which, CAS was the first agent of this group to be approved for therapeutic use 3 . Echinocandin drugs are non-competitive inhibitors of the β-1,3-glucan synthase (GS), an enzyme that is required for the formation of the essential polymer β-1,3-glucan found in the cell wall of several fungal pathogens 4 . Echinocandin resistance has been increasing in recent years 5 , which is of great concern because echinocandin drugs are the main course of therapy for patients with invasive candidiasis or aspergillosis. The resistance to echinocandins fungal pathogens exhibit has been attributed to two mechanisms: mutations that alter the target of the enzyme, the Fks1 catalytic subunit of the GS; and the activation of the signalling pathways required to maintain the cell wall integrity, which leads to an increase in the synthesis of the cell wall components, particularly chitin, which compensates for the inhibition of β-1,3-glucan s...
In search for natural products with antimicrobial properties for use in the prevention and treatment of peri-implantitis, the purpose of this investigation was to evaluate the antimicrobial activity of curcumin and xanthohumol, using an in vitro multi-species dynamic biofilm model including Streptococcus oralis, Actinomyces naeslundii, Veillonella parvula, Fusobacterium nucleatum, Porphyromonas gingivalis and Aggregatibacter actinomycetemcomitans. The antimicrobial activities of curcumin (5 mM) and xanthohumol (100 μM) extracts, and the respective controls, were evaluated with 72-h biofilms formed over dental implants by their submersion for 60 seconds. The evaluation was assessed by quantitative polymerase chain reaction (qPCR), confocal laser scanning microscopy (CLSM) and scanning electron microscopy (SEM). For the data analysis, comparisons were tested applying ANOVA tests with post-hoc Bonferroni corrections to evaluate the antimicrobial activity of both extracts. With qPCR, statistically significant reductions in bacterial counts were observed for curcumin and xanthohumol, when compared to the negative control. The results with CLSM and SEM were consistent with those reported with qPCR. It was concluded that both curcumin and xanthohumol have demonstrated antimicrobial activity against the six bacterial species included in the dynamic in vitro biofilm model used.
<span>Se evaluó la mutagenicidad del agua del</span><br /><span>río Cauca debida a la presencia de metales</span><br /><span>pesados en la zona urbana de la Ciudad</span><br /><span>de Santiago de Cali, a partir de muestras</span><br /><span>tomadas en la temporada seca y lluviosa</span><br /><span>en el año 2013. Los metales se extrajeron</span><br /><span>pasando el agua por la resina Amberlite</span><br /><span>XAD-16. Las concentraciones de los metales</span><br /><span>pesados se midieron por absorción atómica</span><br /><span>y la mutagenicidad se evaluó por medio</span><br /><span>del test de Ames, con las cepas TA98 y</span><br /><span>TA100 de Salmonella typhimurium con y</span><br /><span>sin activador enzimático S9. Los resultados</span><br /><span>mostraron índices de mutagenicidad (IM)</span><br /><span>positivos (IM > 2,0) para muestras colectadas</span><br /><span>en temporada lluviosa en tres de los cinco</span><br /><span>puntos evaluados: puente El Hormiguero</span><br /><span>(IM = 3,6), desembocadura del Canal</span><br /><span>Colector Sur (IM = 2,9) y desembocadura</span><br /><span>del río Cali (IM = 2,7), todos con la cepa</span><br /><span>TA98 sin S9. Estos sitios presentaron a su</span><br /><span>vez las mayores concentraciones totales de</span><br /><span>metales pesados en sus extractos. El análisis</span><br /><span>de la variación espacio-temporal del índice</span><br /><span>mutagénico se realizó haciendo un análisis de</span><br /><span>varianza multifactorial del IM. Los resultados</span><br /><span>encontrados indican que la época de</span><br /><span>muestreo contribuye significativamente a la</span><br /><span>variabilidad del IM , mientras que los puntos</span><br /><span>de muestreo no.</span>
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