Antibacterial activity and mode of action of selected glucosinolate hydrolysis products against bacterial pathogens

Plants contain numerous components that are important sources of new bioactive molecules with antimicrobial properties. Isothiocyanates (ITCs) are plant secondary metabolites found in cruciferous vegetables that are arising as promising antimicrobial agents in food industry. The aim of this study was to assess the antibacterial activity of two isothiocyanates (ITCs), allylisothiocyanate (AITC) and 2-phenylethylisothiocyanate (PEITC) against Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus and Listeria monocytogenes. The antibacterial mode of action was also characterized by the assessment of different physiological indices: membrane integrity, intracellular potassium release, physicochemical surface properties and surface charge. The minimum inhibitory concentration (MIC) of AITC and PEITC was 100 g/mL for all bacteria. The minimum bactericidal concentration (MBC) of the ITCs was at least 10 times higher than the MIC. Both AITC and PEITC changed the membrane properties of the bacteria decreasing their surface charge and compromising the integrity of the cytoplasmatic membrane with consequent potassium leakage and propidium iodide uptake. The surface hydrophobicity was also non-specifically altered (E. coli and L. monocytogenes become less hydrophilic; P. aeruginosa and S. aureus become more hydrophilic). This study shows that AITC and PEITC have strong antimicrobial potential against the bacteria tested, through the disruption of the bacterial cell membranes. Moreover, phytochemicals are highlighted as a valuable sustainable source of new bioactive products.This work was supported by the Operational Programme for Competitiveness Factors - COMPETE and by the Portuguese Foundation for Science and Technology through Project Phytodisinfectants - PTDC/DTP-SAP/1078/2012 (COMPETE: FCOMP-01-0124-FEDER-028765), the PhD grant awarded to Ana Abreu (SFRH/BD/84393/2012), and the post-doctoral grants awarded to Anabela Borges (SFRH/BPD/98684/2013) and Lucia C. Simoes (SFRH/BPD/81982/2011). Also, this work was undertaken as part of the European Research Project SUSCLEAN (Contract no FP7-KBBE-2011-5, project number: 287514) and the COST Action FA1202. The authors are solely responsible for this work. It does not represent the opinion of the European Community, and the Community is not responsible for any use that might be made of data appearing herein

Analysis of the Promoters Involved in Enterocin AS-48 Expression

The enterocin AS-48 is the best characterized antibacterial circular protein in prokaryotes. It is a hydrophobic and cationic bacteriocin, which is ribosomally synthesized by enterococcal cells and post-translationally cyclized by a head-to-tail peptide bond. The production of and immunity towards AS-48 depend upon the coordinated expression of ten genes organized in two operons, as-48ABC (where genes encoding enzymes with processing, secretion, and immunity functions are adjacent to the structural as-48A gene) and as-48C1DD1EFGH. The current study describes the identification of the promoters involved in AS-48 expression. Seven putative promoters have been here amplified, and separately inserted into the promoter-probe vector pTLR1, to create transcriptional fusions with the mCherry gene used as a reporter. The activity of these promoter regions was assessed measuring the expression of the fluorescent mCherry protein using the constitutive pneumococcal promoter PX as a reference. Our results revealed that only three promoters PA, P2(2) and PD1 were recognized in Enterococcus faecalis, Lactococcus lactis and Escherichia coli, in the conditions tested. The maximal fluorescence was obtained with PX in all the strains, followed by the P2(2) promoter, which level of fluorescence was 2-fold compared to PA and 4-fold compared to PD1. Analysis of putative factors influencing the promoter activity in single and double transformants in E. faecalis JH2-2 demonstrated that, in general, a better expression was achieved in presence of pAM401-81. In addition, the P2(2) promoter could be regulated in a negative fashion by genes existing in the native pMB-2 plasmid other than those of the as-48 cluster, while the pH seems to affect differently the as-48 promoter expression.This work was supported in part by the Ministerio de Ciencia e Innovación project BIO2008-01708, the Plan Propio from the University of Granada (Spain) and by the Research Plan Group (BIO 160)

Control of Propionibacterium acnes by natural antimicrobial substances: Role of the bacteriocin AS-48 and lysozyme

We report the high susceptibility of several clinical isolates of Propionibacterium acnes from different sources (skin, bone, wound exudates, abscess or blood contamination) to the head-to-tail cyclized bacteriocin AS-48. This peptide is a feasible candidate for further pharmacological development against this bacterium, due to its physicochemical and biological characteristics, even when it is growing in a biofilm. Thus, the treatment of pre-formed biofilms with AS-48 resulted in a dose- and time-dependent disruption of the biofilm architecture beside the decrease of bacterial viability. Furthermore, we demonstrated the potential of lysozyme to bolster the inhibitory activity of AS-48 against P. acnes, rendering high reductions in the MIC values, even in matrix-growing cultures, according to the results obtained using a range of microscopy and bioassay techniques. The improvement of the activity of AS-48 through its co-formulation with lysozyme may be considered an alternative in the control of P. acnes, especially after proving the absence of cytotoxicity demonstrated by these natural compounds on relevant human skin cell lines. In summary, this study supports that compositions comprising the bacteriocin AS-48 plus lysozyme must be considered as promising candidates for topical applications with medical and pharmaceutical purposes against dermatological diseases such as acne vulgaris.This research was funded by a grant from the Spanish Ministry of Economy and Competitiveness (SAF2013-48971-C2-1-R that included funds from European Regional Development, ERDF), and the Research Group General (BIO160, UGR)

Liquidity Regulation and Bank Risk

In this paper, we investigate the impact of liquidity requirements on bank risk. We take advantage of the implementation of the Liquidity Balance Rule (LBR) in the Netherlands in 2003 and analyze its impact on bank default risk. The LBR was imposed on Dutch banks only and did not apply to other banks operating elsewhere within the Eurozone. Using this differential regulatory treatment to overcome identification concerns, we find that following the introduction of the LBR, the risk of Dutch banks declined relatively to counterparts not affected by the rule. Concomitantly, despite the lower cost of funding driven by the LBR, the profitability of Dutch banks decreased in comparison with other banks in Europe, as a result of a decrease in income accruing from interest-bearing activities. Our findings also indicate that relatively to unaffected banks, Dutch banks might not have actively tried to offset their loss in interest income by increasing interest rates of loans. However, better financing conditions allowed Dutch banks to increase the shares of deposits and capital on the liability side of their balance sheets

Liquidity regulation and bank lending

Bank liquidity shortages during the global financial crisis of 2007-2009 led to the introduction of liquidity regulations, the impact of which has attracted the attention of academics and policymakers. In this paper, we investigate the impact of liquidity regulation on bank lending. As a setting, we use the Netherlands, where a Liquidity Balance Rule (LBR) was introduced in 2003. The LBR was imposed on Dutch banks only and did not apply to other banks operating elsewhere within the Eurozone. Using this differential regulatory treatment to overcome identification concerns and a difference-in-differences approach, we find that the LBR increased the volume of lending by Dutch banks relative to other banks located in the Eurozone. Increased equity, an inflow of retail deposits and subsequent increase in balance sheet size allowed Dutch banks to increase lending despite having to meet the LBR requirements. The LBR also affected loan composition (with corporate and retail lending increasing more than mortgage lending) and the maturity profile of loan portfolios. Our results have relevance for policymakers tasked with monitoring the impact of liquidity regulations on banks and the real economy.PostprintPeer reviewe

Bank Liquidity Regulation and Risk

We investigate the impact of liquidity requirements on bank risk. Using the Netherlands as a setting, we investigate the impact of the 2003 liquidity balance rule (LBR) on individual bank default risk and systemic risk. Using the differential regulatory treatment of Dutch and other Eurozone banks (that were not subject to the LBR) to overcome identification concerns, we find that following the introduction of the LBR, the individual risk and systemic risk of Dutch banks declines relative to unaffected counterparts. The observed decline in risk is at the expense of lower profitability (driven by a decline in interest income), which is not offset by the lower funding costs of treated banks following the enactment of the LBR. Our findings also suggest that better financing conditions allow Dutch banks to increase the balance sheet shares of deposits and capital

Liquidity regulation and bank lending

Bank liquidity shortages during the global financial crisis of 2007-2009 led to the introduction of liquidity regulations, the impact of which has attracted the attention of academics and policymakers. In this paper, we investigate the impact of liquidity regulation on bank lending. As a setting, we use the Netherlands, where a Liquidity Balance Rule (LBR) was introduced in 2003. The LBR was imposed on Dutch banks only and did not apply to other banks operating elsewhere within the Eurozone. Using this differential regulatory treatment to overcome identification concerns and a difference-in-differences approach, we find that the LBR increased the volume of lending by Dutch banks relative to other banks located in the Eurozone. Increased equity, an inflow of retail deposits and subsequent increase in balance sheet size allowed Dutch banks to increase lending despite having to meet the LBR requirements. The LBR also affected loan composition (with corporate and retail lending increasing more than mortgage lending) and the maturity profile of loan portfolios. Our results have relevance for policymakers tasked with monitoring the impact of liquidity regulations on banks and the real economy